WO2008044027A2 - Pharmaceutical compounds having hsp90 inhibitory or modulating activity - Google Patents

Pharmaceutical compounds having hsp90 inhibitory or modulating activity Download PDF

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WO2008044027A2
WO2008044027A2 PCT/GB2007/003860 GB2007003860W WO2008044027A2 WO 2008044027 A2 WO2008044027 A2 WO 2008044027A2 GB 2007003860 W GB2007003860 W GB 2007003860W WO 2008044027 A2 WO2008044027 A2 WO 2008044027A2
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WO2008044027A3 (en
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Miles Stuart Congreve
Lyndsey Helen Fazal
Martyn Frederickson
Christopher William Murray
Michael Alistair O'brien
Andrew James Woodhead
John Francis Lyons
Neil Thomas Thompson
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Astex Therapeutics Limited
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Priority to US12/445,154 priority Critical patent/US8779132B2/en
Priority to EP07824114A priority patent/EP2081891A2/en
Priority to JP2009531912A priority patent/JP5410285B2/en
Publication of WO2008044027A2 publication Critical patent/WO2008044027A2/en
Publication of WO2008044027A3 publication Critical patent/WO2008044027A3/en

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Definitions

  • This invention relates to pro-drugs of compounds that inhibit or modulate the activity of the heat shock protein Hsp90, to the use of the pro-drugs in the treatment or prophylaxis of disease states or conditions mediated by Hsp90, and to novel pro-drugs that break down in vivo to give compounds having Hsp90 inhibitory or modulating activity. Also provided are pharmaceutical compositions containing the pro-drugs.
  • Hsps heat shock proteins
  • Most heat shock proteins act as molecular chaperones. Chaperones bind and stabilize proteins at intermediate stages of folding and allow proteins to fold to their functional states.
  • Hsp90 is the most abundant cytosolic Hsp under normal conditions. There are two human isoforms of Hsp90, a major inducible form Hsp90 ⁇ and minor constitutively expressed form Hsp90 ⁇ and two other closely related chaperones which are restricted in their intracellular location (Endoplasmic reticulum GP96/GRP94; mitochondrial TRAP1).
  • HSP90 as used here includes all these analogues unless stated. Hsp90 binds proteins at a late stage of folding and is distinguished from other Hsps in that most of its protein substrates are involved in signal transduction. Hsp90 has a distinct ATP binding site, including a Bergerat fold characteristic of bacterial gyrase, topoisomerases and histidine kinases. It has been shown that ATP bound at the N-terminal pocket of Hsp90 is hydrolysed. This ATPase activity results in a conformational change in Hsp90 that is required to enable conformational changes in the client protein.
  • This complex then associates with Hsp90 via Hop.
  • ADP is replaced by ATP
  • the conformation of Hsp90 is altered, Hop and Hsp70 are released and a different set of co-chaperones is recruited including p50cdc37 and p23.
  • ATP hydrolysis results in the release of these co-chaperones and the client protein from the mature complex.
  • Ansamycin antibiotics herbimycin, geldanamycin (GA) and 17-allylamino-17-desmethoxygeldanamycin (17- AAG) are ATP binding site inhibitors that block the binding of ATP and prevent conversion to the mature complex (Grenert et. a/., 1997. J Biol Chem., 272:23834-23850).
  • Hsp90 being ubiquitously expressed, GA has a higher binding affinity for Hsp90 derived from tumour vs. normal cell lines (Kamal et. a/., Nature 2003; 425: 407-410). GA also shows more potent cytotoxic activity in tumour cells and is sequestered at higher concentrations within tumours in xenograft mouse models (Brazidec J. Med. Chem. 2004, 47, 3865-3873). Furthermore the ATP- ase activity of Hsp90 is elevated in cancer cells and is an indication of the increased level of stress in these cells. Hsp90 gene amplification has also been reported to occur in the later stages of cancer (Jolly and Morimoto JNCI Vol. 92, No. 19, 1564-1572, 2000).
  • Hsp90 is found primarily within "activated” multichaperone complexes in the tumour cells as opposed to “latent” complexes in normal cells.
  • One component of the multichaperone complex is the cdc37 co-chaperone.
  • Cdc37 binds Hsp90 at the base of the ATP binding site and could affect the off rates of inhibitors bound to Hsp90 in the "activated” state (Roe et. a/., Cell 116, (2004), pp. 87-98).
  • the client protein bound to the Hsp90-Hsp70 form of the chaperone complex is believed to be more susceptible to ubiquitination and targeting to the proteasome for degradation.
  • E3 ubiquitin ligases have been identified with chaperone interacting motifs and one of these (CHIP) was shown to promote the ubiquitination and degradation of Hsp90 client proteins (Connell et al., 2001. Xu et al., 2002).
  • Hsp90 client proteins The number of reported Hsp90 client proteins now exceeds 100. Since many of its client proteins are involved in cell signalling proliferation and survival, Hsp90 has received major interest as an oncology target. Two groups of client proteins, cell signalling protein kinases and transcription factors, in particular suggest Hsp90 regulation may have potential benefit as an anticancer therapy.
  • Hsp90 protein kinase client proteins implicated in cell proliferation and survival include the following: c-Src Cellular Src (c-Src) is a receptor tyrosine kinase, required for mitogenesis initiated by multiple growth factor receptors, including the receptors for epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR), colony stimulating factor- 1 (CSF-1 R), and the basic fibroblast growth factor (bFGFR). C-Src is also overexpressed and activated in many of the same human carcinomas that overexpress EGFR and ErbB2. Src is also required for the maintenance of normal bone homeostasis through its regulation of osteoclast function.
  • EGFR epidermal growth factor receptor
  • PDGFR platelet-derived growth factor receptor
  • CSF-1 R colony stimulating factor- 1
  • bFGFR basic fibroblast growth factor
  • C-Src is also overexpressed and activated in many of the same human carcinomas that overexpress
  • ErbB2 (Her2/neu) is a receptor tyrosine kinase overexpressed in a variety of malignancies including breast, ovarian, prostate, and gastric cancers. ErbB2 was originally identified as an oncogene and inhibition of Hsp90 results in the polyubiquitination and degradation of erbB2.
  • Polo mitotic kinase Polo-like kinases are important regulators of cell cycle progression during M-phase. Plks are involved in the assembly of the mitotic spindle apparatus and in the activation of CDK/cyclin complexes. PIkI regulates tyrosine dephosphorylation of CDKs through phosphorylation and activation of Cdc25C. CDK1 activation in turn leads to spindle formation and entry into M phase.
  • Akt Akt
  • PLB Akt
  • Hsp90 inhibition by ansamycins results in a reduction in the Akt half life through ubiquitination and proteasomal degradation.
  • Binding of cdc37 to Hsp90 is also required for the down-regulation of Akt.
  • cancer cells arrest in the G2/M phase of the cell cycle 24 hours after treatment and proceed to apoptosis 24-48 hours later. Normal cells also arrest 24 hours after ansamycin treatment, but do not proceed on to apoptosis.
  • the RAS-RAF-MEK-ERK-MAP kinase pathway mediates cellular responses to growth signals.
  • RAS is mutated to an oncogenic form in approximately 15% of human cancers.
  • the three RAF genes are serine/threonine kinases that are regulated by binding RAS.
  • EGFR EGFR
  • EGFR epidermal growth factor receptor
  • FMS-like tyrosine kinase 3 is a receptor tyrosine kinase involved in cell proliferation, differentiation and apoptosis. Flt3 activation also leads to the activation of phosphatidylinositol 3- kinase (PI3K) and RAS signal-transduction cascades.
  • c-Met c-met is a receptor tyrosine kinase which binds hepatocyte growth factor (HGF) and regulates both cell motility and cell growth, c-met is overexpressed in tumours, including thyroid, stomach, pancreatic and colon cancer. HGF is also detected around the tumours, including liver metastases. This suggests that c-met and HGF play an important role in invasion and metastasis.
  • HGF hepatocyte growth factor
  • Cdk1. Cdk2. Cdk4. Cdk6 Cdk1, Cdk2, Cdk4, and Cdk6 drive the cell cycle.
  • the activity of CDKs is regulated by their binding to specific subunits such as cyclins, inhibitory and assembly factors.
  • the substrate specificity and timing of CDK activities is dictated by their interaction with specific cyclins.
  • Cdk4/cyclin D and Cdk6/cyclin D are active in the G1 phase, Cdk2/cyclin E and Cdk2/cyclin A in S phase, and Cdc2/cyclin A and Cdc2/cyclin B in G2/M phase.
  • Cyclin-dependent kinase type 4 plays a key role in allowing cells to traverse G1 to S- phase transition of the cell cycle and is constitutively activated in many human cancers.
  • the CDK4 activator, cyclin D1 is overexpressed and a CDK4 inhibitor, p16, is deleted in a variety of human tumours.
  • Cdk1/Cdk2 inhibitors have been developed which reversibly block normal cells in either the G1/S- phase or at the G2/M border. G2/M arrest is generally less well tolerated by the cells and consequently, they undergo apoptotic cell death. Since Hsp90 also is known to affect cell survival pathways this effect may be further amplified with an Hsp90 inhibitor.
  • Wee-1 The Wee-1 protein kinase carries out the inhibitory phosphorylation of CDC2 on tyrosine 15
  • Hsp90 transcription factors implicated in cell proliferation and survival include the following: Mutant p53
  • P53 is a tumour suppressor protein that causes cell cycle arrest and induces apoptosis. P53 is mutated in approximately half of all cancers. Mutant p53 associates with Hsp90 and is down- regulated in cancer lines treated with Hsp90 inhibitors, while wild type p53 levels were unaffected.
  • Hsp90 hormone-response elements
  • HREs hormone-response elements
  • Prostate cancers are hormone- dependent malignancies that respond to therapeutic interventions which reduce circulating levels of testosterone or prevent testosterone binding to the androgen receptor. Although patients initally respond to these treatments most subsequently develop resistance via restoration of signalling via the androgen receptor.
  • Hsp90 and other co-chaperones including p23 and immunophilins.
  • Hif-1a Mutated steroid hormone receptors that have arisen for example during anti-hormone therapy and which might be resistant to such therapies are likely to have a greater dependence on HSP90 for their stability and hormone binding function.
  • Hypoxia inducible factor-la is a transcription factor that controls the expression of genes which play a role in angiogenesis.
  • HIF-Ia is expressed in the majority of metastases and is known to associate with Hsp90.
  • Ansamycin treatment of renal carcinoma cell lines leads to the ubiquitination and proteasomal degradation of HIF-Ia.
  • Hsp90 inhibitors are capable of affecting a large number of targets significant to signal transduction in tumour cell proliferation. Signal transduction inhibitors which regulate the activities of a single target, may not be as efficacious due to signalling pathway redundancy and the rapid development of resistance.
  • HSP90 inhibitors By regulating multiple targets involved in cell signalling and cell proliferation HSP90 inhibitors may prove beneficial in the treatment of a wide spectrum of proliferative disorders.
  • ZAP-70 a member of the Syk-ZAP-70 protein tyrosine kinase family, is normally expressed in T cells and natural killer cells and has a critical role in the initiation of T-cell signaling. However, it is also expressed aberrantly in approximately 50% of cases of CLL, usually in those cases with unmutated B-cell receptor genes.
  • the mutational status of immunoglobulin heavy-chain variable- region (lgV H ) genes in the leukemic cells of chronic lymphocytic leukemia (CLL) is an important prognostic factor.
  • the expression of ZAP-70 in CLL cells correlates with lgV H mutational status, disease progression, and survival.
  • ZAP-70 positive CLL is more aggressive than ZAP-70 negative CLL indicating that ZAP-70 may be a key driver of malignancy in this disease.
  • ZAP-70 is physically associated with HSP90 in B-CLL lymphoblasts thus the inhibition of Hsp90 may sensitise these cells to existing chemotherapy or monoclonal antibody therapy.
  • HSP90 Inhibitors as anti-fungal, anti-protozoal and anti-parasitic agents are HSP90 Inhibitors as anti-fungal, anti-protozoal and anti-parasitic agents
  • HSP90 is conserved across evolution being found in bacteria (e.g. HTPG in E.coli) and yeast (e.g. HSC82 and HSP82). Although clients have not been formally identified for the E.coli form, in yeast and all higher organisms the HSP90 family has been shown to function as a chaperone for many essential proteins as described above.
  • HSP90 Infection by a range of pathogens is associated with an antibody response to HSP90.
  • the 47kDa C-terminal fragment of HSP90 is an immunodominant epitope.
  • this antibody response is associated with good prognosis suggesting a protective effect against infection.
  • Recombinant antibodies to an epitope in this polypeptide are also protective against infection in mouse models of invasive candidiasis. (See Mathews ef a/ Antimicrobial Agents and Chemotherapy 2003 vol 47, 2208 - 2216 and references therein).
  • HSP90 Likewise surface expressed HSP90 serves as an antigen in Chagas' disease, ascariasis, leishmaniasis, toxoplamosis and infection due to Schistosoma mansoni and it has been postulated that antibodies to HSP90 convey protection against plamodiutn infection and Malaria.
  • Mycograb (NeuTec Pharma/Novartis) is a human recombinant monoclonal antibody against heat shock protein 90 that is being developed as a treatment for Candida and has shown significant responses in early trials. Furthermore, the natural product HSP90 inhibitors Geldanamycin, Herbimycin and Radicicol were originally identified by their anti-fungal activity. Key essential proteins have been identified as HSP90 clients in several human pathogens (see Cowen and
  • HSP90 can play an important role in the growth of pathogens such as Candida species, and HSP90 inhibitors can be useful as treatments for a range of infectious diseases including candidiasis.
  • Hsp90 increases the capacity of fungi to develop antifungal drug resistance (see Cowen LE, Lindquist S. "Hsp90 potentiates the rapid evolution of new traits: drug resistance in diverse fungi”. Science. 2005 Sep 30;309 (5744):2185-9). Therefore, coadministration of an Hsp90 inhibitor with an antifungal drug may enhance the efficacy of the antifungal drug and reduce resistance by preventing the emergence of resistant phenotypes.
  • HSP90 Inhibitors in the treatment of pain, neuropathic conditions and stroke Cdk5 is a member of the Cdk family of serine/threonine kinases, most of which are key regulators of the cell cycle. Cdk5 activity is regulated through association with its neuron-specific activators, p35 and p39. Recent evidence suggests that CDK5 can phosphorylate tau protein and a number of other neuronal proteins such as NUDE-1 , synapsini , DARPP32 and the Munc18/Syntaxin1A complex.
  • the p35 protein which acts as a regulator of CDK5 activity has recently been identified as a client protein for HSP90 and therefore the activity of CDK5 can be regulated by changes in the level and activity of HSP90.
  • inhibition of HSP90 can lead to loss of p35, an inhibition of CDK5, a reduction of phosphorylated tau protein in susceptible individuals and will bring benefit to sufferers of Alzheimers Disease.
  • Cdk5 has also been shown to have a role in mediating pain signalling. Both Cdk5 and p35 have been shown to be expressed in nociceptive neurons. In p35 knockout mice, which show substantially reduced Cdk ⁇ activity, the response to painful thermal stimuli is delayed (Pareek, T. K., et al., Proceedings of the National Academy of Sciences., 103:791-796 (2006). Additionally administration of the cyclin-dependent kinase 5 (Cdk ⁇ ) inhibitor roscovitine has been shown to attenuate the formalin-induced nociceptive responses in rats (Wang, Cheng-haung, et al., Acta Pharmacologica Sinica., 26:46- ⁇ O (200 ⁇ ).
  • Cdk ⁇ cyclin-dependent kinase 5
  • Activation of calpain is calcium dependent and is known to affected by activation of the NMDA receptor calcium channel (Amadoro, G; Proceedings of the National Academy of Sciences of the United States of America, 103, 2892-2897 (2006)).
  • NMDA receptor antagonists are know to be clinically effective against neuropathic pain conditions (Christoph, T; et al., Neuropharmacology, 51 ,12-17 (2006)). This efficacy may be linked to the effect of NMDA receptor related calcium influx on calpain activity and its subsequent effect on the activity of Cdk ⁇ . As such compounds modulating Cdk ⁇ activity are expected to be useful for the treatment or prevention of pain and thus modulation of the CDK ⁇ regulator p3 ⁇ by HSP90 inhibition could lead to inhibition of CDK ⁇ .
  • an agent for the palliative treatment of pain i.e. the direct relief of pain in addition to the relief of pain as the result of amelioration of the underlying disease or medical condition, which is the cause of the pain.
  • Cdk's especially Cdk's 4, 5 & 6 have been shown to be involved with or mediate neuronal death following hypoxic or ischemic insult (Rashidan, J.; et al.; Proceedings of the National Academy of Sciences., 102:14080-14086 (2005).
  • Cdk inhibitor flavopiridol has been shown to significantly reduce neuronal death in a rat model of focal cerebral ischemia (Osuga, H.; et al.; Proceedings of the National Academy of Sciences., 97:10254-10259 (2000).
  • Cdk5 inhibitors have been shown to have protective effects in both necrotic and apoptotic paradigms of neuronal cell death (Weisberger, J.; et al.; Molecular and Cellular Neuroscience., 24:489-502 (2003).
  • Stroke is a cerebrovascular event, which occurs when the normal bloodflow to the brain is disrupted, and the brain receives too much or too little blood. Stroke is one of the leading causes of death worldwide, and is also one of the most common causes of neurologic disability.
  • Ischemic stroke which is the most common type of stroke, results from insufficient cerebral circulation of blood caused by obstruction of the inflow of arterial blood. Normally, adequate cerebral blood supply is ensured by a system of arteries within the brain.
  • various disorders including inflammation and atherosclerosis, can cause a thrombus, i.e., a blood clot that forms in a blood vessel.
  • the thrombus may interrupt arterial blood flow, causing brain ischemia and consequent neurologic symptoms.
  • Ischemic stroke may also be caused by the lodging of an embolus (an air bubble) from the heart in an intracranial vessel, causing decreased perfusion pressure or increased blood viscosity with inadequate cerebral blood flow.
  • An embolus may be caused by various disorders, including atrial fibrillation and atherosclerosis.
  • a second type of stroke involves a hemorrhage or rupture of an artery leading to the brain.
  • Hemorrhagic stroke results in bleeding into brain tissue, including the epidural, subdural, or subarachnoid space of the brain.
  • a hemorrhagic stroke typically results from the rupture of an arteriosclerotic vessel that has been exposed to arterial hypertension or to thrombosis.
  • One opportunity for intervention in stroke is the prevention or reduction of risk of stroke in patients at risk for stroke.
  • risk factors for stroke including vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation.
  • At risk patients have been treated with agents to control blood pressure or manage blood lipid level, and have been treated with antiplatelet agents (such as clopidrogel) and anticoagulants.
  • a second opportunity is the treatment of acute stroke.
  • current pharmacologic therapies for treating acute stroke are limited to restoring blood flow within a narrow therapeutic time window of less than three hours after stroke. There remains a need for agents which are effective within a longer therapeutic time window.
  • Another opportunity is recovery or restoration after the acute stroke period, i.e. the reduction or prevention of secondary cell damage in the penumbra. There remains a need for agents which are effective in reducing or preventing secondary cell damage after stroke.
  • Such an agent may be administered to patients at risk for stroke, and also may be administered to patients suffering from acute stroke, or patients undergoing treatment for recovery or restoration after the acute stroke period. Such an agent may also target more than one distinct mechanism in the biochemical cascade of stroke. HSP90 inhibitors and the treatment of Hepatitis C and other viral diseases
  • HSP induction may be to assist in the stabilization and folding of the high levels of 'foreign' protein generated in preparation for virus replication.
  • HSP90 is required for stable production of functional NS2/3 protease in Hepatitis C (HCV) replicon infected cells.
  • HSP 90 inhibitors have also been demonstrated to block viral replication in in vitro systems. (Nagkagawa, S, Umehara T, Matsuda C, et al Biochem. Biophys. Res Commun. 353 (2007) 882-888; Waxman L, Witney, M et al PNAS 98 (2001) 13931 - 13935).
  • Hsps heat-shock proteins
  • modulators or inhibitors of stress proteins in general represent a class of chemotherapeutics with the unique ability to inhibit multiple aberrant signalling pathways simultaneously. They can therefore exert antitumour effects whilst eliminating (or reducing the incidence of) resistance relative to other treatment paradigms.
  • modulators or inhibitors of stress protein function in general represent a class of chemotherapeutics with the potential for: (i) sensitizing malignant cells to anticancer drugs and/or treatments; (ii) alleviating or reducing the incidence of resistance to anticancer drugs and/or treatments; (iii) reversing resistance to anticancer drugs and/or treatments; (iv) potentiating the activity of anticancer drugs and/or treatments; (v) delaying or preventing the onset of resistance to anticancer drugs and/or treatments.
  • Pro-Drugs are generally recognised as being chemical compounds that have little or no pharmacological activity themselves but which undergo biotransformation to a therapeutically active metabolite (see for example Bernard Testa, Biochemical Pharmacology, 68 (2004), 2097- 2106, and "Design of Prodrugs” (Bundgaard H. ed.) 1985 Elsevier Science Publishers B. V. (Biomedical Division).
  • Pro-drugs are used for a variety of reasons. For example, they may be used inter alia to:
  • ⁇ provide selective targetting of a drug ⁇ provide in situ activation of a cytotoxic agent
  • WO 99/29705 disclose a class of glycomimetic compounds having a number of possible uses including the treatment of cancer.
  • One compound specifically disclosed in WO 99/29705 is the compound 2-(2-hydroxy-benzoyl)-1 ,2,3,4-tetrahydro-isoquinoline-3-carboxylic acid.
  • the present invention provides pro-drug compounds containing functional groups that are modified or removed in vivo to give compounds having Hsp90 inhibiting or modulating activity. It is envisaged that the pro-drug compounds will be useful in preventing or treating disease states or conditions mediated by Hsp90.
  • the compounds of the invention will be useful in alleviating or reducing the incidence of cancer.
  • the invention provides a compound for use in medicine, the compound being a compound of the formula (Vl 0 ):
  • n 0, 1 , 2 or 3;
  • R 1 is hydrogen, hydroxy, or O-R z ;
  • R 2a is hydroxy, methoxy or O-R z ; provided that at least one of R 1 and R 2a is O-R z ;
  • R z is L p -R p1 ; SO 3 H; a glucuronide residue; a mono-, di- or tripeptide residue; or
  • R 3 is selected from hydrogen; halogen; cyano; C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy; wherein the C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy, amino, mono- and di-Ci -2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members;
  • R 4a is selected from hydrogen, fluorine, chlorine and methoxy
  • R 8 is selected from hydrogen and fluorine
  • R 10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or di-C 1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group R a -R b ; wherein:
  • R a is a bond, O, CO, X 1 C(X 2 ), C(X 2 )X 1 , X 1 C(X 2 )X 1 , S, SO, SO 2 , NR 0 , SO 2 NR 0 or NR 0 SO 2 ;
  • R b is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C 1-12 hydrocarbyl (such as C 1-10 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C 1-8 non-aromatic hydrocarbylamino (e.g. mono- or di-C 1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C 1-12 hydrocarbyl group (or C 1-10 hydrocarbyl group) may optionally be replaced by O, S, SO,
  • the invention provides a compound of the formula (Vl):
  • n 0, 1 , 2 or 3;
  • R 1 is hydrogen, hydroxy, or O-R z ;
  • R 2a is hydroxy, methoxy or O-R z ; provided that at least one of R 1 and R 2a is O-R z ;
  • R z is L p -R p1 ; SO 3 H; a glucuronide residue; a mono-, di- or tripeptide residue; or
  • R p2 and R p3 are the same or different and each is a group R p1 ;
  • R 3 is selected from hydrogen; halogen; cyano; C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy; wherein the C 1-5 hydrocarbyl and Ci -5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy, amino, mono- and di-C 1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R 4a is selected from hydrogen, fluorine, chlorine and methoxy; R 8 is selected from hydrogen and fluorine; and R 10 is selected from: halogen; hydroxy; trifluoromethyl; cya ⁇ o; nitro; carboxy; amino; mono- or di-C 1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group R a -R b ; wherein:
  • R a is a bond, O, CO, X 1 C(X 2 ), C(X 2 )X ⁇ X 1 C(X 2 )X 1 , S, SO, SO 2 , NR 0 , SO 2 NR 0 or NR 0 SO 2 ;
  • R b is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C 1 - I2 hydrocarbyl (such as C 1-I0 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C ⁇ s non-aromatic hydrocarbylamino (e.g. mono- or di-C 1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C 1-12 hydrocarbyl group (or C 1-I0 hydrocarbyl group) may optionally be replaced by O, S, SO,
  • R 0 is selected from R b , hydrogen and C 1-4 hydrocarbyl
  • the invention provides a compound for use in medicine, the compound being a compound of the formula (I 0 );
  • R 1 is hydrogen, hydroxy, or O-R z ;
  • R 2 is hydrogen, hydroxy, methoxy or O-R z ; provided that at least one of R 1 and R 2 is O-R z ; R 2 is L p -R p1 ; SO 3 H; a glucuronide residue; a mono-, di- or tripeptide residue; or
  • R 3 is selected from hydrogen; halogen; cyano; C 1-5 hydrocarbyl and Ci -5 hydrocarbyloxy; wherein the C 1-5 hydrocarbyl and Ci -5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, Ci -2 alkoxy, amino, mono- and di-C 1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members;
  • R 4 is selected from hydrogen; a group -(O) n -R 7 where n is O or 1 and R 7 is an acyclic C 1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-Ci -5 hydrocarbyl-amino, wherein the acyclic C 1-5 hydrocarbyl group and the mono and di-C 1-5 hydrocarbylamino moieties in each instance
  • R 8 is selected from hydrogen and fluorine
  • R 10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or di-C 1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group R a -R b ; wherein:
  • R a is a bond, O, CO, X 1 C(X 2 ), C(X 2 )X 1 , X 1 C(X 2 )X 1 , S, SO, SO 2 , NR C , SO 2 NR 0 or NR 0 SO 2 ; and R b is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C 1-12 hydrocarbyl (such as C 1-10 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C 1-8 non-aromatic hydrocarbylamino (e.g.
  • C 1-12 hydrocarbyl group or C 1-10 hydrocarbyl group
  • one or more carbon atoms of the C 1-12 hydrocarbyl group (or C 1-10 hydrocarbyl group) may optionally be replaced by O, S, SO, SO 2 , NR C , X 1 C(X 2 ), C(X 2 )X 1 or X 1 C(X 2 )X 1 ;
  • the invention provides a compound of the formula (I):
  • R 1 is hydrogen, hydroxy, or O-R z ;
  • R 2 is hydrogen, hydroxy, methoxy or 0-R 2 ; provided that at least one of R 1 and R 2 is O-R z ; R z is L p -R p1 ; SO 3 H; a glucuronide residue; a mono-, di- or tripeptide residue; or
  • R 8 is selected from hydrogen and fluorine; and R 10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or di-C 1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group R a -R b ; wherein:
  • R a is a bond, O, CO, X 1 C(X 2 ), C(X 2 )X 1 , X 1 C(X 2 )X 1 , S, SO, SO 2 , NR C , SO 2 NR 0 or NR 0 SO 2 ; and R b is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and Ci -12 hydrocarbyl (such as C 1-10 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C 1-8 non-aromatic hydrocarbylamino (e.g.
  • C 1- - I2 hydrocarbyl group may optionally be replaced by O, S, SO, SO 2 , NR C , X 1 C(X 2 ), C(X 2 )X 1 or X 1 C(X 2 )X 1 ;
  • the invention provides:
  • Hsp90 which method comprises administering to a subject in need thereof a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
  • a method for treating a disease or condition comprising or arising from abnormal cell growth in a mammal comprises administering to the mammal a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (VI), (Via), (VIl), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein in an amount effective in inhibiting abnormal cell growth.
  • Vila (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for the manufacture of a medicament for for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal.
  • a method for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal comprises administering to the mammal a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein in an amount effective in inhibiting abnormal cell growth.
  • a method for treating a disease or condition comprising or arising from abnormal cell growth in a mammal comprising administering to the mammal a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VIl), (Vila), (VIIb) or (VlII) or any sub-groups or examples thereof as defined herein in an amount effective to inhibit Hsp90 activity.
  • a method for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal comprising administering to the mammal a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein in an amount effective to inhibit Hsp90 activity.
  • a method of inhibiting Hsp90 which method comprises contacting the Hsp90 with an Hsp ⁇ O-inhibiting compound of the formula (I 0 ), (I), (II), (111), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
  • an Hsp ⁇ O-inhibiting compound of the formula (I 0 ), (I), (II), (111), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
  • a method of modulating a cellular process by inhibiting the activity of Hsp90 using a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
  • a pharmaceutical composition comprising a compound of the formula (I 0 ), (I), (II), (III),
  • Vl 0 (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII)) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier.
  • a pharmaceutical composition comprising a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier in a form suitable for oral administration.
  • a pharmaceutical composition comprising a compound of the formula (I 0 ), (I), (II), (III),
  • Vl 0 (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier in a form suitable for parenteral administration, for example by intravenous (i.v.) administration.
  • a pharmaceutically acceptable carrier in a form suitable for parenteral administration, for example by intravenous (i.v.) administration.
  • a pharmaceutical composition comprising a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier in a form suitable for intravenous (i.v.) administration by injection or infusion.
  • Hsp90 which method comprises (i) screening a patient to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against Hsp90; and (ii) where it is indicated that the disease or condition from which the patient is thus susceptible, thereafter administering to the patient a compound of the formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
  • the compounds of the invention are pro-drugs of the hydroxybenzoic acid amide Hsp90 inhibitors disclosed in our earlier international application PCT/GB2006/001382.
  • the compounds of PCT/GB2006/001382 may be referred to variously in this application as “the parent compounds” or “the phenolic compounds”.
  • References to the pro-drugs being "derived” from the “parent compounds” or “phenolic compounds” is only intended to imply a structural relationship and is not intended to imply any particular method of preparing the prodrugs.
  • the pro-drugs may be prepared from the compounds of PCT/GB2006/001382 as described below, or they may be prepared by methods that do not involve the intermediacy of a phenolic compound of PCT/GB2006/001382.
  • references to a compound of formula (I) includes formula (I 0 ), formula (Vl 0 ) and all subgroups of formula (I) as defined herein, including formulae (II), (III), (IV), (V), (Vl), (Via), (VII), (Vila) and (VIIb) and the term 'subgroups' includes all preferences, embodiments, examples and particular compounds defined herein.
  • a reference to formula (Vl) includes formula (Vl 0 ) and all sub-groups, preferences, embodiments, examples and particular compounds within formula (Vl).
  • a reference to a compound of formula (I 0 ), (I), (II), (III), (Vl 0 ), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) and sub-groups thereof includes ionic forms, salts, solvates, isomers, tautomers, N-oxides, esters, isotopes and protected forms thereof, as discussed below:- preferably, the salts or tautomers or isomers or N-oxides or solvates thereof:- and more preferably, the salts or tautomers or N-oxides or solvates thereof.
  • the term encompasses situations where pain is already being experienced by a subject or patient, as well as situations where pain is not currently being experienced but is expected to arise.
  • treatment also cover both complete and partial pain reduction or prevention.
  • the compounds of the invention may prevent existing pain from worsening, or they reduce or even eliminate pain.
  • the compounds may prevent any pain from developing or they may lessen the extent of pain that may develop.
  • the term “modulation”, as applied to the activity of the heat shock protein Hsp90 is intended to define a change in the level of biological activity of the heat shock protein.
  • modulation encompasses physiological changes which effect an increase or decrease in the relevant heat shock protein activity.
  • the modulation may be described as "inhibition".
  • the modulation may arise directly or indirectly, and may be mediated by any mechanism and at any physiological level, including for example at the level of gene expression (including for example transcription, translation and/or post-translational modification), at the level of expression of genes encoding regulatory elements which act directly or indirectly on the levels of heat shock protein activity.
  • modulation may imply elevated/suppressed expression or over- or under-expression of the heat shock protein, including gene amplification (i.e. multiple gene copies) and/or increased or decreased expression by a transcriptional effect, as well as hyper- (or hypo-)activity and (de)activation of the heat shock protein (including (de)activation) by mutation(s).
  • the terms “modulated”, “modulating” and “modulate” are to be interpreted accordingly.
  • the term "mediated”, as used e.g. in conjunction with the heat shock protein as described herein (and applied for example to various physiological processes, diseases, states, conditions, therapies, treatments or interventions) is intended to operate limitatively so that the various processes, diseases, states, conditions, treatments and interventions to which the term is applied are those in which heat shock protein Hsp90 plays a biological role.
  • the biological role played by heat shock protein Hsp90 may be direct or indirect and may be necessary and/or sufficient for the manifestation of the symptoms of the disease, state or condition (or its aetiology or progression).
  • heat shock protein Hsp90 activity (and in particular aberrant levels of heat shock protein Hsp90 activity, e.g. Hsp90 over-expression) need not necessarily be the proximal cause of the disease, state or condition: rather, it is contemplated that the heat shock protein Hsp90 mediated diseases, states or conditions include those having multifactorial aetiologies and complex progressions in which Hsp90 is only partially involved. In cases where the term is applied to treatment, prophylaxis or intervention (e.g.
  • Hsp90-mediated treatments and “Hsp90-mediated prophylaxis” of the invention
  • the role played by Hsp90 may be direct or indirect and may be necessary and/or sufficient for the operation of the treatment, prophylaxis or outcome of the intervention.
  • a disease state or condition mediated by Hsp90 includes the development of resistance to any particular cancer drug or treatment (including in particular resiatnce to one or more of the signalling inhibitors described herein).
  • modulation As used herein, the term “modulation”, as applied to the activity of cyclin dependent kinase 5 (CDK5), is intended to define a change in the level of biological activity of the kinase(s). Thus, modulation encompasses physiological changes which effect an increase or decrease in the relevant kinase activity. In the latter case, the modulation may be described as "inhibition”.
  • the modulation may arise directly or indirectly, and may be mediated by any mechanism and at any physiological level, including for example at the level of gene expression (including for example transcription, translation and/or post-translational modification), at the level of expression of genes encoding regulatory elements which act directly or indirectly on the levels of cyclin dependent kinase 5 (CDK5), or at the level of enzyme (e.g. cyclin dependent kinase 5 (CDK5) activity (for example by allosteric mechanisms, competitive inhibition, active-site inactivation, perturbation of feedback inhibitory pathways etc.).
  • CDK5 cyclin dependent kinase 5
  • modulation may imply elevated/suppressed expression or over- or under-expression of the cyclin dependent kinase 5 (CDK5) including gene amplification (i.e.
  • CDK5 cyclin dependent kinase 5
  • the term "mediated”, as used e.g. in conjunction with the cyclin dependent kinase 5 (CDK5) as described herein (and applied for example to various physiological processes, diseases, states, conditions, therapies, treatments or interventions) is intended to operate limitatively so that the various processes, diseases, states, conditions, treatments and interventions to which the term is applied are those in which cyclin dependent kinase 5 (CDK5) plays a biological role, in cases where the term is applied to a disease, state or condition, the biological role played by cyclin dependent kinase 5 (CDK5) may be direct or indirect and may be necessary and/or sufficient for the manifestation of the symptoms of the disease, state or condition (or its aetiology or progression).
  • cyclin dependent kinase 5 (CDK5) activity (and in particular aberrant levels of cyclin dependent kinase 5 (CDK5) activity, e.g. cyclin dependent kinase 5 (CDK5) over-expression) need not necessarily be the proximal cause of the disease, state or condition: rather, it is contemplated that the CDK5-mediated diseases, states or conditions include those having multifactorial aetiologies and complex progressions in which CDK5.
  • the role played by CDK5 may be direct or indirect and may be necessary and/or sufficient for the operation of the treatment, prophylaxis or outcome of the intervention.
  • intervention is a term of art used herein to define any agency which effects a physiological change at any level.
  • the intervention may comprise the induction or repression of any physiological process, event, biochemical pathway or cellular/biochemical event.
  • the interventions of the invention typically effect (or contribute to) the therapy, treatment or prophylaxis of a disease or condition.
  • the terms “combined” and “combining” in this context are to be interpreted accordingly.
  • association of the two or more compounds/agents in a combination may be physical or non- physical.
  • Examples of physically associated combined compounds/agents include:
  • compositions e.g. unitary formulations
  • two or more compounds/agents in admixture (for example within the same unit dose);
  • compositions comprising material in which the two or more compounds/agents are chemically/physicochemically linked (for example by crosslinking, molecular agglomeration or binding to a common vehicle moiety);
  • compositions comprising material in which the two or more compounds/agents are chemically/physicochemically co-packaged (for example, disposed on or within lipid vesicles, particles (e.g. micro- or nanoparticles) or emulsion droplets);
  • material e.g. a non-unitary formulation
  • material comprising at least one of the two or more compounds/agents together with instructions for the extemporaneous association of the at least one compound to form a physical association of the two or more compounds/agents
  • material e.g. a non-unitary formulation
  • material comprising at least one of the two or more compounds/agents together with instructions for combination therapy with the two or more compounds/agents
  • the term "in combination" may refer to compounds/agents that are administered as part of the same overall treatment regimen.
  • the posology of each of the two or more compounds/agents may differ: each may be administered at the same time or at different times. It will therefore be appreciated that the compounds/agents of the combination may be administered sequentially (e.g. before or after) or simultaneously, either in the same pharmaceutical formulation (i.e. together), or in different pharmaceutical formulations (i.e. separately). Simultaneously in the same formulation is as a unitary formulation whereas simultaneously in different pharmaceutical formulations is non-unitary.
  • the posologies of each of the two or more compounds/agents in a combination therapy may also differ with respect to the route of administration.
  • the term "pharmaceutical kit” defines an array of one or more unit doses of a pharmaceutical composition together with dosing means (e.g. measuring device) and/or delivery means (e.g. inhaler or syringe), optionally all contained within common outer packaging.
  • dosing means e.g. measuring device
  • delivery means e.g. inhaler or syringe
  • the individual compounds/agents may unitary or non-unitary formulations.
  • the unit dose(s) may be contained within a blister pack.
  • the pharmaceutical kit may optionally further comprise instructions for use.
  • the term "pharmaceutical pack” defines an array of one or more unit doses of a pharmaceutical composition, optionally contained within common outer packaging.
  • pharmaceutical packs comprising a combination of two or more compounds/agents
  • the individual compounds/agents may unitary or non-unitary formulations.
  • the unit dose(s) may be contained within a blister pack.
  • the pharmaceutical pack may optionally further comprise instructions for use.
  • patient pack defines a package, prescribed to a patient, which contains pharmaceutical compositions for the whole course of treatment.
  • Patient packs usually contain one or more blister pack(s).
  • Patient packs have an advantage over traditional prescriptions, where a pharmacist divides a patient's supply of a pharmaceutical from a bulk supply, in that the patient always has access to the package insert contained in the patient pack, normally missing in patient prescriptions. The inclusion of a package insert has been shown to improve patient compliance with the physician's instructions.
  • glucuronide residue as used herein is used to denote a group of the formula: where the asterisk denotes the point of attachment to the oxygen atom of the group O-R z .
  • glucuronide residue is linked by a glycosidic bond to the benzene ring to which the moieties R 1 , R 8 , R 2a , R 3 and R 4a are attached.
  • Compounds of the invention containing a glucuronide group may be referred to herein as “glucuronides” or “a glucuronide”, “glucuronide derivatives” or “ a “glucuronide” derivative”.
  • the glucuronides of the invention can exist in either the ⁇ - or ⁇ -anomeric forms and can be provided in a solid form in which one anomer predominates or as mixtures of the two anomers.
  • mono-, di or tri-peptide residue refers to an acyl residue of an an amino acid or a dipeptide or tripeptide.
  • the mono-peptide residue is the group:
  • the di-peptide residue is the group:
  • non-terminal carbon atoms refers to a carbon atom which (i) has at least two other carbon atoms attached to it or (ii) has at least one other carbon atom attached to it and is also attached directly to the group L p , the oxygen atom in the group O-R z (when L p is a bond) or an
  • references to "carbocyclic” and “heterocyclic” groups as used herein shall, unless the context indicates otherwise, include both aromatic and non-aromatic ring systems.
  • the term “carbocyclic and heterocyclic groups” includes within its scope aromatic, non-aromatic, unsaturated, partially saturated and fully saturated carbocyclic and heterocyclic ring systems.
  • such groups may be monocyclic or bicyclic and may contain, for example, 3 to 12 ring members, more usually 5 to 10 ring members.
  • Examples of monocyclic groups are groups containing 3, 4, 5, 6, 7, and 8 ring members, more usually 3 to 7, for example 5 to 7, and preferably 5 or 6 ring members.
  • Examples of bicyclic groups are those containing 8, 9, 10, 11 and 12 ring members, and more usually 9 or 10 ring members.
  • bicyclic refers to groups that have two rings joined together in such as way that at least one ring member is shared by both rings.
  • the bicyclic group can be a fused ring (two ring members shared by both rings), spirocyclic (one ring member shared by both rings) or a bridged ring (three or more ring members shared by both rings).
  • the carbocyclic or heterocyclic groups can be aryl or heteroaryl groups having from 5 to 12 ring members, more usually from 5 to 10 ring members.
  • aryl refers to a carbocyclic group having aromatic character and the term “heteroaryl” is used herein to denote a heterocyclic group having aromatic character.
  • the terms “aryl” and “heteroaryl” embrace polycyclic (e.g. bicyclic) ring systems wherein one or more rings are non-aromatic, provided that at least one ring is aromatic. In such polycyclic systems, the group may be attached by the aromatic ring, or by a non-aromatic ring.
  • the aryl or heteroaryl groups can be monocyclic or bicyclic groups and can be unsubstituted or substituted with one or more substituents, for example one or more groups R 10 as defined herein.
  • non-aromatic group embraces unsaturated ring systems without aromatic character, partially saturated and fully saturated carbocyclic and heterocyclic ring systems.
  • fully saturated and “saturated” refer to rings where there are no multiple bonds between ring atoms.
  • Saturated carbocyclic groups include cycloalkyl groups as defined below.
  • Partially saturated carbocyclic groups include cycloalkenyl groups as defined below, for example cyclopentenyl, cycloheptenyl and cyclooctenyl.
  • a further example of a cycloalkenyl group is cyclohexenyl.
  • heteroaryl groups are monocyclic and bicyclic groups containing from five to twelve ring members, and more usually from five to ten ring members.
  • the heteroaryl group can be, for example, a five membered or six membered monocyclic ring or a bicyclic structure formed from fused five and six membered rings or two fused six membered rings or, by way of a further example, two fused five membered rings.
  • Each ring may contain up to about four heteroatoms typically selected from nitrogen, sulphur and oxygen.
  • the heteroaryl ring will contain up to 4 heteroatoms, more typically up to 3 heteroatoms, more usually up to 2, for example a single heteroatom.
  • the heteroaryl ring contains at least one ring nitrogen atom.
  • the nitrogen atoms in the heteroaryl rings can be basic, as in the case of an imidazole or pyridine, or essentially non-basic as in the case of an indole or pyrrole nitrogen. In general the number of basic nitrogen atoms present in the heteroaryl group, including any amino group substituents of the ring, will be less than five.
  • Examples of five membered heteroaryl groups include but are not limited to pyrrole, furan, thiophene, imidazole, furazan, oxazole, oxadiazole, oxatriazole, isoxazole, thiazole, isothiazole, pyrazole, triazole and tetrazole groups.
  • Examples of six membered heteroaryl groups include but are not limited to pyridine, pyrazine, pyridazine, pyrimidine and triazine.
  • a bicyclic heteroaryl group may be, for example, a group selected from: a) a benzene ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; b) a pyridine ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; c) a pyrimidine ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; d) a pyrrole ring fused to a a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; e) a pyrazole ring fused to a a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; f) a pyrazine ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; g) an imidazole ring fuse
  • a thiophene ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms m) a furan ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; n) a cyclohexyl ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; and o) a cyclopentyl ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms.
  • One sub-group of bicyclic heteroaryl groups consists of groups (a) to (e) and (g) to (o) above.
  • bicyclic heteroaryl groups containing a five membered ring fused to another five membered ring include but are not limited to imidazothiazole (e.g. imidazo[2,1 -b]thiazole) and imidazoimidazole (e.g. imidazo[1 ,2-a]imidazole).
  • imidazothiazole e.g. imidazo[2,1 -b]thiazole
  • imidazoimidazole e.g. imidazo[1 ,2-a]imidazole
  • bicyclic heteroaryl groups containing a six membered ring fused to a five membered ring include but are not limited to benzfuran, benzthiophene, benzimidazole, benzoxazole, isobenzoxazole, benzisoxazole, benzthiazole, benzisothiazole, isobenzofuran, indole, isoindole, indolizine, indoline, isoindoline, purine (e.g., adenine, guanine), indazole, pyrazolopyrimidine (e.g. pyrazolo[1 ,5-a]pyrimidine), triazolopyrimidine (e.g. [1 ,2,4]triazolo[1 ,5- a]pyrimidine), benzodioxole and pyrazolopyridine (e.g. pyrazolo[1 ,5-a]pyridine) groups.
  • bicyclic heteroaryl groups containing two fused six membered rings include but are not limited to quinoline, isoquinoline, chroman, thiochroman, chromene, isochromene, chroman, isochroman, benzodioxan, quinolizine, benzoxazine, benzodiazine, pyridopyridine, quinoxaline, quinazoline, cinnoline, phthalazine, naphthyridine and pteridine groups.
  • One sub-group of heteroaryl groups comprises pyridyl, pyrrolyl, furanyl, thienyl, imidazolyl, oxazolyl, oxadiazolyl, oxatriazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, pyrazinyl, pyridazinyl, pyrimidinyl, triazinyl, triazolyl, tetrazolyl, quinolinyl, isoquinolinyl, benzfuranyl, benzthienyl, chromanyl, thiochromanyl, benzimidazolyl, benzoxazolyl, benzisoxazole, benzthiazolyl and benzisothiazole, isobenzofuranyl, indolyl, isoindolyl, indolizinyl, indolinyl, isoindolinyl, pur
  • polycyclic aryl and heteroaryl groups containing an aromatic ring and a non-aromatic ring examples include tetrahydronaphthalene, tetrahydroisoquinoline, tetrahydroquinoline, dihydrobenzthiene, dihydrobenzfuran, 2,3-dihydro-benzo[1 ,4]dioxine, benzo[1 ,3]dioxole, 4,5,6,7- tetrahydrobenzofuran, indoline and indane groups.
  • Examples of carbocyclic aryl groups include phenyl, naphthyl, indenyl, and tetrahydronaphthyl groups.
  • Examples of non-aromatic heterocyclic groups include unsubstituted or substituted (by one or more groups R 10 ) heterocyclic groups having from 3 to 12 ring members, typically 4 to 12 ring members, and more usually from 5 to 10 ring members.
  • Such groups can be monocyclic or bicyclic, for example, and typically have from 1 to 5 heteroatom ring members (more usually 1 ,2,3 or 4 heteroatom ring members) typically selected from nitrogen, oxygen and sulphur. When sulphur is present, it may, where the nature of the adjacent atoms and groups permits, exist as -S-, -S(O)- Or -S(O) 2 -.
  • the heterocylic groups can contain, for example, cyclic ether moieties (e.g. as in tetrahydrofuran and dioxane), cyclic thioether moieties (e.g. as in tetrahydrothiophene and dithiane), cyclic amine moieties (e.g. as in pyrrolidine), cyclic amide moieties (e.g. as in pyrrolidone), cyclic thioamides, cyclic thioesters, cyclic ester moieties (e.g. as in butyrolactone), cyclic sulphones (e.g.
  • heterocyclic groups are those containing a cyclic urea moiety (e.g. as in imidazolidin-2-one),
  • the heterocyclic groups contain cyclic ether moieties (e.g as in tetrahydrofuran and dioxane), cyclic thioether moieties (e.g. as in tetrahydrothiophene and dithiane), cyclic amine moieties (e.g. as in pyrrolidine), cyclic sulphones (e.g. as in sulpholane and sulpholene), cyclic sulphoxides, cyclic sulphonamides and combinations thereof (e.g. thiomorpholine).
  • cyclic ether moieties e.g as in tetrahydrofuran and dioxane
  • cyclic thioether moieties e.g. as in tetrahydrothiophene and dithiane
  • cyclic amine moieties e.g. as in pyrrolidine
  • cyclic sulphones e.g. as in sul
  • Examples of monocyclic non-aromatic heterocyclic groups include 5-, 6-and 7-membered monocyclic heterocyclic groups.
  • Particular examples include morpholine, piperidine (e.g. 1- piperidinyl, 2-piperidinyl, 3-piperidinyl and 4-piperidinyl), pyrrolidine (e.g.
  • thiomorpholine and its S-oxide and S,S-dioxide particularly thiomorpholine
  • Still further examples include azetidine, piperidone, piperazone, and N-alkyl piperidines such as N- methyl piperidine.
  • non-aromatic heterocyclic groups consists of saturated groups such as azetidine, pyrrolidine, piperidine, morpholine, thiomorpholine, thiomorpholine S,S-dioxide, piperazine, N-alkyl piperazines, and N-alkyl piperidines.
  • Another sub-set of non-aromatic heterocyclic groups consists of pyrrolidine, piperidine, morpholine, thiomorpholine, thiomorpholine S,S-dioxide, piperazine and N-alkyl piperazines such as N-methyl piperazine.
  • heterocyclic groups consists of pyrrolidine, piperidine, morpholine and N- alkyl piperazines (e.g. N-methyl piperazine), and optionally thiomorpholine.
  • non-aromatic carbocyclic groups include cycloalkane groups such as cyclohexyl and cyclopentyl, cycloalkenyl groups such as cyclopentenyl, cyclohexenyl, cycloheptenyl and cyclooctenyl, as well as cyclohexadienyl, cyclooctatetraene, tetrahydronaphthenyl and decalinyl.
  • Preferred non-aromatic carbocyclic groups are monocyclic rings and most preferably saturated monocyclic rings. Typical examples are three, four, five and six membered saturated carbocyclic rings, e.g. optionally substituted cyclopentyl and cyclohexyl rings.
  • Non-aromatic carboyclic groups includes unsubstituted or substituted (by one or more groups R 10 ) monocyclic groups and particularly saturated monocyclic groups, e.g. cycloalkyl groups.
  • cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl; more typically cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl, particularly cyclohexyl.
  • non-aromatic cyclic groups include bridged ring systems such as bicycloalkanes and azabicycloalkanes although such bridged ring systems are generally less preferred.
  • bridged ring systems is meant ring systems in which two rings share more than two atoms, see for example Advanced Organic Chemistry, by Jerry March, 4 th Edition, Wiley Interscience, pages 131-133, 1992.
  • bridged ring systems examples include bicyclo[2.2.1]heptane, aza-bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, aza-bicyclo[2.2.2]octane, bicyclo[3.2.1]octane and aza-bicyclo[3.2.1]octane.
  • a particular example of a bridged ring system is the 1-aza-bicyclo[2.2.2]octan-3-yl group.
  • the carbocyclic or heterocyclic ring can, unless the context indicates otherwise, be unsubstituted or substituted by one or more substituent groups R 10 selected from halogen, hydroxy, trifluoromethyl, cyano, nitro, carboxy, amino, mono- or di-C 1-4 hydrocarbylamino, carbocyclic and heterocyclic groups having from 3 to 12 ring members; a group R a -R b wherein R a is a bond, O, CO, X 1 C(X 2 ), C(X 2 )X 1 , X 1 C(X 2 )X 1 , S, SO, SO 2 , NR C , SO 2 NR 0 or NR 0 SO 2 ; and R b is selected from hydrogen, carbocyclic and heterocyclic groups having from 3 to 12 ring members, and a C 1-12 hydrocarbyl group (such as a C 1-10 hydrocarbyl group) optionally
  • R° is selected from R b , hydrogen and C 1-4 hydrocarbyl
  • substituent group R 10 comprises or includes a carbocyclic or heterocyclic group
  • the said carbocyclic or heterocyclic group may be unsubstituted or may itself be substituted with one or more further substituent groups R 10 .
  • such further substituent groups R 10 may include carbocyclic or heterocyclic groups, which are typically not themselves further substituted.
  • the said further substituents do not include carbocyclic or heterocyclic groups but are otherwise selected from the groups listed above in the definition of R 10 .
  • the substituents R 10 may be selected such that they contain no more than 20 non-hydrogen atoms, for example, no more than 15 non-hydrogen atoms, e.g. no more than 12, or 11, or 10, or 9, or 8, or 7, or 6, or 5 non-hydrogen atoms.
  • the two substituents may be linked so as to form a cyclic group.
  • two adjacent groups R 10 together with the carbon atoms or heteroatoms to which they are attached may form a 5-membered heteroaryl ring or a 5- or 6-membered non-aromatic carbocyclic or heterocyclic ring, wherein the said heteroaryl and heterocyclic groups contain up to 3 heteroatom ring members selected from N, O and S.
  • an adjacent pair of substituents on adjacent carbon atoms of a ring may be linked via one or more heteroatoms and optionally substituted alkylene groups to form a fused oxa-, dioxa-, aza-, diaza- or oxa-aza-cycloalkyl group.
  • halogen substituents include fluorine, chlorine, bromine and iodine. Fluorine and chlorine are particularly preferred.
  • hydrocarbyl is a generic term encompassing aliphatic, alicyclic and aromatic groups having an all-carbon backbone and consisting of carbon and hydrogen atoms, except where otherwise stated.
  • one or more of the carbon atoms making up the carbon backbone may be replaced by a specified atom or group of atoms.
  • hydrocarbyl groups include alky], cycloalkyl, cycloalkenyl, carbocyclic aryl, alkenyl, alkynyl, cycloalkylalkyl, cycloalkenylalkyl, and carbocyclic aralkyl, aralkenyl and aralkynyl groups. Such groups can be unsubstituted or, where stated, substituted by one or more substituents as defined herein.
  • the examples and preferences expressed below apply to each of the hydrocarbyl substituent groups or hydrocarbyl-containing substituent groups referred to in the various definitions of substituents for compounds of the formula (I) unless the context indicates otherwise.
  • Ci x-y (where x and y are integers) as used herein refers to the number of carbon atoms in a given group.
  • a Ci -4 hydrocarbyl group contains from 1 to 4 carbon atoms
  • a C 3 . s cycloalkyl group contains from 3 to 6 carbon atoms, and so on.
  • acyclic hydrocarbyl (e.g. as in “acyclic C 1-5 hydrocarbyl”) as used herein refers to non- cyclic hydrocarbyl groups and in particular to alkyl, alkenyl and alkynyl groups as defined herein.
  • mono- or CJi-C 1-5 hydrocarbylamino refers to a monosubstituted or disubsituted amine group bearing either one or two hydrocarbyl substituent groups that each contain from 1 to 5 carbon atoms.
  • Preferred non-aromatic hydrocarbyl groups are saturated groups such as alkyl and cycloalkyl groups.
  • the hydrocarbyl groups can have up to ten carbon atoms (and more typically up to eight carbon atoms), unless the context requires otherwise.
  • C 1-8 hydrocarbyl groups or C 1-6 hydrocarbyl groups such as Ci -4 hydrocarbyl groups (e.g.
  • C 1-3 hydrocarbyl groups or C 1-2 hydrocarbyl groups or C 2-3 hydrocarbyl groups or C 2-4 hydrocarbyl groups specific examples being any individual value or combination of values selected from C 1 , C 2 , C 3 , C 4 , C 5 , C 6 , C 7 , C 8 , C 9 and C 10 hydrocarbyl groups.
  • alkyl covers both straight chain and branched chain alkyl groups.
  • alkyl groups include methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, 2-pentyl, 3- pentyl, 2-methyl butyl, 3-methyl butyl, and n-hexyl and its isomers.
  • C 1-6 alkyl groups such as C 1-4 alkyl groups (e.g. C 1-3 alkyl groups or C 1-2 alkyl groups or C 2-3 alkyl groups or C 2-4 alkyl groups).
  • cycloalkyl groups are those derived from cyclopropane, cyclobutane, cyclopentane, cyclohexane and cycloheptane. Within the sub-set of cycloalkyl groups the cycloalkyl group will have from 3 to 10 carbon atoms, more typically 3 to 8 carbon atoms, particular examples being C 3 . 6 cycloalkyl groups.
  • alkenyl groups include, but are not limited to, ethenyl (vinyl), 1-propenyl, 2-propenyl (allyl), isopropenyl, butenyl, buta-1 ,4-dienyl, pentenyl, and hexenyl.
  • alkenyl groups will have 2 to 10 carbon atoms, more typically 2 to 8 carbon atoms, particular examples being C 2-6 alkenyl groups, such as C 2-4 alkenyl groups.
  • cycloalkenyl groups include, but are not limited to, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclopentadienyl and cyclohexenyl. Within the sub-set of cycloalkenyl groups the cycloalkenyl groups have from 3 to 10 carbon atoms, more typically 3 to 8 carbon atoms, and particular examples are C 3-6 cycloalkenyl groups.
  • alkynyl groups include, but are not limited to, ethynyl and 2-propynyl (propargyl) groups. Within the sub-set of alkynyl groups having 2 to 10 carbon atoms, more typically 2 to 8 carbon atoms, particular examples are C 2-6 alkynyl groups, such as C 2-4 alkynyl groups.
  • Examples of carbocyclic aryl groups include substituted and unsubstituted phenyl groups.
  • Examples of cycloalkylalkyl, cycloalkenylalkyl, carbocyclic aralkyl, aralkenyl and aralkynyl groups include phenethyl, benzyl, styryl, phenylethynyl, cyclohexylmethyl, cyclopentylmethyl, cyclobutylmethyl, cyclopropylmethyl and cyclopentenylmethyl groups.
  • C 1-12 hydrocarbyl, C 1- - I0 hydrocarbyl and C 1-8 hydrocarbyl as used herein encompasses alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, phenyl, benzyl and phenylethyl groups wherein the preferences for and examples of each of the aforesaid groups are as defined above.
  • particular hydrocarbyl groups are alkyl, cycloalkyl, phenyl, benzyl and phenylethyl (e.g.
  • 1-phenylethyl or 2-phenylethyl) groups one subset of hydrocarbyl groups consisting of alkyl and cycloalkyl groups and in particular C 1-4 alkyl and cycloalkyl groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, cyclopropyl and cyclobutyl.
  • C 1 - S hydrocarbyl as used herein encompasses alkyl, alkenyl, alkynyl, cycloalkyl and cycloalkenyl groups wherein the preferences for and examples of the aforesaid groups are as defined above.
  • particular Ci -5 hydrocarbyl groups are saturated C 1-5 hydrocarbyl groups, namely alkyl and cycloalkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, 3-methylbutyl, 2-methylbutyl, pent-2-yl, pent-3-yl, 3-methylbut- 2-yl, 2,2-dimethylpropyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclopropylmethyl, cyclopropylethyl, cyclobutylmethyl, methylcyclopropyl, ethylcyclopropyl, dimethylcyclopropyl and methylcyclobutyl.
  • alkyl and cycloalkyl groups such as methyl, ethyl, n-propyl, isopropyl, n-buty
  • C 1-5 hydrocarbyl as used herein are unsaturated C 1-5 hydrocarbyl groups, i.e. alkene, cycloalkene and alkyne groups such as vinyl, 1-propenyl, isopropenyl, 1- butenyl, 2-butenyl, 1-pentenyl, 2-pentenyl, allyl, methylallyl, dimethylallyl, acetylenyl, propargyl, cyclobutenyl and cyclopentenyl.
  • alkene alkene
  • cycloalkene and alkyne groups such as vinyl, 1-propenyl, isopropenyl, 1- butenyl, 2-butenyl, 1-pentenyl, 2-pentenyl, allyl, methylallyl, dimethylallyl, acetylenyl, propargyl, cyclobutenyl and cyclopentenyl.
  • C 1-4 hydrocarbyl as used herein encompasses alky], alkenyl, alkynyl, cycloalkyl and cycloalkenyl groups wherein the preferences for and examples of the aforesaid groups are as defined above.
  • particular C1-4 hydrocarbyl groups are alkyl and cycloalkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, cyclopropyl and cyclobutyl.
  • a hydrocarbyl group can be optionally substituted by one or more substituents selected from hydroxy, oxo, alkoxy, carboxy, halogen, cyano, nitro, amino, mono- or di-C 1-4 hydrocarbylamino, and monocyclic or bicyclic carbocyclic and heterocyclic groups having from 3 to 12 (typically 3 to 10 and more usually 5 to 10) ring members.
  • substituents include halogen such as fluorine.
  • the substituted hydrocarbyl group can be a partially fluorinated or perfluorinated group such as difluoromethyl or trifluoromethyl.
  • preferred substituents include monocyclic carbocyclic and heterocyclic groups having 3-7 ring members, more usually 3, 4, 5 or 6 ring members.
  • one or more carbon atoms of a hydrocarbyl group may optionally be replaced by O, S, SO, SO 2 , NR C , X 1 C(X 2 ), C(X 2 )X 1 or X 1 C(X 2 )X 1 (or a sub-group thereof) wherein X 1 and X 2 are as hereinbefore defined, provided that at least one carbon atom of the hydrocarbyl group remains.
  • 1 , 2, 3 or 4 carbon atoms of the hydrocarbyl group may be replaced by one of the atoms or groups listed, and the replacing atoms or groups may be the same or different.
  • the number of linear or backbone carbon atoms replaced will correspond to the number of linear or backbone atoms in the group replacing them.
  • groups in which one or more carbon atom of the hydrocarbyl group have been replaced by a replacement atom or group as defined above include ethers and thioethers (C replaced by O or S), amides, esters, thioamides and thioesters (C-C replaced by X 1 C(X 2 ) or C(X 2 )X 1 ), sulphones and sulphoxides (C replaced by SO or SO 2 ), amines (C replaced by NR 0 ).
  • ureas, carbonates and carbamates C-C-C replaced by X 1 C(X 2 )X 1 ).
  • an amino group may, together with the nitrogen atom to which they are attached, and optionally with another heteroatom such as nitrogen, sulphur, or oxygen, link to form a ring structure of 4 to 7 ring members, more usually 5 to 6 ring members.
  • aza-cycloalkyl refers to a cycloalkyl group in which one of the carbon ring members has been replaced by a nitrogen atom.
  • examples of aza-cycloalkyl groups include piperidine and pyrrolidine.
  • oxa-cycloalkyl refers to a cycloalkyl group in which one of the carbon ring members has been replaced by an oxygen atom.
  • examples of oxa-cycloalkyl groups include tetrahydrofuran and tetrahydropyran.
  • diaza-cycloalkyl refers respectively to cycloalkyl groups in which two carbon ring members have been replaced by two nitrogen atoms, or by two oxygen atoms, or by one nitrogen atom and one oxygen atom.
  • oxa-C 4 . 6 cycloalkyl group there will be from 3 to 5 carbon ring members and an oxygen ring member.
  • an oxa-cyclohexyl group is a tetrahydropyranyl group.
  • R a -R b as used herein, either with regard to substituents present on a carbocyclic or heterocyclic moiety, or with regard to other substituents present at other locations on the compounds of the formula (I), includes inter alia compounds wherein R a is selected from a bond, O, CO, OC(O), SC(O), NR 0 C(O), OC(S), SC(S), NR 0 C(S), OC(NR 0 ), SC(NR 0 ), NR 0 C(NR 0 ), C(O)O, C(O)S, C(O)NR 0 , C(S)O, C(S)S, C(S) NR 0 , C(NR°)0, C(NR°)S, C(NR°)NR°, OC(O)O, SC(O)O, NR 0 C(O)O, OC(S)O, SC(O)O, NR 0 C(O)O, OC(S)O
  • R b can be hydrogen or it can be a group selected from carbocyclic and heterocyclic groups having from 3 to 12 ring members (typically 3 to 10 and more usually from 5 to 10), and a C- I-8 hydrocarbyl group optionally substituted as hereinbefore defined. Examples of hydrocarbyl, carbocyclic and heterocyclic groups are as set out above.
  • hydrocarbyloxy groups include saturated hydrocarbyloxy such as alkoxy (e.g. C 1- 6 alkoxy, more usually C 1-4 alkoxy such as ethoxy and methoxy, particularly methoxy), cycloalkoxy (e.g. C 3-6 cycloalkoxy such as cyclopropyloxy, cyclobutyloxy, cyclopentyloxy and cyclohexyloxy) and cycloalkylalkoxy (e.g. C 3-6 cycloalkyl-C 1-2 alkoxy such as cyclopropylmethoxy).
  • alkoxy e.g. C 1- 6 alkoxy, more usually C 1-4 alkoxy such as ethoxy and methoxy, particularly methoxy
  • cycloalkoxy e.g. C 3-6 cycloalkoxy such as cyclopropyloxy, cyclobutyloxy, cyclopentyloxy and cyclohexyloxy
  • the hydrocarbyloxy groups can be substituted by various substituents as defined herein.
  • the alkoxy groups can be substituted by halogen (e.g. as in difluoromethoxy and trifluoromethoxy), hydroxy (e.g. as in hydroxyethoxy), C- I-2 alkoxy (e.g. as in methoxyethoxy), hydroxy-C 1-2 alkyl (as in hydroxyethoxyethoxy) or a cyclic group (e.g. a cycloalkyl group or non- aromatic heterocyclic group as hereinbefore defined).
  • halogen e.g. as in difluoromethoxy and trifluoromethoxy
  • hydroxy e.g. as in hydroxyethoxy
  • C- I-2 alkoxy e.g. as in methoxyethoxy
  • hydroxy-C 1-2 alkyl as in hydroxyethoxyethoxy
  • a cyclic group e.g. a cyclo
  • alkoxy groups bearing a non- aromatic heterocyclic group as a substituent are those in which the heterocyclic group is a saturated cyclic amine such as morpholine, piperidine, pyrrolidine, piperazine, C 1 . 4 -alkyl- piperazines, C 3-7 -cycloalkyl-piperazines, tetrahydropyran or tetrahydrofuran and the alkoxy group is a C- I-4 alkoxy group, more typically a C- I-3 alkoxy group such as methoxy, ethoxy or n-propoxy.
  • the heterocyclic group is a saturated cyclic amine such as morpholine, piperidine, pyrrolidine, piperazine, C 1 . 4 -alkyl- piperazines, C 3-7 -cycloalkyl-piperazines, tetrahydropyran or tetrahydrofuran
  • the alkoxy group is a C- I-4 alkoxy group, more
  • Alkoxy groups may be substituted by a monocyclic group such as pyrrolidine, piperidine, morpholine and piperazine and N-substituted derivatives thereof such as N-benzyl, N-C 1-4 acyl and N-C 1-4 alkoxycarbonyl.
  • a monocyclic group such as pyrrolidine, piperidine, morpholine and piperazine and N-substituted derivatives thereof such as N-benzyl, N-C 1-4 acyl and N-C 1-4 alkoxycarbonyl.
  • Particular examples include pyrrolidinoethoxy, piperidinoethoxy and piperazinoethoxy.
  • hydrocarbyl groups R a -R b are as hereinbefore defined.
  • the hydrocarbyl groups may be saturated groups such as cycloalkyl and alkyl and particular examples of such groups include methyl, ethyl and cyclopropyl.
  • the hydrocarbyl (e.g. alkyl) groups can be substituted by various groups and atoms as defined herein. Examples of substituted alkyl groups include alkyl groups substituted by one or more halogen atoms such as fluorine and chlorine (particular examples including bromoethyl, chloroethyl and trifluoromethyl), or hydroxy (e.g.
  • acyloxy e.g. acetoxymethyl and benzyloxymethyl
  • amino and mono- and dialkylamino e.g. aminoethyl, methylaminoethyl, dimethylaminomethyl, dimethylaminoethyl and ferf-butylaminomethyl
  • alkoxy e.g. C 1-2 alkoxy such as methoxy - as in methoxyethyl
  • cyclic groups such as cycloalkyl groups, aryl groups, heteroaryl groups and non-aromatic heterocyclic groups as hereinbefore defined).
  • alkyl groups substituted by a cyclic group are those wherein the cyclic group is a saturated cyclic amine such as morpholine, piperidine, pyrrolidine, piperazine, C ⁇ -alkyl- piperazines, C 3 . 7 -cycloalkyl-piperazines, tetrahydropyran or tetrahydrofuran and the alkyl group is a C 1-4 alkyl group, more typically a C 1-3 alkyl group such as methyl, ethyl or n-propyl.
  • a saturated cyclic amine such as morpholine, piperidine, pyrrolidine, piperazine, C ⁇ -alkyl- piperazines, C 3 . 7 -cycloalkyl-piperazines, tetrahydropyran or tetrahydrofuran
  • the alkyl group is a C 1-4 alkyl group, more typically a C 1-3 alkyl group such as methyl, eth
  • alkyl groups substituted by a cyclic group include pyrrolidinomethyl, pyrrolidinopropyl, morpholinomethyl, morpholinoethyl, morpholinopropyl, piperidinylmethyl, piperazinomethyl and N- substituted forms thereof as defined herein.
  • alkyl groups substituted by aryl groups and heteroaryl groups include benzyl and pyridylmethyl groups.
  • R a is SO 2 NR 0
  • R b can be, for example, hydrogen or an optionally substituted C 1-8 hydrocarbyl group, or a carbocyclic or heterocyclic group.
  • R a -R b where R a is SO 2 NR 0 include aminosulphonyl, C- ⁇ alkylaminosulphonyl and di-C 1-4 alkylaminosulphonyl groups, and sulphonamides formed from a cyclic amino group such as piperidine, morpholine, pyrrolidine, or an optionally N-substituted piperazine such as N-methyl piperazine.
  • groups R a -R b where R a is SO 2 include alkylsulphonyl, heteroarylsulphonyl and arylsulphonyl groups, particularly monocyclic aryl and heteroaryl sulphonyl groups. Particular examples include methylsulphonyl, phenylsulphonyl and toluenesulphonyl.
  • R b can be, for example, hydrogen or an optionally substituted C 1-I0 hydrocarbyl group, or a carbocyclic or heterocyclic group.
  • R a -R b where R a is NR 0 include amino, C 1-4 alkylamino (e.g. methylamino, ethylamino, propylamino, isopropylamino, fe/f-butylamino), di- C 1-4 alkylamino (e.g. dimethylamino and diethylamino) and cycloalkylamino (e.g. cyclopropylamino, cyclopentylamino and cyclohexylamino).
  • C 1-4 alkylamino e.g. methylamino, ethylamino, propylamino, isopropylamino, fe/f-butylamino
  • di- C 1-4 alkylamino e.g
  • R 1 is hydrogen, hydroxy, or O-R z ; and R 2a is hydroxy, methoxy or O-R z ; provided that at least one of R 1 and R 2a is O-R z .
  • R 1 is hydrogen, hydroxy, or O-R z ; and R 2 is hydrogen, hydroxy, methoxy or O-R z ; provided that at least one of R 1 and R 2 is O-R z .
  • R 1 is hydroxy or O-R z and R 2a is hydroxy or O-R 1p ; provided that at least one of R 1 and R 2a is O-R z .
  • both R 1 and R 2a are O-R z .
  • both R 1 and R 2 are O-R z .
  • R z is L p -R p1 ; SO 3 H; a glucuronide v residue; a mono-, di- or tripeptide residue; or
  • R p2 and R p3 are the same or different and each is a group R p1 .
  • R z is as defined above in respect of (Vl 0 ), (Vl) and (I 0 ), except that the formula excludes compounds wherein L p is a bond and R p1 is an unsubstituted benzyl group or a methyl group.
  • R p1 is an unsubstituted benzyl group or a methyl group.
  • R p1 is hydrogen or an optionally substituted C 1-25 hydrocarbyl group as defined herein except that the optional substituents for the C 1-25 hydrocarbyl group do not include hydroxy.
  • hydrocarbyl as used in the context of formulae (Vl 0 ) (Vl), (I 0 ) and (I) and sub-groups thereof is as defined in the General Preferences and Definitions section above.
  • the term covers inter alia alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, cycloalkylalkenyl, cycloalkylalkynyl, cycloalkenyl, cycloalkenylalkyl, cycloalkenylalkenyl, cycloalkenylalkynyl, phenyl, phenylalkyl, phenylalkenyl, phenylalkynyl, naphthyl, naphthylalkyl, naphthylalkenylyl, naphthylalkynyl, indanyl, indanylalkyl, indanylalkenyl, ind
  • the carbocyclic group is typically a cycloalkyl group or a phenyl or naphthyl group.
  • the phenyl or naphthyl groups are typically optionally substituted by relatively lipophilic groups such as halogen or C 1-4 alkoxy or C 1 - 4 alkyl groups wherein the C 1-4 alkoxy or C 14 alkyl groups are each optionally substituted by two or more fluorine atoms.
  • the phenyl or naphthyl groups may be substituted by hydroxy.
  • C 1-2S hydrocarbyl groups R p1 containing a carbocyclic group are cycloalkyl, cycloalkylalkyl and phenylalkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl, cyclohexylmethyl, benzyl and phenylethyl groups.
  • R p1 is a Ci -2S hydrocarbyl group, it can contain 0, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds.
  • the carbon-carbon multiple bonds can be double bonds or triple bonds or a combination of double and triple bonds.
  • R p1 is acyclic, i.e. it contains no carbocyclic groups, it can take the form of a short chain (up to 8 carbon atoms) alkyl, alkenyl or alkynyl group, or it can take the form of a longer chain fatty acid residue (8-25 carbon atoms) which may be saturated or unsaturated.
  • the group R p1 can contain one or more ether linkages and/or an ester linkage.
  • R p1 is hydrogen or a C 1-2S hydrocarbyl group containing 0 or 1 carbocyclic rings and 0, 1, 2 or 3 carbon-carbon multiple bonds, the C 1-25 hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-Ci.
  • one sub-set of compounds consists of the compounds wherein R p1 is hydrogen, a C 1 ⁇ alkyl group, a C 3 . 6 cycloalkyl group or a benzyl group wherein the phenyl and benzyl groups are optionally substituted by halogen or Ci -4 alkoxy or Ci -4 alkyl groups wherein the C 1-4 alkoxy or C 1 . 4 alkyl groups are each optionally substituted by two or more fluorine atoms.
  • L p is a bond.
  • one group of compounds is the group wherein R p1 is a C 1-4 alkyl group or a C 1-6 alkanoyloxy-Ci -2 -alkyl group. In one sub-group, R p1 may be other than a methyl group.
  • the C 1-6 alkanoyloxy-C 1-2 -alkyl group can be a C 1-6 alkanoyloxymethyl group or a C 1-6 alkanoyloxyethyl group.
  • the C 1-6 alkanoyloxy moiety can be, for example, acetoxy, propanoyloxy, ⁇ -butanoyloxy, 2-methylpropanoyloxy, 3-methylbutanoyloxy and pivaloyloxy.
  • R p1 is C 1-6 alkyl optionally substituted by mono- or di-C 1-s alkylamino or a C 1-6 acylamino group.
  • R p1 can be C 1-4 alkyl (e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or te/t-butyl group) optionally substituted by mono- or di-C 1-2 alkylamino or a C 1-4 acylamino group.
  • R p1 is preferably a C 1-6 alkyl group, e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or te/t-butyl group.
  • the two groups R p1 can be the same or different and each can be, for example, hydrogen or a C 1-6 alkyl group (e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or fert-butyl group) or a C 3-6 cyclic group such as cyclopropyl, cyclobutyl, cyclopentyl, cylcohexyl or phenyl group.
  • L p is SO m NR p1 and hence the moiety L p -NR p1 is a group SO m N(R p1 ) 2 .
  • the two groups R p1 can be the same or different and each can be, for example, hydrogen or a C 1-6 alkyl group (e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or fe/f-butyl group) or a C 3-6 cyclic group such as cyclopropyl, cyclobutyl, cyclopentyl, cylcohexyl or phenyl group.
  • both R p1 groups are C 1-4 alkyl groups such as methyl and the moiety L p -NR p1 is, for example, SOp 1 NMe 2 .
  • the integer "m" can be 1 or 2. In one preferred subgroup of compounds, m is 2.
  • R z is SO 3 H.
  • R 1 is OSO 3 H and R 2a is hydroxy;
  • R 2a is OSO 3 H and R 1 is hydroxy;
  • R 1 and R 2a are both OSO 3 H.
  • R z is:
  • R p2 and R p3 are the same or different and each is a group R p1 .
  • R p1 are hydrogen and C 1-4 alkyl groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl and fert-butyl groups.
  • R p2 and R p3 are both hydrogen.
  • R p2 is hydrogen and R p3 is methyl, ethyl, n- propyl, isopropyl, n-butyl, isobutyl or ferf-butyl
  • R p2 and R p3 are both independently selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl and tert- butyl groups.
  • R 1 and R 2a is a group O-R z where R z is a glucuronide residue as defined herein.
  • R 1 is hydroxy and R 2a is a group O-R z where R z is a glucuronide residue.
  • R 2a is hydroxy and R 1 is a group O-R z where R z is a glucuronide residue.
  • R z is a mono-, di- or tripeptide residue as defined herein.
  • the amino acids making up the mono-, di- and tri-peptide residues can be any of the naturally occurring amino acids.
  • the amino acids may be selected from alanine (Ala, A), valine (VaI, V), leucine (Leu, L), lsoleucine (He, I), proline (Pro, P), phenylalanine (Phe, F), tryptophan (T rp, W), methionine (Met, M), glycine (GIy, G), serine (Ser, S), threonine (Thr, T), cysteine (Cys, C), tyrosine (Tyr, Y), asparagine (Asn, N), glutamine (GIn, Q), aspartic acid (Asp, D), glutamic acid (GIu, E), lysine (Lys, K), arginine (Arg, R) and histidine (His, H).
  • amino acids making up the mono-, di or tri-peptide residues in the compounds of the invention also include the D- isomeric forms of the naturally occurring amino acids as well a the L-isomeric forms.
  • Examples of mono-peptide pro-drugs are the compounds wherein R z is an L-GIy, L-AIa or L-Lys or L-VaI residue.
  • R z is a di- or tripeptide residue
  • hPEPTI intestinal human peptide transporter
  • R z can be a di- or tripeptide residue selected from VaI-VaI, Tyr-Gly, GIy-VaI, GIy-GIy, Gly-Tyr, Val-Tyr, Tyr-Val, D-GIu-AIa and D-Asp-Ala.
  • Tri-peptide prodrugs may be constructed so that they are activated by the protease plasminogen activator found in elevated concentrations in many types of malignant cells and human tumours (see PNAS (1980) vol. 77, no. 4, pp 2224-2228).
  • One such pro-drug is formed with the tri-peptide D-Val-Leu-Lys wherein the lysine moiety is bonded to the oxygen atom of the group O-R z .
  • R is selected from hydrogen and fluorine. Preferably R is hydrogen. tf
  • R 3 is selected from hydrogen, halogen, cyano, C 1-S hydrocarbyl and C 1-5 hydrocarbyloxy; wherein the C 1-5 hydrocarbyl and Ci -5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy, amino, mono- and di-C 1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members.
  • R 3 is selected from hydrogen, halogen, cyano, C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy; wherein the C 1-5 hydrocarbyl moiety in each instance is optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy and amino.
  • R 3 is selected from halogen (e.g. chlorine or bromine), C 1-5 alkyl and C 3-4 cycloalkyl. More typically, R 3 is selected from hydrogen, chlorine, C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy.
  • halogen e.g. chlorine or bromine
  • C 1-5 alkyl and C 3-4 cycloalkyl More typically, R 3 is selected from hydrogen, chlorine, C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy.
  • Particular groups R 3 include hydrogen, C 1-5 alkyl, C 2-5 alkenyl and C 3-4 cycloalkyl groups, preferably secondary alkyl and alkenyl groups such as isopropyl, sec-butyl, fe/t-butyl, 1 ,2-dimethylallyl and 1 ,2-dimethylpropyl, or cycloalkyl groups such as cyclopropyl.
  • a further sub-group of substituents R 3 consists of C 1-5 alkyl, C 2-5 alkenyl and C 3 . 4 cycloalkyl groups, preferably secondary alkyl and alkenyl groups such as isopropyl, sec-butyl, fe/t-butyl, 1 ,2- dimethylallyl and 1 ,2-dimethylpropyl, or cycloalkyl groups such as cyclopropyl.
  • R 3 may be other than hydrogen.
  • R 1 and R 2 are both hydroxy and R 3 is hydrogen.
  • R 3 is selected from isopropyl and te/t-butyl. In one general embodiment, R 3 is other than halogen.
  • R 3 may be other than fluorine.
  • R 3 may be other than fluorine or methoxy.
  • R 4 is selected from hydrogen; a group -(O) n -R 7 where n is 0 or 1 and R 7 is an acyclic C 1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C 1-5 hydrocarbylamino, wherein the acyclic C 1-5 hydrocarbyl group and the mono and CU-C 1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy, amino, mono- and di-Ci -2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members.
  • R 4 is selected from hydrogen; a group -(O) n -R 7 where n is 0 or 1 and R 7 is an acyclic Ci -5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C 1-5 hydrocarbylamino, wherein the Ci -5 hydrocarbyl moiety in each instance is optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy and amino.
  • R 4 is more typically selected from hydrogen, methoxy, halogen (e.g. fluorine or chlorine), cyano; hydroxy; amino and C 3-6 cycloalkyl. More particularly, R 4 can be selected from a sub-set R 4a wherein the sub-set R 4a consists of hydrogen, methoxy, fluorine and chlorine.
  • halogen e.g. fluorine or chlorine
  • R 4 is hydrogen
  • R 3 and R 4 together form a carbocyclic or heterocyclic ring of 5 to 7 ring members.
  • the carbocyclic and heterocyclic groups can be any of the groups listed above in General Definitions and Preferences section but one particular group is a group wherein R 3 and R 4 together with the phenyl ring form a dihydrobenzofuran group.
  • phenyl ring containing the moieties R 1 , R 2 , R 3 and R 4 are as set out in Table 1.
  • the point of attachment to the carbonyl group is indicated by means of an asterisk.
  • the phenyl moiety is selected from groups A1 to A21.
  • the phenyl moiety is selected from groups A1 to A18.
  • Preferred phenyl moieties include groups A5, A7, A11 , A13, A14, A15, A16, A17 and A18.
  • Particularly preferred phenyl moieties are A5, A7, A13, A14 and A17.
  • Particularly preferred phenyl moieties are A11 and A13.
  • One particularly preferred phenyl moiety is group A13.
  • each of the structures shown in Table 1 may have one O-R z group and one hydroxy group.
  • R 5 & R 6 R s and R 6 together with the nitrogen atom to which they are attached form a bicyclic heterocyclic group having up to 12 ring members of which up to 5 ring members are heteroatoms selected from oxygen, nitrogen and sulphur.
  • the bicyclic groups can be any of the groups listed above in the General Preferences and Definitions section or listed below in the Particular and Preferred Sub-groups section, and such groups may be unsubstituted or substituted by one or more substituents R 10 as defined herein.
  • the bicyclic heterocyclic group is typically a fused ring bicyclic group or a spirocylic group and more typically is a fused ring bicyclic group.
  • Particular fused ring systems of interest in the context of the invention are 5.6 and 6.6 fused ring systems.
  • one of the rings may be a heterocyclic ring and the other may be a carbocyclic ring, or both rings may be heterocyclic.
  • one of the rings of the bicyclic heterocyclic group is non-aromatic and other is aromatic.
  • the nitrogen atom of the group NR 5 R 6 forms part of the non- aromatic ring.
  • Particular examples of such groups are dihydroindole, dihydroisoindole, tetrahydroquinoline and tetrahydroisoquinoline groups.
  • Such groups are dihydroindole, dihydroisoindole, tetrahydroquinoline and tetrahydroisoquinoline groups, but wherein the tetrahydroisoquinoline group bears no substituent groups on the non-aromatic ring thereof.
  • the bicyclic heterocyclic rings are optionally substituted by one or more substituent groups R 10 as defined herein.
  • the bicyclic heterocyclic ring is subsitituted by 1 , 2 or 3 substituent groups R 10 as defined herein.
  • the bicyclic heterocyclic ring is subsitituted by 1 or 2 substituent groups R 10 as defined herein.
  • the substituent group or groups R 10 may be attached to either or both of the two rings making up the bicyclic heterocyclic group.
  • the ring containing the nitrogen atom of the group NR 5 R 6 does not bear any substituents R 10 .
  • the ring containing the nitrogen atom of the group NR 5 R 6 bears a substituent R 10 but the substituent is other than a carboxylic acid group.
  • the bicyclic heterocyclic group is unsubstituted or is substituted by one, two or three (preferably one or two) substituents selected from a group R 1Oa consisting of halogen, hydroxy, amino and a group R a -R b
  • R a is selected from a bond, O, CO, C(O)O, C(O)NR 0 , NR C C(O), NR 0 C(O)O, NR 0 , SO, SO 2 , SONR 0 , and SO 2 NR 0
  • R b is selected from hydrogen; carbocyclic and heterocyclic groups having 5 or 6 ring members; and Ci -10 hydrocarbyl (e.g.
  • C 1-8 hydrocarbyl such as C 1-8 alkyl or C 3 . 7 cycloalkyl) optionally substituted by one or more substituents selected from hydroxy, oxo, amino, mono- or CJi-C 1 ⁇ 8 non-aromatic hydrocarbylamino, (e.g. mono- or di-C 1-4 hydrocarbylamino), carboxy, and carbocyclic and heterocyclic groups having from 3 to 7 ring members, and wherein one or more of the carbon atoms of the C 1-8 hydrocarbyl group may optionally be replaced by O, S, C(O)O, C(O)NR 0 or NR 0 .
  • R b when R b is a carbocyclic or heterocyclic group, the carbocyclic or heterocyclic group may be substituted by one or more substituents R 10 as defined herein.
  • R 10 substituents
  • the carbocyclic or heterocyclic group when R b is a carbocyclic or heterocyclic group, the carbocyclic or heterocyclic group may be substituted by one or more substituents selected from CO 2 R 14 wherein R 14 is hydrogen or C 1-6 alky!;
  • the bicyclic ring is unsubstituted or is substituted by 1, 2 or 3 (e.g. 1 or 2, for example 1) groups R 100 where R 10 ° is a group [sol], CH 2 [SoI] or OCH 2 CH 2 [SoI] where [sol] is selected from the following groups
  • R 1Qc is optionally further selected from a group OCH 2 CH 2 CH 2 [SoI] and/or (ii) [sol] is further selected from NHR 11 wherein R 11 is COR 12 or R 12 and R 12 is C 1-4 alkyl, aryl or aryl-C-,- 4 alkyl.
  • the bicyclic ring is unsubstituted or is substituted by one or two substituents R 1Occ where R 1Occ is selected from: halogen;
  • R 14 wherein R 14 is hydrogen or C 1-5 alkyl
  • R 11 is hydrogen, COR 12 , C(O)OR 12 or R 12 ;
  • R 12 is C 1-6 alkyl, C 3-6 cycloalkyl, aryl, aryl-C 1-6 alkyl or CH 2 R 15 ; and
  • R 15 is selected from hydrogen, C 1-6 alkyl, C 3-6 cycloalkyl, hydroxy-Ci -6 alkyl, piperidine, N-C 1-6 alkylpiperazine, piperazine, morpholine,
  • the bicyclic ring is unsubstituted or is substituted by one or two substituents R 1Occc where R 10occ is selected from: a group [sol] or CH 2 [sol] where [sol] is selected from the following groups:
  • R 1 1 1 1 i-s hydrogen, COR i12 , C(O)OR 1 " or R j1 J 2 Z .; D R1 1 2 ⁇ is
  • R 10b or R 10c or R 10cc is a group [sol], CH 2 [SoI], OCH 2 CH 2 [SoI] or OCH 2 CH 2 CH 2 [SoI] and [sol] contains a primary or secondary amine group
  • the primary or secondary amine group can be derivatised to form an acyl derivative such as an amide, carbamate or urea.
  • the amine group can be derivatised to form a carbamate such as a C ⁇ alkyloxycarbonylamino group, or a benzyloxycarbonylamino group.
  • R 5 and R 6 together with the nitrogen atom to which they are attached form an optionally substituted dihydroisoindole group wherein the optional substituents aarree sseelleecctteedd ffrroom groups R 10 , R 1Oa , R 1Ob , R 1Oc and R 1Occ and sub-groups and examples thereof as defined herein.
  • One set of preferred groups NR 5 R 6 consists of or includes groups B8 and B30. Another preferred group NR 5 R 6 is group B8.
  • a further set of preferred groups NR 5 R 6 consists of groups B8, B35, B36, B37, B38, B39, B40, B41 , B42, B43, B45, B46, B48, B53, B54, B55, B55, B57, B58, B59, B60, B61 and B62.
  • a further set of preferred groups NR 5 R 6 consists of groups B8, B35, B36, B37, B38, B39, B40, B41 , B42, B43, B45, B46, B48, B53, B54, B55, B56, B57, B58, B59, B60, B61 and B62
  • Another set of preferred groups consists of B8, B35, B36, B37, B38, B39, B40, B41 , B42, B43, B45, B46, B48, B53, B54, B55, B56, B57, B58, B59, B60, B61 , B62, B71 , B72, B74, B75, B76, B77, B78, B79, B80, B81, B82, B83, B85, B86, B87, B93, B94, B95, B97, B98, B99, B100 and B101.
  • a further sub-set of groups NR 5 R 6 consists of B43, B46, B48, B76, B82, B89, B91 and B96.
  • more preferred groups are groups B43, B46, B48, B76, B82, B89 and B91 , with B76, B82 and B89 being particularly preferred.
  • R z is as defined herein;
  • R 3a is selected from hydrogen, halogen, cyano, Ci -5 hydrocarbyl and Ci -5 hydrocarbyloxy; wherein the Ci -5 hydrocarbyl and Ci -5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, Ci -2 alkoxy, amino, mono- and di-C 1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R 4 is selected from hydrogen; a group -(O) n -R 7 where n is 0 or 1 and R 7 is an acyclic C 1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C 1-5 hydrocarbyl-amino, wherein the acyclic Ci -5 hydrocarbyl group and the mono and di-C 1-5 hydrocarbylamino moieties in each instance are
  • R z is as defined herein;
  • R 3b is selected from hydrogen, halogen, cyano, C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy; wherein the C 1-5 hydrocarbyl and C 1-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy, amino, mono- and di-C 1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R 4 is selected from hydrogen; a group -(O) n -R 7 where n is 0 or 1 and R 7 is an acyclic C 1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-Ci -5 hydrocarbyl-amino, wherein the acyclic C 1-5 hydrocarbyl group and the mono and di-C 1-5 hydrocarbylamino moieties in each instance
  • a further sub-group of novel compounds of the invention can be represented by the formula (IV): or salts, tautomers, solvates and N-oxides thereof; wherein:
  • R 1 and R 2a are as defined herein;
  • R 3c is selected from hydrogen, halogen, cyano, Ci -5 hydrocarbyl and Ci -5 hydrocarbyloxy; wherein the C 1-5 hydrocarbyl and Ci -5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy, amino, mono- and di-C 1-2 alkylamino, and ary] and heteroaryl groups of 5 to 12 ring members;
  • R 4 is selected from hydrogen; a group -(O) n -R 7 where n is 0 or 1 and R 7 is an acyclic C 1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C 1-5 hydrocarbyl-amino, wherein the acyclic C 1-5 hydrocarbyl group and the mono and di-C 1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, Ci -2 alkoxy, amino, mono- and di-C 1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; or R 3c and R 4 together form a monocyclic carbocyclic or heterocyclic ring of 5 to 7 ring members; R 5 and R 6 together with the nitrogen atom to which they are attached form a bicyclic group having up
  • R 8 is selected from hydrogen and fluorine.
  • NR 5 R 6 forms a bicyclic ring of up 10 ring members (e.g. 9 or 10 ring members, preferably 9 ring members) of which up to 5 ring members are heteroatoms selected from O, N and S, the monocylic or bicyclic ring being optionally substituted by up to three substituent groups R 10 , R 1Oa , R 10b , R 1Oc and R 1Occ as defined herein, more typically up to two substituents, for example up to one substituent.
  • up 10 ring members e.g. 9 or 10 ring members, preferably 9 ring members
  • up to 5 ring members are heteroatoms selected from O, N and S
  • R 10 , R 1Oa , R 10b , R 1Oc and R 1Occ as defined herein, more typically up to two substituents, for example up to one substituent.
  • substituents for the bicyclic heterocyclic group NR 5 R 8 are those forming part of a sub-group R 10d which consists of the members of sub-group R 1Oc and fluoro, chloro, bromo, trifluoromethyl, difluoromethoxy, trifluoromethoxy, cyano, methyl, ethyl, cyclopropyl, hydroxy, methylsuiphonyl, amino, methylamino, dimethylamino, methoxy, ethoxy, hydroxy methyl, hydroxyethyl, ethoxycarbonyl, methoxycarbonyl, aminocarbonyl, oxo, methoxymethyl, carboxy, phenyl, C 1-2 alkoxycarbonyl, aminocarbonyl, acetyl, methylsuiphonyl and pyridyl.
  • one sub-set of substituents includes methyl, ethyl, chloro, fluoro, hydroxy, methylsuiphonyl, amino, methylamino, dimethylamino, cyano, methoxy, ethoxy, hydroxymethyl, cyclopropyl, hydroxyethyl, ethoxycarbonyl, methoxycarbonyl, aminocarbonyl, oxo, methoxymethyl and acetyl.
  • NR 5 R 6 can form a 5.6 or 6.6 fused bicyclic ring of 9 or ten ring members of which 1 to 3 are heteroatoms, the bicyclic ring being optionally substituted by one or more substituents R 10 or R 1Oa or R 1Qb or R 10c or R 10 ⁇ or R 10d and sub-groups (sub-sets) and examples thereof as defined herein.
  • fused bicyclic rings are those in which a non-aromatic ring such as a pyrrolidine, piperidine, piperazine or morpholine ring is fused to a 6-membered aryl or heteroaryl ring such as a benzene or pyridine ring, and wherein a nitrogen atom present in the non-aromatic ring is bonded to the carbonyl group in formulae (II), (IiI) or (IV).
  • a non-aromatic ring such as a pyrrolidine, piperidine, piperazine or morpholine ring
  • a 6-membered aryl or heteroaryl ring such as a benzene or pyridine ring
  • fused bicyclic rings include dihydroindole, dihydroisoindole, tetrahydroquinoline and tetrahydroisoquinoline, and aza-analogues thereof in which one or two carbon ring members in the aromatic ring are replaced by nitrogen.
  • One sub-group of bicyclic heterocyclic groups formed by NR 5 R 6 consists of dihydroisoindole optionally substituted by one or more (e.g. 1 , 2 or 3) optional substituents selected from groups R 10 , R 1Oa , R 1Ob and R 10o or R 10co and or R 10d and sub-groups (sub-sets) and examples thereof as defined herein.
  • Preferred compounds are those wherein the group R 3a or R 3b or R 30 is selected from hydrogen, halogen and C 1-5 alkyl; wherein the C 1-5 alkyl moiety in each instance is optionally substituted by one or more substituents selected from hydroxy, halogen, C ⁇ 2 alkoxy and amino.
  • the group R 3a or R 3b or R 3c is hydrogen or a C 3-5 alkyl group optionally substituted by one or more substituents selected from hydroxy, halogen, C 1-2 alkoxy and amino.
  • the group R 3a or R 3b or R 3c is selected from hydrogen and isopropyl, sec-butyl, ferf-butyl and 1,2- dimethylpropyl groups.
  • R 1 and R 2a are as defined herein;
  • R 3d is selected from ethyl and secondary and tertiary alkyl groups of 3 to 6 carbon atoms;
  • R 4a is selected from hydrogen, fluorine, chlorine and methoxy;
  • R 5 , R s and R 8 are as defined herein; provided that when R 1 and R 2 are both hydroxy, then R 3d can additionally be selected from hydrogen.
  • R 3d when one of R 1 and R 2a is hydroxy and the other is O-R z , or when R 1 and R 2a is both O-R z , then R 3d is hydrogen.
  • ⁇ ,3d
  • R is ethyl or a secondary or tertiary alkyl group.
  • Particularly preferred alkyl groups R 3d are ethyl, isopropyl and ferf-butyl, and in particular isopropyl.
  • preferred groups NR 5 R 6 are dihydroisoindole groups which may be substituted or unsubstituted by one, two or three groups R 10 , R 1Oa or R 1Ob or R 100 or R 10 ⁇ or R 10d and sub-groups (sub-sets) and examples thereof as defined herein but, in one particular embodiment, are unsubstituted.
  • R 1 and R 2a are as defined herein;
  • ring B is an aromatic ring containing up to two (and preferably 0 or 1) nitrogen heteroatom ring members;
  • T is a group (CHR 10b ) j and Q is a group (CHR 10b ) k where j and k are each 0, 1 , 2 or 3 provided that the sum of j and k is 2 or 3;
  • n is 0, 1 , 2 or 3 and R , R 3 4a , O R8 and R 10 U b D are as defined herein.
  • R 1 is- hydrogen.
  • R 5 1 i,s. hydroxy or a group O-R z .
  • Preferrred groups are groups C1 , C5 and C6
  • R 10 can be a group R 10 as hereinbefore defined or can be a group R 10b , R 100 , R 1Occ or R 1Occc as defined herein.
  • n is preferably 1 , 2 or 3, and more preferably is 1 or 2, e.g. 1.
  • a currently preferred group is group C1.
  • R 1 , R 2a , R 3 R 4a , R 8 and R 10b are as defined herein and n is 0, 1 2 or 3 (more preferably 0, 1 or 2, e.g. 0 or 1), and provided that at least one of R 1 and R 2a is O-R z .
  • R 1 and R 2a are both hydroxy.
  • R 3 is selected from hydrogen, halogen, C 1-5 alkyl, C 2-5 alkenyl and C 3-4 cycloalkyl groups;
  • R ⁇ 4a is selected from hydrogen, fluorine, chlorine and methoxy; R is hydrogen or fluorine; n is 0, 1 2 or 3; and R 1 , R 2a and R 10 are each as defined herein.
  • R 10 can be selected from, for example, one, two or three groups R 1Oa or R 10b or R 10o or R 1Occ or R 10d and sub-groups (sub-sets) and examples thereof as defined herein.
  • R 1 and R 2a are each as defined herein, R 3 is selected from hydrogen, halogen, C 1-5 alkyl, C 2-5 alkenyl and C 3-4 cycloalkyl groups; R 4a is selected from hydrogen, fluorine, chlorine and methoxy; R 8 is hydrogen or fluorine; n is 0, 1 2 or 3; and R 1Occ is selected from: halogen;
  • R 14 is hydrogen or C 1-6 alkyl; C 1-4 alkyl optionally substituted by hydroxy or C 1-2 alkoxy; C 1-4 alkoxy optionally substituted by hydroxy or C 1-2 alkoxy; or a group [sol], CH 2 [SoI], C(O)[sol], OCH 2 CH 2 [SoI] or OCH 2 CH 2 CH 2 [SoI] where [sol] is selected from the following groups
  • R 12 is C 1-6 alkyl, C 3-6 cycloalkyl, aryl, aryl-Ci -6 alkyl or CH 2 R 15 ; and R 15 is selected from hydrogen, Ci -6 alkyl, C 3 .
  • B cycloalkyl, hydroxy-C 1-6 alkyl, piperidine, N-Ci -6 alkylpiperazine, piperazine, morpholine, COR 13 or C(O)OR 13 ; and R 13 is C 1 * alkyl.
  • the compound can be an aza- or diaza- analogue of the the compounds of formulae (Vl), (VII) and (Vila) as defined herein wherein one or two of the carbon atoms of the benzene ring attached to the five membered ring is replaced by nitrogen.
  • n is preferably 1 , 2 or 3, and more preferably is 1 or 2. In one embodiment, n is 1.
  • Q 1 is a bond or a group CH 2 and R 1 and R 2a are as defined herein.
  • Specific compounds of the invention include pro-drugs of the following compounds: (5-chloro-2-hydroxy-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone; (3-tert-butyl-4-hydroxy-phenyl)-(2,3-dihydro-indoI-1-yl)-methanone; (3-tert-butyl-4-hydroxy-phenyl)-(3,4-dihydro-2H-quinolin-1-yl)-methanone; (3,4-dihydro-1 H-isoquinolin-2-yl)-(4-hydroxy-3-isopropyl-phenyl)-methanone; (1 ,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone;
  • Preferred individual compounds of the formula (I) are:
  • a particularly preferred set of individual compounds consists of pro-drugs of: (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]- methanone;
  • each general and specific preference, embodiment and example of the group R 1 may be combined with each general and specific preference, embodiment and example of the groups R 2 or R 2a and/or R 3 and/or R 4 and/or R 4 and/or R 5 and/or R 6 and/or R 10 and/or Q and/or T and/or sub-groups thereof as defined herein and that all such combinations are embraced by this application.
  • the various functional groups and substituents making up the compounds of the formula (I) are typically chosen such that the molecular weight of the compound of the formula (I) does not exceed 1000. More usually, the molecular weight of the compound will be less than 750, for example less than 700, or less than 650, or less than 600, or less than 550. More preferably, the molecular weight is less than 525 and, for example, is 500 or less.
  • a reference to a compound of the formulae (V! 0 ), (Vl), (I 0 ), (I) and sub-groups thereof also includes ionic forms, salts, solvates, isomers, tautomers, N-oxides, esters, isotopes and protected forms thereof, for example, as discussed below; preferably, the salts or tautomers or isomers or N-oxides or solvates thereof; and more preferably, the salts or tautomers or N-oxides or solvates thereof
  • the salts of the present invention can be synthesized from the parent compound that contains a basic or acidic moiety by conventional chemical methods such as methods described in Pharmaceutical Salts: Properties, Selection, and Use, P. Heinrich Stahl (Editor), Camille G. Wermuth (Editor), ISBN: 3-90639-026-8, Hardcover, 388 pages, August 2002.
  • such salts can be prepared by reacting the free acid or base forms of these compounds with the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media such as ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are used.
  • Acid addition salts may be formed with a wide variety of acids, both inorganic and organic.
  • acid addition salts include salts formed with an acid selected from the group consisting of acetic, 2,2-dichloroacetic, adipic, alginic, ascorbic (e.g.
  • a salt may be formed with a suitable cation.
  • suitable inorganic cations include, but are not limited to, alkali metal ions such as Na + and K + , alkaline earth metal cations such as Ca 2+ and Mg 2+ , and other cations such as Al 3+ .
  • suitable organic cations include, but are not limited to, ammonium ion (i.e., NH 4 + ) and substituted ammonium ions (e.g., NH 3 R + , NH 2 Ra + , NHR 3 + , NR 4 + ).
  • Examples of some suitable substituted ammonium ions are those derived from: ethylamine, diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids, such as lysine and arginine.
  • An example of a common quaternary ammonium ion is N(CH 3 )/.
  • the salt forms of the compounds of the invention are typically pharmaceutically acceptable salts, and examples of pharmaceutically acceptable salts are discussed in Berge ef a/., 1977, "Pharmaceutically Acceptable Salts," J. Pharm. Sci., Vol. 66, pp. 1-19. However, salts that are not pharmaceutically acceptable may also be prepared as intermediate forms which may then be converted into pharmaceutically acceptable salts. Such non-pharmaceutically acceptable salts forms, which may be useful, for example, in the purification or separation of the compounds of the invention, also form part of the invention.
  • Compounds of the formulae (Vl 0 ), (Vl), (I 0 ), (I) and sub-groups thereof containing an amine function may also form N-oxides.
  • a reference herein to a compound of any of the formulae (Vl 0 ), (Vl), (1°), (I) and sub-groups thereof that contains an amine function also includes the N-oxide.
  • N-oxides are the N-oxides of a tertiary amine or a nitrogen atom of a nitrogen-containing heterocycle.
  • N-Oxides can be formed by treatment of the corresponding amine with an oxidizing agent such as hydrogen peroxide or a per-acid (e.g. a peroxycarboxylic acid), see for example Advanced Organic Chemistry, by Jerry March, 4 th Edition, Wiley Interscience, pages. More particularly, N-oxides can be made by the procedure of L. W. Deady (Syn. Comm. 1977, 7, 509-514) in which the amine compound is reacted with m-chloroperoxybenzoic acid (MCPBA), for example, in an inert solvent such as dichloromethane.
  • MCPBA m-chloroperoxybenzoic acid
  • tautomeric forms include, for example, keto-, enol-, and enolate-forms, as in, for example, the following tautomeric pairs: keto/enol (illustrated below), imine/enamine, amide/imino alcohol, amidine/amidine, nitroso/oxime, thioketone/enethiol, and nitro/aci-nitro.
  • references to compounds of the formulae (Vl 0 ), (Vl), (I 0 ), (I) and sub-groups thereof include all optical isomeric forms thereof (e.g. enantiomers, epimers and diastereoisomers), either as individual optical isomers, or mixtures (e.g. racemic mixtures) or two or more optical isomers, unless the context requires otherwise.
  • optical isomers may be characterised and identified by their optical activity (i.e. as + and - isomers, or d and / isomers) or they may be characterised in terms of their absolute stereochemistry using the "R and S" nomenclature developed by Cahn, lngold and Prelog, see Advanced Organic Chemistry by Jerry March, 4 th Edition, John Wiley & Sons, New York, 1992, pages 109-114, and see also Cahn, lngold & Prelog, Angew. Chem. Int. Ed. Engl., 1966, 5, 385- 415.
  • Optical isomers can be separated by a number of techniques including chiral chromatography (chromatography on a chiral support) and such techniques are well known to the person skilled in the art.
  • optical isomers can be separated by forming diastereoisomeric salts with chiral acids such as (+)-tartaric acid, (-)-pyroglutamic acid, (-)-di- toluoyl-L-tartaric acid, (+)-mandelic acid, (-)-malic acid, and (-)-camphorsulphonic, separating the diastereoisomers by preferential crystallisation, and then dissociating the salts to give the individual enantiomer of the free base.
  • chiral acids such as (+)-tartaric acid, (-)-pyroglutamic acid, (-)-di- toluoyl-L-tartaric acid, (+)-mandelic acid, (-)-malic acid, and (-)-camphorsulphonic
  • the compound of the formula (I) is present as a single optical isomer (e.g. enantiomer or diastereoisomer). In one general embodiment, 99% or more (e.g. substantially all) of the total amount of the compound of the formula (I) may be present as a single optical isomer (e.g. enantiomer or diastereoisomer).
  • the compounds of the invention include compounds with one or more isotopic substitutions, and a reference to a particular element includes within its scope all isotopes of the element.
  • a reference to hydrogen includes within its scope 1 H, 2 H (D), and 3 H (T).
  • references to carbon and oxygen include within their scope respectively 12 C, 13 C and 14 C and 16 O and 18 O.
  • the isotopes may be radioactive or non-radioactive.
  • the compounds contain no radioactive isotopes. Such compounds are preferred for therapeutic use.
  • the compound may contain one or more radioisotopes. Compounds containing such radioisotopes may be useful in a diagnostic context.
  • Esters such as carboxylic acid esters and acyloxy esters of the compounds of formulae (Vl 0 ), (Vl), (I 0 ), (I) and sub-groups thereof bearing a carboxylic acid group or a hydroxyl group are also embraced by formulae (Vl 0 ), (Vl), (I 0 ), (I) and sub-groups thereof.
  • formulae (Vl 0 ), (Vl), (I 0 ), (I) and sub-formulae, sub-groups, preferences and examples thereof do not cover esters such as carboxylic acid esters and acyloxy esters.
  • formulae (Vl 0 ), (Vl), (I 0 ), (I) and sub-groups thereof are any polymorphic forms of the compounds, solvates (e.g. hydrates) and complexes (e.g. inclusion complexes or clathrates with compounds such as cyclodextrins, or complexes with metals) of the compounds.
  • the compounds of the formulae (Vl 0 ), (VI), (I 0 ), (I) and sub-groups thereof are pro-drugs of compounds that are inhibitors of Hsp90
  • the pro-drug compounds of the invention have a number of advantages with respect to their parent compounds. For example, they may provide improved oral bioavailability, e.g. by virtue of improved intestinal absorption. Furthermore, by forming a pro-drug, conjugation and/or metabolism of the parent compound may be substantially reduced.
  • the pro-drug compounds of the invention are expected to be beneficial in the treatment of wide spectrum of proliferative disorders.
  • proliferative disorders include, but are not limited to, a carcinoma, for example a carcinoma of the bladder, breast, colon (e.g. colorectal carcinomas such as colon adenocarcinoma and colon adenoma), kidney, epidermis, liver, lung, for example adenocarcinoma, small cell lung cancer and non-small cell lung carcinomas, oesophagus, gall bladder, ovary, pancreas e.g. exocrine pancreatic carcinoma, stomach, cervix, thyroid, prostate, gastrointestinal system, e.g.
  • gastrointestinal stromal tumours or skin, for example squamous cell carcinoma
  • a hematopoieitic tumour of lymphoid lineage for example leukaemia, acute lymphocytic leukaemia, chronic lymphocytic leukaemia, B-cell lymphoma (such as diffuse large B cell lymphoma), T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, hairy cell lymphoma, or Burkitt's lymphoma
  • a hematopoieitic tumour of myeloid lineage including acute myeloid leukaemia, chronic myeloid leukaemias, myelogenous leukaemias, and lmatinib sensitive and refractory chronic myelogenous leukaemias, myelodysplastic syndrome, Bortezomib sensitive and refractory multiple myeloma, myeloproliferative disease or promy
  • the cancers may be cancers which are sensitive to Hsp90 inhibition, and such cancers may be determined by a method as set out in the section headed "Methods of Diagnosis".
  • One group of cancers includes human breast cancers (e.g. primary breast tumours, node-negative breast cancer, invasive duct adenocarcinomas of the breast, non-endometrioid breast cancers); and mantle cell lymphomas.
  • other cancers are colorectal and endometrial cancers.
  • Another sub-set of cancers includes hematopoietic tumours of both lymphoid and myeloid lineage, for example acute lymphoblastic leukemia, chronic lymphocytic leukaemia (Both T and B cell), acute myeloid leukaemia, chronic myeloid leukaemia, mantle cell lymphoma and B-cell lymphoma (such as diffuse large B cell lymphoma) and optionally further includes chronic myelogenous leukaemia and multiple myeloma.
  • acute lymphoblastic leukemia chronic lymphocytic leukaemia (Both T and B cell)
  • acute myeloid leukaemia chronic myeloid leukaemia
  • chronic myeloid leukaemia chronic myeloid leukaemia
  • mantle cell lymphoma such as diffuse large B cell lymphoma
  • B-cell lymphoma such as diffuse large B cell lymphoma
  • chronic myelogenous leukaemia and multiple myeloma optionally
  • a preferred sub-set of cancers consists of ErbB2-positive breast, prostate, lung, and gastric cancer; chronic myeloid leukemia; androgen receptor dependent prostate cancer; Flt3-dependent acute myeloid leukaemia; melanoma associated with Braf mutation; multiple myeloma; velcade refractory multiple myeloma; and gastrointestinal stromal tumours (GIST).
  • cancers are multiple myelomas and velcade refractory tumour types as defined herein.
  • Another preferred sub-set of cancers consists of hormone refractory prostate cancer, metastatic melanoma, HER2 positive breast cancer, mutant EGFR positive non-small cell lung carcinoma and Gleevec resistant gastrointestinal stromal tumours.
  • Hsp90 inhibitors could also be used to treat other conditions such as viral infections, parasitic disease, autoimmune diseases (e.g. multiple sclerosis and lupus erythematosus), neuro- degenerative disorders (e.g. Alzheimer's disease), inflammation, Type I and Il diabetes, atherosclerosis and cardiac disease.
  • autoimmune diseases e.g. multiple sclerosis and lupus erythematosus
  • neuro- degenerative disorders e.g. Alzheimer's disease
  • inflammation Type I and Il diabetes
  • atherosclerosis and cardiac disease e.g. Alzheimer's disease
  • Hsp90 inhibitors could also have clinical benefit in transplantation and immunosuppression.
  • Hsp90 inhibitors may also have clinical benefit in the previously described diseases when used in combination with existing or new therapeutic agents. Based on the activities of Hsp90 client proteins and experimental evidence, the following disorders may be particularly sensitive to treatment by Hsp90 inhibitors.
  • ErbB2 Overexpression of ErbB2 (HER-2) occurs in approximately 30 % of breast cancers and ErbB2 receptor down-regulation by herceptin sensitized cells to Taxol. ErbB2 over-expression is linked to poor prognosis and drug resistance (Tsugawa et. al., 1993. Oncology 1993; 50: 418).
  • Ansamycin antibiotics, such as geldanamycin potently inhibit heat shock protein 90 (Hsp90), promoting ubiquitin-mediated degradation of oncogenic kinases that require the chaperone for proper conformational folding.
  • the aberrant BCR-AbI protein is created through a chromosomal translocation and results in a constitutively active AbI kinase domain. This translocation event has been shown to be causal for CML.
  • P210BcrAbl is a known client protein for Hsp90.
  • Treatment of the BCR-AbI positive cell line K562 with an hsp90 inhibitor induced apoptosis.
  • the Bcr-Abl inhibitor Gleevec ® also induces apoptosis in K562 cells; however Gleevec ® resistant K562 cells still retain sensitivity towards Hsp90 inhibitors (Gorre et. al. 2002, Blood 100: 3041-3044).
  • the androgen receptor kinase is an Hsp90 client protein. Testosterone deprivation is usually adopted where surgery is not curative. The cancer may become refractory to hormone manipulation through receptor mutation. Hsp90 regulation of the receptor would still be viable post-mutation.
  • the Hsp90 inhibitor 17-AAG potently inhibits proliferation of Bortezomib refractory multiple myeloma cell lines.
  • Cell surface levels of IGF-1 R and IL-6R were also diminished in 17-aag treated MM-1 cells (Mitsiades et. al., Blood 107:1092-1100, 2006).
  • Autocrine stimulation of multiple myeloma cells, as well as paracrine stimulation of bone marrow stromal cells with IL-6 is also diminished through downregulation of the Hsp90 client IKK.
  • Compounds of the present invention can be used in the treatment of velcade refractory tumour types including treatment of patients with second line mantle cell lymphoma, indolent non- Hodgkin's lymphoma, stage NIB and IV Bronchioloalveolar carcinoma, advanced non-small cell lung cancer, breast, prostate and ovarian cancers and non-Hodgkin's lymphoma.
  • GIST Gastrointestinal stromal tumours
  • GIST disease particularly disease dependent on growth factor activation or overexpression (e.g. c- kit).
  • B-CLL ZAP-70 is an Hsp90 client protein and the requirement for Hsp90 by ZAP-70 is limited to CLL cells and is not observed in T cells where this kinase is normally expressed. Hence, ZAP-70 is unique among identified Hsp90 clients as its chaperone dependency is conditional on the type of cell in which it is expressed.
  • Hsp90 inhibitor may be of clinical benefit.
  • Huntington's disease is a progressive neurodegenerative disorder with no effective treatment. GA inhibition of Hsp90 and the resulting up-regulation of Hsps are effective in preventing huntington protein aggregation in neuronal cells. (Sittler et. al., 2001 , Human Molecular Genetics, Vol. 10, No. 12 1307-1315). Up-regulation of HSP may also be of clinical benefit in other diseases of protein misfolding e.g., CJD and Alzheimer's.
  • Inflammatory disease including Rheumatoid arthritis, Asthma, Chronic obstructive pulmonary disease, and inflammatory bowel disease
  • GA has been shown to dissociate HSF-1 from Hsp90 leading to the activation and nuclear translocation of HSF-1.
  • HSF-1 subsequently acts as a transcription factor to induce HSP90 and Hsp70.
  • the induction of Hsp70 has been implicated in the resolution of inflammation in an induced mouse model of edema (lanaro et al., 2004 Human Molecular Genetics, 2001 , Vol. 10, No. 12 1307-1315).
  • GA treatment inhibited IkappaB kinase (IKK) activation by TNF-a or PMA.
  • IkBa is a regulator of Nf-kB and Ap- 1. (Broemer et al. 2004 ).
  • Ap-1 and Nf-kB is a major transcription factor leading to the production of pro-inflammatory cytokines (Yeo et. al., 2004 Biochem Biophys Res Commun. 30; 320(3):816-24).
  • the stability of pro-inflammatory cytokine transcripts is also regulated through inhibition of p38 MapK (Wax et. al., 2003. Rheumatism Vol. 48, No. 2, pp 541-550).
  • Atherosclerosis It is known that inflammatory and immune cells play a central role in the initiation and progression of human atherosclerosis (Rigan ⁇ et al., Ann. N. Y. Acad. ScL, 2007, 1107:1-1010) and it has been proposed that Hsp90 acts as an autoantigen in carotid atherosclerosis. Rigan ⁇ et al. found specific antibodies and cells against Hsp90 in the sera of 60% of patients tested who were suffering from carotid atherosclerotic plaques but no specific antibodies and T cells against Hsp90 in the sera of healthy patients. Therefore, inhibitors of Hsp90 should be useful in the treatment or prevention of atherosclerosis.
  • Angioqenesis related disease including but not limited to: tumour anqioqenesis, psoriasis, rheumatoid arthritis, and diabetic retinopathy
  • Induction of angiogenesis is regulated by Hsp90 client proteins eNOS and Akt in endothelial cells (Sun and Liao, 2004 Arterioscler Thromb Vase Biol. 24(12):2238-44). Suppression of hypoxia- inducible factor (HIF)-Ia can also impair the growth, angiogenesis and vessel maturation of gastric tumours in a mouse model. (Stoeltzing et. al., 2004 J Natl Cancer Inst; 96:946-956.).
  • HIF hypoxia- inducible factor
  • Type I and type Il diabetes Hsp90 inhibition has a profound effect on Akt signalling as well as e-nos. These are two key regulators in high glucose induced endothelial cell apoptosis in type I diabetes (Lin et. al., 2005 J Cell Biochem. 1 ; 94(1): 194-201) and the development of hypertension in type Il diabetes (Kobayashi et. al., 2004 Hypertension. 44(6):956-62.).
  • Hsps Cardiac ischemic is the most common cause of death in the western world. Hsps, and notably Hsp70 (induced by radicicol treatment) have demonstrated cardioprotective activity in rat cardiomyocytes (Griffin ef. a/., 2004). Inhibition of Hsp90 results in the release of HSF-1 from the chaperone complex and its subsequent activation of Hsp genes. Inhibition of Hsp90 also leads to the down-regulation of HIF-1 , which has been implicated in the pathogenesis of ischemic heart disease and stroke.
  • Hepatits C viral NS2/3 protease is an Hsp90 client protein and Hsp90 activity is required for viral processing and replication (Whitney et. al., 2001. Proc Natl Acad Sci U S A. 20;98(24):13931-5.).
  • GA has reported antimalarial activity against an Hsp90 ortholog of Plasmodium falciparum. Plasmodium growth was inhibited with GA at an IC 50 similar to that observed with chloroquine. GA was also effective against chloroquine resistant strains of Plasmodium falciparum (Kamar et. al., 2003 . Malar J.15; 2(1):30).
  • modulators or inhibitors of stress protein function in general represent a class of chemotherapeutics with the potential for: (i) sensitizing malignant cells to anticancer drugs and/or treatments; (ii) alleviating or reducing the incidence of resistance to anticancer drugs and/or treatments; (iii) reversing resistance to anticancer drugs and/or treatments; (iv) potentiating the activity of anticancer drugs and/or treatments; (v) delaying or preventing the onset of resistance to anticancer drugs and/or treatments. Accordingly, the invention further provides:
  • a method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90 which method comprises administering to a subject in need thereof a compound of the invention, wherein the disease state or condition mediated by Hsp90 is the development of resistance to a cancer drug.
  • a method for the treatment of a cancer which method comprises administering to a subject in need thereof a compound of the invention, which method is characterized by the absence of drug resistance.
  • a method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90 in a subject undergoing treatment with a therapeutic agent such as an anti-cancer agent
  • a therapeutic agent such as an anti-cancer agent
  • a method for the treatment of a cancer in a subject undergoing treatment with an anticancer agent which method comprises administering to a subject in need thereof a compound of the invention, which method is characterized by the absence of drug resistance to the anti-cancer agent.
  • the biological activity (e.g. as inhibitors of Hsp90) of the phenolic compounds from which the prodrug compounds of the invention are derived can be measured using the assays set forth in the examples below, for example the isothermal titration calorimetry (ITC) experiments described in Example 86 and the anti-proliferative activity assays described in Example 87.
  • the level of activity exhibited by a given compound in the ITC assay can be defined in terms of the K d value, and preferred compounds of the present invention are compounds having a K d value of less than 1 micromolar, more preferably less than 0.1 micromolar.
  • the level of activity exhibited by a given compound in an assay can be defined in terms of the IC 50 value, and preferred compounds of the present invention are compounds having an IC 50 value of less than 1 micromolar, more preferably less than 0.1 micromolar.
  • the hERG channel is one of a family of potassium ion channels the first member of which was identified in the late 1980s in a mutant Drosophila melanogaster fruitfly (see Jan, L. Y. and Jan, Y.N. (1990). A Superfamily of Ion Channels. Nature, 345(6277):672).
  • the biophysical properties of the hERG potassium ion channel are described in Sanguinetti, M.
  • HERG encodes the lkr potassium channel. Cell, 81 :299-307, and Trudeau, M. C 1 Warmke, J.W., Ganetzky, B., and Robertson, G.A. (1995). HERG, a Human Inward Rectifier in the Voltage-Gated Potassium Channel Family. Science, 269:92-95.
  • Preferred pro-drug compounds are the pro-drug compounds of phenolic compounds having mean IC 50 values against hERG that are greater than 30 times, or greater than 40 times, or greater than 50 times the IC 50 values of the compounds in cellular proliferation assays.
  • Preferred pro-drugs are the pro-drugs of phenolic compounds having mean IC 50 values against hERG that are greater than 5 ⁇ M, more particularly greater than 10 ⁇ M, and more preferably greater than 15 ⁇ M.
  • Some phenolic compounds from which the pro-drug compounds of the invention are derived have mean IC 50 values against hERG that are greater than 50 ⁇ M.
  • Compounds of the invention have advantageous ADME properties and in particular better oral bioavailability than the phenolic parent compounds.
  • the compounds of the invention have Hsp90 inhibiting or modulating activity and hence are useful in for use in treating, alleviating or preventing certain cdk5 mediated diseases and conditions.
  • the invention provides the use of a compound of the invention as defined herein for the manufacture of a medicament for the treatment of pain.
  • the invention provides the use of a compound of the invention as defined herein thereof for the manufacture of a medicament for the prophylaxis or treatment of stroke.
  • the invention provides the use of a compound of the invention as defined herein for the manufacture of a medicament for use as a neuroprotective agent.
  • the invention provides:
  • a compound of the invention as defined herein for use in the reduction or elimination of pain in a patient e.g. a mammal such as a human
  • a patient e.g. a mammal such as a human
  • a compound of the invention as defined herein for the manufacture of a medicament for use in the reduction or elimination of pain in a patient (e.g. a mammal such as a human) suffering from pain.
  • a compound of the invention as defined herein for the manufacture of a medicament for the treatment of any one or more of nociception, somatic pain, visceral pain, acute pain, chronic pain, hyperalgesia, allodynia, post operative pain, pain due to hypersensivity, headache, inflammatory pain (rheumatic, dental, dysmenorrhoea or infection), neurological pain, musculoskeletal pain, cancer related pain or vascular pain.
  • a method for the reduction or elimination of pain in a patient (e.g. a mammal such as a human) suffering from pain which method comprises administering to the patient an effective pain-reducing or pain-eliminating amount of a compound of the invention as defined herein.
  • a method for the prophylaxis or treatment of stroke in a patient such as a mammal (e.g. human), which method comprises administering to the patient a therapeutically effective amount of a compound of the invention as defined herein.
  • a compound of the invention as defined herein for use as a neuroprotective agent for use as a neuroprotective agent.
  • a method of preventing or reducing neuronal damage in a patient suffering from stroke comprises administering to the patient an effective neuroprotective amount of a compound of the invention as defined herein.
  • a compound of the invention as defined herein for the manufacture of a medicament for the prevention or reduction of risk of stroke in patients at risk for stroke, for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation.
  • a compound of the invention as defined herein for the prevention or reduction of risk of stroke in patients at risk for stroke for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation.
  • a method for the prevention or reduction of risk of stroke in patients at risk for stroke for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation, which method comprises administering to the patient an effective therapeutic amount of a compound of the invention as defined herein.
  • a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a disease state or condition mediated by cdk5 or p35.
  • ⁇ A method of prophylaxis or treatment of a disease state or condition mediated by cdk ⁇ or p35, said disease state or condition being other than Alzheimer's disease, Huntington's disease or Creuzfeldt-Jacob disease, which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein.
  • ⁇ A method of prophylaxis or treatment of a disease state or condition mediated by cdk5 or p35, said disease state or condition being other than a neurodegenerative disease, which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein.
  • a method of prophylaxis or treatment of a disease state or condition characterised by elevated levels of cdk5 or p35 which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein.
  • a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a neuropathy, such as a peripheral neuropathy, other than Alzheimer's disease, Huntington's disease or Creuzfeldt-Jacob disease.
  • a neuropathy such as a peripheral neuropathy, other than Alzheimer's disease, Huntington's disease or Creuzfeldt-Jacob disease.
  • a method of prophylaxis or treatment of a neuropathy such as a peripheral neuropathy, other than Alzheimer's disease, Huntington's disease or Creuzfeldt-Jacob disease, which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein.
  • Compounds of the present invention and their acid addition salts and crystalline forms thereof have antifungal activity, anti-protozoal activity and anti-parasitic activity.
  • compounds of the invention are useful in treating infection by pathogenic fungi, protozoa and parasites where infection by the pathogen is normally associated with an antibody response to HSP90.
  • the invention provides compounds of the formula (I) and sub-groups thereof as defined herein for use as anti-fungal agents.
  • fungi examples include those that are pathogenic in man and other animals, for example:
  • Candida species such as Candida albicans and Candida tropicalis
  • Cryptococcus species such as Cryptococcus neoformans and Cryptococcal meningitis
  • Aspergillus species such as Aspergillus fumigatus, Aspergillus flavus and Aspergillus niger,
  • Microsporum species such as Microsporum oanis and Microsporum gypseum;
  • Trichophyton species such as Trichophyton equinum, Trichophyton mentagrophytes and Trichophyton rubrum;
  • Fusarium species such as Fusarium solani; ' Sporoth ⁇ x schenckii;
  • Penicillium species such as Penicillium rubrum; ⁇ Altermaria species;
  • Paecilomyces va ⁇ otti ⁇ yeasts, for example Saccharomyces cerevisiae and Pityrosporum species such as Pityrosporum orbiculare and Pityrosporum ovale;
  • Histoplasma species such as Histoplasma capsulatum
  • the invention provides compounds of the formula (!) and sub-groups thereof as defined herein for use as anti-protozoal agents.
  • protozoa examples include:
  • L chagasi the L mexicana complex (3 main species - L. mexicana, L amazonensis, and L. venezuelensis); L tropica; L major, L aethiopica; and the subgenus Viannia with four main species (L. (V.) braziliensis, L (V.) guyanensis, L (V.) panamensis, and L (V.) peruviana); ⁇ Toxoplasma gondii; and
  • the invention provides compounds of the formula (I) and sub-groups thereof as defined herein for use as anti-parasitic agents.
  • parasites examples include parasitic worms such as: ⁇ parasitic roundworms such as Ascaris lumbricoides;
  • parasitic flatworms such as the parasitic trematode worms, e.g. Schistosoma mansoni
  • the invention also provides inter alia:
  • a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a fungal, protozoal or parasitic disease state or condition for example a disease state or condition characterised by an antibody response to Hsp90.
  • a method for the prophylaxis or treatment of a fungal, protozoal or parasitic disease state or condition (other than a disease state or condition due to Plasmodium falciparum), for example a disease state or condition characterised by an antibody response to Hsp90, which method comprises administering to a subject in need thereof a compound of the invention as defined herein.
  • a compound of the invention as defined herein for use in the prophylaxis or treatment of a fungal disease state or condition for example a disease state or condition characterised by an antibody response to Hsp90.
  • a method for the prophylaxis or treatment of a fungal disease state or condition for example a disease state or condition characterised by an antibody response to Hsp90, which method comprises administering to a subject in need thereof a compound of the invention as defined herein.
  • a compound of the invention as defined herein for the manufacture of a medicament for preventing, arresting or reversing the infection of an animal (such as a mammal, e.g. a human) by pathogenic fungi.
  • a method for preventing, arresting or reversing the infection of an animal (such as a mammal, e.g. a human) by pathogenic fungi which method comprises administering to a subject in need thereof a compound of the invention as defined herein.
  • a combination of a compound of the invention as defined herein with an ancilliary compound which is an antifungal agent e.g. an azole antifungal agent.
  • a pharmaceutical composition comprising a compound of the invention as defined herein with an ancilliary compound which is an antifungal agent (e.g. an azole antifungal agent).
  • a method of preventing or reducing development of resistance to an anti-fungal agent in a patient which method comprises administering to the patient a combination of an anti-fungal agent, anti-protozoal agent or anti-parasitic agent (preferably an anti-fungal agent) and a compound of the invention as defined herein.
  • a method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90 which method comprises administering to a subject in need thereof a combination of a compound of the invention as defined herein with an anti-fungal, anti-protozoal or anti-parasitic drug, wherein the disease state or condition mediated by Hsp90 is the development of resistance to the anti-fungal, anti- protozoal or anti-parasitic drug.
  • a method for the treatment of a fungal, protozoal or parasitic disease or condition which method comprises administering to a subject in need thereof a combination of compound of the invention as defined herein with an anti-fungal, anti-protozoal or anti-parasitic drug, which method is characterized by the absence of drug resistance.
  • a method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90 in a subject undergoing treatment with an anti-fungal, anti-protozoal or anti-parasitic drug which method comprises administering to the subject a compound of the invention as defined herein, wherein the disease state or condition mediated by Hsp90 is the development of resistance to said anti-fungal, antiprotozoal or anti-parasitic drug.
  • a method for the treatment of a fungal, protozoal or parasitic disease in a subject undergoing treatment with an anti-fungal, anti-protozoal or anti-parasitic drug comprises administering to a subject in need thereof a compound of the invention as defined herein, which method is characterized by the absence of drug resistance e.g. to said anti-fungal, anti-protozoal or anti-parasitic drug).
  • compounds having Hsp90 inhibitory activity have been found to exhibit potent anti-fungal activity and prevent the development of resistance to anti-fungals and in particular Hsp90 dependent resistance to anti-fungals .
  • inhibition of Hsp90 activity can reduce the development of resistance to commonly used anti-fungal drugs such as the azoles. It is therefore envisaged that the compounds of the invention will be useful in the prophylaxis or treatment of a range of fungal diseases and conditions and will also be useful, when coadminstered with other anti-fungal drugs such as the azoles, in enhancing the activity of the anti-fungal drugs.
  • the antifungal activity of ,the compounds of the present invention may be evaluated by determining the minimum fungistatic (inhibition) concentration (m.i. ⁇ ). This test is usually performed by preparing a series of plates or tubes containing a suitable nutrient medium, each plate or tube also containing a different concentration of the test compound and then inoculating the medium with the fungal species. After an incubation period the plates are examined visually for the presence or absence of fungal growth. The m.i.c. is the minimum concentration required to prevent fungal growth.
  • the compounds may be used in animal medicine (for example in the treatment of mammals such as humans).
  • Subcutaneous mycoses i.e. fungal infections involving the dermis, subcutaneous tissues, muscle, and fascia; ⁇ Systemic mycoses due to primary pathogens (these typically originate primarily in the lungs and may spread to other organ systems); and
  • fungal disease states for which compounds of the invention as defined herein may be used include:
  • Dermatophyte infections such as tinea versiColour (a superficial fungal infection of the skin), tinea pedis (Athletes' Foot), tinea capitis (superficial fungal infection on the head), tinea barbae (fungal infection of bearded areas), tinea corporis (fungal infection of smooth skin areas).
  • Mucosal Candidiasis such as Oral Candidiasis, esophagitis and Vaginal candidiasis.
  • Invasive or deep organ candidiasis e.g., fungemia, endocarditis, and endophthalmitis.
  • Blastomycosis a fungal infection of the lungs and occasionally the skin.
  • ⁇ Aspergilloses such as Allergic Bronchopulmonary Aspergillosis.
  • Compounds of the invention also have anti-protozoal activity and anti-parasitic activity.
  • the antiprotozoal activity of the compounds of the present invention may be assessed by conventional methods, for example by determining the minimum inhibition concentration (m.i.c.) or 50% inhibition level (IC 50 ). Examples of protozoal and parasitic diseases or conditions for which compounds of the invention may prove useful include:
  • Chagas disease (trypanosomiasis) - an infection caused by the parasite Trypanosoma cruzi. * Ascariasis - a human disease caused by the parasitic roundworm Ascaris lumbricoides.
  • Leishmaniasis a disease caused by parasites of the genus Leishmania.
  • Toxoplasmosis a parasitic disease caused by the protozoan Toxoplasma gondii.
  • Schistosomiasis (Bilharzia) - a disease caused by the parasite Schistoma mansoni.
  • Trichomoniasis a sexually transmitted disease caused by the parasitic protozoan Trichomonas vaginalis.
  • HSP 90 inhibitors can block viral replication. Accordingly, the compounds of the invention are useful in combatting viral infections, for example by blocking or inhibiting viral replication.
  • the invention provides a compound of the invention as defined herein for use in the prophylaxis or treatment of a viral infection (or viral disease).
  • the invention provides:
  • a compound of the invention as defined herein for use in blocking or inhibiting viral replication in a host organism e.g. an animal such as a mammal (e.g. human)
  • a host organism e.g. an animal such as a mammal (e.g. human)
  • a compound of the invention as defined herein for the manufacture of a medicament for use in blocking or inhibiting viral replication in a host organism (e.g. an animal such as a mammal (e.g. human)).
  • a method of blocking or inhibiting viral replication in a host organism e.g. an animal such as a mammal (e.g. human)
  • which method comprises administering to the host organism a compound of the invention as defined herein.
  • viral infections that may be treated with the compounds of the invention include infections due to any one or more of the following viruses: o Picomaviruses such as rhinoviruses (common cold virus), Coxsackie virus (e.g. Coxsackie
  • Hepatitis viruses such as hepatitis A virus (HAV), hepatitis B virus (HBV), hepatitis C virus
  • HCV hepatitis D virus
  • HAV hepatitis E virus
  • Coronaviruses e.g. common cold virus and Severe acute respiratory syndrome (SARS) virus
  • Adenoviruses such as Human Adenoviruses (a cause of respiratory and conjunctival infections)
  • Astroviruses a cause of flu-like symptoms
  • Flaviviruses such as the Yellow Fever virus
  • Orthomyxoviruses such as influenza viruses (e.g.
  • influenza A, B and C viruses o Parainfluenza viruses; o Respiratory syncytial virus; o Enteroviruses such as Poliovirus (Poliomyelitis virus); o Paramyxoviruses such as the Measles (rubeola) virus, mumps virus, respiratory syncytial virus (RSV) and canine distemper virus (CDV); o Togaviruses such as the Rubella (German Measles) virus and Sindbis virus; o Herpes viruses such as:
  • Herpes simplex virus for example HSV-1 which causes fever blisters (cold sores), gingivostomatitis, herpes keratitis, eczema herpeticum and HSV encephalitis); and HSV-2 which causes genital lesions, neonatal infections, HSV meningitis, HSV proctitis; » Varicella zoster virus (VZV), which causes chickenpox, congenital varicella syndrome and shingles; ⁇ Epstein-Barr Virus (EBV), which causes infectious mononucleosis, Burkitt's lymphoma and nasopharyngeal cancer;
  • HSV Herpes simplex virus
  • HSV-1 which causes fever blisters (cold sores), gingivostomatitis, herpes keratitis, eczema herpeticum and HSV encephalitis
  • HSV-2 which causes genital lesions, neonatal infections, HSV meningitis,
  • CMV Cytomegalovirus
  • HCMV human cytomegalovirus
  • HHV-6 Human herpes virus 6
  • HHV-8 Human herpes virus 8
  • KSHV Human immunodefficiency virus
  • HPV human papilloma virus
  • HTLV Human T-lymphotrophic virus
  • Particular viral infections against which the compounds of the invention may be used include herpes virus, pox virus, Epstein-Barr virus, Sindbis virus, adenovirus, HIV (for prevention of AIDS development in HIV-infected individuals), HPV, HCV and HCMV viruses.
  • the viral infection may be other than an infection with hepatitis C virus (HCV).
  • HCV hepatitis C virus
  • the activity of the compounds of the invention as agents for blocking or preventing viral replication in host organisms or host cells can be determined in accordance with standard procedures well known to the skilled person.
  • the compounds of the invention may be used as the sole antiviral agent or they may be used in conjunction with other anti-viral agents such as acyclovir, ganciclovir, oseltamavir (Tamiflu®) and zanamavir (Relenza®), amantidine, rimantadine, adefovir dipivoxil, interferons (e.g.
  • interferon alfa- 2b and pegylated interferon alfa-2a lamivudine, entecavir, ribavirin, famciclovir, valcicylovir, valacyclovir, azidothymidine (AZT - Retrovir®) , atazanavir, fosamprenavir, lamivudine, lamivudine + abacavir, tenofovir disoproxil fumarate, tenofovir disoproxil fumarate + emtricitabine, tipranavir, nelfinavir, indinavir, raltegravir, ritonavir, lopinavir + ritonavir, darunavir, amprenavir, enfuvirtide, saquinavir, hydroxyurea, VGV-1 and anti-viral vaccines.
  • the invention further provides:
  • a pharmaceutical composition comprising a compound of the invention as defined herein with an ancilliary compound which is an antiviral agent.
  • references to Formula (I) also include Formula (I 0 ), Formula (Vl 0 ) and all sub-groups (e.g. formula (Vl) and examples of Formula (I) as defined herein.
  • Formula (I 0 ) also include Formula (I 0 ), Formula (Vl 0 ) and all sub-groups (e.g. formula (Vl) and examples of Formula (I) as defined herein.
  • R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 10 or any other "R" group the definition of the group in question is as set out above and as set out in the following sections of this application unless the context requires otherwise.
  • R 1 is hydrogen or hydroxy
  • R 2 is hydrogen, hydroxy or methoxy; provided that at least one of R 1' and R 2' is hydroxy.
  • the compounds of formula (I) of the present invention can be prepared by the reaction of a compound of the formula (IX) with a compound of the formula R Z -LG, where LG is a leaving group, in the presence of a base, and typically with heating.
  • the leaving group will depend on the nature of the group L p .
  • the reaction is typically carried out with heating, for example to a temperature in excess of 100 0 C, e.g. a temperature of approximately 120 0 C.
  • Compounds of the formula (I) wherein L p is a bond can be prepared by standard and well known methods of ether synthesis.
  • the compound of formula (IX) can be reacted with an alkali metal hydride such as sodium hydride in a polar non-protic solvent to form a phenolate anion which is then reacted with a compound of the formula R p1 -LG where LG is a halide leaving group such as chlorine or bromine to give the compound of formula (I).
  • the compound of the formula (IX) can be reacted with a compound of the formula R p1 -OH under Mitsunobu alkylation conditions, e.g.
  • the Mitsunobu reaction is typically carried out in a polar non-protic solvent such as tetrahydrofuran at a moderate temperature such as ambient temperature.
  • the reaction is typically carried out in an inert polar solvent such as N,N-dimethylformamide, N 1 N- dimethylacetamide or N-methylpyrrolidinone either at room temperature or at an elevated temperature such as 100 0 C or above, see for example the methods described by Leese et al [Bioorg. Med. Chem. Lett, 2004, 14, 3135], Okada et al [Tetrahedron Lett, 2000, 41 , 7047], Patel et al [Bioorg. Med. Chem. Lett, 2004, 14, 605] or Chu et al. [Bioorg. Med. Chem. Lett, 1999, 9, 141].
  • an inert polar solvent such as N,N-dimethylformamide, N 1 N- dimethylacetamide or N-methylpyrrolidinone
  • LG is a leaving group such as a halogen (e.g. chlorine or bromine) or an alkyl carbonate (e.g. fe/t-
  • the reaction is typically carried out in an inert polar solvent such as dichloromethane, ethyl acetate, tetrahydrofuran or N,N-dimethylformamide, either at room temperature or at an elevated temperature such as 100 0 C or above, see for example the methods described by Yajima et al [Chem. Pharm. Bull., 1970, 18, 2574], Ramesh et al [J. Org. Chem., 2005, 70, 837], Wu et al [Synth. Commun., 2002, 32, 1615] or Bhat et a ⁇ [Tetrahedron Lett, 2002, 43, 2467].
  • an inert polar solvent such as dichloromethane, ethyl acetate, tetrahydrofuran or N,N-dimethylformamide
  • a non-interfering base such as pyridine or triethylamine.
  • Compounds of the formula (I) wherein R z is a glucuronide residue can be prepared by synthetic chemical methods well known the the skilled person or by bio-transformation process, for example by incubating the compound of the formula (IX) with animal microsomes, such as dog microsomes or rat microsomes. By using microsomes from different animal species, different glucuronide isomers or mixtures of isomers can be obtained as described in the examples below.
  • Compounds of the formula (I) wherein R z is a mono-, di- or tripeptide residue can be prepared by the reaction of a compound of the formula (IX) with a suitably protected acid chloride of an amino acid, dipeptide or tripeptide in accordance with known methods, followed by removal of any protecting groups.
  • the reaction is typically carried out in a polar solvent in the presence of a non- interfering base such as triethylamine or pyridine; see for example the method described by Brenner et al, HeIv. Chim. Acta, 1957, 40, 1933. Examples of deprotection of the protecting groups can be found in Green et al (see below).
  • the alcohol (IX) can be converted to the ester (I) by reaction of a suitably protected carboxylic acid of an amino acid, dipeptide or tripeptide in the presence of ester coupling reagents, followed by removal of any protecting groups.
  • ester coupling reagents include 1 ,3-dicyclohexylcarbodiimide (DCC) (Johnson et al, JCS Chem. Comm., 1968, 73), DCC and 1-hydroxybenzotriazole (HOBt) (Kessler et al, Tet.
  • amines of the formula HNR 5 R 6 are either commercially available or can be made using methods well known to the skilled person, see for example, Advanced Organic Chemistry, by Jerry March, 4 th edition, 119, Wiley Interscience, New York; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN: 0-471-31192-8).
  • the carboxylic acid (X) can be converted to an amide of the formula (IX) by first forming an acid chloride by treatment of the carboxylic acid with thionyl chloride, or by reaction with oxalyl chloride in the presence of a catalytic amount of dimethyl formamide, or by reaction of a potassium salt of the acid with oxalyl chloride.
  • the acid chloride can then be reacted with the amine HNR 5 R 6 in the presence of a non-interfering base such as triethylamine.
  • the reaction may be carried out at around room temperature in a polar solvent such as dioxan.
  • the carboxylic acid (X) can be converted to the amide (IX) by reaction with the amine HNR 5 R 6 in the presence of amide coupling reagents of the type commonly used in the formation of amide or peptide linkages.
  • amide coupling reagents include 1 ,1 '-carbonyldiimidazole (CDI), I.S-dicyclohexylcarbodiimide (DCC) (Sheehan et al, J. Amer. Chem Soc.
  • uronium-based coupling agents such as O-(7-azabenzotriazol-1-yl)- ⁇ / J ⁇ /, ⁇ /', ⁇ /'-tetramethyluronium hexafluorophosphate (HATU) and phosphonium-based coupling agents such as 1-benzo-triazolyloxytris-(pyrrolidino)phosphonium hexafluorophosphate (PyBOP) (Castro et al, Tetrahedron Letters, 1990, 31, 205).
  • Carbodiimide- based coupling agents are advantageously used in combination with 1-hydroxy-7-azabenzotriazole (HOAt) (L. A. Carpino, J. Amer.
  • Preferred coupling reagents include EDC (EDAC) and DCC in combination with HOAt or HOBt.
  • One particular coupling reagent comprises EDC in combination with HOBt.
  • a preferred coupling agent is 1 ,1 '-carbonyldiimidazole (CDI).
  • the coupling reaction is typically carried out in a non-aqueous, non-protic solvent such as acetonitrile, dioxan, dimethylsulphoxide, dichloromethane, dimethylformamide or N- methylpyrrolidine, or in an aqueous solvent optionally together with one or more miscible co- solvents.
  • a non-aqueous, non-protic solvent such as acetonitrile, dioxan, dimethylsulphoxide, dichloromethane, dimethylformamide or N- methylpyrrolidine
  • an aqueous solvent optionally together with one or more miscible co- solvents.
  • the reaction can be carried out at room temperature or, where the reactants are less reactive (for example in the case of electron-poor anilines bearing electron withdrawing groups such as sulphonamide groups) at an appropriately elevated temperature.
  • the reaction may be carried out in the presence of a non-interfering base, for example a tertiary amine such as triethyl
  • Compounds of formula (IX) can also be made according to the method shown in Scheme 2.
  • the starting material for the synthetic route shown in Scheme 2 is 4-ethyl anisole, which can be obtained commercially.
  • Conversion to the carboxylic acid can be carried out by lithiation at low temperature, followed by quenching of the resulting anion with solid carbon dioxide.
  • the carboxylic acid may be coupled with various amines, using standard amide coupling reagents of the type commonly used in the formation of peptide linkages as described above.
  • Deprotection of the methyl ether can be effected using using boron tribromide (e.g. by the method described in Synthesis 1991 , 469) to give the compound of formula (IX).
  • the method illustrated in Scheme 2 can be used to prepare other simple 2-hydroxy-5-substituted benzoic acids which can then be coupled to an appropriate amine to give the compounds of formula (IX).
  • the process of coupling intermediates acids with amines, anilines or amino-heterocyclic compounds, followed by removal of any protecting groups, is straightforward and is suitable for the synthesis of large combinatorial libraries of molecules, useful for this invention. Examples of combinatorial libraries are described in Solid-Phase Synthesis and Combinatorial Technologies by Pierfausto Seneci. Wiley-lnterscience, New York. 2000. xii + 637 pp. ISBN 0471331953).
  • Friedel-Crafts acylation of a phenol is carried out by treatment of the phenol with an acylating agent (such as an acid chloride or acid anhydride) in the presence of a Lewis acid catalyst (such as boron trifluoride or aluminium chloride) either at room temperature or at more elevated temperatures (60-120 0 C).
  • an acylating agent such as an acid chloride or acid anhydride
  • a Lewis acid catalyst such as boron trifluoride or aluminium chloride
  • the 4- hydroxy group of the 2,4-dihydroxybenzoic acid methyl ester can be O-acylated using an acid anhydride such as acetic anhydride in the presence of a base such as 4-dimethylaminopyridine (4- DMAP) to give a ' 2-hydroxy-4-acyloxy-benzoic acid methyl ester.
  • an acid anhydride such as acetic anhydride
  • 4-dimethylaminopyridine (4- DMAP) 4-dimethylaminopyridine
  • Treatment of the 2-hydroxy-4- acyloxy-benzoic acid methyl ester with trifluoromethanesulphonic acid in the presence of an acyl chloride such as acetyl chloride then gives the 2,4-dihydroxy-5-acylbenzoic acid methyl ester.
  • the Wittig reaction can be carried out in an inert polar solvent (such as tetrahydrofuran) and can involve treatment of an aldehyde or ketone with a phosphorus ylide species that may be prepared by the reaction of a phosphonium salt with a base (such as butyl lithium or potassium tert- butoxide).
  • a base such as butyl lithium or potassium tert- butoxide.
  • the ester hydrolysis to the carboxylic acid is usually carried out by treatment with an aqueous alkali metal hydroxide such sodium hydroxide.
  • the saponification reaction may be carried out using an organic co-solvent such as an alcohol (e.g. methanol) and the reaction mixture is typically heated to a non-extreme temperature, for example up to about 50-60 0 C.
  • the intermediate compound 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (XXVII) in Scheme 4 can be made using a variety of methods well known to the skilled person.
  • compound (XXVII) can be made by the synthetic route illustrated in Scheme 4A.
  • the palladium compound can be a palladium (0) compound such as Pd(PPh 3 ) 4 or a palladium (II) compound such as [1 ,1'-bis(diphenyl ⁇ phosphino)ferrocene]- dichloropalladium(ll).
  • the base can be an organic base such as n-butylamine or an inorganic base such as a metal carbonate, e.g. caesium carbonate.
  • the reaction with potassium isoprenyl trifluoroborate is typically carried out at reflux temperature for a prolonged period, for example 15 hours or more.
  • the resulting isopropenyl bis-benzyloxy ester (XXXI) is then hydrolysed to give the carboxylic acid (XXVII) using, for example, an alkali metal hydroxide such as lithium hydroxide, typically with heating to a non-extreme temperature.
  • an alkali metal hydroxide such as lithium hydroxide
  • A- Hydroxy-3-(1',2'-dimethyl-propyl)-benzoic acid amides can be prepared by amide coupling using standard coupling agents (as outlined above) from the alkyl substituted acid.
  • the olefinic acid itself can be prepared by Claisen rearrangement of a precursor ether, as shown in the scheme, by thermal rearrangement in anisole, followed by saponification, which in this case can yield more than one isomer of the olefin, the major one being shown in the scheme.
  • Claisen reactions are well known in the literature, e.g. see J. Chem. Soc, Perkin Trans 1 1981 , 897.
  • the ether itself can be prepared by simple alkylation of commercially available 4-hydroxy benzoic acid ethyl ester.
  • the alkylation and saponification reactions are simple modifications that can be carried out under various conditions (for example see, Advanced Organic Chemistry, by Jerry March, 4 th edition, 119, Wiley Interscience, New York; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN; 0-471-31192-8)). It is to be understood that other 4- hydroxy-3-substituted benzoic acids could be made using this procedure to synthesise different examples of compounds of formula 1 not specifically exemplified herein.
  • the reaction may be carried out in an aqueous solvent system, for example aqueous ethanol, and the reaction mixture is typically subjected to heating, for example to a temperature in excess of 100 0 C
  • aqueous solvent system for example aqueous ethanol
  • the reaction mixture is typically subjected to heating, for example to a temperature in excess of 100 0 C
  • boronates suitable for use in preparing compounds of the invention are commercially available, for example from Boron Molecular Limited of Noble Park, Australia, or from Combi-Blocks Inc, of San Diego, USA. Where the boronates are not commercially available, they can be prepared by methods known in the art, for example as described in the review article by N. Miyaura and A. Suzuki, Chem. Rev. 1995, 95, 2457.
  • boronates can be prepared by reacting the corresponding bromo-compound with an alkyl lithium such as butyl lithium and then reacting with a borate ester.
  • the resulting boronate ester derivative can, if desired, be hydrolysed to give the corresponding boronic acid.
  • the final products of the reaction sequence illustrated in Scheme 6 are formed by catalytic hydrogenation (as outlined above) to remove the benzyl protecting groups and to reduce the olefin, formed in the Suzuki reaction to the alkyl substituent. It is to be understood that other 2,4-dihydroxy-5-substituted benzoic acids could be made using this procedure to synthesise different examples of compounds of formula I not specifically exemplified herein.
  • an optionally substituted 1 ,2-dimethylbenzene (Xl) is heated with N- bromosuccinimide in the presence of dibenzoyl peroxide to give the dibromo-compound (XII).
  • the reaction is typically carried out in carbon tetrachloride with heating at reflux.
  • the dibromo- compound (XlI) is then reacted with a compound PG-NH 2 where PG is a protecting group such as tosyl or para-methoxybenzyl in the presence of a base such as a metal hydride (e.g. sodium hydride), when PG is a tosyl group, or an alkali metal carbonate (e.g.
  • PG para-methoxybenzyl
  • the protecting group PG can then be removed to give the amine (XIV).
  • a tosyl group can be removed by heating with a mxture of phenol, hydrobromic acid and propanoic acid, whereas a para-methoxybenzyl group can be removed in standard manner using trifluoroacetic acid and anisole.
  • the amine (XIV) is then coupled with a carboxylic acid of the formula (X) as described above.
  • one or more functional groups R 10b present in the protected isoindoline (XIII) or the deprotected isoindoline compound (XIV) can be converted into other groups R 1Ob .
  • the group R 10b in compound (XIV) is a nitro group, it can be reduced to give the corresponding amino group, for example by catalytic hydrogenation in the presence of a palladium on charcoal catalyst.
  • R 1Ob in the compound (XIII) is an ester group (e.g. CO 2 Me), it can be hydrolysed to give a carboxylic acid which can then be reacted with an amine such as morpholine to give the corresponding amide.
  • Further functional group interconversions may subsequently be carried out (for example reduction of the amide to the corresponding aminomethyl compound with lithium aluminium hydride) before removal of the protecting group PG.
  • the ester group can be reduced to a hydroxymethyl group which may then be converted to an aldehyde, and the aldehyde group may subsequently be used as a substrate for further conversions, for example a reductive amination reaction with an amine such as morpholine, piperidine or N-methyl piperazine.
  • Debenzylation of the hydroxymethyldihydroisoindole intermediate (LIII) is then carried out by hydrogenation over palladium on charcoal catalyst in an alcohol (e.g. ethanol) solvent at a mildly elevated temperature (e.g. up to about 50 °) to give the intermediate (LIV).
  • Intermediate (LIV) is then converted to intermediate (LV) by reaction with a reagent suitable for introducing a benzyloxycarbonyl ("Z") group onto the nitrogen atom of the dihydroisoindole ring.
  • the intermediate (LIV) can be reacted with benzyl chloroformate in a polar non-protic solvent such as THF in the presence of a non-interfering base such as triethylamine to give intermediate (LV).
  • the starting material for Scheme 8 is the ortho diester (XV) which is hydrolysed to the corresponding dicarboxylic acid (XVI) using an alkali metal hydroxide such as potassium hydroxide before being subjected to cyclisation to the phthalic anhydride (XVII) by reaction with acetic anhydride.
  • the phthalic anhydride (XVII) can be converted to the corresponding phthalimide (XVIII) by reaction with formamide at an elevated temperature (e.g. approximately 210 0 C).
  • the phthalimide (XVIiI) can then be reduced to the isoindoline (XIV) using a suitable reducing agent such as borane in tetrahydrofuran.
  • Isoindoline intermediate compounds can also be prepared be prepared by the synthetic route shown in Scheme 9.
  • the mesyl compound (LXI) is reacted with alkylpiperazine (LXII) in acetone solution to give the Z-protected isoindoline (LXIII). Removal of the benzyloxycarbonyl group to give the unprotected isoindoline compound (LXIV) is then accomplished by hydrogenation over a palladium on charcoal catalyst.
  • n, R 3 , R 4a , R 8 and R 1Occ are as defined herein, under amide forming conditions as described above and in the examples.
  • n is 0 or 1 ; M is N or CHOH and R is hydrogen or methyl; provided that when n is 0 and R 25 is methyl, then M is CHOH.
  • intermediates of formula (XXI) can be made by methods well known to the skilled person or methods analogous thereto.
  • intermediate XXII can be prepared by lithium-halogen exchange of a suitably N-protected 5-bromoisoindoline, quenching with 1-methyl-4-piperidone and subsequent deprotection.
  • Intermediate XXII can be prepared by Buchwald palladium coupling of 4-BOC-piperazine and a suitably N-protected 5-bromoisoindoline followed by subsequent deprotection.
  • One method of preparation for intermediate XXIV is from a suitably N-protected isoindoline-5-carboxylic acid, Weinreb amide formation, reduction to the aldehyde, followed by reductive amination and subsequent deprotection.
  • the above synthetic routes involve the formation of a phenolic compound of the formula (IX) followed by the subsequent reaction of the phenolic group in the compound of formula (IX) with a reagent suitable for introducing the group R z .
  • the group R z can be introduced into the benzoic acid of the formula (X), or a protected derivative thereof, to give a compound of the formula (XXV) or a protected derivative thereof:
  • R 1 and R 2 are as defined herein.
  • the reactions used to introduce the group R z may be analogous to the reactions described above for introducing the group R z into compounds of the formula (IX), except that the carboxylic acid group may require protection (e.g. as an ester) under some of the reaction conditions, and then subsequent deprotection.
  • protection e.g. as an ester
  • one compound of the formula (I), or a protected form thereof can be converted into another compound of the formula (I).
  • R 3 is bromine
  • the bromine atom can be replaced by trifluoromethyl by reaction with a trifluoroacetate salt (e.g. sodium trifluoroacetate), and copper (I) iodide in a polar solvent such as dimethylformamide.
  • a trifluoroacetate salt e.g. sodium trifluoroacetate
  • copper (I) iodide in a polar solvent such as dimethylformamide.
  • compounds of the formula (I) wherein R 8 is fluorine can be prepared from compounds of the formula (I) where R 8 is hydrogen by electrophilic fluorination.
  • Electrophilic fluorination can be carried out using a fluorinating agent such as 1-(chloromethyl)-4-fluoro-1 ,4- diazoniabicyclo[2.2.2]octane bis(tetrafluoroborate) or similar N-fluoro-diazonia compounds.
  • the hydroxy group is a phenolic hydroxy group, for example in compounds of the formula (I) wherein R 1 and/or R 2 are hydroxy, a preferred protecting group is a benzyl group.
  • the aldehyde or ketone group is readily regenerated by hydrolysis using a large excess of water in the presence of acid.
  • An amine group may be protected, for example, as an amide (-NRCO-R) or a urethane (-NRCO-OR), for example, as: a methyl amide (-NHCO-CH 3 ); a benzyloxy amide (-NHCO-OCH 2 C 6 H 5 , -NH-Cbz); as a t-butoxy amide (-NHCO-OC(CH 3 ) 3 , -NH-Boc); a 2-biphenyl-2-propoxy amide (-NHCO-OC(CH 3 ) 2 C S H 4 C 6 H 5 , -NH-Bpoc), as a 9-fluorenylmethoxy amide (-NH-Fmoc), as a 6-nitroveratryloxy amide (-NH-Nvoc), as a 2-trimethylsilylethyloxy amide (-NH-Teoc), as a 2,2,2-trichloroethyloxy amide (-NH-Troc), as an
  • protecting groups for amines such as cyclic amines and heterocyclic N-H groups, include toluenesulphonyl (tosyl) and methanesulphonyl (mesyl) groups and benzyl groups such as a para- methoxybenzyl (PMB) group.
  • tosyl toluenesulphonyl
  • methanesulphonyl meyl
  • benzyl groups such as a para- methoxybenzyl (PMB) group.
  • a carboxylic acid group may be protected as an ester for example, as: an C 1-7 alkyl ester (e.g., a methyl ester; a t-butyl ester); a C 1-7 haloalkyl ester (e.g., a Ci -7 trihaloalkyl ester); a triC 1-7 alkylsilyl-C 1-7 alkyl ester; or a C 5 . 2 o aryl-C 1-7 alkyl ester (e.g., a benzyl ester; a nitrobenzyl ester); or as an amide, for example, as a methyl amide.
  • an C 1-7 alkyl ester e.g., a methyl ester; a t-butyl ester
  • a C 1-7 haloalkyl ester e.g., a Ci -7 trihaloalkyl ester
  • the compounds may be isolated and purified by a number of methods well known to those skilled in the art and examples of such methods include chromatographic techniques such as column chromatography (e.g. flash chromatography) and HPLC.
  • Preparative LC-MS is a standard and effective method used for the purification of small organic molecules such as the compounds described herein.
  • the methods for the liquid chromatography (LC) and mass spectrometry (MS) can be varied to provide better separation of the crude materials and improved detection of the samples by MS.
  • Optimisation of the preparative gradient LC method will involve varying columns, volatile eluents and modifiers, and gradients. Methods are well known in the art for optimising preparative LC-MS methods and then using them to purify compounds.
  • the active compound While it is possible for the active compound to be administered alone, it is preferable to present it as a pharmaceutical composition (e.g. formulation) comprising at least one active compound of the invention together with one or more pharmaceutically acceptable carriers, adjuvants, excipients, diluents, fillers, buffers, stabilisers, preservatives, lubricants, or other materials well known to those skilled in the art and optionally other therapeutic or prophylactic agents; for example agents that reduce or alleviate some of the side effects associated with chemotherapy.
  • a pharmaceutical composition e.g. formulation
  • a pharmaceutical composition comprising at least one active compound of the invention together with one or more pharmaceutically acceptable carriers, adjuvants, excipients, diluents, fillers, buffers, stabilisers, preservatives, lubricants, or other materials well known to those skilled in the art and optionally other therapeutic or prophylactic agents; for example agents that reduce or alleviate some of the side effects associated with chemotherapy.
  • agents include anti-emetic agents and agents that prevent or decrease the duration of chemotherapy-associated neutropenia and prevent complications that arise from reduced levels of red blood cells or white blood cells, for example erythropoietin (EPO), granulocyte macrophage- colony stimulating factor (GM-CSF), and granulocyte-colony stimulating factor (G-CSF).
  • EPO erythropoietin
  • GM-CSF granulocyte macrophage- colony stimulating factor
  • G-CSF granulocyte-colony stimulating factor
  • the present invention further provides pharmaceutical compositions, as defined above, and methods of making a pharmaceutical composition
  • a pharmaceutical composition comprising admixing at least one active compound, as defined above, together with one or more pharmaceutically acceptable carriers, excipients, buffers, adjuvants, stabilizers, or other materials, as described herein.
  • pharmaceutically acceptable as used herein pertains to compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of a subject (e.g. human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
  • Each carrier, excipient, etc. must also be “acceptable” in the sense of being compatible with the other ingredients of the formulation.
  • the invention provides compounds of the formula (I) and subgroups thereof as defined herein in the form of pharmaceutical compositions.
  • compositions can be in any form suitable for oral, parenteral, topical, intranasal, ophthalmic, otic, rectal, intra-vaginal, or transdermal administration.
  • compositions are intended for parenteral administration, they can be formulated for intravenous, intramuscular, intraperitoneal, subcutaneous administration or for direct delivery into a target organ or tissue by injection, infusion or other means of delivery.
  • the delivery can be by bolus injection, short term infusion or longer term infusion and can be via passive delivery or through the utilisation of a suitable infusion pump.
  • compositions adapted for parenteral administration include aqueous and nonaqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats, co- solvents, organic solvent mixtures, cyclodextrin complexation agents, emulsifying agents (for forming and stabilizing emulsion formulations), liposome components for forming liposomes, gellable polymers for forming polymeric gels, lyophilisation protectants and combinations of agents for, inter alia, stabilising the active ingredient in a soluble form and rendering the formulation isotonic with the blood of the intended recipient.
  • aqueous and nonaqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats, co- solvents, organic solvent mixtures, cyclodextrin complexation agents, emulsifying agents (for forming and stabilizing emulsion formulations), liposome components for forming liposomes, gellable polymers for forming polymeric gels, lyophilisation protect
  • compositions for parenteral administration may also take the form of aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents (R. G. Strickly, Solubilizing Excipients in oral and injectable formulations, Pharmaceutical Research, VoI 21(2) 2004, p 201-230).
  • a drug molecule that is ionizable can be solubilized to the desired concentration by pH adjustment if the drug's pK a is sufficiently away from the formulation pH value.
  • the acceptable range is pH 2-12 for intravenous and intramuscular administration, but subcutaneously the range is pH 2.7-9.0.
  • the solution pH is controlled by either the salt form of the drug, strong acids/bases such as hydrochloric acid or sodium hydroxide, or by solutions of buffers which include but are not limited to buffering solutions formed from glycine, citrate, acetate, maleate, succinate, histidine, phosphate, tris(hydroxymethyl)- aminomethane (TRIS), or carbonate.
  • the combination of an aqueous solution and a water-soluble organic solvent/surfactant is often used in injectable formulations.
  • the water-soluble organic solvents and surfactants used in injectable formulations include but are not limited to propylene glycol, ethanol, polyethylene glycol 300, polyethylene glycol 400, glycerin, dimethylacetamide (DMA), N-methyl-2-pyrrolidone (NMP; Pharmasolve), dimethylsulphoxide (DMSO), Solutol HS 15, Cremophor EL, Cremophor RH 60, and polysorbate 80.
  • Such formulations can usually be, but are not always, diluted prior to injection.
  • Propylene glycol, PEG 300, ethanol, Cremophor EL, Cremophor RH 60, and polysorbate 80 are the entirely organic water-miscible solvents and surfactants used in commercially available injectable formulations and can be used in combinations with each other.
  • the resulting organic formulations are usually diluted at least 2-fold prior to IV bolus or IV infusion. Alternatively increased water solubility can be achieved through molecular complexation with cyclodextrins.
  • Liposomes are closed spherical vesicles composed of outer lipid bilayer membranes and an inner aqueous core and with an overall diameter of ⁇ 100 ⁇ m.
  • moderately hydrophobic drugs can be solubilized by liposomes if the drug becomes encapsulated or intercalated within the liposome.
  • Hydrophobic drugs can also be solubilized by liposomes if the drug molecule becomes an integral part of the lipid bilayer membrane, and in this case, the hydrophobic drug is dissolved in the lipid portion of the lipid bilayer.
  • a typical liposome formulation contains water with phospholipid at 5-20 mg/ml, an isotonicifier, a pH 5-8 buffer, and optionally cholesterol.
  • the formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use.
  • the pharmaceutical formulation can be prepared by lyophilising a compound of Formula (I) or acid addition salt thereof. Lyophilisation refers to the procedure of freeze-drying a composition. Freeze-drying and lyophilisation are therefore used herein as synonyms. A typical process is to solubilise the compound and the resulting formulation is clarified, sterile filtered and aseptically transferred to containers appropriate for lyophilisation (e.g. vials).
  • vials they are partially stoppered with lyo-stoppers.
  • the formulation can be cooled to freezing and subjected to lyophilisation under standard conditions and then hermetically capped forming a stable, dry lyophile formulation.
  • the composition will typically have a low residual water content, e.g. less than 5% e.g. less than 1 % by weight based on weight of the lyophile.
  • the lyophilisation formulation may contain other excipients for example, thickening agents, dispersing agents, buffers, antioxidants, preservatives, and tonicity adjusters.
  • Typical buffers include phosphate, acetate, citrate and glycine.
  • antioxidants include ascorbic acid, sodium bisulphite, sodium metabisulphite, monothioglycerol, thiourea, butylated hydroxytoluene, butylated hydroxyl anisole, and ethylenediaminetetraacetic acid salts.
  • Preservatives may include benzoic acid and its salts, sorbic acid and its salts, alkyl esters of para-hydroxybenzoic acid, phenol, chlorobutanol, benzyl alcohol, thimerosal, benzalkonium chloride and cetylpyridinium chloride.
  • the buffers mentioned previously, as well as dextrose and sodium chloride, can be used for tonicity adjustment if necessary.
  • Bulking agents are generally used in lyophilisation technology for facilitating the process and/or providing bulk and/or mechanical integrity to the lyophilized cake.
  • Bulking agent means a freely water soluble, solid particulate diluent that when co-lyophilised with the compound or salt thereof, provides a physically stable lyophilized cake, a more optimal freeze-drying process and rapid and complete reconstitution.
  • the bulking agent may also be utilised to make the solution isotonic.
  • the water-soluble bulking agent can be any of the pharmaceutically acceptable inert solid materials typically used for lyophilisation.
  • Such bulking agents include, for example, sugars such as glucose, maltose, sucrose, and lactose; polyalcohols such as sorbitol or mannitol; amino acids such as glycine; polymers such as polyvinylpyrrolidine; and polysaccharides such as dextran.
  • the ratio of the weight of the bulking agent to the weight of active compound is typically within the range from about 1 to about 5, for example of about 1 to about 3, e.g. in the range of about 1 to 2.
  • dosage forms may be via filtration or by autoclaving of the vials and their contents at appropriate stages of the formulation process.
  • the supplied formulation may require further dilution or preparation before delivery for example dilution into suitable sterile infusion packs.
  • Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets.
  • the pharmaceutical composition is in a form suitable for i.v. administration, for example by injection or infusion.
  • the pharmaceutical composition is in a form suitable for subcutaneous (s.c.) administration.
  • Pharmaceutical dosage forms suitable for oral administration include tablets, capsules, caplets, pills, lozenges, syrups, solutions, powders, granules, elixirs and suspensions, sublingual tablets, wafers or patches and buccal patches.
  • compositions containing compounds of the formula (I) can be formulated in accordance with known techniques, see for example, Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, PA, USA.
  • tablet compositions can contain a unit dosage of active compound together with an inert diluent or carrier such as a sugar or sugar alcohol, eg; lactose, sucrose, sorbitol or mannitol; and/or a non-sugar derived diluent such as sodium carbonate, calcium phosphate, calcium carbonate, or a cellulose or derivative thereof such as methyl cellulose, ethyl cellulose, hydroxypropyl methyl cellulose, and starches such as corn starch. Tablets may also contain such standard ingredients as binding and granulating agents such as polyvinylpyrrolidone, disintegrants (e.g.
  • swellable crosslinked polymers such as crosslinked carboxymethylcellulose
  • lubricating agents e.g. stearates
  • preservatives e.g. parabens
  • antioxidants e.g. BHT
  • buffering agents for example phosphate or citrate buffers
  • effervescent agents such as citrate/bicarbonate mixtures.
  • Capsule formulations may be of the hard gelatin or soft gelatin variety and can contain the active component in solid, semi-solid, or liquid form.
  • Gelatin capsules can be formed from animal gelatin or synthetic or plant derived equivalents thereof.
  • the solid dosage forms can be coated or un-coated, but typically have a coating, for example a protective film coating (e.g. a wax or varnish) or a release controlling coating.
  • the coating e.g. a Eudragit TM type polymer
  • the coating can be designed to release the active component at a desired location within the gastro-intestinal tract.
  • the coating can be selected so as to degrade under certain pH conditions within the gastrointestinal tract, thereby selectively release the compound in the stomach or in the ileum or duodenum.
  • the coating can be used as a taste masking agent to mask unpleasant tastes such as bitter tasting drugs.
  • the coating may contain sugar or other agents that assist in masking unpleasant tastes.
  • the drug can be presented in a solid matrix comprising a release controlling agent, for example a release delaying agent which may be adapted to selectively release the compound under conditions of varying acidity or alkalinity in the gastrointestinal tract.
  • a release controlling agent for example a release delaying agent which may be adapted to selectively release the compound under conditions of varying acidity or alkalinity in the gastrointestinal tract.
  • the matrix material or release retarding coating can take the form of an erodible polymer (e.g. a maleic anhydride polymer) which is substantially continuously eroded as the dosage form passes through the gastrointestinal tract.
  • the active compound can be formulated in a delivery system that provides osmotic control of the release of the compound. Osmotic release and other delayed release or sustained release formulations may be prepared in accordance with methods well known to those skilled in the art.
  • the pharmaceutical formulations may be presented to a patient in "patient packs" containing an entire course of treatment in a single package, usually a blister pack.
  • Patient packs have an advantage over traditional prescriptions, where a pharmacist divides a patient's supply of a pharmaceutical from a bulk supply, in that the patient always has access to the package insert contained in the patient pack, normally missing in patient prescriptions.
  • the inclusion of a package insert has been shown to improve patient compliance with the physician's instructions.
  • compositions for topical use include ointments, creams, sprays, patches, gels, liquid drops and inserts (for example intraocular inserts). Such compositions can be formulated in accordance with known methods.
  • compositions for parenteral administration are typically presented as sterile aqueous or oily solutions or fine suspensions, or may be provided in finely divided sterile powder form for making up extemporaneously with sterile water for injection.
  • formulations for rectal or intra-vaginal administration include pessaries and suppositories which may be, for example, formed from a shaped moldable or waxy material containing the active compound.
  • unit-dose suppositories or pessaries may be prepared by admixture of the active ingredient with one or more conventional solid carriers, for example coca butter, and shaping the resulting mixture.
  • mouldable waxy materials include polymers such as high molecular weight polyalkylene glycols, e.g. high molecular weight polyethylene glycols.
  • the formulation may be presented as a tampon impregnated with the active ingredients and optionally one or more excipients or diluents.
  • Other formulations suitable for rectal and vaginal administration include creams, gels, foams, pastes and sprays.
  • topical compositions include dressings such as bandages and adhesive plasters impregnated with active ingredients and optionally one or more excipients or diluents.
  • Carriers which may be used include e.g. polyhydric alcohols such as polyethylene glycols, propylene glycol or glycerol. Suitable excipients are those known in the art to be appropriate.
  • compositions for administration by inhalation may take the form of inhalable powder compositions or liquid or powder sprays, and can be administrated in standard form using powder inhaler devices or aerosol dispensing devices. Such devices are well known.
  • the powdered formulations typically comprise the active compound together with an inert solid powdered diluent such as lactose.
  • the compounds of the formula (I) will generally be presented in unit dosage form and, as such, will typically contain sufficient compound to provide a desired level of biological activity.
  • a formulation may contain from 1 nanogram to 2 grams of active ingredient, e.g. from 1 nanogram to 2 milligrams of active ingredient.
  • particular sub-ranges of compound are 0.1 milligrams to 2 grams of active ingredient (more usually from 10 milligrams to 1 gram, e.g. 50 milligrams to 500 milligrams), or 1 microgram to 20 milligrams (for example 1 microgram to 10 milligrams, e.g. 0.1 milligrams to 2 milligrams of active ingredient).
  • a unit dosage form may contain from 1 milligram to 2 grams, more typically 10 milligrams to 1 gram, for example 50 milligrams to 1 gram, e.g. 100 miligrams to 1 gram, of active compound.
  • the active compound will be administered to a patient in need thereof (for example a human or animal patient) in an amount sufficient to achieve the desired therapeutic effect.
  • the compounds are generally administered to a subject in need of such administration, for example a human or animal patient, preferably a human.
  • the compounds will typically be administered in amounts that are therapeutically or > prophylactically useful and which generally are non-toxic.
  • the benefits of administering a compound of the formula (I) may outweigh the disadvantages of any toxic effects or side effects, in which case it may be considered desirable to administer compounds in amounts that are associated with a degree of toxicity.
  • the compounds may be administered over a prolonged term to maintain beneficial therapeutic effects or may be administered for a short period only. Alternatively they may be administered in a continuous manner or in a manner that provides persistent intermittent dosing (e.g. a pulsatile manner).
  • a typical daily dose of the compound of formula (I) can be in the range from 100 picograms to 100 milligrams per kilogram of body weight, more typically 5 nanograms to 25 milligrams per kilogram of bodyweight, and more usually 10 nanograms to 15 milligrams per kilogram (e.g. 10 nanograms to 10 milligrams, and more typically 1 microgram per kilogram to 20 milligrams per kilogram, for example 1 microgram to 10 milligrams per kilogram) per kilogram of bodyweight although higher or lower doses may be administered where required.
  • the compound can be administered on a daily basis or on a repeat basis every 2, or 3, or 4, or 5, or 6, or 7, or 10 or 14, or 21 , or 28 days for example.
  • a patient will be given an infusion of a compound for periods of one hour to 4 hours daily for up to ten days in particular up to two days for one week, every two weeks in three, and the treatment repeated at a desired interval such as three to six weeks, in particular every three weeks. More particularly, a patient may be given an infusion of a compound for periods of one hour daily twice a week for two weeks in three weeks and the treatment repeated every three weeks.
  • a patient may be given an infusion of a compound for periods of one hour daily twice a week for three weeks in four weeks and the treatment repeated every four weeks.
  • a patient will be given an infusion of a compound for periods of one hour daily for up to ten days in particular up to five days for one week, and the treatment repeated at a desired interval such as two to four weeks, in particular every three weeks.
  • a patient may be given an infusion of a compound for periods of one hour daily for 5 days and the treatment repeated every three weeks.
  • a patient is given an infusion over 30 minutes to 1 hour followed by maintenance infusions of variable duration, for example 1 to 5 hours, e.g. 3 hours.
  • a patient is given a continuous infusion for a period of 12 hours to 5 days, an in particular a continuous infusion of 24 hours to 72 hours.
  • the quantity of compound administered and the type of composition used will be commensurate with the nature of the disease or physiological condition being treated and will be at the discretion of the physician.
  • the compounds as defined herein can be administered as the sole therapeutic agent or they can be administered in combination therapy with one of more other compounds for treatment of a particular disease state, for example a neoplastic disease such as a cancer as hereinbefore defined.
  • a neoplastic disease such as a cancer as hereinbefore defined.
  • other therapeutic agents or treatments that may be administered together (whether concurrently or at different time intervals) with the compounds of the formula (I) include but are not limited to:
  • Chromatin targeted therapies e.g. HDAC or HAT modulators
  • Radiotherapy and • Other therapeutic or prophylactic agents; for example agents that reduce or alleviate some of the side effects associated with chemotherapy.
  • agents include anti-emetic agents and agents that prevent or decrease the duration of chemotherapy-associated neutropenia and prevent complications that arise from reduced levels of red blood cells or white blood cells, for example erythropoietin (EPO), granulocyte macrophage-colony stimulating factor (GM-CSF), and granulocyte-colony stimulating factor (G-CSF).
  • EPO erythropoietin
  • GM-CSF granulocyte macrophage-colony stimulating factor
  • G-CSF granulocyte-colony stimulating factor
  • agents that inhibit bone resorption such as bisphosphonate agents e.g.
  • zoledronate, pamidronate and ibandronate agents that suppress inflammatory responses (such as dexamethazone, prednisone, and prednisolone) and agents used to reduce blood levels of growth hormone and IGF-I in acromegaly patients such as synthetic forms of the brain hormone somatostatin, which includes octreotide acetate which is a long-acting octapeptide with pharmacologic properties mimicking those of the natural hormone somatostatin.
  • agents that suppress inflammatory responses such as dexamethazone, prednisone, and prednisolone
  • agents used to reduce blood levels of growth hormone and IGF-I in acromegaly patients such as synthetic forms of the brain hormone somatostatin, which includes octreotide acetate which is a long-acting octapeptide with pharmacologic properties mimicking those of the natural hormone somatostatin.
  • agents such as leucovorin, which is used as an antidote to drugs that decrease levels of folic acid, or folinic acid itself and agents such as megestrol acetate which can be used for the treatment of side-effects including oedema and thromboembolic episodes.
  • the two or more treatments may be given in individually varying dose schedules and via different routes.
  • the compounds can be administered simultaneously or sequentially.
  • they can be administered at closely spaced intervals (for example over a period of 5-10 minutes) or at longer intervals (for example 1 , 2, 3, 4 or more hours apart, or even longer periods apart where required), the precise dosage regimen being commensurate with the properties of the therapeutic agent(s).
  • the compounds of the invention may also be administered in conjunction with non- chemotherapeutic treatments such as radiotherapy, photodynamic therapy, gene therapy; surgery and controlled diets.
  • non- chemotherapeutic treatments such as radiotherapy, photodynamic therapy, gene therapy; surgery and controlled diets.
  • the compound and one, two, three, four or more other therapeutic agents can be, for example, formulated together in a dosage form containing two, three, four or more therapeutic agents.
  • the individual therapeutic agents may be formulated separately and presented together in the form of a kit, optionally with instructions for their use.
  • a patient Prior to administration of a compound, a patient may be screened to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against Hsp90. For example, a biological sample taken from a patient may be analysed to determine whether a condition or disease, such as cancer, that the patient is or may be suffering from is one which is characterised by a genetic abnormality or abnormal protein expression which leads to the mutation or over-activation of an Hsp90 client protein. Examples of such abnormalities that result in activation of Hsp90 client proteins include; Bcr-ABL translocation, Flt-3 internal duplication, and mutation of Braf, or over-expression of ErbB2.
  • the patient may be subjected to a diagnostic test to detect a marker characteristic of up- regulation.
  • diagnosis includes screening.
  • marker we include genetic markers including, for example, the measurement of DNA composition to identify mutations of Braf, BCR- abl, and Flt3 or other affected client proteins.
  • the term marker also includes proteins such as ErbB2, including levels or concentrations of the protein or some fragments or degradation product and for enzymes the enzymic activity.
  • the protein (e.g. phosphorylated or not) and mRNA levels of the aforementioned proteins could also be assessed to characterise a change in activity.
  • the level of phosphorylated AKT can be an indicator of sensitivity to HSP90 inhibitors
  • the diagnostic tests are typically conducted on a biological sample selected from for example tumour biopsy samples, blood samples (isolation and enrichment of shed tumour cells), stool biopsies, sputum, chromosome analysis, pleural fluid, peritoneal fluid, buccal spears or biopsy or from urine.
  • a biological sample selected from for example tumour biopsy samples, blood samples (isolation and enrichment of shed tumour cells), stool biopsies, sputum, chromosome analysis, pleural fluid, peritoneal fluid, buccal spears or biopsy or from urine.
  • the screening process will typically involve direct sequencing, oligonucleotide or protein microarray analysis, proteomic analysis by mass spectrometry, immunohistochemical techniques or detection using a specific antibody.
  • RT-PCR reverse-transcriptase polymerase chain reaction
  • in-situ hybridisation immunoblotting.
  • RT-PCR reverse-transcriptase polymerase chain reaction
  • the level of mRNA in the tumour is assessed by creating a cDNA copy of the mRNA followed by amplification of the cDNA by PCR.
  • Methods of PCR amplification, the selection of primers, and conditions for amplification are known to a person skilled in the art. Nucleic acid manipulations and PCR are carried out by standard methods, as described for example in Ausubel, F. M. et al., eds.
  • kits for RT-PCR for example Roche Molecular Biochemicals
  • methodology as set forth in United States patents 4,666,828; 4,683,202; 4,801,531; 5,192,659, 5,272,057, 5,882,864, and 6,218,529 and incorporated herein by reference
  • FISH fluorescence in-situ hybridisation
  • in situ hybridization comprises the following major steps: (1) fixation of tissue to be analyzed; (2) prehybridization treatment of the sample to increase accessibility of target nucleic acid, and to reduce non-specific binding; (3) hybridization of the mixture of nucleic acids to the nucleic acid in the biological structure or tissue; (4) post-hybridization washes to remove nucleic acid fragments not bound in the hybridization, and (5) detection of the hybridized nucleic acid fragments.
  • the probes used in such applications are typically labelled, for example, with radioisotopes or fluorescent reporters.
  • Preferred probes are sufficiently long, for example, from about 50, 100, or 200 nucleotides to about 1000 or more nucleotides, to enable specific hybridization with the target nucleic acid(s) under stringent conditions.
  • Commercially available FISH probes also exist for cytogenetic detection of chromosome rearrangemrnts, which can be used to detect Flt3 and Bcr-Abl translocations within leukeamia cell populations. Standard methods for carrying out FISH are described in Ausubel, F. M. et al., eds. Current Protocols in Molecular Biology, 2004, John Wiley & Sons lnc and Fluorescence In Situ Hybridization: Technical Overview by John M. S. Bartlett in Molecular Diagnosis of Cancer, Methods and Protocols, 2nd ed.; ISBN: 1-59259-760-2; March 2004, pps. 077-088; Series: Methods in Molecular Medicine.
  • double-stranded cDNA is synthesized from total RNA Using a (dT)24 oligomer for priming first-strand cDNA synthesis, followed by second strand cDNA synthesis with random hexamer primers.
  • the double-stranded cDNA is used as a template for in vitro transcription of cRNA using biotinylated ribonucleotides.
  • cRNA is chemically fragmented according to protocols described by Affymetrix (Santa Clara, CA, USA), and then hybridized overnight on Human Genome Arrays.
  • the protein products expressed from the mRNAs may be assayed by immunohistochemistry of tumour samples, solid phase immunoassay with microtiter plates,
  • DMAW90 Solvent mixture DCM: MeOH, AcOH, H 2 O (90:18:3:2)
  • DMAW120 Solvent mixture DCM: MeOH, AcOH, H 2 O (120:18:3:2)
  • DMAW240 Solvent mixture DCM: MeOH, AcOH, H 2 O (240:20:3:2)
  • the mass quoted for the compound is the monoisotopic mass (i.e. 35 CI; 79 Br etc.).
  • Different systems were used, as described below, and these were equipped with, and were set up to run under, closely similar operating conditions. The operating conditions used are also described below.
  • Scan Range 125-800 amu lonisation Mode: ElectroSpray Positive, Negative or Positive & Negative
  • Fragmentor/Gain 150 / 1 Drying gas Temp/flow: 350 0 C / 13.0 Lmin "1 Nebuliser pressure: 50 psig
  • Anisole or benzyl ether dealkylation (BBr 2 method) A stirred solution of the anisole or benzyl ether (1 equivalent) in dichloromethane at 0 0 C was treated dropwise with a 1 M solution of boron tribromide in dichloromethane (1.5 equivalents per group to be deprotected) and the mixture was stirred for 2 hours. The reaction was quenched by the addition of water and saturated aqueous sodium bicarbonate, the organic layer was separated and the solvent was removed in vacuo. The pure products were obtained either by trituration with diethyl ether or ethyl acetate or by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether).
  • the crude product may be purified using a three step procedure involving di-benzylation [according to the conditions outlined below in Preparation B5 for the synthesis of methyl 5-acetyl-2,4-bis-benzyloxybenzoate (BnBr, K 2 CO 3 , MeCN, reflux)], column chromatography on silica to remove highly coloured impurites (eluting with 3-5% ethyl acetate in petroleum ether) and catalytic hydrogenation [according to Method A5 outlined above (10% Pd/C, EtOH, H 2 )] to afford 4-hydroxy-3-isopropylbenzoic acid as a colourless solid.
  • n-Butyl lithium (2.5M in hexanes, 38.5 m , 100.0 mmol) was added dropwise under a nitrogen atomosphere to a stirred solution of 4-ethylanisole (11.7 g, 86.0 mmol) and ⁇ /, ⁇ /, ⁇ /', ⁇ /'- tetramethylethylenediamine (10 ml, 88.0 mmol) in anhydrous diethyl ether (100 ml) and the mixture was stirred and held at 3O 0 C for 16 hours. The mixture was cooled and poured slowly in to a mixture of excess solid carbon dioxide in anhydrous diethyl ether.
  • 1-(2,4-Bis-benzyloxy-5-chloro-phenyl)-ethanone [prepared as per WO 2004/0500087] (1.10 g, 3.0 mmol) was added to a stirred solution of sodium hydroxide (1.20 g, 30.0 mmol) in water (10 ml) and dioxane (10 ml). Bromine (1.44 g, 9.0 mmol) was added dropwise and the mixture stirred at room temperature for 3 hours. The dioxane was removed by evaporation in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid.
  • Acetic anhydride (3.06 g, 30.0 mmol) was added to methyl 2,4-dihydroxybenzoate (5.04 g, 30.0 mmol) in boron trifluoride diethyl etherate (7.6 ml) and the mixture was stirred and held at reflux for 3 hours and then allowed to cool to room temperature. Water (80 ml) was added and the mixture stirred at room temperature for 30 minutes. The resulting yellow solid was removed by filtration and sucked as dry as possible under vacuum.
  • Methyl 3-bromo-4-hydroxybenzoate [prepared as per Tetrahedron, 2003, 59, 9173] (3.47 g, 15.0 mmol) was dissolved in acetonitrile (50 ml), anhydrous potassium carbonate (3.11 g, 22.5 mmol) was added and the stirred mixture was treated with benzyl bromide (3.08 g, 18.0 mmol) and held at reflux for 5 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between water and dichloromethane. The organic layer was separated, the solvent removed in vacuo and the residue subjected to column chromatography on silica.

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Abstract

The invention provides a compound for use in medicine, the compound being a compound of the formula (Vl0) or a salt, solvate, tautomer or N-oxide thereof: wherein the bicyclic group: is selected from the structures C1, C5 and C6: wherein n is 0, 1, 2 or 3; R 1 : is„ hydrogen, hydroxy, or O-Rz; R2a is hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2a is O-Rz; Rz is Lp-Rp1; SO3H; a glucuronide residue; a mono-, di- or tripeptide residue; or; Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or S(O)xNRp1; x is 1 or 2; Rp1 is hydrogen or a an optionally substituted C1-25 hydrocarbyl group containing 0, 1 or 2 carbocyclic rings and 0, 1, 2, 3, 4, 5 or 6 carbon-carbon multiple bonds, provided that Rp1 is not hydrogen when Lp is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an 0-0 moiety; and excluding compounds wherein R1 is hydroxy and R2a is methoxy; Rp2 and Rp3 are the same or different and each is a group Rp1; and R3, R4a, R8 and R10 are defined in the claims. The compounds of formula (Vl0) are pro-drugs of parent compounds wherein R1 and/or R2a are hydroxy, wherein the parent compounds have Hsp90 inhibiting activity.

Description

PHARMACEUTICAL COMPOUNDS
This invention relates to pro-drugs of compounds that inhibit or modulate the activity of the heat shock protein Hsp90, to the use of the pro-drugs in the treatment or prophylaxis of disease states or conditions mediated by Hsp90, and to novel pro-drugs that break down in vivo to give compounds having Hsp90 inhibitory or modulating activity. Also provided are pharmaceutical compositions containing the pro-drugs.
Background of the Invention
In response to cellular stresses including heat, toxins, radiation, infection, inflammation, and oxidants, all cells produce a common set of heat shock proteins (Hsps) (Macario & de Macario 2000). Most heat shock proteins act as molecular chaperones. Chaperones bind and stabilize proteins at intermediate stages of folding and allow proteins to fold to their functional states. Hsp90 is the most abundant cytosolic Hsp under normal conditions. There are two human isoforms of Hsp90, a major inducible form Hsp90α and minor constitutively expressed form Hsp90β and two other closely related chaperones which are restricted in their intracellular location (Endoplasmic reticulum GP96/GRP94; mitochondrial TRAP1). The term HSP90 as used here includes all these analogues unless stated. Hsp90 binds proteins at a late stage of folding and is distinguished from other Hsps in that most of its protein substrates are involved in signal transduction. Hsp90 has a distinct ATP binding site, including a Bergerat fold characteristic of bacterial gyrase, topoisomerases and histidine kinases. It has been shown that ATP bound at the N-terminal pocket of Hsp90 is hydrolysed. This ATPase activity results in a conformational change in Hsp90 that is required to enable conformational changes in the client protein.
A dimerization domain and a second ATP binding site, which may regulate ATPase activity, is found near the c-terminus of Hsp90. Dimerization of HSP90 appears critical for ATP hydrolysis. Activation of Hsp90 is further regulated through interactions with a variety of other chaperone proteins and can be isolated in complex with other chaperones including Hsp70, Hip, Hop, p23, and p50cdc37. Many other co-chaperone proteins have also been demonstrated to bind HSP90. A simplified model has emerged in which ATP binding to the amino terminal pocket alters Hsp90 conformation to allow association with a multichaperone complex. First the client protein is bound to an Hsp70/Hsp40 complex. This complex then associates with Hsp90 via Hop. When ADP is replaced by ATP, the conformation of Hsp90 is altered, Hop and Hsp70 are released and a different set of co-chaperones is recruited including p50cdc37 and p23. ATP hydrolysis results in the release of these co-chaperones and the client protein from the mature complex. Ansamycin antibiotics herbimycin, geldanamycin (GA) and 17-allylamino-17-desmethoxygeldanamycin (17- AAG) are ATP binding site inhibitors that block the binding of ATP and prevent conversion to the mature complex (Grenert et. a/., 1997. J Biol Chem., 272:23834-23850).
Despite Hsp90 being ubiquitously expressed, GA has a higher binding affinity for Hsp90 derived from tumour vs. normal cell lines (Kamal et. a/., Nature 2003; 425: 407-410). GA also shows more potent cytotoxic activity in tumour cells and is sequestered at higher concentrations within tumours in xenograft mouse models (Brazidec J. Med. Chem. 2004, 47, 3865-3873). Furthermore the ATP- ase activity of Hsp90 is elevated in cancer cells and is an indication of the increased level of stress in these cells. Hsp90 gene amplification has also been reported to occur in the later stages of cancer (Jolly and Morimoto JNCI Vol. 92, No. 19, 1564-1572, 2000).
Increased genetic instability associated with the cancer phenotype leads to an increase in the production of non-native or mutant proteins. The ubiquitin pathway also serves to protect the cell from non-native or misfolded proteins, by targeting these proteins for proteasomal degradation. Mutant proteins are by their nature not native and therefore have the potential to show structural instability and an increased requirement for the chaperone system. (Giannini et a/., MoI Cell Biol. 2004; 24(13):5667-76).
There is some evidence that Hsp90 is found primarily within "activated" multichaperone complexes in the tumour cells as opposed to "latent" complexes in normal cells. One component of the multichaperone complex is the cdc37 co-chaperone. Cdc37 binds Hsp90 at the base of the ATP binding site and could affect the off rates of inhibitors bound to Hsp90 in the "activated" state (Roe et. a/., Cell 116, (2004), pp. 87-98). The client protein bound to the Hsp90-Hsp70 form of the chaperone complex is believed to be more susceptible to ubiquitination and targeting to the proteasome for degradation. E3 ubiquitin ligases have been identified with chaperone interacting motifs and one of these (CHIP) was shown to promote the ubiquitination and degradation of Hsp90 client proteins (Connell et al., 2001. Xu et al., 2002).
Hsp90 client proteins
The number of reported Hsp90 client proteins now exceeds 100. Since many of its client proteins are involved in cell signalling proliferation and survival, Hsp90 has received major interest as an oncology target. Two groups of client proteins, cell signalling protein kinases and transcription factors, in particular suggest Hsp90 regulation may have potential benefit as an anticancer therapy.
Hsp90 protein kinase client proteins implicated in cell proliferation and survival include the following: c-Src Cellular Src (c-Src) is a receptor tyrosine kinase, required for mitogenesis initiated by multiple growth factor receptors, including the receptors for epidermal growth factor receptor (EGFR), platelet-derived growth factor receptor (PDGFR), colony stimulating factor- 1 (CSF-1 R), and the basic fibroblast growth factor (bFGFR). C-Src is also overexpressed and activated in many of the same human carcinomas that overexpress EGFR and ErbB2. Src is also required for the maintenance of normal bone homeostasis through its regulation of osteoclast function. p185erbB2 ErbB2 (Her2/neu) is a receptor tyrosine kinase overexpressed in a variety of malignancies including breast, ovarian, prostate, and gastric cancers. ErbB2 was originally identified as an oncogene and inhibition of Hsp90 results in the polyubiquitination and degradation of erbB2.
Polo mitotic kinase Polo-like kinases (Plks) are important regulators of cell cycle progression during M-phase. Plks are involved in the assembly of the mitotic spindle apparatus and in the activation of CDK/cyclin complexes. PIkI regulates tyrosine dephosphorylation of CDKs through phosphorylation and activation of Cdc25C. CDK1 activation in turn leads to spindle formation and entry into M phase.
Akt (PKB) Akt is involved in pathways that regulate cell growth by stimulating cell proliferation and suppressing apoptosis. Hsp90 inhibition by ansamycins results in a reduction in the Akt half life through ubiquitination and proteasomal degradation. Binding of cdc37 to Hsp90 is also required for the down-regulation of Akt. Following ansamycin treatment cancer cells arrest in the G2/M phase of the cell cycle 24 hours after treatment and proceed to apoptosis 24-48 hours later. Normal cells also arrest 24 hours after ansamycin treatment, but do not proceed on to apoptosis. c-Raf, B-RAF, Mek
The RAS-RAF-MEK-ERK-MAP kinase pathway mediates cellular responses to growth signals. RAS is mutated to an oncogenic form in approximately 15% of human cancers. The three RAF genes are serine/threonine kinases that are regulated by binding RAS. EGFR
The epidermal growth factor receptor (EGFR) is implicated in cell growth, differentiation, proliferation, survival, apoptosis, and migration. Overexpression of EGFR has been found in many different cancers and activating mutations of its kinase domain appear to be pathogenic in a subset of adenocarcinoams of the lung. Flt3
FMS-like tyrosine kinase 3 (FLT3) is a receptor tyrosine kinase involved in cell proliferation, differentiation and apoptosis. Flt3 activation also leads to the activation of phosphatidylinositol 3- kinase (PI3K) and RAS signal-transduction cascades. c-Met c-met is a receptor tyrosine kinase which binds hepatocyte growth factor (HGF) and regulates both cell motility and cell growth, c-met is overexpressed in tumours, including thyroid, stomach, pancreatic and colon cancer. HGF is also detected around the tumours, including liver metastases. This suggests that c-met and HGF play an important role in invasion and metastasis.
Cdk1. Cdk2. Cdk4. Cdk6 Cdk1, Cdk2, Cdk4, and Cdk6 drive the cell cycle. The activity of CDKs is regulated by their binding to specific subunits such as cyclins, inhibitory and assembly factors. The substrate specificity and timing of CDK activities is dictated by their interaction with specific cyclins. Cdk4/cyclin D and Cdk6/cyclin D are active in the G1 phase, Cdk2/cyclin E and Cdk2/cyclin A in S phase, and Cdc2/cyclin A and Cdc2/cyclin B in G2/M phase.
Cyclin-dependent kinase type 4 (CDK4), plays a key role in allowing cells to traverse G1 to S- phase transition of the cell cycle and is constitutively activated in many human cancers. The CDK4 activator, cyclin D1 , is overexpressed and a CDK4 inhibitor, p16, is deleted in a variety of human tumours. Cdk1/Cdk2 inhibitors have been developed which reversibly block normal cells in either the G1/S- phase or at the G2/M border. G2/M arrest is generally less well tolerated by the cells and consequently, they undergo apoptotic cell death. Since Hsp90 also is known to affect cell survival pathways this effect may be further amplified with an Hsp90 inhibitor.
Wee-1 The Wee-1 protein kinase carries out the inhibitory phosphorylation of CDC2 on tyrosine 15
(TyM 5). This is required for activation of the G2-phase checkpoint in response to DNA damage.
Hsp90 transcription factors implicated in cell proliferation and survival include the following: Mutant p53
P53 is a tumour suppressor protein that causes cell cycle arrest and induces apoptosis. P53 is mutated in approximately half of all cancers. Mutant p53 associates with Hsp90 and is down- regulated in cancer lines treated with Hsp90 inhibitors, while wild type p53 levels were unaffected.
Estrogen receptor/ Androgen receptor
Approximately 70% of post-menopausal women who develop breast cancer have tumours that express the estrogen receptor. The first line treatment of these patients is directed at preventing signalling through this pathway and thus inhibiting tumour growth. This can be done by ovarian ablation, treatment with gonadotrophin releasing hormone agonists, aromatase inhibition or treatment with specific agonists which bind to the estrogen receptor but prevent further signalling. Ultimately patients develop resistance to these interventions often as a consequence of crosstalk between the estrogen receptor and growth factor receptors located on the cell membrane. In the unliganded state estrogen receptors are complexed with Hsp90 which facilitates hormone binding. Following binding to the mature receptor Hsp90 complex the liganded receptor can bind to hormone-response elements (HREs) within the regulatory regions of target genes involved in maintaining cell proliferation. Inhibition of Hsp90 initiates proteosomal degradation of the estrogen receptor thus preventing further growth signalling via this pathway. Prostate cancers are hormone- dependent malignancies that respond to therapeutic interventions which reduce circulating levels of testosterone or prevent testosterone binding to the androgen receptor. Although patients initally respond to these treatments most subsequently develop resistance via restoration of signalling via the androgen receptor. Prior to ligand binding the androgen receptor exists in a complex with Hsp90 and other co-chaperones including p23 and immunophilins. This interaction maintains the androgen receptor in a high-affinity ligand binding conformation. Inhibition of Hsp90 leads to proteosomal degradation of the androgen receptor and other co-chaperones which may sensitise the tumour to further hormonal therapies.
Mutated steroid hormone receptors that have arisen for example during anti-hormone therapy and which might be resistant to such therapies are likely to have a greater dependence on HSP90 for their stability and hormone binding function. Hif-1a
Hypoxia inducible factor-la (HIF-Ia) is a transcription factor that controls the expression of genes which play a role in angiogenesis. HIF-Ia is expressed in the majority of metastases and is known to associate with Hsp90. Ansamycin treatment of renal carcinoma cell lines leads to the ubiquitination and proteasomal degradation of HIF-Ia. Hsp90 inhibitors are capable of affecting a large number of targets significant to signal transduction in tumour cell proliferation. Signal transduction inhibitors which regulate the activities of a single target, may not be as efficacious due to signalling pathway redundancy and the rapid development of resistance.
By regulating multiple targets involved in cell signalling and cell proliferation HSP90 inhibitors may prove beneficial in the treatment of a wide spectrum of proliferative disorders.
ZAP70
ZAP-70, a member of the Syk-ZAP-70 protein tyrosine kinase family, is normally expressed in T cells and natural killer cells and has a critical role in the initiation of T-cell signaling. However, it is also expressed aberrantly in approximately 50% of cases of CLL, usually in those cases with unmutated B-cell receptor genes. The mutational status of immunoglobulin heavy-chain variable- region (lgVH) genes in the leukemic cells of chronic lymphocytic leukemia (CLL) is an important prognostic factor. The expression of ZAP-70 in CLL cells correlates with lgVH mutational status, disease progression, and survival. ZAP-70 positive CLL is more aggressive than ZAP-70 negative CLL indicating that ZAP-70 may be a key driver of malignancy in this disease. ZAP-70 is physically associated with HSP90 in B-CLL lymphoblasts thus the inhibition of Hsp90 may sensitise these cells to existing chemotherapy or monoclonal antibody therapy.
HSP90 Inhibitors as anti-fungal, anti-protozoal and anti-parasitic agents
Fungal infections have become a major cause for concern in recent years due to the limited number of antifungal agents available, and the the increasing incidence of species that are resistant to established antifungal agents such as the azoles. In addition, the growing population of immunocompromised patients (e.g. patients such as organ transplant patients, cancer patients undergoing chemotherapy, burn patients, AIDS patients, or patients with diabetic ketoacidosis) has given rise to an increase in the incidence of opportunistic fungal infections by fungal agents such as Candida, Cryptoccocus and Aspergillus species and, on occasion, Fusarium, Trichosporon and Dreschlera species. Consequently, there is a need for new anti-fungal agents that can be used to treat the growing numbers of patients with fungal infections and in particular infections due to fungi that have become resistant to existing antifungal drugs.
HSP90 is conserved across evolution being found in bacteria (e.g. HTPG in E.coli) and yeast (e.g. HSC82 and HSP82). Although clients have not been formally identified for the E.coli form, in yeast and all higher organisms the HSP90 family has been shown to function as a chaperone for many essential proteins as described above.
Infection by a range of pathogens is associated with an antibody response to HSP90. For example in Candida albicans infected patients the 47kDa C-terminal fragment of HSP90 is an immunodominant epitope. Furthermore this antibody response is associated with good prognosis suggesting a protective effect against infection. Recombinant antibodies to an epitope in this polypeptide are also protective against infection in mouse models of invasive candidiasis. (See Mathews ef a/ Antimicrobial Agents and Chemotherapy 2003 vol 47, 2208 - 2216 and references therein). Likewise surface expressed HSP90 serves as an antigen in Chagas' disease, ascariasis, leishmaniasis, toxoplamosis and infection due to Schistosoma mansoni and it has been postulated that antibodies to HSP90 convey protection against plamodiutn infection and Malaria.
Mycograb (NeuTec Pharma/Novartis) is a human recombinant monoclonal antibody against heat shock protein 90 that is being developed as a treatment for Candida and has shown significant responses in early trials. Furthermore, the natural product HSP90 inhibitors Geldanamycin, Herbimycin and Radicicol were originally identified by their anti-fungal activity. Key essential proteins have been identified as HSP90 clients in several human pathogens (see Cowen and
Lindquist, Science. 2005 Sep 30;309(5744):2175-6.) Thus HSP90 can play an important role in the growth of pathogens such as Candida species, and HSP90 inhibitors can be useful as treatments for a range of infectious diseases including candidiasis.
It has also been found that Hsp90 increases the capacity of fungi to develop antifungal drug resistance (see Cowen LE, Lindquist S. "Hsp90 potentiates the rapid evolution of new traits: drug resistance in diverse fungi". Science. 2005 Sep 30;309 (5744):2185-9). Therefore, coadministration of an Hsp90 inhibitor with an antifungal drug may enhance the efficacy of the antifungal drug and reduce resistance by preventing the emergence of resistant phenotypes.
HSP90 Inhibitors in the treatment of pain, neuropathic conditions and stroke Cdk5 is a member of the Cdk family of serine/threonine kinases, most of which are key regulators of the cell cycle. Cdk5 activity is regulated through association with its neuron-specific activators, p35 and p39. Recent evidence suggests that CDK5 can phosphorylate tau protein and a number of other neuronal proteins such as NUDE-1 , synapsini , DARPP32 and the Munc18/Syntaxin1A complex. The evidence also suggests that aberrant Cdk5 activity induced by the conversion of p35 to p25 plays a role in the pathogenesis of neurodegenerative diseases such as Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS) and Niemann's Pick type-C disease (NPD). Abnormal hyperphosphorylation of tau after AuSi-42 treatment destabilizes microtubules, contributing to neurite degeneration and the formation of paired helical filaments (PHFs) containing neurofibrillary tangles (NFTs), one of the principal lesions of AD. It has furher been found that cdkδ is necessary for correct neuronal development
The p35 protein which acts as a regulator of CDK5 activity has recently been identified as a client protein for HSP90 and therefore the activity of CDK5 can be regulated by changes in the level and activity of HSP90. Thus inhibition of HSP90 can lead to loss of p35, an inhibition of CDK5, a reduction of phosphorylated tau protein in susceptible individuals and will bring benefit to sufferers of Alzheimers Disease.
Additionally inhibition of HSP90 using known agents has been shown to reduce the accumulation of tau protein aggregates in cellular systems in vitro. (Dickey et al Curr Alzheimer Res. 2005 Apr;2(2):231-8).
Cdk5 has also been shown to have a role in mediating pain signalling. Both Cdk5 and p35 have been shown to be expressed in nociceptive neurons. In p35 knockout mice, which show substantially reduced Cdkδ activity, the response to painful thermal stimuli is delayed (Pareek, T. K., et al., Proceedings of the National Academy of Sciences., 103:791-796 (2006). Additionally administration of the cyclin-dependent kinase 5 (Cdkδ) inhibitor roscovitine has been shown to attenuate the formalin-induced nociceptive responses in rats (Wang, Cheng-haung, et al., Acta Pharmacologica Sinica., 26:46-δO (200δ). Activation of calpain is calcium dependent and is known to affected by activation of the NMDA receptor calcium channel (Amadoro, G; Proceedings of the National Academy of Sciences of the United States of America, 103, 2892-2897 (2006)). NMDA receptor antagonists are know to be clinically effective against neuropathic pain conditions (Christoph, T; et al., Neuropharmacology, 51 ,12-17 (2006)). This efficacy may be linked to the effect of NMDA receptor related calcium influx on calpain activity and its subsequent effect on the activity of Cdkδ. As such compounds modulating Cdkδ activity are expected to be useful for the treatment or prevention of pain and thus modulation of the CDKδ regulator p3δ by HSP90 inhibition could lead to inhibition of CDKδ.
It is desirable to have an agent for the palliative treatment of pain, i.e. the direct relief of pain in addition to the relief of pain as the result of amelioration of the underlying disease or medical condition, which is the cause of the pain. Various Cdk's (especially Cdk's 4, 5 & 6) have been shown to be involved with or mediate neuronal death following hypoxic or ischemic insult (Rashidan, J.; et al.; Proceedings of the National Academy of Sciences., 102:14080-14086 (2005). Furthermore the Cdk inhibitor flavopiridol has been shown to significantly reduce neuronal death in a rat model of focal cerebral ischemia (Osuga, H.; et al.; Proceedings of the National Academy of Sciences., 97:10254-10259 (2000). Cdk5 inhibitors have been shown to have protective effects in both necrotic and apoptotic paradigms of neuronal cell death (Weishaupt, J.; et al.; Molecular and Cellular Neuroscience., 24:489-502 (2003). Stroke is a cerebrovascular event, which occurs when the normal bloodflow to the brain is disrupted, and the brain receives too much or too little blood. Stroke is one of the leading causes of death worldwide, and is also one of the most common causes of neurologic disability.
Ischemic stroke, which is the most common type of stroke, results from insufficient cerebral circulation of blood caused by obstruction of the inflow of arterial blood. Normally, adequate cerebral blood supply is ensured by a system of arteries within the brain. However, various disorders, including inflammation and atherosclerosis, can cause a thrombus, i.e., a blood clot that forms in a blood vessel. The thrombus may interrupt arterial blood flow, causing brain ischemia and consequent neurologic symptoms. Ischemic stroke may also be caused by the lodging of an embolus (an air bubble) from the heart in an intracranial vessel, causing decreased perfusion pressure or increased blood viscosity with inadequate cerebral blood flow. An embolus may be caused by various disorders, including atrial fibrillation and atherosclerosis.
A second type of stroke, hemorrhagic stroke, involves a hemorrhage or rupture of an artery leading to the brain. Hemorrhagic stroke results in bleeding into brain tissue, including the epidural, subdural, or subarachnoid space of the brain. A hemorrhagic stroke typically results from the rupture of an arteriosclerotic vessel that has been exposed to arterial hypertension or to thrombosis.
One opportunity for intervention in stroke is the prevention or reduction of risk of stroke in patients at risk for stroke. There are many known risk factors for stroke, including vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation. At risk patients have been treated with agents to control blood pressure or manage blood lipid level, and have been treated with antiplatelet agents (such as clopidrogel) and anticoagulants. A second opportunity is the treatment of acute stroke. However, current pharmacologic therapies for treating acute stroke are limited to restoring blood flow within a narrow therapeutic time window of less than three hours after stroke. There remains a need for agents which are effective within a longer therapeutic time window. Another opportunity is recovery or restoration after the acute stroke period, i.e. the reduction or prevention of secondary cell damage in the penumbra. There remains a need for agents which are effective in reducing or preventing secondary cell damage after stroke.
It would be desirable to obtain a single pharmaceutical agent which can be used in more than one of the above-mentioned opportunities for treating stroke. Such an agent may be administered to patients at risk for stroke, and also may be administered to patients suffering from acute stroke, or patients undergoing treatment for recovery or restoration after the acute stroke period. Such an agent may also target more than one distinct mechanism in the biochemical cascade of stroke. HSP90 inhibitors and the treatment of Hepatitis C and other viral diseases
Infection of a host cell with viral RNA/DNA results in a substantial redirection of cellular protein sysnthesis towards key viral proteins encoded by the viral nucleic acid. The increased protein synthetic burden places a stress on the cell as a consequence of increased demand for energy and synthetic precursers. Upregulation of heat shock proteins is frequently a consequence of viral infection at least in part due to this stress. One function of the HSP induction may be to assist in the stabilization and folding of the high levels of 'foreign' protein generated in preparation for virus replication. In particular recent work has suggested that HSP90 is required for stable production of functional NS2/3 protease in Hepatitis C (HCV) replicon infected cells. HSP 90 inhibitors have also been demonstrated to block viral replication in in vitro systems. (Nagkagawa, S, Umehara T, Matsuda C, et al Biochem. Biophys. Res Commun. 353 (2007) 882-888; Waxman L, Witney, M et al PNAS 98 (2001) 13931 - 13935).
Heat Shock Proteins and antitumour drug resistance
It has long been recognized that the native tertiary conformation of any given polypeptide is determined by its primary (amino acid) sequence. However, as explained above, it is now clear that the proper folding of many proteins in vivo requires the assistance of heat-shock proteins (Hsps) acting as molecular chaperones. While this chaperone function is important to normal cellular function under all conditions, it becomes crucial in cells which are stressed (for example by heat, hypoxia or acidosis). Such conditions typically prevail in tumour cells, which exist in a hostile host environment. The upregulation of Hsps often seen in such cells is therefore likely to represent a mechanism by which malignant cells maintain the integrity of their proteomes under conditions which compromise protein folding. Thus, modulators or inhibitors of stress proteins in general (and Hsp90 in particular) represent a class of chemotherapeutics with the unique ability to inhibit multiple aberrant signalling pathways simultaneously. They can therefore exert antitumour effects whilst eliminating (or reducing the incidence of) resistance relative to other treatment paradigms.
Moreover, therapeutic anticancer interventions of all types necessarily increase the stresses imposed on the target tumour cells. In mitigating the deleterious effects of such stresses, Hsps are directly implicated in resisting the effects of cancer drugs and treatment regimens. Thus, modulators or inhibitors of stress protein function in general (and Hsp90 in particular) represent a class of chemotherapeutics with the potential for: (i) sensitizing malignant cells to anticancer drugs and/or treatments; (ii) alleviating or reducing the incidence of resistance to anticancer drugs and/or treatments; (iii) reversing resistance to anticancer drugs and/or treatments; (iv) potentiating the activity of anticancer drugs and/or treatments; (v) delaying or preventing the onset of resistance to anticancer drugs and/or treatments.
Pro-Drugs Pro-drugs are generally recognised as being chemical compounds that have little or no pharmacological activity themselves but which undergo biotransformation to a therapeutically active metabolite (see for example Bernard Testa, Biochemical Pharmacology, 68 (2004), 2097- 2106, and "Design of Prodrugs" (Bundgaard H. ed.) 1985 Elsevier Science Publishers B. V. (Biomedical Division).
Pro-drugs are used for a variety of reasons. For example, they may be used inter alia to:
■ impart better solubility to an otherwise insoluble or poorly soluble drug
■ improve chemical stability
■ improve the organoleptic properties of a drug ■ alter the pharmacokinetics of a drug
■ reduce pre-systemic (first pass) metabolism
■ reduce the extent of conjugation of a drug
■ improve oral absorption
■ provide selective targetting of a drug ■ provide in situ activation of a cytotoxic agent
WO 99/29705 (Glycomed et al) disclose a class of glycomimetic compounds having a number of possible uses including the treatment of cancer. One compound specifically disclosed in WO 99/29705 is the compound 2-(2-hydroxy-benzoyl)-1 ,2,3,4-tetrahydro-isoquinoline-3-carboxylic acid.
Our earlier International application PCT/GB2006/001382 discloses hydroxybenzoic acid amides as Hsp90 inhibitors.
Summary of the Invention
The present invention provides pro-drug compounds containing functional groups that are modified or removed in vivo to give compounds having Hsp90 inhibiting or modulating activity. It is envisaged that the pro-drug compounds will be useful in preventing or treating disease states or conditions mediated by Hsp90.
Thus, for example, it is envisaged that the compounds of the invention will be useful in alleviating or reducing the incidence of cancer.
Accordingly, in a first aspect, the invention provides a compound for use in medicine, the compound being a compound of the formula (Vl0):
Figure imgf000012_0001
or a salt, solvate, tautomer or N-oxide thereof; wherein the bicyclic group:
Figure imgf000012_0002
is selected from the structures C1 , C5 and C6:
Figure imgf000012_0003
wherein n is 0, 1 , 2 or 3;
R1 is hydrogen, hydroxy, or O-Rz;
R2a is hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2a is O-Rz;
Rz is Lp-Rp1; SO3H; a glucuronide residue; a mono-, di- or tripeptide residue; or
Figure imgf000012_0004
Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or S(O)xNRp1; x is 1 or 2;
Rp1 is hydrogen or a C1-2S hydrocarbyl group containing O, 1 or 2 carbocyclic rings and O, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds, the C1-2S hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or CIi-C1-6 alkylamino, C1-6 acylamino, halogen or Ci-4 alkoxy or C1-4 alkyl groups wherein the Ci-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 3 non-terminal carbon atoms in the Ci-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or O(C=O); provided that Rp1 is not hydrogen when Lp is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an 0-0 moiety; and excluding compounds wherein R1 is hydroxy and R2a is methoxy; Rp2 and Rp3 are the same or different and each is a group Rp1;
R3 is selected from hydrogen; halogen; cyano; C1-5 hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and C1-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-Ci-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members;
R4a is selected from hydrogen, fluorine, chlorine and methoxy;
R8 is selected from hydrogen and fluorine; and
R10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or di-C1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group Ra-Rb; wherein:
Ra is a bond, O, CO, X1C(X2), C(X2)X1, X1C(X2)X1, S, SO, SO2, NR0, SO2NR0 or NR0SO2; and
Rb is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C1-12 hydrocarbyl (such as C1-10 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino (e.g. mono- or di-C1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C1-12 hydrocarbyl group (or C1-10 hydrocarbyl group) may optionally be replaced by O, S, SO,
SO2, NR0, X1C(X2), C(X2)X1 or X1C(X2)X1;
R° is selected from Rb, hydrogen and C1-4 hydrocarbyl; and X1 is O, S or NR0 and X2 is =0, =S or =NRC
In another aspect, the invention provides a compound of the formula (Vl):
Figure imgf000013_0001
or a salt, solvate, tautomer or N-oxide thereof; wherein the bicyclic group:
Figure imgf000014_0001
is selected from the structures C1 , C5 and C6:
Figure imgf000014_0002
wherein n is 0, 1 , 2 or 3;
R1 is hydrogen, hydroxy, or O-Rz;
R2a is hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2a is O-Rz;
Rz is Lp-Rp1; SO3H; a glucuronide residue; a mono-, di- or tripeptide residue; or
Figure imgf000014_0003
Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or S(O)xNRp1; x is 1 or 2;
Rp1 is hydrogen or a C1-2S hydrocarbyl group containing 0, 1 or 2 carbocyclic rings and 0, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds, the C1-2S hydrocarbyl group being optionally substituted by amino, mono- or di-C1-6 alkylamino, C1-6 acylamino, halogen or Ci-4 alkoxy or Ci-4 alkyl groups wherein the Ci-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 3 non-terminal carbon atoms in the C1-2S hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O); provided that Rp1 is not hydrogen when Lp is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an O-O moiety; and excluding compounds wherein R1 is hydroxy and R2a is methoxy;
Rp2 and Rp3 are the same or different and each is a group Rp1;
R3 is selected from hydrogen; halogen; cyano; C1-5 hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and Ci-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R4a is selected from hydrogen, fluorine, chlorine and methoxy; R8 is selected from hydrogen and fluorine; and R10 is selected from: halogen; hydroxy; trifluoromethyl; cyaπo; nitro; carboxy; amino; mono- or di-C1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group Ra-Rb; wherein:
Ra is a bond, O, CO, X1C(X2), C(X2)X\ X1C(X2)X1, S, SO, SO2, NR0, SO2NR0 or NR0SO2; and
Rb is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C1-I2 hydrocarbyl (such as C1-I0 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C^s non-aromatic hydrocarbylamino (e.g. mono- or di-C1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C1-12 hydrocarbyl group (or C1-I0 hydrocarbyl group) may optionally be replaced by O, S, SO,
SO2, NR0, X1C(X2), C(X2)X1 or X1C(X2)X1; R0 is selected from Rb, hydrogen and C1-4 hydrocarbyl; and
X1 is O, S or NR0 and X2 is =0, =S or =NR°.
In another aspect, the invention provides a compound for use in medicine, the compound being a compound of the formula (I0);
Figure imgf000015_0001
or salts, tautomers, solvates and N-oxides thereof; wherein: R1 is hydrogen, hydroxy, or O-Rz;
R2 is hydrogen, hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2 is O-Rz; R2 is Lp-Rp1 ; SO3H; a glucuronide residue; a mono-, di- or tripeptide residue; or
Figure imgf000015_0002
Lp is a bond, C=O, (C=O)O, (C=0)NRp1 or S(0)xNRp1; x is 1 or 2; Rp1 is hydrogen or a C-ι.25 hydrocarbyl group containing 0, 1 or 2 carbocyclic rings and 0, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds, the C1-25 hydrocarbyl group being optionally substituted by amino, mono- or di-C^e alkylamino, C1-6 acylamino, halogen or C1-4 alkoxy or C1-4 alkyl groups wherein the C1-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 3 non-terminal carbon atoms in the Ci-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O); provided that Rp1 is not hydrogen when Lp is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an 0-0 moiety; and excluding compounds wherein R1 is hydroxy and R2a is methoxy; Rp2 and Rp3 are the same or different and each is a group Rp1;
R3 is selected from hydrogen; halogen; cyano; C1-5 hydrocarbyl and Ci-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and Ci-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, Ci-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R4 is selected from hydrogen; a group -(O)n-R7 where n is O or 1 and R7 is an acyclic C1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-Ci-5 hydrocarbyl-amino, wherein the acyclic C1-5 hydrocarbyl group and the mono and di-C1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, Ci-2 alkoxy, amino, mono- and di-Ci-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; or R3 and R4 together form a monocyclic carbocyclic or heterocyclic ring of 5 to 7 ring members; R5 and R6 together with the nitrogen atom to which they are attached form a bicyclic heterocyclic group having 8 to 12 ring members of which up to 5 ring members are heteroatoms selected from oxygen, nitrogen and sulphur; wherein the bicyclic heterocyclic group is optionally substituted by one or more substituents R10;
R8 is selected from hydrogen and fluorine; and
R10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or di-C1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group Ra-Rb; wherein:
Ra is a bond, O, CO, X1C(X2), C(X2)X1, X1C(X2)X1, S, SO, SO2, NRC, SO2NR0 or NR0SO2; and Rb is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C1-12 hydrocarbyl (such as C1-10 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino (e.g. mono- or di-C1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C1-12 hydrocarbyl group (or C1-10 hydrocarbyl group) may optionally be replaced by O, S, SO, SO2, NRC, X1C(X2), C(X2)X1 or X1C(X2)X1;
Rc is selected from Rb, hydrogen and C1-4 hydrocarbyl; and X1 is O, S or NRC and X2 is =0, =S or =NR°; but excluding the compound 2-(2-hydroxy-benzoyl)-1 ,2,3,4-tetrahydro-isoquinoline-3-carboxylic acid.
In a further aspect, the invention provides a compound of the formula (I):
Figure imgf000017_0001
or salts, tautomers, solvates and N-oxides thereof; wherein: R1 is hydrogen, hydroxy, or O-Rz;
R2 is hydrogen, hydroxy, methoxy or 0-R2; provided that at least one of R1 and R2 is O-Rz; Rz is Lp-Rp1; SO3H; a glucuronide residue; a mono-, di- or tripeptide residue; or
Figure imgf000017_0002
Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or S(O)xNR"1; x is 1 or 2;
Rp1 is hydrogen or a C1-25 hydrocarbyl group containing O, 1 or 2 carbocyclic rings and O, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds, the C1-25 hydrocarbyl group being optionally substituted by amino, mono- or di-C1-6 alkylamino, C1-6 acylamino, halogen or C1-4 alkoxy or C1-4 alkyl groups wherein the C1-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 3 non-terminal carbon atoms in the C1-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=0)0 or 0(C=O); provided that Rp1 is not hydrogen when l_p is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an 0-0 moiety; but excluding compounds wherein Lp is a bond and Rp1 is an unsubstituted benzyl group or a methyl group; Rp2 and Rp3 are the same or different and each is a group Rp1; R3 is selected from hydrogen; halogen; cyano; C1-5 hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and Ci-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, Ci-2 alkoxy, amino, mono- and di-Ci-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R4 is selected from hydrogen; a group -(O)n-R7 where n is 0 or 1 and R7 is an acyclic Ci-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C1-5 hydrocarbyl-amino, wherein the acyclic C1-5 hydrocarbyl group and the mono and di-C1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C-ι-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; or R3 and R4 together form a monocyclic carbocyclic or heterocyclic ring of 5 to 7 ring members; R5 and R6 together with the nitrogen atom to which they are attached form a bicyclic heterocyclic group having 8 to 12 ring members of which up to 5 ring members are heteroatoms selected from oxygen, nitrogen and sulphur; wherein the bicyclic heterocyclic group is optionally substituted by one or more substituents R10;
R8 is selected from hydrogen and fluorine; and R10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or di-C1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group Ra-Rb; wherein:
Ra is a bond, O, CO, X1C(X2), C(X2)X1, X1C(X2)X1, S, SO, SO2, NRC, SO2NR0 or NR0SO2; and Rb is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and Ci-12 hydrocarbyl (such as C1-10 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino (e.g. mono- or CIi-C1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C1--I2 hydrocarbyl group (or C1--I0 hydrocarbyl group) may optionally be replaced by O, S, SO, SO2, NRC, X1C(X2), C(X2)X1 or X1C(X2)X1;
R° is selected from Rb, hydrogen and C1-4 hydrocarbyl; and X1 is O, S or NR0 and X2 is =0, =S or =NR°; but excluding the compound 2-(2-hydroxy-benzoyl)-1 ,2,3,4-tetrahydro-isoquinoline-3-carboxylic acid.
Compounds of the formulae (Vl0), (Vl), (I0) and (I) and sub-groups thereof as defined herein are pro-drugs of the compounds disclosed in our earlier application PCT/GB2006/001382. Thus, it is envisaged that compounds of the formulae (Vl0), (Vl), (I0) and (I) and sub-groups thereof will be converted in vivo to give compounds wherein O-Rz is OH.
In further aspects, the invention provides:
• A compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use in the prophylaxis or treatment of a disease state or condition mediated by Hsp.90.
• The use of a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a disease state or condition mediated by Hsp90. • A method for the prophylaxis or treatment of a disease state or condition mediated by
Hsp90, which method comprises administering to a subject in need thereof a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
• A compound of the formula (I0), (1), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use in alleviating or reducing the incidence of a disease state or condition mediated by Hsp90.
• The use of a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
• A compound of the formula (I0), (I), (II), (III), Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use in treating a disease or condition comprising or arising from abnormal cell growth in a mammal.
• The use of a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for the manufacture of a medicament for treating a disease or condition comprising or arising from abnormal cell growth in a mammal.
• A method for treating a disease or condition comprising or arising from abnormal cell growth in a mammal, which method comprises administering to the mammal a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (VI), (Via), (VIl), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein in an amount effective in inhibiting abnormal cell growth. • A compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use in alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal. . The use of a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII),
(Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for the manufacture of a medicament for for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal.
• A method for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal, which method comprises administering to the mammal a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein in an amount effective in inhibiting abnormal cell growth.
• A method for treating a disease or condition comprising or arising from abnormal cell growth in a mammal, the method comprising administering to the mammal a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VIl), (Vila), (VIIb) or (VlII) or any sub-groups or examples thereof as defined herein in an amount effective to inhibit Hsp90 activity.
• A method for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal, the method comprising administering to the mammal a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein in an amount effective to inhibit Hsp90 activity.
. A compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use as an inhibitor of
Hsp90.
• A method of inhibiting Hsp90, which method comprises contacting the Hsp90 with an HspΘO-inhibiting compound of the formula (I0), (I), (II), (111), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein. • A compound of the formula (1°), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or
(VIII) or any sub-groups or examples thereof as defined herein for use in modulating a cellular process (for example cell division) by inhibiting the activity of Hsp90.
• A method of modulating a cellular process (for example cell division) by inhibiting the activity of Hsp90 using a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein. • A compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use in the prophylaxis or treatment of a disease state as described herein.
• The use of a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for the manufacture of a medicament, wherein the medicament is for any one or more of the uses defined herein.
• A pharmaceutical composition comprising a compound of the formula (I0), (I), (II), (III),
Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII)) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier.
• A pharmaceutical composition comprising a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier in a form suitable for oral administration. • A pharmaceutical composition comprising a compound of the formula (I0), (I), (II), (III),
(Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier in a form suitable for parenteral administration, for example by intravenous (i.v.) administration.
• A pharmaceutical composition comprising a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier in a form suitable for intravenous (i.v.) administration by injection or infusion.
• A compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use in medicine. • A compound as defined herein for any of the uses and methods set forth above, and as described elsewhere herein.
• A compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for use in treatment or prophylaxis of a disease state or condition in a patient who has been screened and has been determined as suffering from, or being at risk of suffering from, a disease or condition which would be susceptible to treatment with a compound having activity against Hsp90.
• The use of a compound of the formula (1°), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein for the manufacture of a medicament for the treatment or prophylaxis of a disease state or condition in a patient who has been screened and has been determined as suffering from, or being at risk of suffering from, a disease or condition which would be susceptible to treatment with a compound having activity against Hsp90.
• A method for the diagnosis and treatment of a disease state or condition mediated by
Hsp90, which method comprises (i) screening a patient to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against Hsp90; and (ii) where it is indicated that the disease or condition from which the patient is thus susceptible, thereafter administering to the patient a compound of the formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein.
General Preferences and Definitions
The compounds of the invention are pro-drugs of the hydroxybenzoic acid amide Hsp90 inhibitors disclosed in our earlier international application PCT/GB2006/001382. For convenience, the compounds of PCT/GB2006/001382 may be referred to variously in this application as "the parent compounds" or "the phenolic compounds". References to the pro-drugs being "derived" from the "parent compounds" or "phenolic compounds" is only intended to imply a structural relationship and is not intended to imply any particular method of preparing the prodrugs. Thus, the pro-drugs may be prepared from the compounds of PCT/GB2006/001382 as described below, or they may be prepared by methods that do not involve the intermediacy of a phenolic compound of PCT/GB2006/001382.
In this section, as in all other sections of this application, unless the context indicates otherwise, references to a compound of formula (I) includes formula (I0), formula (Vl0) and all subgroups of formula (I) as defined herein, including formulae (II), (III), (IV), (V), (Vl), (Via), (VII), (Vila) and (VIIb) and the term 'subgroups' includes all preferences, embodiments, examples and particular compounds defined herein. Similarly, a reference to formula (Vl) includes formula (Vl0) and all sub-groups, preferences, embodiments, examples and particular compounds within formula (Vl).
Moreover, a reference to a compound of formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) and sub-groups thereof includes ionic forms, salts, solvates, isomers, tautomers, N-oxides, esters, isotopes and protected forms thereof, as discussed below:- preferably, the salts or tautomers or isomers or N-oxides or solvates thereof:- and more preferably, the salts or tautomers or N-oxides or solvates thereof.
The compounds of formula (I0), (I), (II), (III), (Vl0), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) and sub-groups thereof, including ionic forms, salts, solvates, isomers, tautomers, N-oxides, esters, isotopes and protected forms thereof as defined herein may be referred to for convenience as "the compounds of the invention" or, in the singular, "a compound of the invention". As used herein, the term "treatment" and the related terms "treat" and "treating" refer to both prophylactic or preventative treatment as well as curative or palliative treatment of pain. Thus, the term encompasses situations where pain is already being experienced by a subject or patient, as well as situations where pain is not currently being experienced but is expected to arise. The term "treatment", "treat", "treating" and related terms also cover both complete and partial pain reduction or prevention. Thus, for example, the compounds of the invention may prevent existing pain from worsening, or they reduce or even eliminate pain. When used in a prophylactic sense, the compounds may prevent any pain from developing or they may lessen the extent of pain that may develop. As used herein, the term "modulation", as applied to the activity of the heat shock protein Hsp90, is intended to define a change in the level of biological activity of the heat shock protein. Thus, modulation encompasses physiological changes which effect an increase or decrease in the relevant heat shock protein activity. In the latter case, the modulation may be described as "inhibition". The modulation may arise directly or indirectly, and may be mediated by any mechanism and at any physiological level, including for example at the level of gene expression (including for example transcription, translation and/or post-translational modification), at the level of expression of genes encoding regulatory elements which act directly or indirectly on the levels of heat shock protein activity. Thus, modulation may imply elevated/suppressed expression or over- or under-expression of the heat shock protein, including gene amplification (i.e. multiple gene copies) and/or increased or decreased expression by a transcriptional effect, as well as hyper- (or hypo-)activity and (de)activation of the heat shock protein (including (de)activation) by mutation(s). The terms "modulated", "modulating" and "modulate" are to be interpreted accordingly.
As used herein, the term "mediated", as used e.g. in conjunction with the heat shock protein as described herein (and applied for example to various physiological processes, diseases, states, conditions, therapies, treatments or interventions) is intended to operate limitatively so that the various processes, diseases, states, conditions, treatments and interventions to which the term is applied are those in which heat shock protein Hsp90 plays a biological role. In cases where the term is applied to a disease, state or condition, the biological role played by heat shock protein Hsp90 may be direct or indirect and may be necessary and/or sufficient for the manifestation of the symptoms of the disease, state or condition (or its aetiology or progression). Thus, heat shock protein Hsp90 activity (and in particular aberrant levels of heat shock protein Hsp90 activity, e.g. Hsp90 over-expression) need not necessarily be the proximal cause of the disease, state or condition: rather, it is contemplated that the heat shock protein Hsp90 mediated diseases, states or conditions include those having multifactorial aetiologies and complex progressions in which Hsp90 is only partially involved. In cases where the term is applied to treatment, prophylaxis or intervention (e.g. in the "Hsp90-mediated treatments" and "Hsp90-mediated prophylaxis" of the invention), the role played by Hsp90 may be direct or indirect and may be necessary and/or sufficient for the operation of the treatment, prophylaxis or outcome of the intervention. Thus, a disease state or condition mediated by Hsp90 includes the development of resistance to any particular cancer drug or treatment (including in particular resiatnce to one or more of the signalling inhibitors described herein).
As used herein, the term "modulation", as applied to the activity of cyclin dependent kinase 5 (CDK5), is intended to define a change in the level of biological activity of the kinase(s). Thus, modulation encompasses physiological changes which effect an increase or decrease in the relevant kinase activity. In the latter case, the modulation may be described as "inhibition". The modulation may arise directly or indirectly, and may be mediated by any mechanism and at any physiological level, including for example at the level of gene expression (including for example transcription, translation and/or post-translational modification), at the level of expression of genes encoding regulatory elements which act directly or indirectly on the levels of cyclin dependent kinase 5 (CDK5), or at the level of enzyme (e.g. cyclin dependent kinase 5 (CDK5) activity (for example by allosteric mechanisms, competitive inhibition, active-site inactivation, perturbation of feedback inhibitory pathways etc.). Thus, modulation may imply elevated/suppressed expression or over- or under-expression of the cyclin dependent kinase 5 (CDK5) including gene amplification (i.e. multiple gene copies) and/or increased or decreased expression by a transcriptional effect, as well as hyper- (or hypo-)activity and (de)activation of the cyclin dependent kinase 5 (CDK5) including (de)activation) by mutation(s). The terms "modulated", "modulating" and "modulate" are to be interpreted accordingly.
As used herein, the term "mediated", as used e.g. in conjunction with the cyclin dependent kinase 5 (CDK5) as described herein (and applied for example to various physiological processes, diseases, states, conditions, therapies, treatments or interventions) is intended to operate limitatively so that the various processes, diseases, states, conditions, treatments and interventions to which the term is applied are those in which cyclin dependent kinase 5 (CDK5) plays a biological role, in cases where the term is applied to a disease, state or condition, the biological role played by cyclin dependent kinase 5 (CDK5) may be direct or indirect and may be necessary and/or sufficient for the manifestation of the symptoms of the disease, state or condition (or its aetiology or progression). Thus, cyclin dependent kinase 5 (CDK5) activity (and in particular aberrant levels of cyclin dependent kinase 5 (CDK5) activity, e.g. cyclin dependent kinase 5 (CDK5) over-expression) need not necessarily be the proximal cause of the disease, state or condition: rather, it is contemplated that the CDK5-mediated diseases, states or conditions include those having multifactorial aetiologies and complex progressions in which CDK5. In cases where the term is applied to treatment, prophylaxis or intervention (e.g. in the "CDK5-mediated treatments" of the invention), the role played by CDK5 may be direct or indirect and may be necessary and/or sufficient for the operation of the treatment, prophylaxis or outcome of the intervention.
The term "intervention" is a term of art used herein to define any agency which effects a physiological change at any level. Thus, the intervention may comprise the induction or repression of any physiological process, event, biochemical pathway or cellular/biochemical event. The interventions of the invention typically effect (or contribute to) the therapy, treatment or prophylaxis of a disease or condition.
As used herein, the term "combination", as applied to two or more compounds and/or agents (also referred to herein as the components), is intended to define material in which the two or more compounds/agents are associated. The terms "combined" and "combining" in this context are to be interpreted accordingly.
The association of the two or more compounds/agents in a combination may be physical or non- physical. Examples of physically associated combined compounds/agents include:
• compositions (e.g. unitary formulations) comprising the two or more compounds/agents in admixture (for example within the same unit dose);
• compositions comprising material in which the two or more compounds/agents are chemically/physicochemically linked (for example by crosslinking, molecular agglomeration or binding to a common vehicle moiety);
• compositions comprising material in which the two or more compounds/agents are chemically/physicochemically co-packaged (for example, disposed on or within lipid vesicles, particles (e.g. micro- or nanoparticles) or emulsion droplets);
• pharmaceutical kits, pharmaceutical packs or patient packs in which the two or more compounds/agents are co-packaged or co-presented (e.g. as part of an array of unit doses); Examples of non-physically associated combined compounds/agents include:
• material (e.g. a non-unitary formulation) comprising at least one of the two or more compounds/agents together with instructions for the extemporaneous association of the at least one compound to form a physical association of the two or more compounds/agents;
• material (e.g. a non-unitary formulation) comprising at least one of the two or more compounds/agents together with instructions for combination therapy with the two or more compounds/agents;
• material comprising at least one of the two or more compounds/agents together with instructions for administration to a patient population in which the other(s) of the two or more compounds/agents have been (or are being) administered; • material comprising at least one of the two or more compounds/agents in an amount or in a form which is specifically adapted for use in combination with the other(s) of the two or more compounds/agents.
As used herein, the term "in combination" may refer to compounds/agents that are administered as part of the same overall treatment regimen. As such, the posology of each of the two or more compounds/agents may differ: each may be administered at the same time or at different times. It will therefore be appreciated that the compounds/agents of the combination may be administered sequentially (e.g. before or after) or simultaneously, either in the same pharmaceutical formulation (i.e. together), or in different pharmaceutical formulations (i.e. separately). Simultaneously in the same formulation is as a unitary formulation whereas simultaneously in different pharmaceutical formulations is non-unitary. The posologies of each of the two or more compounds/agents in a combination therapy may also differ with respect to the route of administration.
As used herein, the term "pharmaceutical kit" defines an array of one or more unit doses of a pharmaceutical composition together with dosing means (e.g. measuring device) and/or delivery means (e.g. inhaler or syringe), optionally all contained within common outer packaging. In pharmaceutical kits comprising a combination of two or more compounds/agents, the individual compounds/agents may unitary or non-unitary formulations. The unit dose(s) may be contained within a blister pack. The pharmaceutical kit may optionally further comprise instructions for use.
As used herein, the term "pharmaceutical pack" defines an array of one or more unit doses of a pharmaceutical composition, optionally contained within common outer packaging. In pharmaceutical packs comprising a combination of two or more compounds/agents, the individual compounds/agents may unitary or non-unitary formulations. The unit dose(s) may be contained within a blister pack. The pharmaceutical pack may optionally further comprise instructions for use.
As used herein, the term "patient pack" defines a package, prescribed to a patient, which contains pharmaceutical compositions for the whole course of treatment. Patient packs usually contain one or more blister pack(s). Patient packs have an advantage over traditional prescriptions, where a pharmacist divides a patient's supply of a pharmaceutical from a bulk supply, in that the patient always has access to the package insert contained in the patient pack, normally missing in patient prescriptions. The inclusion of a package insert has been shown to improve patient compliance with the physician's instructions.
The following general preferences and definitions shall apply to each of R1 to R8, R10, Ra, Rb, Rc, X1 and X2 and their various sub-groups, sub-definitions, examples and embodiments unless the context indicates otherwise.
Any references to formula (I) herein shall also be taken to refer to and any sub-group of compounds within formula (I) and any preferences and examples thereof unless the context requires otherwise.
The term "carbon-carbon multiple bonds" as used herein refers to bonds of the type C=C and C≡C. Up to 6 carbon-carbon multiple bonds may be present in the group Rp1 and this number includes any carbon-carbon multiple bonds that may be present in a carbocyclic group. For example, a benzene ring, for the purposes of this application, has three carbon-carbon multiple (double) bonds) whereas a naphthyl group has five carbon-carbon multiple bonds.
The term "glucuronide residue" as used herein is used to denote a group of the formula:
Figure imgf000027_0001
where the asterisk denotes the point of attachment to the oxygen atom of the group O-Rz.
The glucuronide residue is linked by a glycosidic bond to the benzene ring to which the moieties R1, R8, R2a, R3 and R4a are attached. Compounds of the invention containing a glucuronide group may be referred to herein as "glucuronides" or "a glucuronide", "glucuronide derivatives" or " a "glucuronide" derivative".
The glucuronides of the invention can exist in either the α- or β-anomeric forms and can be provided in a solid form in which one anomer predominates or as mixtures of the two anomers.
The term "mono-, di or tri-peptide residue" as used herein refers to an acyl residue of an an amino acid or a dipeptide or tripeptide. Thus, for example, in the case of a monopeptide in which the amino acid is glycine, the mono-peptide residue is the group:
Figure imgf000027_0004
where the asterisk denotes the point of attachment to the oxygen atom of the group O-Rz. In the case of the di-peptide GIy-GIy, the di-peptide residue is the group:
Figure imgf000027_0002
The term "non-terminal carbon atoms" as used herein refers to a carbon atom which (i) has at least two other carbon atoms attached to it or (ii) has at least one other carbon atom attached to it and is also attached directly to the group Lp, the oxygen atom in the group O-Rz (when Lp is a bond) or an
oxygen atom in the group
Figure imgf000027_0003
References to "carbocyclic" and "heterocyclic" groups as used herein shall, unless the context indicates otherwise, include both aromatic and non-aromatic ring systems. Thus, for example, the term "carbocyclic and heterocyclic groups" includes within its scope aromatic, non-aromatic, unsaturated, partially saturated and fully saturated carbocyclic and heterocyclic ring systems. In general, such groups may be monocyclic or bicyclic and may contain, for example, 3 to 12 ring members, more usually 5 to 10 ring members. Examples of monocyclic groups are groups containing 3, 4, 5, 6, 7, and 8 ring members, more usually 3 to 7, for example 5 to 7, and preferably 5 or 6 ring members. Examples of bicyclic groups are those containing 8, 9, 10, 11 and 12 ring members, and more usually 9 or 10 ring members.
The term "bicyclic" as used herein refers to groups that have two rings joined together in such as way that at least one ring member is shared by both rings. Thus, the bicyclic group can be a fused ring (two ring members shared by both rings), spirocyclic (one ring member shared by both rings) or a bridged ring (three or more ring members shared by both rings).
The carbocyclic or heterocyclic groups can be aryl or heteroaryl groups having from 5 to 12 ring members, more usually from 5 to 10 ring members. The term "aryl" as used herein refers to a carbocyclic group having aromatic character and the term "heteroaryl" is used herein to denote a heterocyclic group having aromatic character. The terms "aryl" and "heteroaryl" embrace polycyclic (e.g. bicyclic) ring systems wherein one or more rings are non-aromatic, provided that at least one ring is aromatic. In such polycyclic systems, the group may be attached by the aromatic ring, or by a non-aromatic ring. The aryl or heteroaryl groups can be monocyclic or bicyclic groups and can be unsubstituted or substituted with one or more substituents, for example one or more groups R10 as defined herein.
The term "non-aromatic group" embraces unsaturated ring systems without aromatic character, partially saturated and fully saturated carbocyclic and heterocyclic ring systems. The terms "unsaturated" and "partially saturated" refer to rings wherein the ring structure(s) contains atoms sharing more than one valence bond i.e. the ring contains at least one multiple bond e.g. a C=C, C≡C or N=C bond. The terms "fully saturated" and "saturated" refer to rings where there are no multiple bonds between ring atoms. Saturated carbocyclic groups include cycloalkyl groups as defined below. Partially saturated carbocyclic groups include cycloalkenyl groups as defined below, for example cyclopentenyl, cycloheptenyl and cyclooctenyl. A further example of a cycloalkenyl group is cyclohexenyl. Examples of heteroaryl groups are monocyclic and bicyclic groups containing from five to twelve ring members, and more usually from five to ten ring members. The heteroaryl group can be, for example, a five membered or six membered monocyclic ring or a bicyclic structure formed from fused five and six membered rings or two fused six membered rings or, by way of a further example, two fused five membered rings. Each ring may contain up to about four heteroatoms typically selected from nitrogen, sulphur and oxygen. Typically the heteroaryl ring will contain up to 4 heteroatoms, more typically up to 3 heteroatoms, more usually up to 2, for example a single heteroatom. In one embodiment, the heteroaryl ring contains at least one ring nitrogen atom. The nitrogen atoms in the heteroaryl rings can be basic, as in the case of an imidazole or pyridine, or essentially non-basic as in the case of an indole or pyrrole nitrogen. In general the number of basic nitrogen atoms present in the heteroaryl group, including any amino group substituents of the ring, will be less than five. Examples of five membered heteroaryl groups include but are not limited to pyrrole, furan, thiophene, imidazole, furazan, oxazole, oxadiazole, oxatriazole, isoxazole, thiazole, isothiazole, pyrazole, triazole and tetrazole groups.
Examples of six membered heteroaryl groups include but are not limited to pyridine, pyrazine, pyridazine, pyrimidine and triazine.
A bicyclic heteroaryl group may be, for example, a group selected from: a) a benzene ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; b) a pyridine ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; c) a pyrimidine ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; d) a pyrrole ring fused to a a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; e) a pyrazole ring fused to a a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; f) a pyrazine ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; g) an imidazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; h) an oxazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; i) an isoxazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; j) a thiazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; k) an isothiazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms;
I) a thiophene ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; m) a furan ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; n) a cyclohexyl ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms; and o) a cyclopentyl ring fused to a 5- or 6-membered ring containing 1 , 2 or 3 ring heteroatoms.
One sub-group of bicyclic heteroaryl groups consists of groups (a) to (e) and (g) to (o) above.
Particular examples of bicyclic heteroaryl groups containing a five membered ring fused to another five membered ring include but are not limited to imidazothiazole (e.g. imidazo[2,1 -b]thiazole) and imidazoimidazole (e.g. imidazo[1 ,2-a]imidazole).
Particular examples of bicyclic heteroaryl groups containing a six membered ring fused to a five membered ring include but are not limited to benzfuran, benzthiophene, benzimidazole, benzoxazole, isobenzoxazole, benzisoxazole, benzthiazole, benzisothiazole, isobenzofuran, indole, isoindole, indolizine, indoline, isoindoline, purine (e.g., adenine, guanine), indazole, pyrazolopyrimidine (e.g. pyrazolo[1 ,5-a]pyrimidine), triazolopyrimidine (e.g. [1 ,2,4]triazolo[1 ,5- a]pyrimidine), benzodioxole and pyrazolopyridine (e.g. pyrazolo[1 ,5-a]pyridine) groups.
Particular examples of bicyclic heteroaryl groups containing two fused six membered rings include but are not limited to quinoline, isoquinoline, chroman, thiochroman, chromene, isochromene, chroman, isochroman, benzodioxan, quinolizine, benzoxazine, benzodiazine, pyridopyridine, quinoxaline, quinazoline, cinnoline, phthalazine, naphthyridine and pteridine groups. One sub-group of heteroaryl groups comprises pyridyl, pyrrolyl, furanyl, thienyl, imidazolyl, oxazolyl, oxadiazolyl, oxatriazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, pyrazinyl, pyridazinyl, pyrimidinyl, triazinyl, triazolyl, tetrazolyl, quinolinyl, isoquinolinyl, benzfuranyl, benzthienyl, chromanyl, thiochromanyl, benzimidazolyl, benzoxazolyl, benzisoxazole, benzthiazolyl and benzisothiazole, isobenzofuranyl, indolyl, isoindolyl, indolizinyl, indolinyl, isoindolinyl, purinyl (e.g., adenine, guanine), indazolyl, benzodioxolyl, chromenyl, isochromenyl, isochromanyl, benzodioxanyl, quinolizinyl, benzoxazinyl, benzodiazinyl, pyridopyridinyl, quinoxalinyl, quinazolinyl, cinnolinyl, phthalazinyl, naphthyridinyl and pteridinyl groups.
Examples of polycyclic aryl and heteroaryl groups containing an aromatic ring and a non-aromatic ring include tetrahydronaphthalene, tetrahydroisoquinoline, tetrahydroquinoline, dihydrobenzthiene, dihydrobenzfuran, 2,3-dihydro-benzo[1 ,4]dioxine, benzo[1 ,3]dioxole, 4,5,6,7- tetrahydrobenzofuran, indoline and indane groups.
Examples of carbocyclic aryl groups include phenyl, naphthyl, indenyl, and tetrahydronaphthyl groups. Examples of non-aromatic heterocyclic groups include unsubstituted or substituted (by one or more groups R10) heterocyclic groups having from 3 to 12 ring members, typically 4 to 12 ring members, and more usually from 5 to 10 ring members. Such groups can be monocyclic or bicyclic, for example, and typically have from 1 to 5 heteroatom ring members (more usually 1 ,2,3 or 4 heteroatom ring members) typically selected from nitrogen, oxygen and sulphur. When sulphur is present, it may, where the nature of the adjacent atoms and groups permits, exist as -S-, -S(O)- Or -S(O)2-.
The heterocylic groups can contain, for example, cyclic ether moieties (e.g. as in tetrahydrofuran and dioxane), cyclic thioether moieties (e.g. as in tetrahydrothiophene and dithiane), cyclic amine moieties (e.g. as in pyrrolidine), cyclic amide moieties (e.g. as in pyrrolidone), cyclic thioamides, cyclic thioesters, cyclic ester moieties (e.g. as in butyrolactone), cyclic sulphones (e.g. as in sulpholane and sulpholene), cyclic sulphoxides, cyclic sulphonamides and combinations thereof (e.g. morpholine and thiomorpholine and its S-oxide and S,S-dioxide). Further examples of heterocyclic groups are those containing a cyclic urea moiety (e.g. as in imidazolidin-2-one),
In one sub-set of heterocyclic groups, the heterocyclic groups contain cyclic ether moieties (e.g as in tetrahydrofuran and dioxane), cyclic thioether moieties (e.g. as in tetrahydrothiophene and dithiane), cyclic amine moieties (e.g. as in pyrrolidine), cyclic sulphones (e.g. as in sulpholane and sulpholene), cyclic sulphoxides, cyclic sulphonamides and combinations thereof (e.g. thiomorpholine).
Examples of monocyclic non-aromatic heterocyclic groups include 5-, 6-and 7-membered monocyclic heterocyclic groups. Particular examples include morpholine, piperidine (e.g. 1- piperidinyl, 2-piperidinyl, 3-piperidinyl and 4-piperidinyl), pyrrolidine (e.g. 1-pyrrolidinyl, 2- pyrrolidinyl and 3-pyrrolidinyl), pyrrolidone, pyran (2H-pyran or 4H-pyran), dihydrothiophene, dihydropyran, dihydrofuran, dihydrothiazole, tetrahydrofuran, tetrahydrothiophene, dioxane, tetrahydropyran (e.g. 4-tetrahydro pyranyl), imidazoline, imidazolidinone, oxazoline, thiazoline, 2- pyrazoline, pyrazolidine, piperazine, and N-alkyl piperazines such as N-methyl piperazine. Further examples include thiomorpholine and its S-oxide and S,S-dioxide (particularly thiomorpholine). Still further examples include azetidine, piperidone, piperazone, and N-alkyl piperidines such as N- methyl piperidine.
One preferred sub-set of non-aromatic heterocyclic groups consists of saturated groups such as azetidine, pyrrolidine, piperidine, morpholine, thiomorpholine, thiomorpholine S,S-dioxide, piperazine, N-alkyl piperazines, and N-alkyl piperidines. Another sub-set of non-aromatic heterocyclic groups consists of pyrrolidine, piperidine, morpholine, thiomorpholine, thiomorpholine S,S-dioxide, piperazine and N-alkyl piperazines such as N-methyl piperazine.
One particular sub-set of heterocyclic groups consists of pyrrolidine, piperidine, morpholine and N- alkyl piperazines (e.g. N-methyl piperazine), and optionally thiomorpholine. Examples of non-aromatic carbocyclic groups include cycloalkane groups such as cyclohexyl and cyclopentyl, cycloalkenyl groups such as cyclopentenyl, cyclohexenyl, cycloheptenyl and cyclooctenyl, as well as cyclohexadienyl, cyclooctatetraene, tetrahydronaphthenyl and decalinyl.
Preferred non-aromatic carbocyclic groups are monocyclic rings and most preferably saturated monocyclic rings. Typical examples are three, four, five and six membered saturated carbocyclic rings, e.g. optionally substituted cyclopentyl and cyclohexyl rings.
One sub-set of non-aromatic carboyclic groups includes unsubstituted or substituted (by one or more groups R10) monocyclic groups and particularly saturated monocyclic groups, e.g. cycloalkyl groups. Examples of such cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl; more typically cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl, particularly cyclohexyl.
Further examples of non-aromatic cyclic groups include bridged ring systems such as bicycloalkanes and azabicycloalkanes although such bridged ring systems are generally less preferred. By "bridged ring systems" is meant ring systems in which two rings share more than two atoms, see for example Advanced Organic Chemistry, by Jerry March, 4th Edition, Wiley Interscience, pages 131-133, 1992. Examples of bridged ring systems include bicyclo[2.2.1]heptane, aza-bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, aza-bicyclo[2.2.2]octane, bicyclo[3.2.1]octane and aza-bicyclo[3.2.1]octane. A particular example of a bridged ring system is the 1-aza-bicyclo[2.2.2]octan-3-yl group. Where reference is made herein to carbocyclic and heterocyclic groups, the carbocyclic or heterocyclic ring can, unless the context indicates otherwise, be unsubstituted or substituted by one or more substituent groups R10 selected from halogen, hydroxy, trifluoromethyl, cyano, nitro, carboxy, amino, mono- or di-C1-4 hydrocarbylamino, carbocyclic and heterocyclic groups having from 3 to 12 ring members; a group Ra-Rb wherein Ra is a bond, O, CO, X1C(X2), C(X2)X1, X1C(X2)X1, S, SO, SO2, NRC, SO2NR0 or NR0SO2; and Rb is selected from hydrogen, carbocyclic and heterocyclic groups having from 3 to 12 ring members, and a C1-12 hydrocarbyl group (such as a C1-10 hydrocarbyl group) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C^ non-aromatic hydrocarbylamino (e.g. mono- or di-C1-4 hydrocarbylamino), carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C1-12 hydrocarbyl group (or C1-10 hydrocarbyl group) may optionally be replaced by O, S, SO, SO2, NR0, X1C(X2), C(X2)X1 or X1C(X2)X1;
R° is selected from Rb, hydrogen and C1-4 hydrocarbyl; and
X1 is O, S or NR0 and X2 is =0, =S or =NR°.
Where the substituent group R10 comprises or includes a carbocyclic or heterocyclic group, the said carbocyclic or heterocyclic group may be unsubstituted or may itself be substituted with one or more further substituent groups R10. In one sub-group of compounds of the formula (I), such further substituent groups R10 may include carbocyclic or heterocyclic groups, which are typically not themselves further substituted. In another sub-group of compounds of the formula (I), the said further substituents do not include carbocyclic or heterocyclic groups but are otherwise selected from the groups listed above in the definition of R10.
The substituents R10 may be selected such that they contain no more than 20 non-hydrogen atoms, for example, no more than 15 non-hydrogen atoms, e.g. no more than 12, or 11, or 10, or 9, or 8, or 7, or 6, or 5 non-hydrogen atoms.
Where the carbocyclic and heterocyclic groups have a pair of substituents on the same or adjacent ring atoms, the two substituents may be linked so as to form a cyclic group. Thus, two adjacent groups R10, together with the carbon atoms or heteroatoms to which they are attached may form a 5-membered heteroaryl ring or a 5- or 6-membered non-aromatic carbocyclic or heterocyclic ring, wherein the said heteroaryl and heterocyclic groups contain up to 3 heteroatom ring members selected from N, O and S. For example, an adjacent pair of substituents on adjacent carbon atoms of a ring may be linked via one or more heteroatoms and optionally substituted alkylene groups to form a fused oxa-, dioxa-, aza-, diaza- or oxa-aza-cycloalkyl group.
Examples of such linked substituent groups include:
Figure imgf000032_0001
Figure imgf000033_0001
Examples of halogen substituents include fluorine, chlorine, bromine and iodine. Fluorine and chlorine are particularly preferred.
In the definition of the compounds of the formula (I) above and as used hereinafter, the term "hydrocarbyl" is a generic term encompassing aliphatic, alicyclic and aromatic groups having an all-carbon backbone and consisting of carbon and hydrogen atoms, except where otherwise stated.
In certain cases, as defined herein, one or more of the carbon atoms making up the carbon backbone may be replaced by a specified atom or group of atoms.
Examples of hydrocarbyl groups include alky], cycloalkyl, cycloalkenyl, carbocyclic aryl, alkenyl, alkynyl, cycloalkylalkyl, cycloalkenylalkyl, and carbocyclic aralkyl, aralkenyl and aralkynyl groups. Such groups can be unsubstituted or, where stated, substituted by one or more substituents as defined herein. The examples and preferences expressed below apply to each of the hydrocarbyl substituent groups or hydrocarbyl-containing substituent groups referred to in the various definitions of substituents for compounds of the formula (I) unless the context indicates otherwise. The prefix "Cx-y" (where x and y are integers) as used herein refers to the number of carbon atoms in a given group. Thus, a Ci-4 hydrocarbyl group contains from 1 to 4 carbon atoms, and a C3.s cycloalkyl group contains from 3 to 6 carbon atoms, and so on.
The term "acyclic hydrocarbyl" (e.g. as in "acyclic C1-5 hydrocarbyl") as used herein refers to non- cyclic hydrocarbyl groups and in particular to alkyl, alkenyl and alkynyl groups as defined herein. The term "mono- or CJi-C1-5 hydrocarbylamino" as used herein refers to a monosubstituted or disubsituted amine group bearing either one or two hydrocarbyl substituent groups that each contain from 1 to 5 carbon atoms.
Preferred non-aromatic hydrocarbyl groups are saturated groups such as alkyl and cycloalkyl groups. Generally by way of example, the hydrocarbyl groups can have up to ten carbon atoms (and more typically up to eight carbon atoms), unless the context requires otherwise. Within the sub-set of hydrocarbyl groups having 1 to 10 carbon atoms, particular examples are C1-8 hydrocarbyl groups or C1-6 hydrocarbyl groups, such as Ci-4 hydrocarbyl groups (e.g. C1-3 hydrocarbyl groups or C1-2 hydrocarbyl groups or C2-3 hydrocarbyl groups or C2-4 hydrocarbyl groups), specific examples being any individual value or combination of values selected from C1, C2, C3, C4, C5, C6, C7, C8, C9 and C10 hydrocarbyl groups.
The term "alkyl" covers both straight chain and branched chain alkyl groups. Examples of alkyl groups include methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, 2-pentyl, 3- pentyl, 2-methyl butyl, 3-methyl butyl, and n-hexyl and its isomers. Within the sub-set of alkyl groups having 1 to 8 carbon atoms, particular examples are C1-6 alkyl groups, such as C1-4 alkyl groups (e.g. C1-3 alkyl groups or C1-2 alkyl groups or C2-3 alkyl groups or C2-4 alkyl groups).
Examples of cycloalkyl groups are those derived from cyclopropane, cyclobutane, cyclopentane, cyclohexane and cycloheptane. Within the sub-set of cycloalkyl groups the cycloalkyl group will have from 3 to 10 carbon atoms, more typically 3 to 8 carbon atoms, particular examples being C3. 6 cycloalkyl groups.
Examples of alkenyl groups include, but are not limited to, ethenyl (vinyl), 1-propenyl, 2-propenyl (allyl), isopropenyl, butenyl, buta-1 ,4-dienyl, pentenyl, and hexenyl. Within the sub-set of alkenyl groups the alkenyl group will have 2 to 10 carbon atoms, more typically 2 to 8 carbon atoms, particular examples being C2-6 alkenyl groups, such as C2-4 alkenyl groups.
Examples of cycloalkenyl groups include, but are not limited to, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclopentadienyl and cyclohexenyl. Within the sub-set of cycloalkenyl groups the cycloalkenyl groups have from 3 to 10 carbon atoms, more typically 3 to 8 carbon atoms, and particular examples are C3-6 cycloalkenyl groups.
Examples of alkynyl groups include, but are not limited to, ethynyl and 2-propynyl (propargyl) groups. Within the sub-set of alkynyl groups having 2 to 10 carbon atoms, more typically 2 to 8 carbon atoms, particular examples are C2-6 alkynyl groups, such as C2-4 alkynyl groups.
Examples of carbocyclic aryl groups include substituted and unsubstituted phenyl groups. Examples of cycloalkylalkyl, cycloalkenylalkyl, carbocyclic aralkyl, aralkenyl and aralkynyl groups include phenethyl, benzyl, styryl, phenylethynyl, cyclohexylmethyl, cyclopentylmethyl, cyclobutylmethyl, cyclopropylmethyl and cyclopentenylmethyl groups.
The terms C1-12 hydrocarbyl, C1--I0 hydrocarbyl and C1-8 hydrocarbyl as used herein encompasses alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, phenyl, benzyl and phenylethyl groups wherein the preferences for and examples of each of the aforesaid groups are as defined above. Within this definition, particular hydrocarbyl groups are alkyl, cycloalkyl, phenyl, benzyl and phenylethyl (e.g. 1-phenylethyl or 2-phenylethyl) groups, one subset of hydrocarbyl groups consisting of alkyl and cycloalkyl groups and in particular C1-4 alkyl and cycloalkyl groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, cyclopropyl and cyclobutyl. The term C1-S hydrocarbyl as used herein encompasses alkyl, alkenyl, alkynyl, cycloalkyl and cycloalkenyl groups wherein the preferences for and examples of the aforesaid groups are as defined above. Within this definition, particular Ci-5 hydrocarbyl groups are saturated C1-5 hydrocarbyl groups, namely alkyl and cycloalkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, 3-methylbutyl, 2-methylbutyl, pent-2-yl, pent-3-yl, 3-methylbut- 2-yl, 2,2-dimethylpropyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclopropylmethyl, cyclopropylethyl, cyclobutylmethyl, methylcyclopropyl, ethylcyclopropyl, dimethylcyclopropyl and methylcyclobutyl. Also within the scope of the term C1-5 hydrocarbyl as used herein are unsaturated C1-5 hydrocarbyl groups, i.e. alkene, cycloalkene and alkyne groups such as vinyl, 1-propenyl, isopropenyl, 1- butenyl, 2-butenyl, 1-pentenyl, 2-pentenyl, allyl, methylallyl, dimethylallyl, acetylenyl, propargyl, cyclobutenyl and cyclopentenyl. The term C1-4 hydrocarbyl as used herein encompasses alky], alkenyl, alkynyl, cycloalkyl and cycloalkenyl groups wherein the preferences for and examples of the aforesaid groups are as defined above. Within this definition, particular C1-4 hydrocarbyl groups are alkyl and cycloalkyl groups, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, cyclopropyl and cyclobutyl. When present, and where stated, a hydrocarbyl group can be optionally substituted by one or more substituents selected from hydroxy, oxo, alkoxy, carboxy, halogen, cyano, nitro, amino, mono- or di-C1-4 hydrocarbylamino, and monocyclic or bicyclic carbocyclic and heterocyclic groups having from 3 to 12 (typically 3 to 10 and more usually 5 to 10) ring members. Preferred substituents include halogen such as fluorine. Thus, for example, the substituted hydrocarbyl group can be a partially fluorinated or perfluorinated group such as difluoromethyl or trifluoromethyl. In one embodiment preferred substituents include monocyclic carbocyclic and heterocyclic groups having 3-7 ring members, more usually 3, 4, 5 or 6 ring members.
Where stated, one or more carbon atoms of a hydrocarbyl group may optionally be replaced by O, S, SO, SO2, NRC, X1C(X2), C(X2)X1 or X1C(X2)X1 (or a sub-group thereof) wherein X1 and X2 are as hereinbefore defined, provided that at least one carbon atom of the hydrocarbyl group remains. For example, 1 , 2, 3 or 4 carbon atoms of the hydrocarbyl group may be replaced by one of the atoms or groups listed, and the replacing atoms or groups may be the same or different. In general, the number of linear or backbone carbon atoms replaced will correspond to the number of linear or backbone atoms in the group replacing them. Examples of groups in which one or more carbon atom of the hydrocarbyl group have been replaced by a replacement atom or group as defined above include ethers and thioethers (C replaced by O or S), amides, esters, thioamides and thioesters (C-C replaced by X1C(X2) or C(X2)X1), sulphones and sulphoxides (C replaced by SO or SO2), amines (C replaced by NR0). Further examples include ureas, carbonates and carbamates (C-C-C replaced by X1C(X2)X1). Where an amino group has two hydrocarbyl substituents, they may, together with the nitrogen atom to which they are attached, and optionally with another heteroatom such as nitrogen, sulphur, or oxygen, link to form a ring structure of 4 to 7 ring members, more usually 5 to 6 ring members.
The term "aza-cycloalkyl" as used herein refers to a cycloalkyl group in which one of the carbon ring members has been replaced by a nitrogen atom. Thus examples of aza-cycloalkyl groups include piperidine and pyrrolidine. The term "oxa-cycloalkyl" as used herein refers to a cycloalkyl group in which one of the carbon ring members has been replaced by an oxygen atom. Thus examples of oxa-cycloalkyl groups include tetrahydrofuran and tetrahydropyran. In an analogous manner, the terms "diaza-cycloalkyl", "dioxa-cycloalkyl" and "aza-oxa-cycloalkyl" refer respectively to cycloalkyl groups in which two carbon ring members have been replaced by two nitrogen atoms, or by two oxygen atoms, or by one nitrogen atom and one oxygen atom. Thus, in an oxa-C4.6 cycloalkyl group, there will be from 3 to 5 carbon ring members and an oxygen ring member. For example, an oxa-cyclohexyl group is a tetrahydropyranyl group. The definition "Ra-Rb" as used herein, either with regard to substituents present on a carbocyclic or heterocyclic moiety, or with regard to other substituents present at other locations on the compounds of the formula (I), includes inter alia compounds wherein Ra is selected from a bond, O, CO, OC(O), SC(O), NR0C(O), OC(S), SC(S), NR0C(S), OC(NR0), SC(NR0), NR0C(NR0), C(O)O, C(O)S, C(O)NR0, C(S)O, C(S)S, C(S) NR0, C(NR°)0, C(NR°)S, C(NR°)NR°, OC(O)O, SC(O)O, NR0C(O)O, OC(S)O, SC(S)O, NR0C(S)O, OC(NR°)O, SC(NR°)O, NR°C(NR°)O, OC(O)S, SC(O)S, NR0C(O)S, OC(S)S, SC(S)S, NR0C(S)S, OC(NR°)S, SC(NR°)S, NR°C(NR°)S, OC(O)NR0, SC(O)NR0, NR0C(O) NR0, OC(S)NR0, SC(S) NR0, NR0C(S)NR0, OC(NR°)NR°, SC(NR°)NR°, NR0C(NR0NR0, S, SO, SO2 , NR0, SO2NR0 and NR0SO2 wherein R° is as hereinbefore defined.
The moiety Rb can be hydrogen or it can be a group selected from carbocyclic and heterocyclic groups having from 3 to 12 ring members (typically 3 to 10 and more usually from 5 to 10), and a C-I-8 hydrocarbyl group optionally substituted as hereinbefore defined. Examples of hydrocarbyl, carbocyclic and heterocyclic groups are as set out above.
When Ra is O and Rb is a C1--I0 hydrocarbyl group, Ra and Rb together form a hydrocarbyloxy group. Preferred hydrocarbyloxy groups include saturated hydrocarbyloxy such as alkoxy (e.g. C1- 6 alkoxy, more usually C1-4 alkoxy such as ethoxy and methoxy, particularly methoxy), cycloalkoxy (e.g. C3-6 cycloalkoxy such as cyclopropyloxy, cyclobutyloxy, cyclopentyloxy and cyclohexyloxy) and cycloalkylalkoxy (e.g. C3-6 cycloalkyl-C1-2 alkoxy such as cyclopropylmethoxy).
The hydrocarbyloxy groups can be substituted by various substituents as defined herein. For example, the alkoxy groups can be substituted by halogen (e.g. as in difluoromethoxy and trifluoromethoxy), hydroxy (e.g. as in hydroxyethoxy), C-I-2 alkoxy (e.g. as in methoxyethoxy), hydroxy-C1-2 alkyl (as in hydroxyethoxyethoxy) or a cyclic group (e.g. a cycloalkyl group or non- aromatic heterocyclic group as hereinbefore defined). Examples of alkoxy groups bearing a non- aromatic heterocyclic group as a substituent are those in which the heterocyclic group is a saturated cyclic amine such as morpholine, piperidine, pyrrolidine, piperazine, C1.4-alkyl- piperazines, C3-7-cycloalkyl-piperazines, tetrahydropyran or tetrahydrofuran and the alkoxy group is a C-I-4 alkoxy group, more typically a C-I-3 alkoxy group such as methoxy, ethoxy or n-propoxy.
Alkoxy groups may be substituted by a monocyclic group such as pyrrolidine, piperidine, morpholine and piperazine and N-substituted derivatives thereof such as N-benzyl, N-C1-4 acyl and N-C1-4 alkoxycarbonyl. Particular examples include pyrrolidinoethoxy, piperidinoethoxy and piperazinoethoxy.
When Ra is a bond and Rb is a C1-10 hydrocarbyl group, examples of hydrocarbyl groups Ra-Rb are as hereinbefore defined. The hydrocarbyl groups may be saturated groups such as cycloalkyl and alkyl and particular examples of such groups include methyl, ethyl and cyclopropyl. The hydrocarbyl (e.g. alkyl) groups can be substituted by various groups and atoms as defined herein. Examples of substituted alkyl groups include alkyl groups substituted by one or more halogen atoms such as fluorine and chlorine (particular examples including bromoethyl, chloroethyl and trifluoromethyl), or hydroxy (e.g. hydroxymethyl and hydroxyethyl), C1-I0 acyloxy (e.g. acetoxymethyl and benzyloxymethyl), amino and mono- and dialkylamino (e.g. aminoethyl, methylaminoethyl, dimethylaminomethyl, dimethylaminoethyl and ferf-butylaminomethyl), alkoxy (e.g. C1-2 alkoxy such as methoxy - as in methoxyethyl), and cyclic groups such as cycloalkyl groups, aryl groups, heteroaryl groups and non-aromatic heterocyclic groups as hereinbefore defined).
Particular examples of alkyl groups substituted by a cyclic group are those wherein the cyclic group is a saturated cyclic amine such as morpholine, piperidine, pyrrolidine, piperazine, C^-alkyl- piperazines, C3.7-cycloalkyl-piperazines, tetrahydropyran or tetrahydrofuran and the alkyl group is a C1-4 alkyl group, more typically a C1-3 alkyl group such as methyl, ethyl or n-propyl. Specific examples of alkyl groups substituted by a cyclic group include pyrrolidinomethyl, pyrrolidinopropyl, morpholinomethyl, morpholinoethyl, morpholinopropyl, piperidinylmethyl, piperazinomethyl and N- substituted forms thereof as defined herein.
Particular examples of alkyl groups substituted by aryl groups and heteroaryl groups include benzyl and pyridylmethyl groups. When Ra is SO2NR0, Rb can be, for example, hydrogen or an optionally substituted C1-8 hydrocarbyl group, or a carbocyclic or heterocyclic group. Examples of Ra-Rb where Ra is SO2NR0 include aminosulphonyl, C-^ alkylaminosulphonyl and di-C1-4 alkylaminosulphonyl groups, and sulphonamides formed from a cyclic amino group such as piperidine, morpholine, pyrrolidine, or an optionally N-substituted piperazine such as N-methyl piperazine. Examples of groups Ra-Rb where Ra is SO2 include alkylsulphonyl, heteroarylsulphonyl and arylsulphonyl groups, particularly monocyclic aryl and heteroaryl sulphonyl groups. Particular examples include methylsulphonyl, phenylsulphonyl and toluenesulphonyl.
When Ra is NR0, Rb can be, for example, hydrogen or an optionally substituted C1-I0 hydrocarbyl group, or a carbocyclic or heterocyclic group. Examples of Ra-Rb where Ra is NR0 include amino, C1-4 alkylamino (e.g. methylamino, ethylamino, propylamino, isopropylamino, fe/f-butylamino), di- C1-4 alkylamino (e.g. dimethylamino and diethylamino) and cycloalkylamino (e.g. cyclopropylamino, cyclopentylamino and cyclohexylamino).
Specific Embodiments of and Preferences for R1 to R10 R1 & R2/2a In formulae (Vl0) and (Vl), R1 is hydrogen, hydroxy, or O-Rz; and R2a is hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2a is O-Rz. In formulae (I0) and (I), R1 is hydrogen, hydroxy, or O-Rz; and R2 is hydrogen, hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2 is O-Rz.
In formulae (Vl0), (Vl) and (I0) and sub-groups thereof, it is preferred that R1 is hydroxy or O-Rz and R2a is hydroxy or O-R1p; provided that at least one of R1 and R2a is O-Rz.
In one embodiment, both R1 and R2a are O-Rz.
In another embodiment, both R1 and R2 are O-Rz.
In formulae (Vl0), (Vl), (I0) and (I) and sub-groups thereof, Rz is Lp-Rp1; SO3H; a glucuronidev residue; a mono-, di- or tripeptide residue; or
Figure imgf000038_0001
Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or S(O)xNRp1; x is 1 or 2;
Rp1 is hydrogen or a C1-2S hydrocarbyl group containing O, 1 or 2 carbocyclic rings and 0, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds, the C1-2S hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-C1-6 alkylamino, C1-6 acylamino, halogen or C1-4 alkoxy or C1-4 alkyl groups wherein the C1-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 3 non-terminal carbon atoms in the C1-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O); provided that Rp1 is not hydrogen when Lp is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an 0-0 moiety; and excluding compounds wherein R1 is hydroxy and R2a is methoxy;
Rp2 and Rp3 are the same or different and each is a group Rp1.
In formula (I), Rz is as defined above in respect of (Vl0), (Vl) and (I0), except that the formula excludes compounds wherein Lp is a bond and Rp1 is an unsubstituted benzyl group or a methyl group. Compounds wherein Rp1 is an unsubstituted benzyl group or a methyl group are disclosed as synthetic intermediates in our earlier application number PCT/GB2006/001382.
In one embodiment, Rp1 is hydrogen or an optionally substituted C1-25 hydrocarbyl group as defined herein except that the optional substituents for the C1-25 hydrocarbyl group do not include hydroxy.
The term "hydrocarbyl" as used in the context of formulae (Vl0) (Vl), (I0) and (I) and sub-groups thereof is as defined in the General Preferences and Definitions section above. Thus, the term covers inter alia alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, cycloalkylalkenyl, cycloalkylalkynyl, cycloalkenyl, cycloalkenylalkyl, cycloalkenylalkenyl, cycloalkenylalkynyl, phenyl, phenylalkyl, phenylalkenyl, phenylalkynyl, naphthyl, naphthylalkyl, naphthylalkenylyl, naphthylalkynyl, indanyl, indanylalkyl, indanylalkenyl and indanylalkynyl groups. Where a carbocyclic group is present in the G1-25 hydrocarbyl group Rp1, the carbocyclic group is typically a cycloalkyl group or a phenyl or naphthyl group. The phenyl or naphthyl groups are typically optionally substituted by relatively lipophilic groups such as halogen or C1-4 alkoxy or C1-4 alkyl groups wherein the C1-4 alkoxy or C14 alkyl groups are each optionally substituted by two or more fluorine atoms. Alternatively or additionally, the phenyl or naphthyl groups may be substituted by hydroxy. Particular examples of C1-2S hydrocarbyl groups Rp1 containing a carbocyclic group are cycloalkyl, cycloalkylalkyl and phenylalkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl, cyclohexylmethyl, benzyl and phenylethyl groups.
When Rp1 is a Ci-2S hydrocarbyl group, it can contain 0, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds. The carbon-carbon multiple bonds can be double bonds or triple bonds or a combination of double and triple bonds. When the C1-25 hydrocarbyl group Rp1 is acyclic, i.e. it contains no carbocyclic groups, it can take the form of a short chain (up to 8 carbon atoms) alkyl, alkenyl or alkynyl group, or it can take the form of a longer chain fatty acid residue (8-25 carbon atoms) which may be saturated or unsaturated.
Up to 3 non-adjacent non-terminal carbon atoms in the C1-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O). Thus the group Rp1 can contain one or more ether linkages and/or an ester linkage.
In one sub-group of compounds, Rp1 is hydrogen or a C1-2S hydrocarbyl group containing 0 or 1 carbocyclic rings and 0, 1, 2 or 3 carbon-carbon multiple bonds, the C1-25 hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-Ci.6 alkylamino, C1-6 acylamino, halogen or Ci-4 alkoxy or C1-4 alkyl groups wherein the C1-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 2 non-adjacent non-terminal carbon atoms in the C1-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O).
In another sub-group of compounds, Rp1 is hydrogen or a C1-8 hydrocarbyl group containing 0 or 1 carbocyclic rings and 0, 1 , 2 or 3 carbon-carbon multiple bonds, the C-J-8 hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-C1-6 alkylamino, C1-6 acylamino, halogen or C-I-4 alkoxy or C1-4 alkyl groups wherein the C1-4 alkoxy or Ci-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 2 non-adjacent non-terminal carbon atoms in the Ci-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O). Within this sub-group, one sub-set of compounds consists of the compounds wherein Rp1 is hydrogen, a C1^ alkyl group, a C3.6 cycloalkyl group or a benzyl group wherein the phenyl and benzyl groups are optionally substituted by halogen or Ci-4 alkoxy or Ci-4 alkyl groups wherein the C1-4 alkoxy or C1.4 alkyl groups are each optionally substituted by two or more fluorine atoms. In another sub-group of compounds, Rp1 is a C1-10 alkyl group wherein up to 2 non-adjacent nonterminal carbon atoms thereof may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms thereof may optionally be replaced by C(=O)O or 0(C=O). Within this sub-group of compounds, Rp1 may be a C1-6 alkyl group.
In formulae (Vl0), (Vl), (I) and (I0) and sub-groups thereof, Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or SO2NRp1.
In one embodiment, Lp is a bond. Within this embodiment, one group of compounds is the group wherein Rp1 is a C1-4 alkyl group or a C1-6 alkanoyloxy-Ci-2-alkyl group. In one sub-group, Rp1 may be other than a methyl group. The C1-6 alkanoyloxy-C1-2-alkyl group can be a C1-6 alkanoyloxymethyl group or a C1-6 alkanoyloxyethyl group. The C1-6 alkanoyloxy moiety can be, for example, acetoxy, propanoyloxy, π-butanoyloxy, 2-methylpropanoyloxy, 3-methylbutanoyloxy and pivaloyloxy.
In another embodiment, l_p is C=O. Within this embodiment, Rp1 is C1-6 alkyl optionally substituted by mono- or di-C1-s alkylamino or a C1-6 acylamino group. For example, Rp1 can be C1-4 alkyl (e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or te/t-butyl group) optionally substituted by mono- or di-C1-2 alkylamino or a C1-4 acylamino group.
In another embodiment, Lp is (C=O)O. Within this embodiment, Rp1 is preferably a C1-6 alkyl group, e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or te/t-butyl group.
In another embodiment, Lp is (C=O)NRp1 and hence the moiety Lp-NRp1 is a group (C=O)N(Rp1)2. The two groups Rp1 can be the same or different and each can be, for example, hydrogen or a C1-6 alkyl group (e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or fert-butyl group) or a C3-6 cyclic group such as cyclopropyl, cyclobutyl, cyclopentyl, cylcohexyl or phenyl group. In one preferred group of compounds, both Rp1 groups are Ci-4 alkyl groups such as methyl and the moiety Lp-NRp1 is, for example, (C=O)NMe2.
In a further embodiment, Lp is SOmNRp1 and hence the moiety Lp-NRp1 is a group SOmN(Rp1)2.
The two groups Rp1 can be the same or different and each can be, for example, hydrogen or a C1-6 alkyl group (e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or fe/f-butyl group) or a C3-6 cyclic group such as cyclopropyl, cyclobutyl, cyclopentyl, cylcohexyl or phenyl group. In one preferred group of compounds, both Rp1 groups are C1-4 alkyl groups such as methyl and the moiety Lp-NRp1 is, for example, SOp1NMe2. The integer "m" can be 1 or 2. In one preferred subgroup of compounds, m is 2.
In another embodiment, Rz is SO3H. Within this embodiment are the compounds wherein (i) R1 is OSO3H and R2a is hydroxy; (ii) (i) R2a is OSO3H and R1 is hydroxy; and (iii) R1 and R2a are both OSO3H. In another embodiment, Rz is:
Figure imgf000041_0001
wherein Rp2 and Rp3 are the same or different and each is a group Rp1. Within this embodiment, particular examples of Rp1 are hydrogen and C1-4 alkyl groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl and fert-butyl groups. In one sub-group of compounds, Rp2 and Rp3 are both hydrogen. In another sub-group of compounds, Rp2 is hydrogen and Rp3 is methyl, ethyl, n- propyl, isopropyl, n-butyl, isobutyl or ferf-butyl, In another sub-group of compounds, Rp2 and Rp3 are both independently selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl and tert- butyl groups.
In another embodiment, at least one of R1 and R2a is a group O-Rz where Rz is a glucuronide residue as defined herein. In one sub-group of compounds, R1 is hydroxy and R2a is a group O-Rz where Rz is a glucuronide residue. In another sub-group of compounds, R2a is hydroxy and R1 is a group O-Rz where Rz is a glucuronide residue.
In a further embodiment, Rz is a mono-, di- or tripeptide residue as defined herein.
The amino acids making up the mono-, di- and tri-peptide residues can be any of the naturally occurring amino acids. Thus, the amino acids may be selected from alanine (Ala, A), valine (VaI, V), leucine (Leu, L), lsoleucine (He, I), proline (Pro, P), phenylalanine (Phe, F), tryptophan (T rp, W), methionine (Met, M), glycine (GIy, G), serine (Ser, S), threonine (Thr, T), cysteine (Cys, C), tyrosine (Tyr, Y), asparagine (Asn, N), glutamine (GIn, Q), aspartic acid (Asp, D), glutamic acid (GIu, E), lysine (Lys, K), arginine (Arg, R) and histidine (His, H).
The naturally occurring amino acids exist in the L-isomeric form. However, the amino acids making up the mono-, di or tri-peptide residues in the compounds of the invention also include the D- isomeric forms of the naturally occurring amino acids as well a the L-isomeric forms.
Examples of mono-peptide pro-drugs are the compounds wherein Rz is an L-GIy, L-AIa or L-Lys or L-VaI residue.
Compounds wherein Rz is a di- or tripeptide residue may be constructed so that they act as substrates for the intestinal human peptide transporter (hPEPTI) (see Anand et al, The Journal Of Pharmacology And Experimental Therapeutics, vol 304, pp 781-791 (2003) and references therein), thereby facilating their absorption through the intestinal wall. For example, Rz can be a di- or tripeptide residue selected from VaI-VaI, Tyr-Gly, GIy-VaI, GIy-GIy, Gly-Tyr, Val-Tyr, Tyr-Val, D-GIu-AIa and D-Asp-Ala.
Tri-peptide prodrugs may be constructed so that they are activated by the protease plasminogen activator found in elevated concentrations in many types of malignant cells and human tumours (see PNAS (1980) vol. 77, no. 4, pp 2224-2228). One such pro-drug is formed with the tri-peptide D-Val-Leu-Lys wherein the lysine moiety is bonded to the oxygen atom of the group O-Rz.
R is selected from hydrogen and fluorine. Preferably R is hydrogen. tf
R3 is selected from hydrogen, halogen, cyano, C1-S hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and Ci-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members. In one sub-group of compounds, R3 is selected from hydrogen, halogen, cyano, C1-5 hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl moiety in each instance is optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy and amino.
In another sub-group of compounds, R3 is selected from halogen (e.g. chlorine or bromine), C1-5 alkyl and C3-4 cycloalkyl. More typically, R3 is selected from hydrogen, chlorine, C1-5 hydrocarbyl and C1-5 hydrocarbyloxy.
Particular groups R3 include hydrogen, C1-5 alkyl, C2-5 alkenyl and C3-4 cycloalkyl groups, preferably secondary alkyl and alkenyl groups such as isopropyl, sec-butyl, fe/t-butyl, 1 ,2-dimethylallyl and 1 ,2-dimethylpropyl, or cycloalkyl groups such as cyclopropyl.
A further sub-group of substituents R3 consists of C1-5 alkyl, C2-5 alkenyl and C3.4 cycloalkyl groups, preferably secondary alkyl and alkenyl groups such as isopropyl, sec-butyl, fe/t-butyl, 1 ,2- dimethylallyl and 1 ,2-dimethylpropyl, or cycloalkyl groups such as cyclopropyl.
When only one of R1 and R2 is hydroxy, R3 may be other than hydrogen. In one particular embodiment, R1 and R2 are both hydroxy and R3 is hydrogen. In a further particular embodiment, R3 is selected from isopropyl and te/t-butyl. In one general embodiment, R3 is other than halogen.
In another general embodiment, R3 may be other than fluorine.
In a further general embodiment, R3 may be other than fluorine or methoxy.
In one embodiment, R4 is selected from hydrogen; a group -(O)n-R7 where n is 0 or 1 and R7 is an acyclic C1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C1-5 hydrocarbylamino, wherein the acyclic C1-5 hydrocarbyl group and the mono and CU-C1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-Ci-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members.
In one sub-group of compounds, R4 is selected from hydrogen; a group -(O)n-R7 where n is 0 or 1 and R7 is an acyclic Ci-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C1-5 hydrocarbylamino, wherein the Ci-5 hydrocarbyl moiety in each instance is optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy and amino.
Within this sub-group, R4 is more typically selected from hydrogen, methoxy, halogen (e.g. fluorine or chlorine), cyano; hydroxy; amino and C3-6 cycloalkyl. More particularly, R4 can be selected from a sub-set R4a wherein the sub-set R4a consists of hydrogen, methoxy, fluorine and chlorine.
Preferably R4 is hydrogen.
In another embodiment, R3 and R4 together form a carbocyclic or heterocyclic ring of 5 to 7 ring members. The carbocyclic and heterocyclic groups can be any of the groups listed above in General Definitions and Preferences section but one particular group is a group wherein R3 and R4 together with the phenyl ring form a dihydrobenzofuran group.
Particular examples of the phenyl ring containing the moieties R1, R2, R3 and R4 are as set out in Table 1. The point of attachment to the carbonyl group is indicated by means of an asterisk.
Table 1
Figure imgf000043_0001
Figure imgf000044_0001
In one embodiment, the phenyl moiety is selected from groups A1 to A21.
In another embodiment, the phenyl moiety is selected from groups A1 to A18.
Preferred phenyl moieties include groups A5, A7, A11 , A13, A14, A15, A16, A17 and A18.
Particularly preferred phenyl moieties are A5, A7, A13, A14 and A17.
Particularly preferred phenyl moieties are A11 and A13.
One particularly preferred phenyl moiety is group A13.
In many of the structures shown in Table 1 , two O-Rz groups are present. It is to be understood, however, that instead of two O-Rz groups, each of the structures shown may have one O-Rz group and one hydroxy group.
R5 & R6 Rs and R6 together with the nitrogen atom to which they are attached form a bicyclic heterocyclic group having up to 12 ring members of which up to 5 ring members are heteroatoms selected from oxygen, nitrogen and sulphur.
The bicyclic groups can be any of the groups listed above in the General Preferences and Definitions section or listed below in the Particular and Preferred Sub-groups section, and such groups may be unsubstituted or substituted by one or more substituents R10 as defined herein.
The bicyclic heterocyclic group is typically a fused ring bicyclic group or a spirocylic group and more typically is a fused ring bicyclic group. Particular fused ring systems of interest in the context of the invention are 5.6 and 6.6 fused ring systems. In the bicyclic heterocyclic groups, one of the rings may be a heterocyclic ring and the other may be a carbocyclic ring, or both rings may be heterocyclic.
In one sub-group of compounds, one of the rings of the bicyclic heterocyclic group is non-aromatic and other is aromatic. Preferably the nitrogen atom of the group NR5R6 forms part of the non- aromatic ring. Particular examples of such groups are dihydroindole, dihydroisoindole, tetrahydroquinoline and tetrahydroisoquinoline groups.
More particular examples of such groups are dihydroindole, dihydroisoindole, tetrahydroquinoline and tetrahydroisoquinoline groups, but wherein the tetrahydroisoquinoline group bears no substituent groups on the non-aromatic ring thereof.
The bicyclic heterocyclic rings are optionally substituted by one or more substituent groups R10 as defined herein.
In one embodiment, the bicyclic heterocyclic ring is subsitituted by 1 , 2 or 3 substituent groups R10 as defined herein.
In another embodiment, the bicyclic heterocyclic ring is subsitituted by 1 or 2 substituent groups R10 as defined herein. The substituent group or groups R10 may be attached to either or both of the two rings making up the bicyclic heterocyclic group. In one embodiment, the ring containing the nitrogen atom of the group NR5R6 does not bear any substituents R10. In another embodiment, the ring containing the nitrogen atom of the group NR5R6 bears a substituent R10 but the substituent is other than a carboxylic acid group. In one sub-group of compounds, the bicyclic heterocyclic group is unsubstituted or is substituted by one, two or three (preferably one or two) substituents selected from a group R1Oa consisting of halogen, hydroxy, amino and a group Ra-Rb where Ra is selected from a bond, O, CO, C(O)O, C(O)NR0, NRCC(O), NR0C(O)O, NR0, SO, SO2, SONR0, and SO2NR0; and Rb is selected from hydrogen; carbocyclic and heterocyclic groups having 5 or 6 ring members; and Ci-10 hydrocarbyl (e.g. C1-8 hydrocarbyl such as C1-8 alkyl or C3.7 cycloalkyl) optionally substituted by one or more substituents selected from hydroxy, oxo, amino, mono- or CJi-C1^8 non-aromatic hydrocarbylamino, (e.g. mono- or di-C1-4 hydrocarbylamino), carboxy, and carbocyclic and heterocyclic groups having from 3 to 7 ring members, and wherein one or more of the carbon atoms of the C1-8 hydrocarbyl group may optionally be replaced by O, S, C(O)O, C(O)NR0 or NR0.
Within this sub-group of compounds and sub-groups, preferences and examples thereof, where it is stated that one or more of the carbon atoms of the C1-8 hydrocarbyl group may optionally be replaced by O, S, C(O)O, C(O)NR0 or NR0, the orientation of the ester and amide groups may be in either direction unless indicated to the contrary.
In the above sub-groups, when Rb is a carbocyclic or heterocyclic group, the carbocyclic or heterocyclic group may be substituted by one or more substituents R10 as defined herein. For example, when Rb is a carbocyclic or heterocyclic group, the carbocyclic or heterocyclic group may be substituted by one or more substituents selected from CO2R14 wherein R14 is hydrogen or C1-6 alky!;
C1-4 alkyl optionally substituted by hydroxy or C1-2 alkoxy;
C1-4 alkoxy optionally substituted by hydroxy or Ci-2 alkoxy; or a group [sol], CH2[SoI], C(0)[sol], OCH2CH2[SOl] or OCH2CH2CH2[SoI] where [sol] is as defined below.
In a more particular sub-group, the bicyclic heterocyclic group is unsubstituted or is substituted by one, two or three (preferably one or two) substituents selected from a group R10b consisting of halogen, OH, NH2, CH2OH, CH2NH2, 0-C1-6-alkyl, NH-C1-6 alkyl, aryl, heteroaryl, C3-7 cycloalkyl, heterocyclyl, O-heteroaryl, 0-C3-7 cycloalkyl, O-heterocycloalkyl, C(O)C1-6 alkyl, C(=0)0C1.6 alkyl, C(=O)NH2, C(=O)NHC1.6 alkyl, C(=O)N(C1-6alkyl)2, NH(C1-6 alkyl), N(C1-6 alkyl)2, NC(O)C1-6 alkyl, C6 aryl, OC6 aryl, C(O)C6aryl, C(O)0C6aryl, C(O)NH2, C(O)NHC6aryl, C(O)N(C6 aryl)2, NH(C6 aryl), N(C6 aryl)2, NC(O)C6 aryl, C5-6 heterocyclyl, OC5-6 heterocyclyl, C(O)C5-6 heterocyclyl, C(O)OC5-6 heterocyclyl, C(O)NHC5-6 heterocyclyl, C(O)N(C5-6 heterocyclyl)2, NH(C5-6 heterocyclyl), N(C5-6 heterocyclyl)2, NC(O)C5-6 heterocyclyl, C(O)NHC1-6 alkyl, C5-6 aryl, S(O)C1-6 alkyl, S(O)N-C1-6 alkyl and SO2N-C1-6 alkyl; and a group [sol], CH2[SoI] or OCH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000046_0001
Figure imgf000047_0001
In another sub-group of compounds, the bicyclic ring is unsubstituted or is substituted by 1, 2 or 3 (e.g. 1 or 2, for example 1) groups R100 where R10° is a group [sol], CH2[SoI] or OCH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000047_0002
and wherein (i) R1Qc is optionally further selected from a group OCH2CH2CH2[SoI] and/or (ii) [sol] is further selected from NHR11 wherein R11 is COR12 or R12 and R12 is C1-4 alkyl, aryl or aryl-C-,-4 alkyl.
In another sub-group of compounds, the bicyclic ring is unsubstituted or is substituted by one or two substituents R1Occ where R1Occ is selected from: halogen;
CO2R14 wherein R14 is hydrogen or C1-5 alkyl;
C1-4 alkyl optionally substituted by hydroxy or C1-2 alkoxy;
C1-4 alkoxy optionally substituted by hydroxy or C1-2 alkoxy; or a group [sol], CH2[sol], C(O)[SoI]1 OCH2CH2[SoI] or OCH2CH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000048_0001
wherein X4 is NH or O, m is O or 1 , n is 1 , 2 or 3, R11 is hydrogen, COR12, C(O)OR12 or R12; R12 is C1-6 alkyl, C3-6 cycloalkyl, aryl, aryl-C1-6 alkyl or CH2R15; and R15 is selected from hydrogen, C1-6 alkyl, C3-6 cycloalkyl, hydroxy-Ci-6 alkyl, piperidine, N-C1-6 alkylpiperazine, piperazine, morpholine,
COR13 or C(O)OR ,1I3Λ; and R 1'30 is C1-6 alkyl.
In a further sub-group of compounds, the bicyclic ring is unsubstituted or is substituted by one or two substituents R1Occc where R10occ is selected from: a group [sol] or CH2[sol] where [sol] is selected from the following groups:
Figure imgf000048_0002
wherein X4 is NH or O, m is O or 1 , n is 1 , 2 or 3, R 1111 : i-s hydrogen, COR i12 , C(O)OR1" or R j1J2Z.; D R112^ is
C1-6 alkyl, C3-6 cycloalkyl, aryl, aryl-C1-6 alkyl or CH2R j15. -,15 ■ ; and R is selected from hydrogen, C1-6 alky], C3-6 cycloalkyl, hydroxy-C^ alkyl, piperidine, N-C1-6 alkylpiperazine, piperazine, morpholine, COR13 or C(O)OR13; and R13 is C1-6 alkyl.
In another sub-group of compounds, where R10b or R10c or R10cc is a group [sol], CH2[SoI], OCH2CH2[SoI] or OCH2CH2CH2[SoI] and [sol] contains a primary or secondary amine group, the primary or secondary amine group can be derivatised to form an acyl derivative such as an amide, carbamate or urea. For example, the amine group can be derivatised to form a carbamate such as a C^alkyloxycarbonylamino group, or a benzyloxycarbonylamino group.
In one sub-group of compounds, R5 and R6 together with the nitrogen atom to which they are attached form an optionally substituted dihydroisoindole group wherein the optional substituents aarree sseelleecctteedd ffrroom groups R10, R1Oa, R1Ob, R1Ocand R1Occ and sub-groups and examples thereof as defined herein.
Particular examples of the group NR5R6 are shown in Table 2. The point of attachment to the carbonyl group is shown by means of an asterisk. Table 2
Figure imgf000049_0001
Figure imgf000050_0001
Figure imgf000051_0001
Figure imgf000052_0002
One set of preferred groups NR5R6 consists of or includes groups B8 and B30. Another preferred group NR5R6 is group B8.
A further set of preferred groups NR5R6 consists of groups B8, B35, B36, B37, B38, B39, B40, B41 , B42, B43, B45, B46, B48, B53, B54, B55, B55, B57, B58, B59, B60, B61 and B62. A further set of preferred groups NR5R6 consists of groups B8, B35, B36, B37, B38, B39, B40, B41 , B42, B43, B45, B46, B48, B53, B54, B55, B56, B57, B58, B59, B60, B61 and B62
Another set of preferred groups consists of B8, B35, B36, B37, B38, B39, B40, B41 , B42, B43, B45, B46, B48, B53, B54, B55, B56, B57, B58, B59, B60, B61 , B62, B71 , B72, B74, B75, B76, B77, B78, B79, B80, B81, B82, B83, B85, B86, B87, B93, B94, B95, B97, B98, B99, B100 and B101.
A further sub-set of groups NR5R6 consists of B43, B46, B48, B76, B82, B89, B91 and B96. Within this sub-set, more preferred groups are groups B43, B46, B48, B76, B82, B89 and B91 , with B76, B82 and B89 being particularly preferred.
Particular and Preferred Sub-groups One sub-group of novel compounds of the invention can be represented by the general formula (II):
Figure imgf000052_0001
or salts, tautomers, solvates and N-oxides thereof; wherein:
Rz is as defined herein;
R3a is selected from hydrogen, halogen, cyano, Ci-5 hydrocarbyl and Ci-5 hydrocarbyloxy; wherein the Ci-5 hydrocarbyl and Ci-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, Ci-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R4 is selected from hydrogen; a group -(O)n-R7 where n is 0 or 1 and R7 is an acyclic C1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C1-5 hydrocarbyl-amino, wherein the acyclic Ci-5 hydrocarbyl group and the mono and di-C1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; or R3a and R4 together form a monocyclic carbocyclic or heterocyclic ring of 5 to 7 ring members; R5 and R6 together with the nitrogen atom to which they are attached form a bicyclic group having up to 12 ring members (e.g 8-12 ring members or 9-10 ring members) of which up to 5 ring members are heteroatoms selected from oxygen, nitrogen and sulphur; and R8 is selected from hydrogen and fluorine.
Another sub-group of novel compounds of the invention can be represented by the formula (III):
Figure imgf000053_0001
or salts, tautomers, solvates and N-oxides thereof; wherein: Rz is as defined herein;
R3b is selected from hydrogen, halogen, cyano, C1-5 hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and C1-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; R4 is selected from hydrogen; a group -(O)n-R7 where n is 0 or 1 and R7 is an acyclic C1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-Ci-5 hydrocarbyl-amino, wherein the acyclic C1-5 hydrocarbyl group and the mono and di-C1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; or R3b and R4 together form a monocyclic carbocyclic or heterocyclic ring of 5 to 7 ring members; R5 and R6 together with the nitrogen atom to which they are attached form a bicyclic group having from up to 12 ring members (e.g. 8-12 ring members or 9-10 ring members) of which up to 5 ring members are heteroatoms selected from oxygen, nitrogen and sulphur; and R8 is selected from hydrogen and fluorine.
A further sub-group of novel compounds of the invention can be represented by the formula (IV):
Figure imgf000054_0001
or salts, tautomers, solvates and N-oxides thereof; wherein:
R1 and R2a are as defined herein;
R3c is selected from hydrogen, halogen, cyano, Ci-5 hydrocarbyl and Ci-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and Ci-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and ary] and heteroaryl groups of 5 to 12 ring members;
R4 is selected from hydrogen; a group -(O)n-R7 where n is 0 or 1 and R7 is an acyclic C1-5 hydrocarbyl group or a monocyclic carbocyclic or heterocyclic group having 3 to 7 ring members; halogen; cyano; hydroxy; amino; and mono- or di-C1-5 hydrocarbyl-amino, wherein the acyclic C1-5 hydrocarbyl group and the mono and di-C1-5 hydrocarbylamino moieties in each instance are optionally substituted by one or more substituents selected from hydroxy, halogen, Ci-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members; or R3c and R4 together form a monocyclic carbocyclic or heterocyclic ring of 5 to 7 ring members; R5 and R6 together with the nitrogen atom to which they are attached form a bicyclic group having up to 12 ring members (e.g. 8-12 ring members or 9-10 ring members) of which up to 5 ring members are heteroatoms selected from oxygen, nitrogen and sulphur; and
R8 is selected from hydrogen and fluorine.
Within formulae (II), (III) and (IV), particular sub-groups of compounds are those wherein NR5R6 forms a bicyclic ring of up 10 ring members (e.g. 9 or 10 ring members, preferably 9 ring members) of which up to 5 ring members are heteroatoms selected from O, N and S, the monocylic or bicyclic ring being optionally substituted by up to three substituent groups R10, R1Oa, R10b, R1Ocand R1Occ as defined herein, more typically up to two substituents, for example up to one substituent.
More particular substituents for the bicyclic heterocyclic group NR5R8 are those forming part of a sub-group R10d which consists of the members of sub-group R1Oc and fluoro, chloro, bromo, trifluoromethyl, difluoromethoxy, trifluoromethoxy, cyano, methyl, ethyl, cyclopropyl, hydroxy, methylsuiphonyl, amino, methylamino, dimethylamino, methoxy, ethoxy, hydroxy methyl, hydroxyethyl, ethoxycarbonyl, methoxycarbonyl, aminocarbonyl, oxo, methoxymethyl, carboxy, phenyl, C1-2 alkoxycarbonyl, aminocarbonyl, acetyl, methylsuiphonyl and pyridyl. Within this sub- group, one sub-set of substituents includes methyl, ethyl, chloro, fluoro, hydroxy, methylsuiphonyl, amino, methylamino, dimethylamino, cyano, methoxy, ethoxy, hydroxymethyl, cyclopropyl, hydroxyethyl, ethoxycarbonyl, methoxycarbonyl, aminocarbonyl, oxo, methoxymethyl and acetyl. For example, NR5R6 can form a 5.6 or 6.6 fused bicyclic ring of 9 or ten ring members of which 1 to 3 are heteroatoms, the bicyclic ring being optionally substituted by one or more substituents R 10 or R1Oa or R1Qb or R10cor R10∞ or R10d and sub-groups (sub-sets) and examples thereof as defined herein. Within this sub-group, examples of fused bicyclic rings are those in which a non-aromatic ring such as a pyrrolidine, piperidine, piperazine or morpholine ring is fused to a 6-membered aryl or heteroaryl ring such as a benzene or pyridine ring, and wherein a nitrogen atom present in the non-aromatic ring is bonded to the carbonyl group in formulae (II), (IiI) or (IV).
Particular fused bicyclic rings include dihydroindole, dihydroisoindole, tetrahydroquinoline and tetrahydroisoquinoline, and aza-analogues thereof in which one or two carbon ring members in the aromatic ring are replaced by nitrogen.
One sub-group of bicyclic heterocyclic groups formed by NR5R6 consists of dihydroisoindole optionally substituted by one or more (e.g. 1 , 2 or 3) optional substituents selected from groups R10, R1Oa, R1Ob and R10oor R10co and or R10d and sub-groups (sub-sets) and examples thereof as defined herein.
Preferred compounds are those wherein the group R3a or R3b or R30 is selected from hydrogen, halogen and C1-5 alkyl; wherein the C1-5 alkyl moiety in each instance is optionally substituted by one or more substituents selected from hydroxy, halogen, C^2 alkoxy and amino.
More preferably, the group R3a or R3b or R3c is hydrogen or a C3-5 alkyl group optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy and amino. In particular, the group R3a or R3b or R3c is selected from hydrogen and isopropyl, sec-butyl, ferf-butyl and 1,2- dimethylpropyl groups.
Another sub-group of compounds of the invention is represented by formula (V):
Figure imgf000055_0001
or salts, tautomers, solvates and N-oxides thereof; wherein R1 and R2a are as defined herein; R3d is selected from ethyl and secondary and tertiary alkyl groups of 3 to 6 carbon atoms; R4a is selected from hydrogen, fluorine, chlorine and methoxy; and R5, Rs and R8 are as defined herein; provided that when R1 and R2 are both hydroxy, then R3d can additionally be selected from hydrogen. In one embodiment, when one of R1 and R2a is hydroxy and the other is O-Rz, or when R1 and R2a is both O-Rz, then R3d is hydrogen. ■,3d ,
In another embodiment, R is ethyl or a secondary or tertiary alkyl group. Particularly preferred alkyl groups R3d are ethyl, isopropyl and ferf-butyl, and in particular isopropyl.
Within formulae (II) to (V), preferred groups NR5R6 are dihydroisoindole groups which may be substituted or unsubstituted by one, two or three groups R10, R1Oa or R1Ob or R100 or R10∞ or R10d and sub-groups (sub-sets) and examples thereof as defined herein but, in one particular embodiment, are unsubstituted.
Another preferred sub-set of compounds can be represented by formula (Via):
Figure imgf000056_0001
or salts, tautomers, solvates and N-oxides thereof; wherein R1 and R2a are as defined herein; ring B is an aromatic ring containing up to two (and preferably 0 or 1) nitrogen heteroatom ring members; T is a group (CHR10b)j and Q is a group (CHR10b)k where j and k are each 0, 1 , 2 or 3 provided that the sum of j and k is 2 or 3; n is 0, 1 , 2 or 3 and R , R 34a , O R8 and R 10 UbD are as defined herein.
In one sub-group of compounds within formula (VI) or formula (Via), R 1 ; is- hydrogen.
In another subgroup of compounds within formula (Vl) or formula (Via), R 51 : i,s. hydroxy or a group O-Rz.
In formula (Vl), examples of the bicyclic group:
Figure imgf000056_0002
include the groups C1 to C6 below.
Figure imgf000056_0003
C3
Figure imgf000057_0001
Preferrred groups are groups C1 , C5 and C6
In the groups C1 to C6, the moiety R10 can be a group R10 as hereinbefore defined or can be a group R10b, R100, R1Occ or R1Occc as defined herein. In each case, n is preferably 1 , 2 or 3, and more preferably is 1 or 2, e.g. 1.
A currently preferred group is group C1.
Within formula (Vl), one particular group of compounds can be represented by the formula (VII):
Figure imgf000057_0002
or salts, tautomers, solvates and N-oxides thereof; wherein R1, R2a, R3 R4a, R8and R10b are as defined herein and n is 0, 1 2 or 3 (more preferably 0, 1 or 2, e.g. 0 or 1), and provided that at least one of R1 and R2a is O-Rz.
3d
Within formulae (Vl) and (VII), the substituent R is preferably a group R as defined herein and/or the substituent R -,10b is either absent (n=0) or is selected from groups R and R 310d and sub-groups (sub-sets) and examples thereof as defined herein. Preferably R1 and R2a are both hydroxy.
One particular group of compounds of the invention within formula (VII) is represented by the formula (Vila):
Figure imgf000058_0001
or salts, tautomers, solvates and N-oxides thereof; wherein R3 is selected from hydrogen, halogen, C1-5 alkyl, C2-5 alkenyl and C3-4 cycloalkyl groups;
R τ4a is selected from hydrogen, fluorine, chlorine and methoxy; R is hydrogen or fluorine; n is 0, 1 2 or 3; and R1, R2a and R10 are each as defined herein.
Within formula (Vila), R10 can be selected from, for example, one, two or three groups R1Oa or R10b or R10o or R1Occ or R10d and sub-groups (sub-sets) and examples thereof as defined herein.
One preferred group of compounds of the invention within formula (VIl) is represented by the formula (VIIb):
Figure imgf000058_0002
or salts, tautomers, solvates and N-oxides thereof; wherein R1 and R2a are each as defined herein, R3 is selected from hydrogen, halogen, C1-5 alkyl, C2-5 alkenyl and C3-4 cycloalkyl groups; R4a is selected from hydrogen, fluorine, chlorine and methoxy; R8 is hydrogen or fluorine; n is 0, 1 2 or 3; and R1Occ is selected from: halogen;
CO2R14 wherein R14 is hydrogen or C1-6 alkyl; C1-4 alkyl optionally substituted by hydroxy or C1-2 alkoxy; C1-4 alkoxy optionally substituted by hydroxy or C1-2 alkoxy; or a group [sol], CH2[SoI], C(O)[sol], OCH2CH2[SoI] or OCH2CH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000059_0001
,11 wherein X4 is NH or O, m is O or 1 , n is 1 , 2 or 3, R" is hydrogen, COR j1 '2z, C(O)OR !)1 l2z or R12; R12 is C1-6 alkyl, C3-6 cycloalkyl, aryl, aryl-Ci-6 alkyl or CH2R15; and R15 is selected from hydrogen, Ci-6 alkyl, C3.B cycloalkyl, hydroxy-C1-6 alkyl, piperidine, N-Ci-6 alkylpiperazine, piperazine, morpholine, COR13 or C(O)OR13; and R13 is C1* alkyl.
In a further embodiment, the compound can be an aza- or diaza- analogue of the the compounds of formulae (Vl), (VII) and (Vila) as defined herein wherein one or two of the carbon atoms of the benzene ring attached to the five membered ring is replaced by nitrogen.
For example, the group:
Figure imgf000059_0002
in the compound of formula (Vila) may be replaced by:
Figure imgf000060_0001
In each of formulae (Vl), (Via), (VII), (Vila) and (VIIb) and sub-groups thereof as defined herein, n is preferably 1 , 2 or 3, and more preferably is 1 or 2. In one embodiment, n is 1.
A further group of preferred compounds of the invention is represented by the formula (VIII):
Figure imgf000060_0002
wherein Q1 is a bond or a group CH2 and R1 and R2a are as defined herein.
Particular compounds within formula (VIII) are the compounds wherein one of R1 and R2a is hydroxy, O-C(=O)-R20 or O-C(=O)-N(R20)2, and the other of R1 and R2a is O-C(=0)-R20 or O-C(=O)- N(R20)2 wherein R20 is C1-4 alkyl.
Specific compounds of the invention include pro-drugs of the following compounds: (5-chloro-2-hydroxy-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone; (3-tert-butyl-4-hydroxy-phenyl)-(2,3-dihydro-indoI-1-yl)-methanone; (3-tert-butyl-4-hydroxy-phenyl)-(3,4-dihydro-2H-quinolin-1-yl)-methanone; (3,4-dihydro-1 H-isoquinolin-2-yl)-(4-hydroxy-3-isopropyl-phenyl)-methanone; (1 ,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone;
(3-tert-butyl-4-hydroxy-phenyl)-(1 ,4-dioxa-8-aza-spiro[4.5]dec-8-yl)-methanone; (3-tert-butyl-4-hydroxy-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone; (3-tert-butyl-4-hydroxy-phenyl)-pyrrolo[3,2-b]pyridin-1-yl-methanone; 8-(3-tert-butyl-4-hydroxy-benzoyl)-2-methyl-2,8-diaza-spiro[4.5]decan-1-one; (1 ,3-dihydro-isoindol-2-yl)-(4-hydroxy-3-isopropyl-phenyl)-methanone;
(3-tert-butyl-4-hydroxy-phenyl)-(3,4-dihydro-1 H-isoquinolin-2-yl)-methanone; (1 ,3-dihydro-isoindol-2-yl)-(5-ethyl-2,4-dihydroxy-phenyl)-methanone; (δ-cyclopropyl^^-dihydroxy-phenylHI .S-dihydro-isoindol^-yO-methanone;
(5-sec-butyl-2,4-dihydroxy-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone;
(1 ,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-phenyl)-methanone;
(5-chloro-1 ,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone; [5-(3-amino-propoxy)-1 ,3-dihydro-isoindol-2-yl]-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone;
(5-bromo-2,4-dihydroxy-phenyl)-(1,3-dihydro-isoindol-2-yl)-methanone;
(1,3-dihydro-isoindo]-2-yl)-(2,4-dihydroxy-5-trifluoromethyl-phenyl)-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-{4-[2-(2-methoxy-ethoxy)-ethoxy]-1 ,3-dihydro-isoiπdol-2- yl}methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[4-(2-dimethy!amino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1 ,3-dihydro-isoindol-2- yljmethanone;
(3-sec-butyl-4-hydroxy-phenyl)-(1,3-dihydro-isoindoI-2-yl)-methanone; (5-tert-butyl-2,4-dihydroxy-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone;
(5-chloro-2,4-dihydroxy-phenyl)-(1,3-dihydro-isoindol-2-yl)-methanone;
(1 ,3-dihydro-isoindol-2-yl)-(2-hydroxy-5-isopropyI-4-methoxy-phenyl)-methanone;
(4,7-difluoro-1 ,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-fluoro-1 ,3-dihydro-isoindol-2-yl)-methanone; (1 ,3-dihydro-isoindol-2-yl)-(3-fluoro-2,4-dihydroxy-5-isopropyl-phenyl)-methanone;
(1 ,3-dihydro-isoindol-2-yl)-(2-fluoro-4,6-dihydroxy-3-isopropyl-phenyl)-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-(4-fluoro-1 ,3-dihydro-isoindol-2-yl)-methanone hydrochloride;
(5-chloro-6-methoxy-1,3-dihydro-iso-indol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(2-methoxy-ethoxy)-1 ,3-dihydro-isoindol-2-yl]-methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(3-morpholin-4-yl-propoxy)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)- (2-oxa-5-aza-bicyclo[2.2.1]hept-5-yl)-methanone; (3,4-dihydro-1 H-isoquinolin-2-yl)- (2,4-dihydroxy-5-isopropyl-pheny!)-methanone;
(5-amino-1 ,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-pheny])-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-methoxy-1 ,3-dihydro-isoindol-2-yl)-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-morpholin-4-yl-1 ,3-dihydro-isoindol-2-yl)-methanone;
(2,4-dihydroxy-5-isopropyI-phenyl)-[5-(4-methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2-yl]- methanone;
2-(2,4-dihydroxy-5-isopropyl-benzoyI)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methyl ester;
2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-morpholin-4-ylmethyl-1 ,3-dihydro-isoindo!-2-yl)-methanone; {3-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-ciihyclro-1 H-isoindol-5-yloxy]-propyl}-carbamic acicl tert-butyl ester;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-methyl-1 ,3-ciihydro-isoindol-2-yl)-methanone;
(2,4-dihydroxy-5-isopropyI-phenyl)-[5-(2-isopropylamino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]- methanone;
N-{2-[2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindol-5-yloxy]-ethyl}-2-morpholin-4-yl- acetamide;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1 ,3-dihydro-isoindol-2-yl]- methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1 ,3-dihydro- isoindol-2-yi}-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-piperazin-1-yl-phenyl)-1 ,3-dihydro-isoindol-2-yl]- methanone;
2,4-dihydroxy-5-isopropyl-phenyl)-[5-(1-dimethylamino-2-hydroxy-ethyl)-1 ,3-dihydro-isoindol-2-yl]- methanone; dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-1-hydroxy-ethyl)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(piperazine-1-carbonyi)-1 ,3-dihydro-isoindol-2-yl]-methanone hydrochloride; (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1 ,3-dihydro-isoindol-2- yl]methanone;
[5-(2-amino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-(5-hydroxy-1,3-dihydro-isoindol-2-yI)-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-{5-[4-(2-hydroxy-ethyl)-piperazin-1-yl]-1 ,3-dihydro-isoindol-2-yl}- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-morpholin-4-yl-piperidin-1-yl)-1 ,3-dihydro-isoindol-2-yl]- methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(1-methyl-piperidin-4-ylamino)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-isopropyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-piperazin-1-yl-1 ,3-dihydro-isoindol-2-yl)-methanone; 4-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindo]-5-ylamino]-piperidine-1-carboxylic acid tert-butyl ester;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(piperidin-4-ylamino)-1 ,3-dihydro-isoindol-2-yl]-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[4-(4-methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2-yl]- methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[4-(piperidin-4-ylamino)-1 ,3-dihydro-isoindol-2-yl]-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-dimethylaminomethyl-1 ,3dihydroisoindol-2-yl)-methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazine-1-carbonyl)-1 ,3-dihydro-isoindol-2-yl]- methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-{5-[2-(2,2-dimethyl-propylamino)-ethoxy]-1 ,3-dihydro-isoindol-2- yl}-methanone;
[δ^-cyclopentylamino-ethoxyJ-I .S-dihydro-isoindol^-yll^^-dihydroxy-S-isopropyl-phenyl)- methanone;
(2,4-dihydroxy-5-isopropyl-phenyI)-(5-piperidin-1-ylmethyl-1 ,3-dihydro-isoindol-2-yl)-methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxypiperidin-4-yl)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(5-chloro-2,4-dihydroxy-phenyl)-[5-(4-methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2-yl] methanone; and
(5-chloro-6-hydroxy-1 ,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(5-chloro-2,4-dihydroxy-phenyl)-[5-(4-methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2-yl] methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-t5-(2-dimethylamino-ethoxy)-7-methyl-1 ,3-dihydro-isoindol-2- yl]-methanone; and salts, solvates, N-oxides and tautomers thereof.
Preferred individual compounds of the formula (I) are:
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]- methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(1-methyl-piperidin-4-ylamino)-1,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-(5-piperazin-1-yl-1,3-dihydro-isoindol-2-yl)-methanone;
(2,4-dihydroxy-5-isoρropyl-phenyl)-(5-dimethylaminomethyl-1 ,3dihydroisoindol-2-yl)-methanone; or salts, solvates, N-oxides and tautomers thereof; wherein, when only a single phenolic hydroxyl group is present, this is replaced by a group O-Rz as defined herein and, when two phenolic groups are present, one or both phenolic groups are replaced by a group O-Rz as defined herein.
A particularly preferred set of individual compounds consists of pro-drugs of: (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]- methanone;
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-dihydro-isoindol-2-yl]- methanone; (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]- methanone; and
(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(1-methyl-piperidin-4-ylamino)-1,3-dihydro-isoindol-2-yl]- methanone; or salts, solvates or tautomers thereof; wherein one or both of the phenolic hydroxyl groups has been replaced by a group O-Rz as defined herein.
For the avoidance of doubt, it is to be understood that each general and specific preference, embodiment and example of the group R1 may be combined with each general and specific preference, embodiment and example of the groups R2 or R2a and/or R3 and/or R4 and/or R4 and/or R5 and/or R6 and/or R10 and/or Q and/or T and/or sub-groups thereof as defined herein and that all such combinations are embraced by this application.
The various functional groups and substituents making up the compounds of the formula (I) are typically chosen such that the molecular weight of the compound of the formula (I) does not exceed 1000. More usually, the molecular weight of the compound will be less than 750, for example less than 700, or less than 650, or less than 600, or less than 550. More preferably, the molecular weight is less than 525 and, for example, is 500 or less.
Salts, Solvates, Tautomers, Isomers, N-Oxides, Esters and Isotopes
A reference to a compound of the formulae (V!0), (Vl), (I0), (I) and sub-groups thereof also includes ionic forms, salts, solvates, isomers, tautomers, N-oxides, esters, isotopes and protected forms thereof, for example, as discussed below; preferably, the salts or tautomers or isomers or N-oxides or solvates thereof; and more preferably, the salts or tautomers or N-oxides or solvates thereof
Many compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof can exist in the form of salts, for example acid addition salts or, in certain cases salts of organic and inorganic bases such as phenolate, carboxylate, sulphonate and phosphate salts. All such salts are within the scope of this invention, and references to compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof include the salt forms of the compounds.
The salts of the present invention can be synthesized from the parent compound that contains a basic or acidic moiety by conventional chemical methods such as methods described in Pharmaceutical Salts: Properties, Selection, and Use, P. Heinrich Stahl (Editor), Camille G. Wermuth (Editor), ISBN: 3-90639-026-8, Hardcover, 388 pages, August 2002. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media such as ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are used.
Acid addition salts may be formed with a wide variety of acids, both inorganic and organic. Examples of acid addition salts include salts formed with an acid selected from the group consisting of acetic, 2,2-dichloroacetic, adipic, alginic, ascorbic (e.g. L-ascorbic), L-aspartic, benzenesulphonic, benzoic, 4-acetamidobenzoic, butanoic, (+) camphoric, camphor-sulphonic, (+)-(1S)-camphor-10-sulphonic, capric, caproic, caprylic, cinnamic, citric, cyclamic, dodecylsulphuric, ethane-1 ,2-disulphonic, ethanesulphonic, 2-hydroxyethanesulphonic, formic, fumaric, galactaric, gentisic, glucoheptonic, D-gluconic, glucuronic (e.g. D-glucuronic), glutamic (e.g. L-glutamic), α-oxoglutaric, glycolic, hippuric, hydrobromic, hydrochloric, hydriodic, isethionic, (+)-L-lactic, (±)-DL-lactic, lactobionic, maleic, malic, (-)-L-malic, malonic, (±)-DL-mandelic, methanesulphonic, naphthalene-2-sulphonic, naphthalene-1 ,5-disulphonic, 1-hydroxy-2-naphthoic, nicotinic, nitric, oleic, orotic, oxalic, palmitic, pamoic, phosphoric, propionic, L-pyroglutamic, salicylic, 4-amino-salicylic, sebacic, stearic, succinic, sulphuric, tannic, (+)-L-tartaric, thiocyanic, p- toluenesulphonic, undecylenic and valeric acids, as well as acylated amino acids and cation exchange resins.
If the compound is anionic, or has a functional group which may be anionic (e.g., -COOH may be -COO"), then a salt may be formed with a suitable cation. Examples of suitable inorganic cations include, but are not limited to, alkali metal ions such as Na+ and K+, alkaline earth metal cations such as Ca2+ and Mg2+, and other cations such as Al3+. Examples of suitable organic cations include, but are not limited to, ammonium ion (i.e., NH4 +) and substituted ammonium ions (e.g., NH3R+, NH2Ra+, NHR3 +, NR4 +). Examples of some suitable substituted ammonium ions are those derived from: ethylamine, diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids, such as lysine and arginine. An example of a common quaternary ammonium ion is N(CH3)/.
Where the compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof contain an amine function, these may form quaternary ammonium salts, for example by reaction with an alkylating agent according to methods well known to the skilled person. Such quaternary ammonium compounds are within the scope of formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof.
The salt forms of the compounds of the invention are typically pharmaceutically acceptable salts, and examples of pharmaceutically acceptable salts are discussed in Berge ef a/., 1977, "Pharmaceutically Acceptable Salts," J. Pharm. Sci., Vol. 66, pp. 1-19. However, salts that are not pharmaceutically acceptable may also be prepared as intermediate forms which may then be converted into pharmaceutically acceptable salts. Such non-pharmaceutically acceptable salts forms, which may be useful, for example, in the purification or separation of the compounds of the invention, also form part of the invention. Compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof containing an amine function may also form N-oxides. A reference herein to a compound of any of the formulae (Vl0), (Vl), (1°), (I) and sub-groups thereof that contains an amine function also includes the N-oxide.
Where a compound contains several amine functions, one or more than one nitrogen atom may be oxidised to form an N-oxide. Particular examples of N-oxides are the N-oxides of a tertiary amine or a nitrogen atom of a nitrogen-containing heterocycle.
N-Oxides can be formed by treatment of the corresponding amine with an oxidizing agent such as hydrogen peroxide or a per-acid (e.g. a peroxycarboxylic acid), see for example Advanced Organic Chemistry, by Jerry March, 4th Edition, Wiley Interscience, pages. More particularly, N-oxides can be made by the procedure of L. W. Deady (Syn. Comm. 1977, 7, 509-514) in which the amine compound is reacted with m-chloroperoxybenzoic acid (MCPBA), for example, in an inert solvent such as dichloromethane.
Compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof may exist in a number of different geometric isomeric, and tautomeric forms and references to compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof include all such forms. For the avoidance of doubt, where a compound can exist in one of several geometric isomeric or tautomeric forms and only one is specifically described or shown, all others are nevertheless embraced by formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof.
Examples of tautomeric forms include, for example, keto-, enol-, and enolate-forms, as in, for example, the following tautomeric pairs: keto/enol (illustrated below), imine/enamine, amide/imino alcohol, amidine/amidine, nitroso/oxime, thioketone/enethiol, and nitro/aci-nitro.
Figure imgf000066_0001
compounds of the formulae (Vl0), (VI), (I0), (I) and sub-groups thereof contain one or more chiral centres, and can exist in the form of two or more optical isomers, references to compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof include all optical isomeric forms thereof (e.g. enantiomers, epimers and diastereoisomers), either as individual optical isomers, or mixtures (e.g. racemic mixtures) or two or more optical isomers, unless the context requires otherwise.
The optical isomers may be characterised and identified by their optical activity (i.e. as + and - isomers, or d and / isomers) or they may be characterised in terms of their absolute stereochemistry using the "R and S" nomenclature developed by Cahn, lngold and Prelog, see Advanced Organic Chemistry by Jerry March, 4th Edition, John Wiley & Sons, New York, 1992, pages 109-114, and see also Cahn, lngold & Prelog, Angew. Chem. Int. Ed. Engl., 1966, 5, 385- 415. Optical isomers can be separated by a number of techniques including chiral chromatography (chromatography on a chiral support) and such techniques are well known to the person skilled in the art.
As an alternative to chiral chromatography, optical isomers can be separated by forming diastereoisomeric salts with chiral acids such as (+)-tartaric acid, (-)-pyroglutamic acid, (-)-di- toluoyl-L-tartaric acid, (+)-mandelic acid, (-)-malic acid, and (-)-camphorsulphonic, separating the diastereoisomers by preferential crystallisation, and then dissociating the salts to give the individual enantiomer of the free base.
Where compounds of the formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof exist as two or more optical isomeric forms, one enantiomer in a pair of enantiomers may exhibit advantages over the other enantiomer, for example, in terms of biological activity. Thus, in certain circumstances, it may be desirable to use as a therapeutic agent only one of a pair of enantiomers, or only one of a plurality of diastereoisomers. Accordingly, the invention provides compositions containing a compound of the formula (I) having one or more chiral centres, wherein at least 55% (e.g. at least 60%, 65%, 70%, 75%, 80%, 85%, 90% or 95%) of the compound of the formula (I) is present as a single optical isomer (e.g. enantiomer or diastereoisomer). In one general embodiment, 99% or more (e.g. substantially all) of the total amount of the compound of the formula (I) may be present as a single optical isomer (e.g. enantiomer or diastereoisomer).
The compounds of the invention include compounds with one or more isotopic substitutions, and a reference to a particular element includes within its scope all isotopes of the element. For example, a reference to hydrogen includes within its scope 1H, 2H (D), and 3H (T). Similarly, references to carbon and oxygen include within their scope respectively 12C, 13C and 14C and 16O and 18O.
The isotopes may be radioactive or non-radioactive. In one embodiment of the invention, the compounds contain no radioactive isotopes. Such compounds are preferred for therapeutic use. In another embodiment, however, the compound may contain one or more radioisotopes. Compounds containing such radioisotopes may be useful in a diagnostic context.
Esters such as carboxylic acid esters and acyloxy esters of the compounds of formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof bearing a carboxylic acid group or a hydroxyl group are also embraced by formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof. Examples of esters are compounds containing the group -C(=O)OR, wherein R is an ester substituent, for example, a Ci-7 alkyl group, a C3-2O heterocyclyl group, or a C5.2o aryl group, preferably a C1-7 alkyl group. Particular examples of ester groups include, but are not limited to, -C(=O)OCH3, -C(=O)OCH2CH3, -C(=O)OC(CH3)3, and -C(=O)OPh. Examples of acyloxy (reverse ester) groups are represented by -OC(=O)R, wherein R is an acyloxy substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-20 aryl group, preferably a C1-7 alkyl group. Particular examples of acyloxy groups include, but are not limited to, -OC(=O)CH3 (acetoxy), -OC(=O)CH2CH3, -OC(=0)C(CH3)3, -OC(=O)Ph, and -OC(=O)CH2Ph. In one general embodiment, formulae (Vl0), (Vl), (I0), (I) and sub-formulae, sub-groups, preferences and examples thereof do not cover esters such as carboxylic acid esters and acyloxy esters.
Also encompassed by formulae (Vl0), (Vl), (I0), (I) and sub-groups thereof are any polymorphic forms of the compounds, solvates (e.g. hydrates) and complexes (e.g. inclusion complexes or clathrates with compounds such as cyclodextrins, or complexes with metals) of the compounds.
Biological Activity and Therapeutic Uses
The compounds of the formulae (Vl0), (VI), (I0), (I) and sub-groups thereof are pro-drugs of compounds that are inhibitors of Hsp90 The pro-drug compounds of the invention have a number of advantages with respect to their parent compounds. For example, they may provide improved oral bioavailability, e.g. by virtue of improved intestinal absorption. Furthermore, by forming a pro-drug, conjugation and/or metabolism of the parent compound may be substantially reduced.
The pro-drug compounds of the invention are expected to be beneficial in the treatment of wide spectrum of proliferative disorders. Examples of such proliferative disorders include, but are not limited to, a carcinoma, for example a carcinoma of the bladder, breast, colon (e.g. colorectal carcinomas such as colon adenocarcinoma and colon adenoma), kidney, epidermis, liver, lung, for example adenocarcinoma, small cell lung cancer and non-small cell lung carcinomas, oesophagus, gall bladder, ovary, pancreas e.g. exocrine pancreatic carcinoma, stomach, cervix, thyroid, prostate, gastrointestinal system, e.g. gastrointestinal stromal tumours, or skin, for example squamous cell carcinoma; a hematopoieitic tumour of lymphoid lineage, for example leukaemia, acute lymphocytic leukaemia, chronic lymphocytic leukaemia, B-cell lymphoma (such as diffuse large B cell lymphoma), T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, hairy cell lymphoma, or Burkitt's lymphoma; a hematopoieitic tumour of myeloid lineage, including acute myeloid leukaemia, chronic myeloid leukaemias, myelogenous leukaemias, and lmatinib sensitive and refractory chronic myelogenous leukaemias, myelodysplastic syndrome, Bortezomib sensitive and refractory multiple myeloma, myeloproliferative disease or promyelocytic leukaemia; thyroid follicular cancer; a tumour of mesenchymal origin, for example fibrosarcoma or rhabdomyosarcoma; a tumour of the central or peripheral nervous system, for example astrocytoma, neuroblastoma, glioma or schwannoma; melanoma; seminoma; teratocarcinoma; osteosarcoma; xeroderma pigmentosum; keratoacanthoma; thyroid follicular cancer; or Kaposi's sarcoma. A further example of a tumour of mesenchymal origin is Ewing's sarcoma.
The cancers may be cancers which are sensitive to Hsp90 inhibition, and such cancers may be determined by a method as set out in the section headed "Methods of Diagnosis". One group of cancers includes human breast cancers (e.g. primary breast tumours, node-negative breast cancer, invasive duct adenocarcinomas of the breast, non-endometrioid breast cancers); and mantle cell lymphomas. In addition, other cancers are colorectal and endometrial cancers.
Another sub-set of cancers includes hematopoietic tumours of both lymphoid and myeloid lineage, for example acute lymphoblastic leukemia, chronic lymphocytic leukaemia (Both T and B cell), acute myeloid leukaemia, chronic myeloid leukaemia, mantle cell lymphoma and B-cell lymphoma (such as diffuse large B cell lymphoma) and optionally further includes chronic myelogenous leukaemia and multiple myeloma.
A preferred sub-set of cancers consists of ErbB2-positive breast, prostate, lung, and gastric cancer; chronic myeloid leukemia; androgen receptor dependent prostate cancer; Flt3-dependent acute myeloid leukaemia; melanoma associated with Braf mutation; multiple myeloma; velcade refractory multiple myeloma; and gastrointestinal stromal tumours (GIST).
Of these, particularly preferred cancers are multiple myelomas and velcade refractory tumour types as defined herein. Another preferred sub-set of cancers consists of hormone refractory prostate cancer, metastatic melanoma, HER2 positive breast cancer, mutant EGFR positive non-small cell lung carcinoma and Gleevec resistant gastrointestinal stromal tumours.
Hsp90 inhibitors could also be used to treat other conditions such as viral infections, parasitic disease, autoimmune diseases (e.g. multiple sclerosis and lupus erythematosus), neuro- degenerative disorders (e.g. Alzheimer's disease), inflammation, Type I and Il diabetes, atherosclerosis and cardiac disease.
Hsp90 inhibitors could also have clinical benefit in transplantation and immunosuppression.
Hsp90 inhibitors may also have clinical benefit in the previously described diseases when used in combination with existing or new therapeutic agents. Based on the activities of Hsp90 client proteins and experimental evidence, the following disorders may be particularly sensitive to treatment by Hsp90 inhibitors.
ErbB2-positive breast, prostate, lung, and gastric cancer
Overexpression of ErbB2 (HER-2) occurs in approximately 30 % of breast cancers and ErbB2 receptor down-regulation by herceptin sensitized cells to Taxol. ErbB2 over-expression is linked to poor prognosis and drug resistance (Tsugawa et. al., 1993. Oncology 1993; 50: 418).
Mutant EGFR in lung cancer
Somatic mutations in the kinase domain of the epidermal growth factor receptor (EGFR), including L858R and exon 19 deletions, underlie responsiveness to gefitinib and erlotinib in non-small cell lung cancer (NSCLC). Acquired resistance to these tyrosine kinase inhibitors is in some cases mediated by a second mutation, T790M. Ansamycin antibiotics, such as geldanamycin, potently inhibit heat shock protein 90 (Hsp90), promoting ubiquitin-mediated degradation of oncogenic kinases that require the chaperone for proper conformational folding. Exposure of EGFR-mutant cell lines to geldanamycin induced marked depletion of phospho-Akt and cyclin D1 as well as apoptosis. These data suggest mutational activation of EGFR is associated with dependence on Hsp90 for stability and that Hsp90 inhibition may represent a novel strategy for the treatment of EGFR-mutant NSCLC.
Chronic myeloid Leukemia
The aberrant BCR-AbI protein is created through a chromosomal translocation and results in a constitutively active AbI kinase domain. This translocation event has been shown to be causal for CML. P210BcrAbl is a known client protein for Hsp90. Treatment of the BCR-AbI positive cell line K562 with an hsp90 inhibitor induced apoptosis. The Bcr-Abl inhibitor Gleevec® also induces apoptosis in K562 cells; however Gleevec® resistant K562 cells still retain sensitivity towards Hsp90 inhibitors (Gorre et. al. 2002, Blood 100: 3041-3044).
Androgen receptor dependent prostate cancer The androgen receptor kinase is an Hsp90 client protein. Testosterone deprivation is usually adopted where surgery is not curative. The cancer may become refractory to hormone manipulation through receptor mutation. Hsp90 regulation of the receptor would still be viable post-mutation.
The same would apply to estrogen-dependent breast cancers. Flt3-dependent acute myeloid leukaemia
Internal duplication of the tyrosine kinase receptor Flt3 leads to its constitutive activation and oncogenesis. These internal duplications are observed in 20% of all reported cases of AML and are an indication of poor prognosis. Much like the activation of the ABL kinase in CML, this represents another example of a single genetic lesion giving rise to a malignancy. Hsp90 inhibitors are predicted to be of clinical benefit to these patients as Flt3 is an Hsp90 client protein (Bali et. al., 2004 Cancer Res. 64(10):3645-52).
Melanoma associated with Braf mutation
Braf encodes for a serine/threonine kinase which is mutated in 70% of all melanomas. 80% of these represent a single V599E point mutation that confers elevated kinase activity to BRAF. This mutation is also transforming in NIH3T3 cells (Bignell et. al., 2002 Nature. 417(6892):949-54).
Multiple Myeloma
The Hsp90 inhibitor 17-AAG potently inhibits proliferation of Bortezomib refractory multiple myeloma cell lines. Cell surface levels of IGF-1 R and IL-6R were also diminished in 17-aag treated MM-1 cells (Mitsiades et. al., Blood 107:1092-1100, 2006). Autocrine stimulation of multiple myeloma cells, as well as paracrine stimulation of bone marrow stromal cells with IL-6 is also diminished through downregulation of the Hsp90 client IKK. Velcade Refractory Multiple Myeloma
Compounds of the present invention can be used in the treatment of velcade refractory tumour types including treatment of patients with second line mantle cell lymphoma, indolent non- Hodgkin's lymphoma, stage NIB and IV Bronchioloalveolar carcinoma, advanced non-small cell lung cancer, breast, prostate and ovarian cancers and non-Hodgkin's lymphoma.
Gastrointestinal stromal tumours (GIST)
GIST disease particularly disease dependent on growth factor activation or overexpression (e.g. c- kit).
B-CLL ZAP-70 is an Hsp90 client protein and the requirement for Hsp90 by ZAP-70 is limited to CLL cells and is not observed in T cells where this kinase is normally expressed. Hence, ZAP-70 is unique among identified Hsp90 clients as its chaperone dependency is conditional on the type of cell in which it is expressed.
Other conditions or disorders for which an Hsp90 inhibitor may be of clinical benefit include, but are not limited to:
Neurodegenerative disorders
Huntington's disease (HD) is a progressive neurodegenerative disorder with no effective treatment. GA inhibition of Hsp90 and the resulting up-regulation of Hsps are effective in preventing huntington protein aggregation in neuronal cells. (Sittler et. al., 2001 , Human Molecular Genetics, Vol. 10, No. 12 1307-1315). Up-regulation of HSP may also be of clinical benefit in other diseases of protein misfolding e.g., CJD and Alzheimer's.
Inflammatory disease, including Rheumatoid arthritis, Asthma, Chronic obstructive pulmonary disease, and inflammatory bowel disease
GA has been shown to dissociate HSF-1 from Hsp90 leading to the activation and nuclear translocation of HSF-1. HSF-1 subsequently acts as a transcription factor to induce HSP90 and Hsp70. The induction of Hsp70 has been implicated in the resolution of inflammation in an induced mouse model of edema (lanaro et al., 2004 Human Molecular Genetics, 2001 , Vol. 10, No. 12 1307-1315). Additionally GA treatment inhibited IkappaB kinase (IKK) activation by TNF-a or PMA. IkBa is a regulator of Nf-kB and Ap- 1. (Broemer et al. 2004 ). Ap-1 and Nf-kB is a major transcription factor leading to the production of pro-inflammatory cytokines (Yeo et. al., 2004 Biochem Biophys Res Commun. 30; 320(3):816-24). The stability of pro-inflammatory cytokine transcripts is also regulated through inhibition of p38 MapK (Wax et. al., 2003. Rheumatism Vol. 48, No. 2, pp 541-550).
Atherosclerosis It is known that inflammatory and immune cells play a central role in the initiation and progression of human atherosclerosis (Riganό et al., Ann. N. Y. Acad. ScL, 2007, 1107:1-10) and it has been proposed that Hsp90 acts as an autoantigen in carotid atherosclerosis. Riganό et al. found specific antibodies and cells against Hsp90 in the sera of 60% of patients tested who were suffering from carotid atherosclerotic plaques but no specific antibodies and T cells against Hsp90 in the sera of healthy patients. Therefore, inhibitors of Hsp90 should be useful in the treatment or prevention of atherosclerosis.
Angioqenesis related disease, including but not limited to: tumour anqioqenesis, psoriasis, rheumatoid arthritis, and diabetic retinopathy Induction of angiogenesis is regulated by Hsp90 client proteins eNOS and Akt in endothelial cells (Sun and Liao, 2004 Arterioscler Thromb Vase Biol. 24(12):2238-44). Suppression of hypoxia- inducible factor (HIF)-Ia can also impair the growth, angiogenesis and vessel maturation of gastric tumours in a mouse model. (Stoeltzing et. al., 2004 J Natl Cancer Inst; 96:946-956.).
Type I and type Il diabetes Hsp90 inhibition has a profound effect on Akt signalling as well as e-nos. These are two key regulators in high glucose induced endothelial cell apoptosis in type I diabetes (Lin et. al., 2005 J Cell Biochem. 1 ; 94(1): 194-201) and the development of hypertension in type Il diabetes (Kobayashi et. al., 2004 Hypertension. 44(6):956-62.).
Immunosuppression and transplantation Hsp90 inhibition has been shown to down regulate Lck, a T-cell specific tyrosine kinase required for T-cell activation. (Yorgin et. al., 2000 J Immunol. 15; 164(6):2915-23.)
Cardiac disease
Cardiac ischemic is the most common cause of death in the western world. Hsps, and notably Hsp70 (induced by radicicol treatment) have demonstrated cardioprotective activity in rat cardiomyocytes (Griffin ef. a/., 2004). Inhibition of Hsp90 results in the release of HSF-1 from the chaperone complex and its subsequent activation of Hsp genes. Inhibition of Hsp90 also leads to the down-regulation of HIF-1 , which has been implicated in the pathogenesis of ischemic heart disease and stroke.
Infectious disease Hepatits C viral NS2/3 protease is an Hsp90 client protein and Hsp90 activity is required for viral processing and replication (Whitney et. al., 2001. Proc Natl Acad Sci U S A. 20;98(24):13931-5.).
Parasitic disease
GA has reported antimalarial activity against an Hsp90 ortholog of Plasmodium falciparum. Plasmodium growth was inhibited with GA at an IC50 similar to that observed with chloroquine. GA was also effective against chloroquine resistant strains of Plasmodium falciparum (Kamar et. al., 2003 . Malar J.15; 2(1):30).
Inhibition, Prevention or Reversal of the Development of Drug Resistance
As discussed above, modulators or inhibitors of stress protein function in general (and Hsp90 in particular) represent a class of chemotherapeutics with the potential for: (i) sensitizing malignant cells to anticancer drugs and/or treatments; (ii) alleviating or reducing the incidence of resistance to anticancer drugs and/or treatments; (iii) reversing resistance to anticancer drugs and/or treatments; (iv) potentiating the activity of anticancer drugs and/or treatments; (v) delaying or preventing the onset of resistance to anticancer drugs and/or treatments. Accordingly, the invention further provides:
• A method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90, which method comprises administering to a subject in need thereof a compound of the invention, wherein the disease state or condition mediated by Hsp90 is the development of resistance to a cancer drug.
• A method for: (i) sensitizing malignant cells to an anticancer drug; (ii) alleviating or reducing the incidence of resistance to an anticancer drug; (iii) reversing resistance to an anticancer drug; (iv) potentiating the activity of an anticancer drug; (v) delaying or preventing the onset of resistance to an anticancer drug, which method comprises administering to a subject in need thereof a compound of the invention.
• A method for the treatment of a cancer which method comprises administering to a subject in need thereof a compound of the invention, which method is characterized by the absence of drug resistance.
• A method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90 in a subject undergoing treatment with a therapeutic agent (such as an anti-cancer agent), which method comprises administering to the subject a compound of the invention, wherein the disease state or condition mediated by Hsp90 is the development of resistance to the said therapeutic agent.
• A method for: (i) sensitizing malignant cells to an anti-cancer agent; (ii) alleviating or reducing the incidence of resistance to an anti-cancer agent; (iii) reversing resistance to an anti-cancer agent; (iv) potentiating the activity of an anti-cancer agent; (v) delaying or preventing the onset of resistance to an anti-cancer agent, which method comprises administering to a subject undergoing treatment with said anti-cancer agent a compound of the invention. • A method for the treatment of a cancer in a subject undergoing treatment with an anticancer agent, which method comprises administering to a subject in need thereof a compound of the invention, which method is characterized by the absence of drug resistance to the anti-cancer agent.
Biological Activity
The biological activity (e.g. as inhibitors of Hsp90) of the phenolic compounds from which the prodrug compounds of the invention are derived can be measured using the assays set forth in the examples below, for example the isothermal titration calorimetry (ITC) experiments described in Example 86 and the anti-proliferative activity assays described in Example 87. The level of activity exhibited by a given compound in the ITC assay can be defined in terms of the Kd value, and preferred compounds of the present invention are compounds having a Kd value of less than 1 micromolar, more preferably less than 0.1 micromolar. In the anti-proliferative activity assays, the level of activity exhibited by a given compound in an assay can be defined in terms of the IC50 value, and preferred compounds of the present invention are compounds having an IC50 value of less than 1 micromolar, more preferably less than 0.1 micromolar. hERG
In the late 1990s a number of drugs, approved by the US FDA, had to be withdrawn from sale in the US when it was discovered they were implicated in deaths caused by heart malfunction. It was subsequently found that a side effect of these drugs was the development of arrhythmias caused by the blocking of hERG channels in heart cells. The hERG channel is one of a family of potassium ion channels the first member of which was identified in the late 1980s in a mutant Drosophila melanogaster fruitfly (see Jan, L. Y. and Jan, Y.N. (1990). A Superfamily of Ion Channels. Nature, 345(6277):672). The biophysical properties of the hERG potassium ion channel are described in Sanguinetti, M. C1 Jiang, C, Curran, M. E., and Keating, MT. (1995). A Mechanistic Link Between an Inherited and an Acquired Cardiac Arrhythmia: HERG encodes the lkr potassium channel. Cell, 81 :299-307, and Trudeau, M. C1 Warmke, J.W., Ganetzky, B., and Robertson, G.A. (1995). HERG, a Human Inward Rectifier in the Voltage-Gated Potassium Channel Family. Science, 269:92-95.
The elimination of hERG blocking activity remains an important consideration in the development of any new drug.
It has been found that many of the phenolic compounds from which the pro-drug compounds of the invention are derived have low hERG activity and a good separation between Hsp90 inhibitory activity and hERG activity.
Preferred pro-drug compounds are the pro-drug compounds of phenolic compounds having mean IC50 values against hERG that are greater than 30 times, or greater than 40 times, or greater than 50 times the IC50 values of the compounds in cellular proliferation assays. Preferred pro-drugs are the pro-drugs of phenolic compounds having mean IC50 values against hERG that are greater than 5 μM, more particularly greater than 10 μM, and more preferably greater than 15 μM. Some phenolic compounds from which the pro-drug compounds of the invention are derived have mean IC50 values against hERG that are greater than 50 μM.
Compounds of the invention have advantageous ADME properties and in particular better oral bioavailability than the phenolic parent compounds.
Treatment of pain, neuropathies, stroke and related conditions
The compounds of the invention have Hsp90 inhibiting or modulating activity and hence are useful in for use in treating, alleviating or preventing certain cdk5 mediated diseases and conditions.
Acccordingly, in a first aspect, the invention provides the use of a compound of the invention as defined herein for the manufacture of a medicament for the treatment of pain.
In another aspect, the invention provides the use of a compound of the invention as defined herein thereof for the manufacture of a medicament for the prophylaxis or treatment of stroke.
In a further aspect, the invention provides the use of a compound of the invention as defined herein for the manufacture of a medicament for use as a neuroprotective agent. In other aspects, the invention provides:
■ A compound of the invention as defined herein for use in the treatment of pain.
■ A compound of the invention as defined herein for use in the reduction or elimination of pain in a patient (e.g. a mammal such as a human) suffering from pain.
■ The use of a compound of the invention as defined herein for the manufacture of a medicament for use in the reduction or elimination of pain in a patient (e.g. a mammal such as a human) suffering from pain.
■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the treatment of any one or more of nociception, somatic pain, visceral pain, acute pain, chronic pain, hyperalgesia, allodynia, post operative pain, pain due to hypersensivity, headache, inflammatory pain (rheumatic, dental, dysmenorrhoea or infection), neurological pain, musculoskeletal pain, cancer related pain or vascular pain.
■ A compound of the invention as defined herein for use in treating any one or more of nociception, somatic pain, visceral pain, acute pain, chronic pain, hyperalgesia, allodynia, post operative pain, pain due to hypersensivity, headache, inflammatory pain (rheumatic, dental, dysmenorrhoea or infection), neurological pain, musculoskeletal pain, cancer related pain or vascular pain.
» A method of treating pain in a patient such as a mammal (e.g. human), which method comprises administering to the patient a therapeutically effective amount of a compound of the invention as defined herein. ■ A method for the reduction or elimination of pain in a patient (e.g. a mammal such as a human) suffering from pain, which method comprises administering to the patient an effective pain-reducing or pain-eliminating amount of a compound of the invention as defined herein. " A method for the treatment of any one or more of nociception, somatic pain, visceral pain, acute pain, chronic pain, hyperalgesia, allodynia, post operative pain, pain due to hypersensivity, headache, inflammatory pain (rheumatic, dental, dysmenorrhoea or infection), neurological pain, musculoskeletal pain, cancer related pain or vascular pain, which method comprises administering to the patient a therapeutically effective amount of a compound of the invention as defined herein.
■ A compound of the invention as defined herein for use in the prophylaxis or treatment of stroke.
■ A method for the prophylaxis or treatment of stroke in a patient such as a mammal (e.g. human), which method comprises administering to the patient a therapeutically effective amount of a compound of the invention as defined herein.
■ A compound of the invention as defined herein for use as a neuroprotective agent.
■ A method of preventing or reducing neuronal damage in a patient suffering from stroke, which method comprises administering to the patient an effective neuroprotective amount of a compound of the invention as defined herein. ■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the prevention or reduction of risk of stroke in patients at risk for stroke, for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation. ■ A compound of the invention as defined herein for the prevention or reduction of risk of stroke in patients at risk for stroke, for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation.
■ A method for the prevention or reduction of risk of stroke in patients at risk for stroke, for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation, which method comprises administering to the patient an effective therapeutic amount of a compound of the invention as defined herein.
■ A compound of the invention as defined herein for use in the prophylaxis or treatment of a disease state or condition mediated by a cyclin dependent kinase 5. ■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a disease state or condition mediated by a cyclin dependent kinase 5.
■ A method for the prophylaxis or treatment of a disease state or condition mediated by a cyclin dependent kinase 5, which method comprises administering to a subject in need thereof a compound of the invention as defined herein.
■ A method for alleviating or reducing the incidence of a disease state or condition mediated by a cyclin dependent kinase 5, which method comprises administering to a subject in need thereof a compound of the invention as defined herein. ■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a disease state or condition mediated by cdk5 or p35.
■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a disease state or condition mediated by cdk5 or p35, said disease state or condition being other than Alzheimer's disease,
Huntington's disease or Creutzfeldt-Jakob disease.
■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a disease state or condition mediated by cdkδ or p35, said disease state or condition being other than a neurodegenerative disease.
■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a disease state or condition characterised by elevated levels of cdk5 or p35.
■ A compound of the invention as defined herein for use in the prophylaxis or treatment of a disease state or condition mediated by cdk5 or p35, said disease state or condition being other than Alzheimer's disease, Huntington's disease or Creutzfeldt-Jakob disease.
■ A compound of the invention as defined herein for use in the prophylaxis or treatment of a disease state or condition mediated by cdkδ or p35, said disease state or condition being other than a neurodegenerative disease. " A compound of the invention as defined herein for use in the prophylaxis or treatment of a disease state or condition characterised by elevated levels of cdk5 or p35.
■ A method of prophylaxis or treatment of a disease state or condition mediated by cdkδ or p35, said disease state or condition being other than Alzheimer's disease, Huntington's disease or Creuzfeldt-Jacob disease, which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein. ■ A method of prophylaxis or treatment of a disease state or condition mediated by cdk5 or p35, said disease state or condition being other than a neurodegenerative disease, which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein. " A method of prophylaxis or treatment of a disease state or condition characterised by elevated levels of cdk5 or p35, which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein.
■ A compound of the invention as defined herein for use in the prophylaxis or treatment of a neuropathy, such as a peripheral neuropathy, other than Alzheimer's disease,
Huntington's disease or Creutzfeldt-Jakob disease.
■ The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a neuropathy, such as a peripheral neuropathy, other than Alzheimer's disease, Huntington's disease or Creuzfeldt-Jacob disease.
■ A method of prophylaxis or treatment of a neuropathy, such as a peripheral neuropathy, other than Alzheimer's disease, Huntington's disease or Creuzfeldt-Jacob disease, which method comprises administering to a patient in need thereof a therapeutically effective amount of a compound of the invention as defined herein.
Anti-fungal, anti-protozoal, anti-viral and anti-parasitic activity
Compounds of the present invention and their acid addition salts and crystalline forms thereof have antifungal activity, anti-protozoal activity and anti-parasitic activity.
In particular, compounds of the invention are useful in treating infection by pathogenic fungi, protozoa and parasites where infection by the pathogen is normally associated with an antibody response to HSP90.
In one embodiment, the invention provides compounds of the formula (I) and sub-groups thereof as defined herein for use as anti-fungal agents.
Examples of fungi include those that are pathogenic in man and other animals, for example:
■ Candida species such as Candida albicans and Candida tropicalis; ■ Cryptococcus species such as Cryptococcus neoformans and Cryptococcal meningitis;
■ Aspergillus species such as Aspergillus fumigatus, Aspergillus flavus and Aspergillus niger,
■ Microsporum species such as Microsporum oanis and Microsporum gypseum;
■ Epidermophyton species;
■ Trichophyton species such as Trichophyton equinum, Trichophyton mentagrophytes and Trichophyton rubrum;
■ Epidermophyton floccosum; ■ Exophiala werneckii;
■ Fusarium species such as Fusarium solani; ' Sporothήx schenckii;
» Penicillium species such as Penicillium rubrum; ■ Altermaria species;
■ Ceratocystis pilifera;
■ Chrysosporium pruinosum;
■ Helminthsporium species;
■ Paecilomyces vaήotti; ■ yeasts, for example Saccharomyces cerevisiae and Pityrosporum species such as Pityrosporum orbiculare and Pityrosporum ovale;
■ Histoplasma species such as Histoplasma capsulatum;
■ Coccidiodes species;
■ Paracoccidioides species; and ■ Blastomyces species.
In another embodiment, the invention provides compounds of the formula (!) and sub-groups thereof as defined herein for use as anti-protozoal agents.
Examples of protozoa include:
■ Trypanosoma cruzi; ■ Leishmania species; for example the L. donovani complex (L donovani, L infantum, and
L chagasi); the L mexicana complex (3 main species - L. mexicana, L amazonensis, and L. venezuelensis); L tropica; L major, L aethiopica; and the subgenus Viannia with four main species (L. (V.) braziliensis, L (V.) guyanensis, L (V.) panamensis, and L (V.) peruviana); ■ Toxoplasma gondii; and
■ Trichomonas vaginalis.
In a further embodiment, the invention provides compounds of the formula (I) and sub-groups thereof as defined herein for use as anti-parasitic agents.
Examples of parasites include parasitic worms such as: ■ parasitic roundworms such as Ascaris lumbricoides;
' parasitic flatworms such as the parasitic trematode worms, e.g. Schistosoma mansoni
The invention also provides inter alia:
• A compound of the invention as defined herein for use in the prophylaxis or treatment of a fungal, protozoal or parasitic disease state or condition (other than a disease state or condition due to Plasmodium falciparum), for example a disease state or condition characterised by an antibody response to Hsp90. • The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a fungal, protozoal or parasitic disease state or condition (other than a disease state or condition due to Plasmodium falciparum), for example a disease state or condition characterised by an antibody response to Hsp90. • A method for the prophylaxis or treatment of a fungal, protozoal or parasitic disease state or condition (other than a disease state or condition due to Plasmodium falciparum), for example a disease state or condition characterised by an antibody response to Hsp90, which method comprises administering to a subject in need thereof a compound of the invention as defined herein. • A compound of the invention as defined herein for use in the prophylaxis or treatment of a fungal disease state or condition, for example a disease state or condition characterised by an antibody response to Hsp90.
• The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a fungal disease state or condition, for example a disease state or condition characterised by an antibody response to Hsp90.
• A method for the prophylaxis or treatment of a fungal disease state or condition, for example a disease state or condition characterised by an antibody response to Hsp90, which method comprises administering to a subject in need thereof a compound of the invention as defined herein. • A compound of the invention as defined herein for use in preventing, arresting or reversing the infection of an animal (such as a mammal, e.g. a human) by pathogenic fungi.
• The use of a compound of the invention as defined herein for the manufacture of a medicament for preventing, arresting or reversing the infection of an animal (such as a mammal, e.g. a human) by pathogenic fungi. • A method for preventing, arresting or reversing the infection of an animal (such as a mammal, e.g. a human) by pathogenic fungi, which method comprises administering to a subject in need thereof a compound of the invention as defined herein.
• A compound of the invention as defined herein for any of the uses and methods set forth above, and as described elsewhere herein. • The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of any of the disease states or conditions described herein.
• A combination of a compound of the invention as defined herein with an ancilliary compound which is an antifungal agent (e.g. an azole antifungal agent). • A pharmaceutical composition comprising a compound of the invention as defined herein with an ancilliary compound which is an antifungal agent (e.g. an azole antifungal agent).
• A compound of the invention as defined herein for use in preventing, reducing or reversing the development of resistance to an anti-fungal agent, anti-protozoal agent or anti-parasitic agent (preferably an anti-fungal agent) co-administered therewith.
• The use of a compound of the invention as defined herein for the manufacture of a medicament for coadministration with an anti-fungal agent, anti-protozoal agent or antiparasitic agent (preferably an anti-fungal agent) to prevent, reduce or reverse the development of resistance to the anti-fungal agent, anti-protozoal agent or anti-parasitic agent.
• A method of preventing or reducing development of resistance to an anti-fungal agent in a patient (e.g. a human patient), which method comprises administering to the patient a combination of an anti-fungal agent, anti-protozoal agent or anti-parasitic agent (preferably an anti-fungal agent) and a compound of the invention as defined herein.
• A method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90, which method comprises administering to a subject in need thereof a combination of a compound of the invention as defined herein with an anti-fungal, anti-protozoal or anti-parasitic drug, wherein the disease state or condition mediated by Hsp90 is the development of resistance to the anti-fungal, anti- protozoal or anti-parasitic drug.
• A method for: (i) sensitizing fungal, protozoal or parasite cells to an anti-fungal, antiprotozoal or anti-parasitic drug; (ii) alleviating or reducing the incidence of resistance to an anti-fungal, anti-protozoal or anti-parasitic drug ; (iii) reversing resistance to an anti-fungal, anti-protozoal or anti-parasitic drug; (iv) potentiating the activity of an anti-fungal, anti- protozoal or anti-parasitic drug; (v) delaying or preventing the onset of resistance to an anti-fungal, anti-protozoal or anti-parasitic drug, which method comprises administering to a subject in need thereof a combination of a compound of the invention as defined herein with the said anti-fungal, anti-protozoal or anti-parasitic drug.
• A method for the treatment of a fungal, protozoal or parasitic disease or condition, which method comprises administering to a subject in need thereof a combination of compound of the invention as defined herein with an anti-fungal, anti-protozoal or anti-parasitic drug, which method is characterized by the absence of drug resistance.
• A method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90 in a subject undergoing treatment with an anti-fungal, anti-protozoal or anti-parasitic drug, which method comprises administering to the subject a compound of the invention as defined herein, wherein the disease state or condition mediated by Hsp90 is the development of resistance to said anti-fungal, antiprotozoal or anti-parasitic drug.
• A method for: (i) sensitizing fungal, protozoal or parasite cells to an anti-fungal, anti- protozoal or anti-parasitic drug; (ii) alleviating or reducing the incidence of resistance to an anti-fungal, anti-protozoal or anti-parasitic drug (iii) reversing resistance to an anti-fungal, anti-protozoal or anti-parasitic drug; (iv) potentiating the activity of an an anti-fungal, antiprotozoal or anti-parasitic drug; (v) delaying or preventing the onset of resistance to an anti-fungal, anti-protozoal or anti-parasitic drug, which method comprises administering to a subject undergoing treatment with said ancillary compound a compound of the invention as defined herein.
• A method for the treatment of a fungal, protozoal or parasitic disease in a subject undergoing treatment with an anti-fungal, anti-protozoal or anti-parasitic drug, which method comprises administering to a subject in need thereof a compound of the invention as defined herein, which method is characterized by the absence of drug resistance e.g. to said anti-fungal, anti-protozoal or anti-parasitic drug).
As described above in the introductory part of this application, compounds having Hsp90 inhibitory activity have been found to exhibit potent anti-fungal activity and prevent the development of resistance to anti-fungals and in particular Hsp90 dependent resistance to anti-fungals . Moreover, it has been found that inhibition of Hsp90 activity can reduce the development of resistance to commonly used anti-fungal drugs such as the azoles. It is therefore envisaged that the compounds of the invention will be useful in the prophylaxis or treatment of a range of fungal diseases and conditions and will also be useful, when coadminstered with other anti-fungal drugs such as the azoles, in enhancing the activity of the anti-fungal drugs. The antifungal activity of ,the compounds of the present invention may be evaluated by determining the minimum fungistatic (inhibition) concentration (m.i.α). This test is usually performed by preparing a series of plates or tubes containing a suitable nutrient medium, each plate or tube also containing a different concentration of the test compound and then inoculating the medium with the fungal species. After an incubation period the plates are examined visually for the presence or absence of fungal growth. The m.i.c. is the minimum concentration required to prevent fungal growth.
The compounds may be used in animal medicine (for example in the treatment of mammals such as humans).
Fungal infections in animals against which compound of the invention as defined herein may be used include:
■ Superficial mycoses - i.e. fungal infections limited to the outermost layers of the skin and hair; ■ Cutaneous mycoses - i.e. fungal infections that extend deeper into the epidermis but are typically restricted to the keratinized layers of the skin, hair, and nails;
■ Subcutaneous mycoses - i.e. fungal infections involving the dermis, subcutaneous tissues, muscle, and fascia; ■ Systemic mycoses due to primary pathogens (these typically originate primarily in the lungs and may spread to other organ systems); and
■ Systemic mycoses due to opportunistic pathogens (infections of patients with immune deficiencies who would otherwise not be infected).
Particular examples of fungal disease states for which compounds of the invention as defined herein may be used include:
■ Dermatophyte infections such as tinea versiColour (a superficial fungal infection of the skin), tinea pedis (Athletes' Foot), tinea capitis (superficial fungal infection on the head), tinea barbae (fungal infection of bearded areas), tinea corporis (fungal infection of smooth skin areas).
■ Mucosal Candidiasis such as Oral Candidiasis, esophagitis and Vaginal candidiasis. ■ Invasive or deep organ candidiasis (e.g., fungemia, endocarditis, and endophthalmitis).
■ Crytpococcal infections such as Cryptococcal meningitis.
■ Histoplasmosis.
■ Blastomycosis, a fungal infection of the lungs and occasionally the skin.
■ Invasive Fungal Infections in patients with weakened immune systems or under treatment with anti-cancer or anti-AID drugs, for example Invasive Candidiasis and Invasive Aspergillosis.
■ Aspergilloses such as Allergic Bronchopulmonary Aspergillosis.
■ Aspergilloma.
> Intertrigo infections (fungal infections occuring in folds of skin e.g. between the toes or fingers, in the underarm area, or in the groin area). ■ Maduramycosis (fungal invasion of the tissue of the foot, also known as madura foot).
« Coccidioidomycosis.
■ Mucormycosis.
■ Blastomycosis
■ Geotrichosis. ■ Chromoblastomycosis.
■ Conidiosporosis. « Histoplasmosis.
■ Rhinosporidosis.
■ Nocaidiosis. ■ Para-actinomycosis.
• Penicilliosis.
■ Monoliasis.
■ Sporotrichosis.
Fungal infections of particular interest are Candidiasis and Aspergillosis. Compounds of the invention also have anti-protozoal activity and anti-parasitic activity. The antiprotozoal activity of the compounds of the present invention may be assessed by conventional methods, for example by determining the minimum inhibition concentration (m.i.c.) or 50% inhibition level (IC50). Examples of protozoal and parasitic diseases or conditions for which compounds of the invention may prove useful include:
■ Chagas disease ((trypanosomiasis) - an infection caused by the parasite Trypanosoma cruzi. * Ascariasis - a human disease caused by the parasitic roundworm Ascaris lumbricoides.
■ Leishmaniasis - a disease caused by parasites of the genus Leishmania. ■ Toxoplasmosis - a parasitic disease caused by the protozoan Toxoplasma gondii.
■ Schistosomiasis (Bilharzia) - a disease caused by the parasite Schistoma mansoni.
■ Trichomoniasis - a sexually transmitted disease caused by the parasitic protozoan Trichomonas vaginalis.
Anti-viral Activity As discussed above in the introductory sections of this application, infection of a host cell with viral RNA/DNA results in a substantial redirection of cellular protein synthesis towards key viral proteins encoded by the viral nucleic acid, and this frequently gives rise to upregulation of heat shock proteins. It is believed that one function of the HSP induction may be to assist in the stabilization and folding of the high levels of 'foreign' protein generated in preparation for virus replication and it has been shown (Naqkaqawa et al.) that HSP 90 inhibitors can block viral replication. Accordingly, the compounds of the invention are useful in combatting viral infections, for example by blocking or inhibiting viral replication.
Therefore, in another aspect, the invention provides a compound of the invention as defined herein for use in the prophylaxis or treatment of a viral infection (or viral disease). In further aspects, the invention provides:
• The use of a compound of the invention as defined herein for the manufacture of a medicament for the prophylaxis or treatment of a viral infection (or viral disease).
• A method for the prophylaxis or treatment of a viral infection (or viral disease), which method comprises administering to a subject in need thereof a compound of the invention as defined herein.
• A compound of the invention as defined herein for use in blocking or inhibiting viral replication in a host organism (e.g. an animal such as a mammal (e.g. human)).
• The use of a compound of the invention as defined herein for the manufacture of a medicament for use in blocking or inhibiting viral replication in a host organism (e.g. an animal such as a mammal (e.g. human)). • A method of blocking or inhibiting viral replication in a host organism (e.g. an animal such as a mammal (e.g. human)), which method comprises administering to the host organism a compound of the invention as defined herein.
Examples of viral infections that may be treated with the compounds of the invention include infections due to any one or more of the following viruses: o Picomaviruses such as rhinoviruses (common cold virus), Coxsackie virus (e.g. Coxsackie
B virus); and foot and mouth disease virus; o Hepatitis viruses such as hepatitis A virus (HAV), hepatitis B virus (HBV), hepatitis C virus
(HCV), hepatitis D virus (HDV) and hepatitis E virus (HEV), o Coronaviruses (e.g. common cold virus and Severe acute respiratory syndrome (SARS) virus) o Adenoviruses such as Human Adenoviruses (a cause of respiratory and conjunctival infections); o Astroviruses (a cause of flu-like symptoms); o Flaviviruses such as the Yellow Fever virus; o Orthomyxoviruses such as influenza viruses (e.g. influenza A, B and C viruses); o Parainfluenza viruses; o Respiratory syncytial virus; o Enteroviruses such as Poliovirus (Poliomyelitis virus); o Paramyxoviruses such as the Measles (rubeola) virus, mumps virus, respiratory syncytial virus (RSV) and canine distemper virus (CDV); o Togaviruses such as the Rubella (German Measles) virus and Sindbis virus; o Herpes viruses such as:
■ Herpes simplex virus (HSV), for example HSV-1 which causes fever blisters (cold sores), gingivostomatitis, herpes keratitis, eczema herpeticum and HSV encephalitis); and HSV-2 which causes genital lesions, neonatal infections, HSV meningitis, HSV proctitis; » Varicella zoster virus (VZV), which causes chickenpox, congenital varicella syndrome and shingles; ■ Epstein-Barr Virus (EBV), which causes infectious mononucleosis, Burkitt's lymphoma and nasopharyngeal cancer;
■ Cytomegalovirus (CMV), e.g. human cytomegalovirus (HCMV);
■ Human herpes virus 6 (HHV-6), which causes exanthum subitum or roseola infantum ■ Human herpes virus 8 (HHV-8) or Kaposi's sarcoma-associated herpes virus
(KSHV), which is found in the saliva of many AIDS patients and associated with Kaposi's sarcoma; o Papovaviridae such as polyoma virus and human papilloma virus (HPV); o Parvoviruses; o Poxviruses such as Variola virus (human smallpox virus); o Rhabdoviruses such as rabies virus and vesicular stomatitis virus (VSV); and o Retroviruses such as Human immunodefficiency virus (HIV) which is responsible for acquired immune defficiency syndrome (AIDS); and Human T-lymphotrophic virus (HTLV).
Particular viral infections against which the compounds of the invention may be used include herpes virus, pox virus, Epstein-Barr virus, Sindbis virus, adenovirus, HIV (for prevention of AIDS development in HIV-infected individuals), HPV, HCV and HCMV viruses.
The viral infection may be other than an infection with hepatitis C virus (HCV).
The activity of the compounds of the invention as agents for blocking or preventing viral replication in host organisms or host cells can be determined in accordance with standard procedures well known to the skilled person.
The compounds of the invention may be used as the sole antiviral agent or they may be used in conjunction with other anti-viral agents such as acyclovir, ganciclovir, oseltamavir (Tamiflu®) and zanamavir (Relenza®), amantidine, rimantadine, adefovir dipivoxil, interferons (e.g. interferon alfa- 2b and pegylated interferon alfa-2a), lamivudine, entecavir, ribavirin, famciclovir, valcicylovir, valacyclovir, azidothymidine (AZT - Retrovir®) , atazanavir, fosamprenavir, lamivudine, lamivudine + abacavir, tenofovir disoproxil fumarate, tenofovir disoproxil fumarate + emtricitabine, tipranavir, nelfinavir, indinavir, raltegravir, ritonavir, lopinavir + ritonavir, darunavir, amprenavir, enfuvirtide, saquinavir, hydroxyurea, VGV-1 and anti-viral vaccines.
Accordingly, the invention further provides:
• A combination of a compound of the invention as defined herein with an ancilliary compound which is an antiviral agent.
• A pharmaceutical composition comprising a compound of the invention as defined herein with an ancilliary compound which is an antiviral agent.
Methods for the Preparation of Compounds of the Formulae (Vl"), (Vl), (l°), (I) and subgroups thereof
In this section, as in all other sections of this application unless the context indicates otherwise, references to Formula (I) also include Formula (I0), Formula (Vl0) and all sub-groups (e.g. formula (Vl) and examples of Formula (I) as defined herein. Where a reference is made to a group R1, R2, R3, R4, R5, R6, R10 or any other "R" group, the definition of the group in question is as set out above and as set out in the following sections of this application unless the context requires otherwise.
Compounds of the formula (I) can be prepared from the corresponding compounds of the formula (I) in our earlier application PCT/GB2006/001382. The compounds of PCT/GB2006/001382, which will be referred to for convenience in this section of the present application as compounds of the formula (IX), have the formula:
Figure imgf000087_0001
wherein R1 is hydrogen or hydroxy; and R2 is hydrogen, hydroxy or methoxy; provided that at least one of R1' and R2' is hydroxy.
The compounds of formula (I) of the present invention can be prepared by the reaction of a compound of the formula (IX) with a compound of the formula RZ-LG, where LG is a leaving group, in the presence of a base, and typically with heating.
The nature of the leaving group will depend on the nature of the group Lp. For example, where Lp is C=O, and hence the compound of formula (I) is an ester, the leaving group may be a halogen or a carboxylate residue, i.e. the compound Rp1-C(=O)LG may be an acid chloride or an acid anhydride. The reaction of the compound (IX) with a compound of the formula Rp1-C(=O)-LG in an esterification reaction is carried out in the presence of a base, for example an organic base such as pyridine. The reaction is typically carried out with heating, for example to a temperature in excess of 100 0C, e.g. a temperature of approximately 120 0C.
Compounds of the formula (I) wherein Lp is a bond can be prepared by standard and well known methods of ether synthesis. For example, the compound of formula (IX) can be reacted with an alkali metal hydride such as sodium hydride in a polar non-protic solvent to form a phenolate anion which is then reacted with a compound of the formula Rp1-LG where LG is a halide leaving group such as chlorine or bromine to give the compound of formula (I). Alternatively, the compound of the formula (IX) can be reacted with a compound of the formula Rp1-OH under Mitsunobu alkylation conditions, e.g. in the presence of diethyl azodicarboxylate and triphenylphosphine. The Mitsunobu reaction is typically carried out in a polar non-protic solvent such as tetrahydrofuran at a moderate temperature such as ambient temperature.
Compounds of the formula (I) wherein Lp is (C=O)NRp1 can be prepared from compounds of the formula (IX) by reaction with a compound of the formula Rp1N(Rp1)-C(=O)-LG where LG is a halogen such as bromine or chlorine in the presence of a base such as pyridine, typically at an elevated temperature in excess of 100 0C, for example in excess of 130 0C.
Compounds of the formula (I) wherein Lp is S(O)xNRp1 can be prepared from compounds of the formula (IX) by reaction with a compound of the formula Rp1N(Rp1)S(O)x-LG [e.g. H2NS(O)xCI or Me2NS(O)xCI] where LG is a halogen such as bromine or chlorine either in the absence of base or in the presence of a non-interfering base such as sodium hydride, triethylamine or pyridine. The reaction is typically carried out in an inert polar solvent such as N,N-dimethylformamide, N1N- dimethylacetamide or N-methylpyrrolidinone either at room temperature or at an elevated temperature such as 100 0C or above, see for example the methods described by Leese et al [Bioorg. Med. Chem. Lett, 2004, 14, 3135], Okada et al [Tetrahedron Lett, 2000, 41 , 7047], Patel et al [Bioorg. Med. Chem. Lett, 2004, 14, 605] or Chu et al. [Bioorg. Med. Chem. Lett, 1999, 9, 141].
Compounds of the formula (I) wherein Lp is (C=O)O can be prepared from compounds of the formula (IX) by reaction with a compound of the formula Rp1O-(C=O)-LG [e.g. an alkoxycarbonyl halide or an alkoxy carbonic anhydride] where LG is a leaving group such as a halogen (e.g. chlorine or bromine) or an alkyl carbonate (e.g. fe/t-butyl carbonate) in the absence of base or in the presence of a non-interfering base such as sodium hydride, triethylamine or pyridine. The reaction is typically carried out in an inert polar solvent such as dichloromethane, ethyl acetate, tetrahydrofuran or N,N-dimethylformamide, either at room temperature or at an elevated temperature such as 100 0C or above, see for example the methods described by Yajima et al [Chem. Pharm. Bull., 1970, 18, 2574], Ramesh et al [J. Org. Chem., 2005, 70, 837], Wu et al [Synth. Commun., 2002, 32, 1615] or Bhat et a\ [Tetrahedron Lett, 2002, 43, 2467].
Compounds of the formula (I) wherein Rz is
Figure imgf000088_0001
can be prepared by the reaction of a compound of the of the formula (IX) with a compound of the formula:
Figure imgf000088_0002
in the presence of a non-interfering base such as pyridine or triethylamine.
Compounds of the formula (I) wherein Rz is a glucuronide residue can be prepared by synthetic chemical methods well known the the skilled person or by bio-transformation process, for example by incubating the compound of the formula (IX) with animal microsomes, such as dog microsomes or rat microsomes. By using microsomes from different animal species, different glucuronide isomers or mixtures of isomers can be obtained as described in the examples below.
Compounds of the formula (I) wherein Rz is SO3H can be prepared by reaction of a compound of the (IX) with chlorosulphonic acid in N,N-dimethylaniline.
Compounds of the formula (I) wherein Rz is a mono-, di- or tripeptide residue can be prepared by the reaction of a compound of the formula (IX) with a suitably protected acid chloride of an amino acid, dipeptide or tripeptide in accordance with known methods, followed by removal of any protecting groups. The reaction is typically carried out in a polar solvent in the presence of a non- interfering base such as triethylamine or pyridine; see for example the method described by Brenner et al, HeIv. Chim. Acta, 1957, 40, 1933. Examples of deprotection of the protecting groups can be found in Green et al (see below).
As an alternative to using the acid chloride method described above, the alcohol (IX) can be converted to the ester (I) by reaction of a suitably protected carboxylic acid of an amino acid, dipeptide or tripeptide in the presence of ester coupling reagents, followed by removal of any protecting groups. Examples of such reagents include 1 ,3-dicyclohexylcarbodiimide (DCC) (Johnson et al, JCS Chem. Comm., 1968, 73), DCC and 1-hydroxybenzotriazole (HOBt) (Kessler et al, Tet. Lett, 1984, 25, 3971) or by (benzotriazol-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate (BOP) (Castro et al, Synthesis, 1977, 413). Examples of deprotection of the protecting groups can be found in Green et al (see below). The coupling reaction is typically carried out in a non-aqueous, non-protic solvent such as acetonitrile, dioxan, dimethylsulphoxide, dichloromethane, dimethylformamide or N-methylpyrrolidine in the absence of base or in the presence of a non-interfering base such as triethylamine or pyridine.
Compounds of the formula (IX) can be prepared in accordance with synthetic methods well known to the skilled person. For example, compounds of the formula (IX) can be prepared by the reaction of a compound of the formula (X):
Figure imgf000089_0001
or an activated and/or protected form thereof, with an amine of the formula HNR5R6 under conditions suitable for forming an amide bond. The amines of the formula HNR5R6 are either commercially available or can be made using methods well known to the skilled person, see for example, Advanced Organic Chemistry, by Jerry March, 4th edition, 119, Wiley Interscience, New York; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN: 0-471-31192-8). The carboxylic acid (X) can be converted to an amide of the formula (IX) by first forming an acid chloride by treatment of the carboxylic acid with thionyl chloride, or by reaction with oxalyl chloride in the presence of a catalytic amount of dimethyl formamide, or by reaction of a potassium salt of the acid with oxalyl chloride. The acid chloride can then be reacted with the amine HNR5R6 in the presence of a non-interfering base such as triethylamine. The reaction may be carried out at around room temperature in a polar solvent such as dioxan.
As an alternative to using the acid chloride method described above, the carboxylic acid (X) can be converted to the amide (IX) by reaction with the amine HNR5R6 in the presence of amide coupling reagents of the type commonly used in the formation of amide or peptide linkages. Examples of such reagents include 1 ,1 '-carbonyldiimidazole (CDI), I.S-dicyclohexylcarbodiimide (DCC) (Sheehan et al, J. Amer. Chem Soc. 1955, 77, 1067), 1-ethyl-3-(3'-dimethylaminopropyl)- carbodiimide (referred to herein either as EDC or EDAC but also known in the art as EDCI and WSCDI) (Sheehan et al, J. Org. Chem., 1961, 26, 2525), uronium-based coupling agents such as O-(7-azabenzotriazol-1-yl)-Λ/JΛ/,Λ/',Λ/'-tetramethyluronium hexafluorophosphate (HATU) and phosphonium-based coupling agents such as 1-benzo-triazolyloxytris-(pyrrolidino)phosphonium hexafluorophosphate (PyBOP) (Castro et al, Tetrahedron Letters, 1990, 31, 205). Carbodiimide- based coupling agents are advantageously used in combination with 1-hydroxy-7-azabenzotriazole (HOAt) (L. A. Carpino, J. Amer. Chem. Soc, 1993, 115, 4397) or 1-hydroxybenzotriazole (HOBt) (Konig et al, Chem. Ber., 103, 708, 2024-2034). Preferred coupling reagents include EDC (EDAC) and DCC in combination with HOAt or HOBt.
One particular coupling reagent comprises EDC in combination with HOBt. A preferred coupling agent is 1 ,1 '-carbonyldiimidazole (CDI).
The coupling reaction is typically carried out in a non-aqueous, non-protic solvent such as acetonitrile, dioxan, dimethylsulphoxide, dichloromethane, dimethylformamide or N- methylpyrrolidine, or in an aqueous solvent optionally together with one or more miscible co- solvents. The reaction can be carried out at room temperature or, where the reactants are less reactive (for example in the case of electron-poor anilines bearing electron withdrawing groups such as sulphonamide groups) at an appropriately elevated temperature. The reaction may be carried out in the presence of a non-interfering base, for example a tertiary amine such as triethylamine or Λ/,Λ/-diisopropylethylamine.
Illustrative routes to the compounds of formula (IX) are described in more detail below.
Compounds of the formula (IX) in which the benzoyl moiety is derived from a 2-hydroxy-5- substituted benzoic acid can be prepared by the sequence of reactions shown in Scheme 1. The starting material for the synthetic route shown in Scheme 1 is 5-chloro-2-hydroxy benzoic acid, which can be obtained commercially. Conversion to the acid chloride is carried out by heating with thionyl chloride. The acid chloride may be used either in situ and reacted with various amines, or can be isolated as a stable white solid. Other simple 2-hydroxy-5-substituted benzoic acids may be used in this procedure to synthesise other amides of 2-hydroxy-5-substituted benzoic acids.
Figure imgf000091_0001
Scheme 1 : 5-Chloro-2-hydroxybenzoic acid amides
Compounds of formula (IX) can also be made according to the method shown in Scheme 2. The starting material for the synthetic route shown in Scheme 2 is 4-ethyl anisole, which can be obtained commercially. Conversion to the carboxylic acid can be carried out by lithiation at low temperature, followed by quenching of the resulting anion with solid carbon dioxide. The carboxylic acid may be coupled with various amines, using standard amide coupling reagents of the type commonly used in the formation of peptide linkages as described above.
Deprotection of the methyl ether can be effected using using boron tribromide (e.g. by the method described in Synthesis 1991 , 469) to give the compound of formula (IX). The method illustrated in Scheme 2 can be used to prepare other simple 2-hydroxy-5-substituted benzoic acids which can then be coupled to an appropriate amine to give the compounds of formula (IX). The process of coupling intermediates acids with amines, anilines or amino-heterocyclic compounds, followed by removal of any protecting groups, is straightforward and is suitable for the synthesis of large combinatorial libraries of molecules, useful for this invention. Examples of combinatorial libraries are described in Solid-Phase Synthesis and Combinatorial Technologies by Pierfausto Seneci. Wiley-lnterscience, New York. 2000. xii + 637 pp. ISBN 0471331953).
Figure imgf000092_0001
Scheme 2
Compounds of the Formula (IX) can also be made according to the methods desribed in Scheme 3. The starting material 3-fe/t-butyl-4-hydroxybenzoic acid (X = ferf-butyl) is commercially available and can be coupled using the amide coupling agents (as outlined above) with a broad range of amines of the formula HNR5R6 to give compounds of the invention. The other starting material illustrated in Scheme 3, 3-isopropyl-4-hydroxybenzoic acid (X = isopropyl), can be prepared according to a modification of a literature procedure using carbon tetrachloride and copper powder in a Friedel-Crafts type reaction, in which the intermediate species is hydrolysed to the carboxylic acid (J Chem Soc, Chem Commun 1985, 1134). The Friedel Crafts method can be used to prepare other simple 2-hydroxy-3-substituted benzoic acids.
Figure imgf000092_0002
X = isopropyl or tert-butyl
Scheme 3: 3-Alkyl-4-hydroxybenzoic acid amides Compounds of the formula (IX) can also be made according to the method described in Scheme 4. 2,4-Dihydroxy-5-isopropyl-benzoic acid amides can be prepared by amide coupling using coupling reagents (as outlined above) from a bi-benzyl ether protected intermediate, shown in the scheme, followed by catalytic hydrogenation using hydrogen gas and palladium on carbon. The benzoic acid intermediate itself is made by Friedel-Crafts acylation of 2,4-dihydroxybenzoic acid methyl ester (from commercial sources) using a literature procedure (J. Ind. Chem. Soc, 1953, 30, 269). Typically, Friedel-Crafts acylation of a phenol is carried out by treatment of the phenol with an acylating agent (such as an acid chloride or acid anhydride) in the presence of a Lewis acid catalyst (such as boron trifluoride or aluminium chloride) either at room temperature or at more elevated temperatures (60-120 0C). As an alternative to a direct Friedel-Crafts acylation, the 4- hydroxy group of the 2,4-dihydroxybenzoic acid methyl ester can be O-acylated using an acid anhydride such as acetic anhydride in the presence of a base such as 4-dimethylaminopyridine (4- DMAP) to give a'2-hydroxy-4-acyloxy-benzoic acid methyl ester. Treatment of the 2-hydroxy-4- acyloxy-benzoic acid methyl ester with trifluoromethanesulphonic acid in the presence of an acyl chloride such as acetyl chloride then gives the 2,4-dihydroxy-5-acylbenzoic acid methyl ester.
Benzyl protection of the phenol groups, the Wittig reaction of the ketone to the olefin and ester hydrolysis (saponification) can be carried out under standard conditions, well known to those skilled in the art of organic synthesis (for example see, Advanced Organic Chemistry, by Jerry March, 4th edition, 119, Wiley Interscience, New York; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN: 0-471-31192-8). For example, the Wittig reaction can be carried out in an inert polar solvent (such as tetrahydrofuran) and can involve treatment of an aldehyde or ketone with a phosphorus ylide species that may be prepared by the reaction of a phosphonium salt with a base (such as butyl lithium or potassium tert- butoxide). The ester hydrolysis to the carboxylic acid is usually carried out by treatment with an aqueous alkali metal hydroxide such sodium hydroxide. The saponification reaction may be carried out using an organic co-solvent such as an alcohol (e.g. methanol) and the reaction mixture is typically heated to a non-extreme temperature, for example up to about 50-60 0C.
It is to be understood that other 2,4-dihydroxy-5-substituted benzoic acids could be made using this procedure to synthesise different examples of compounds of formula (IX) not specifically exemplified herein.
In Scheme 4, as an alternative to the use of the Wittig reagent MePPH3Br to form the olefin (XXVI), the ketone (XXV) can be reacted with methyl magnesium bromide under standard Grignard reaction conditions to give an intermediate hydroxy compound which is then dehydrated to the olefin by reaction with a suitable reagent such as sodium acetate and acetic acid.
Figure imgf000094_0001
(XXVIII)
Scheme 4: 2,4-Dihydroxy-5-isopropyl-benzoic acid amides
The intermediate compound 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (XXVII) in Scheme 4 can be made using a variety of methods well known to the skilled person. For example, compound (XXVII) can be made by the synthetic route illustrated in Scheme 4A.
Figure imgf000094_0002
Scheme 4A
As shown in Scheme 4A, 5-bromo-2,4-dihydroxybenzoic acid is benzylated using benzyl bromide in the presence of a base such as potassium carbonate to give the bis-benzyloxy-bromobenzoic acid benzyl ester (XXX). The ester (XXX) is then reacted with potassium isoprenyl trifluoroborate in the presence of a palladium (0) or palladium (II) compound and a base to give the isopropenyl- bis benzyl ester (XXXI). The palladium compound can be a palladium (0) compound such as Pd(PPh3)4 or a palladium (II) compound such as [1 ,1'-bis(diphenyl~phosphino)ferrocene]- dichloropalladium(ll). The base can be an organic base such as n-butylamine or an inorganic base such as a metal carbonate, e.g. caesium carbonate. The reaction with potassium isoprenyl trifluoroborate is typically carried out at reflux temperature for a prolonged period, for example 15 hours or more. The resulting isopropenyl bis-benzyloxy ester (XXXI) is then hydrolysed to give the carboxylic acid (XXVII) using, for example, an alkali metal hydroxide such as lithium hydroxide, typically with heating to a non-extreme temperature..
Compounds of the formula (IX) can also be made according to the route illustrated in Scheme 5. A- Hydroxy-3-(1',2'-dimethyl-propyl)-benzoic acid amides can be prepared by amide coupling using standard coupling agents (as outlined above) from the alkyl substituted acid. The olefinic acid itself can be prepared by Claisen rearrangement of a precursor ether, as shown in the scheme, by thermal rearrangement in anisole, followed by saponification, which in this case can yield more than one isomer of the olefin, the major one being shown in the scheme. Such Claisen reactions are well known in the literature, e.g. see J. Chem. Soc, Perkin Trans 1 1981 , 897. The ether itself can be prepared by simple alkylation of commercially available 4-hydroxy benzoic acid ethyl ester. The alkylation and saponification reactions are simple modifications that can be carried out under various conditions (for example see, Advanced Organic Chemistry, by Jerry March, 4th edition, 119, Wiley Interscience, New York; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN; 0-471-31192-8)). It is to be understood that other 4- hydroxy-3-substituted benzoic acids could be made using this procedure to synthesise different examples of compounds of formula 1 not specifically exemplified herein.
Figure imgf000096_0001
Scheme 5: 4-Hydroxy-3-(1>,2'-dimethyl-propyl)-benzoic acid amides
Compounds of the formula (IX) can also be made according to the method shown in Scheme 6. 2,4-Dihydroxy-5-bromobenzoic acid is used as the starting material, which is commercially available. Simple protection and deprotection gives the benzoic acid precursor (for example see, Advanced Organic Chemistry, by Jerry March, 4th edition, 119, Wiley Interscience, New York; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN: 0-471-31192-8)), which can be used in amide coupling reactions with a range of amines (as outlined above). These precursor amides can be subjected to Suzuki cross coupling procedures to make alkyl substituted compounds. A broad range of Suzuki coupling conditions are described in the literature, and the ones used here were taken from J. Am. Chem. Soc. 2003, 11148. Suzuki coupling chemistry is also broadly applicable to synthesis of alkyl-aryl and aryl-aryl compounds. The Suzuki reaction is typically carried out in the presence of a palladium catalyst such as bis(tri-f-butylphosphine)-palladium and a base (e.g. a carbonate such as potassium carbonate). The reaction may be carried out in an aqueous solvent system, for example aqueous ethanol, and the reaction mixture is typically subjected to heating, for example to a temperature in excess of 100 0C, Many boronates suitable for use in preparing compounds of the invention are commercially available, for example from Boron Molecular Limited of Noble Park, Australia, or from Combi-Blocks Inc, of San Diego, USA. Where the boronates are not commercially available, they can be prepared by methods known in the art, for example as described in the review article by N. Miyaura and A. Suzuki, Chem. Rev. 1995, 95, 2457. Thus, boronates can be prepared by reacting the corresponding bromo-compound with an alkyl lithium such as butyl lithium and then reacting with a borate ester. The resulting boronate ester derivative can, if desired, be hydrolysed to give the corresponding boronic acid. The final products of the reaction sequence illustrated in Scheme 6 are formed by catalytic hydrogenation (as outlined above) to remove the benzyl protecting groups and to reduce the olefin, formed in the Suzuki reaction to the alkyl substituent. It is to be understood that other 2,4-dihydroxy-5-substituted benzoic acids could be made using this procedure to synthesise different examples of compounds of formula I not specifically exemplified herein.
Figure imgf000097_0001
Scheme 6: 2,4-Dihydroxy-5-(alkyl)-benzoic acid amides
Compounds of the formula (IX) wherein NR5R6 is an optionally substituted isoindoline group, for example as in compounds of the formulae (VII) and (Vila), can be prepared by the methods illustrated in Scheme 7, or methods analogous thereto.
Figure imgf000098_0001
Scheme 7
As shown in Scheme 7, an optionally substituted 1 ,2-dimethylbenzene (Xl) is heated with N- bromosuccinimide in the presence of dibenzoyl peroxide to give the dibromo-compound (XII). The reaction is typically carried out in carbon tetrachloride with heating at reflux. The dibromo- compound (XlI) is then reacted with a compound PG-NH2 where PG is a protecting group such as tosyl or para-methoxybenzyl in the presence of a base such as a metal hydride (e.g. sodium hydride), when PG is a tosyl group, or an alkali metal carbonate (e.g. sodium carbonate), when PG is para-methoxybenzyl. The protecting group PG can then be removed to give the amine (XIV). Thus, for example, a tosyl group can be removed by heating with a mxture of phenol, hydrobromic acid and propanoic acid, whereas a para-methoxybenzyl group can be removed in standard manner using trifluoroacetic acid and anisole. The amine (XIV) is then coupled with a carboxylic acid of the formula (X) as described above.
In a variation on the reaction sequence of Scheme 7, one or more functional groups R10b present in the protected isoindoline (XIII) or the deprotected isoindoline compound (XIV) can be converted into other groups R1Ob. For example, where the group R10b in compound (XIV) is a nitro group, it can be reduced to give the corresponding amino group, for example by catalytic hydrogenation in the presence of a palladium on charcoal catalyst. In a further example, when R1Ob in the compound (XIII) is an ester group (e.g. CO2Me), it can be hydrolysed to give a carboxylic acid which can then be reacted with an amine such as morpholine to give the corresponding amide. Further functional group interconversions may subsequently be carried out (for example reduction of the amide to the corresponding aminomethyl compound with lithium aluminium hydride) before removal of the protecting group PG. Alternatively, the ester group can be reduced to a hydroxymethyl group which may then be converted to an aldehyde, and the aldehyde group may subsequently be used as a substrate for further conversions, for example a reductive amination reaction with an amine such as morpholine, piperidine or N-methyl piperazine.
A variation on the route illustrated in Scheme 7 is shown in Scheme 7A
Figure imgf000099_0001
Scheme 7A
In Scheme 7A, the commercially available bis-bromomethyl benzoic acid ester ((LI) is reacted with benzylamine in a polar aprotic solvent such as as tetrahydrofuran (THF) in the presence of a non- interfering base such as triethylamine to give the N-benzyl dihydroisoindole intermediate (LII). The ester group in intermediate (LII) is then reduced to the corresponding alcohol using lithium aluminium hydride in THF to give the hydroxymethyldihydroisoindole intermediate (LIII). Debenzylation of the hydroxymethyldihydroisoindole intermediate (LIII) is then carried out by hydrogenation over palladium on charcoal catalyst in an alcohol (e.g. ethanol) solvent at a mildly elevated temperature (e.g. up to about 50 °) to give the intermediate (LIV). Intermediate (LIV) is then converted to intermediate (LV) by reaction with a reagent suitable for introducing a benzyloxycarbonyl ("Z") group onto the nitrogen atom of the dihydroisoindole ring. For example, the intermediate (LIV) can be reacted with benzyl chloroformate in a polar non-protic solvent such as THF in the presence of a non-interfering base such as triethylamine to give intermediate (LV).
An alternative synthesis of the isoindoline compound (XIV) is shown in Scheme 8.
Figure imgf000100_0001
Scheme 8
The starting material for Scheme 8 is the ortho diester (XV) which is hydrolysed to the corresponding dicarboxylic acid (XVI) using an alkali metal hydroxide such as potassium hydroxide before being subjected to cyclisation to the phthalic anhydride (XVII) by reaction with acetic anhydride. The phthalic anhydride (XVII) can be converted to the corresponding phthalimide (XVIII) by reaction with formamide at an elevated temperature (e.g. approximately 210 0C). The phthalimide (XVIiI) can then be reduced to the isoindoline (XIV) using a suitable reducing agent such as borane in tetrahydrofuran.
Isoindoline intermediate compounds can also be prepared be prepared by the synthetic route shown in Scheme 9.
Figure imgf000101_0001
Scheme 9
In Scheme 9, the dipropargylamine (LVI) is reacted with N-(benzyloxycarbonyloxy) succinimide (LVII) in ethyl acetate in the presence of potassium carbonate to give the Z-protected dipropargylamine (LVIII) (the term "Z" referring to a benzyloxycarbonyl group). As an alternative to N-(benzyloxycarbonyloxy) succinimide, benzyl chloroformate may be used to introduce the benzyloxycarbonyl protecting group. Compound (LVIII) is then reacted with propargyl alcohol (LIX) in the presence of Wilkinson's catalyst in a 2+2+2 cycloaddition reaction to give the Z-protected isoindoline (LX). The hydroxymethyl group on the isoindoline (LX) is then converted to a mesyloxy group by reaction with methanesulphonyl chloride in a polar solvent such as THF in the presence of a non-interfering base such as triethylamine to give the mesyl compound (LXI). The mesyl compound (LXI) is reacted with alkylpiperazine (LXII) in acetone solution to give the Z-protected isoindoline (LXIII). Removal of the benzyloxycarbonyl group to give the unprotected isoindoline compound (LXIV) is then accomplished by hydrogenation over a palladium on charcoal catalyst.
A variation on the reaction sequence shown in Scheme 9 is illustrated in Scheme 9A.
Figure imgf000102_0001
Scheme 9A
In Scheme 9A, rather than being converted to the mesylate (LXI), the hydroxymethylindoline (LX) is oxidised to the corresponding aldehyde (LXV) using manganese dioxide in dichloromethane, and the aldehyde is then converted to a compound of the formula (LXIII) by reaction with a compound of the formula (LXII) under reductive amination conditions, e.g. in the presence of sodium triacetoxyborohydride. The Z-group is then removed by hydrogenation as described above in respect of Scheme 9 to give the intermediate (LXIV).
Compounds of the formula (VIIb) as defined herein can be prepared by the reaction of a compound of the formula (XIX) or a protected derivative thereof with a compound of the formula (XX):
Figure imgf000102_0002
wherein n, R3, R4a, R8 and R1Occ are as defined herein, under amide forming conditions as described above and in the examples.
Within formula (XX), particular intermediates of the invention can be represented by the formula (XXI):
Figure imgf000103_0001
D25 wherein n is 0 or 1 ; M is N or CHOH and R is hydrogen or methyl; provided that when n is 0 and R25 is methyl, then M is CHOH.
Particular intermediates within formula (XXI) are the compounds (XXII), (XXIII) and (XXIV) below.
Figure imgf000103_0003
The intermediates of formula (XXI) can be made by methods well known to the skilled person or methods analogous thereto. For example, intermediate XXII can be prepared by lithium-halogen exchange of a suitably N-protected 5-bromoisoindoline, quenching with 1-methyl-4-piperidone and subsequent deprotection. Intermediate XXII can be prepared by Buchwald palladium coupling of 4-BOC-piperazine and a suitably N-protected 5-bromoisoindoline followed by subsequent deprotection. One method of preparation for intermediate XXIV is from a suitably N-protected isoindoline-5-carboxylic acid, Weinreb amide formation, reduction to the aldehyde, followed by reductive amination and subsequent deprotection.
The above synthetic routes involve the formation of a phenolic compound of the formula (IX) followed by the subsequent reaction of the phenolic group in the compound of formula (IX) with a reagent suitable for introducing the group Rz. As an alternative, however, the group Rz can be introduced into the benzoic acid of the formula (X), or a protected derivative thereof, to give a compound of the formula (XXV) or a protected derivative thereof:
Figure imgf000103_0002
where R1 and R2 are as defined herein.
The reactions used to introduce the group Rz may be analogous to the reactions described above for introducing the group Rz into compounds of the formula (IX), except that the carboxylic acid group may require protection (e.g. as an ester) under some of the reaction conditions, and then subsequent deprotection.
Following introduction of the group Rz, the compounds of formula (XXV) can be reacted with an amine of the formula HNR5R6 under conditions analogous to those described elsewhere herein.
Once formed, where the substituent groups permit, one compound of the formula (I), or a protected form thereof, can be converted into another compound of the formula (I). For example, when R3 is bromine, the bromine atom can be replaced by trifluoromethyl by reaction with a trifluoroacetate salt (e.g. sodium trifluoroacetate), and copper (I) iodide in a polar solvent such as dimethylformamide.
In another procedure, compounds of the formula (I) wherein R8 is fluorine can be prepared from compounds of the formula (I) where R8 is hydrogen by electrophilic fluorination. Electrophilic fluorination can be carried out using a fluorinating agent such as 1-(chloromethyl)-4-fluoro-1 ,4- diazoniabicyclo[2.2.2]octane bis(tetrafluoroborate) or similar N-fluoro-diazonia compounds.
Many of the procedures described below and used in this synthesis are well known to those skilled in the art, and examples of alkylations, acylations, functional group interconversions and reagents and conditions for carrying out such conversions can be found in, for example, Advanced Organic Chemistry, by Jerry March, 4th edition, 119, Wiley Interscience, New York; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN: 0-471- 31192-8).
As well as the specific examples, and the methods of preparation outlined below, it is understood that modification to the routes described would allow synthesis of many further examples of compounds claimed in Formula (I). For example, alternative benzoic acid starting materials with differing or additional substitution patterns could be prepared.
In many of the reactions described above, it may be necessary to protect one or more groups to prevent reaction from taking place at an undesirable location on the molecule. Examples of protecting groups, and methods of protecting and deprotecting functional groups, can be found in Protective Groups in Organic Synthesis (T. Green and P. Wuts; 3rd Edition; John Wiley and Sons, 1999).
A hydroxy group may be protected, for example, as an ether (-OR) or an ester (-OC(=O)R), for example, as: a t-butyl ether; a benzyl, benzhydryl (diphenylmethyl), or trityl (triphenylmethyl) ether; a trimethylsilyl or t-butyldimethylsilyl ether; or an acetyl ester (-OC(=O)CH3, -OAc). When the hydroxy group is a phenolic hydroxy group, for example in compounds of the formula (I) wherein R1 and/or R2 are hydroxy, a preferred protecting group is a benzyl group.
An aldehyde or ketone group may be protected, for example, as an acetal (R-CH(OR)2) or ketal (R2C(OR)2), respectively, in which the carbonyl group (>C=O) is converted to a diether (>C(0R)2), by reaction with, for example, a primary alcohol. The aldehyde or ketone group is readily regenerated by hydrolysis using a large excess of water in the presence of acid. An amine group may be protected, for example, as an amide (-NRCO-R) or a urethane (-NRCO-OR), for example, as: a methyl amide (-NHCO-CH3); a benzyloxy amide (-NHCO-OCH2C6H5, -NH-Cbz); as a t-butoxy amide (-NHCO-OC(CH3)3, -NH-Boc); a 2-biphenyl-2-propoxy amide (-NHCO-OC(CH3)2CSH4C6H5, -NH-Bpoc), as a 9-fluorenylmethoxy amide (-NH-Fmoc), as a 6-nitroveratryloxy amide (-NH-Nvoc), as a 2-trimethylsilylethyloxy amide (-NH-Teoc), as a 2,2,2-trichloroethyloxy amide (-NH-Troc), as an allyloxy amide (-NH-Alloc), or as a 2(-phenylsulphonyl)ethyloxy amide (-NH-Psec). Other protecting groups for amines, such as cyclic amines and heterocyclic N-H groups, include toluenesulphonyl (tosyl) and methanesulphonyl (mesyl) groups and benzyl groups such as a para- methoxybenzyl (PMB) group. A carboxylic acid group may be protected as an ester for example, as: an C1-7 alkyl ester (e.g., a methyl ester; a t-butyl ester); a C1-7 haloalkyl ester (e.g., a Ci-7 trihaloalkyl ester); a triC1-7 alkylsilyl-C1-7alkyl ester; or a C5.2o aryl-C1-7 alkyl ester (e.g., a benzyl ester; a nitrobenzyl ester); or as an amide, for example, as a methyl amide. A thiol group may be protected, for example, as a thioether (-SR), for example, as: a benzyl thioether; an acetamidom ethyl ether (-S-CH2NHC(=O)CH3).
Methods of Purification
The compounds may be isolated and purified by a number of methods well known to those skilled in the art and examples of such methods include chromatographic techniques such as column chromatography (e.g. flash chromatography) and HPLC. Preparative LC-MS is a standard and effective method used for the purification of small organic molecules such as the compounds described herein. The methods for the liquid chromatography (LC) and mass spectrometry (MS) can be varied to provide better separation of the crude materials and improved detection of the samples by MS. Optimisation of the preparative gradient LC method will involve varying columns, volatile eluents and modifiers, and gradients. Methods are well known in the art for optimising preparative LC-MS methods and then using them to purify compounds. Such methods are described in Rosentreter U, Huber U.; Optimal fraction collecting in preparative LC/MS; J Comb Chem.; 2004; 6(2), 159-64 and Leister W, Strauss K, Wisnoski D, Zhao Z, Lindsley C, Development of a custom high-throughput preparative liquid chromatography/mass spectrometer platform for the preparative purification and analytical analysis of compound libraries; J Comb Chem.] 2003; 5(3); 322-9.
Alternatively, normal phase preparative LC based methods might be used in place of reverse phase methods. Most preparative LC-MS systems utilise reverse phase LC and volatile acidic modifiers, since the approach is very effective for the purification of small molecules and because the eluents are compatible with positive ion electrospray mass spectrometry. Employing other chromatographic solutions e.g. normal phase LC, alternatively buffered mobile phase, basic modifiers etc as outlined in the analytical methods described above could alternatively be used to purify the compounds. Pharmaceutical Formulations
While it is possible for the active compound to be administered alone, it is preferable to present it as a pharmaceutical composition (e.g. formulation) comprising at least one active compound of the invention together with one or more pharmaceutically acceptable carriers, adjuvants, excipients, diluents, fillers, buffers, stabilisers, preservatives, lubricants, or other materials well known to those skilled in the art and optionally other therapeutic or prophylactic agents; for example agents that reduce or alleviate some of the side effects associated with chemotherapy. Particular examples of such agents include anti-emetic agents and agents that prevent or decrease the duration of chemotherapy-associated neutropenia and prevent complications that arise from reduced levels of red blood cells or white blood cells, for example erythropoietin (EPO), granulocyte macrophage- colony stimulating factor (GM-CSF), and granulocyte-colony stimulating factor (G-CSF).
Thus, the present invention further provides pharmaceutical compositions, as defined above, and methods of making a pharmaceutical composition comprising admixing at least one active compound, as defined above, together with one or more pharmaceutically acceptable carriers, excipients, buffers, adjuvants, stabilizers, or other materials, as described herein. The term "pharmaceutically acceptable" as used herein pertains to compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of a subject (e.g. human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio. Each carrier, excipient, etc. must also be "acceptable" in the sense of being compatible with the other ingredients of the formulation.
Accordingly, in a further aspect, the invention provides compounds of the formula (I) and subgroups thereof as defined herein in the form of pharmaceutical compositions.
The pharmaceutical compositions can be in any form suitable for oral, parenteral, topical, intranasal, ophthalmic, otic, rectal, intra-vaginal, or transdermal administration. Where the compositions are intended for parenteral administration, they can be formulated for intravenous, intramuscular, intraperitoneal, subcutaneous administration or for direct delivery into a target organ or tissue by injection, infusion or other means of delivery. The delivery can be by bolus injection, short term infusion or longer term infusion and can be via passive delivery or through the utilisation of a suitable infusion pump. Pharmaceutical formulations adapted for parenteral administration include aqueous and nonaqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats, co- solvents, organic solvent mixtures, cyclodextrin complexation agents, emulsifying agents (for forming and stabilizing emulsion formulations), liposome components for forming liposomes, gellable polymers for forming polymeric gels, lyophilisation protectants and combinations of agents for, inter alia, stabilising the active ingredient in a soluble form and rendering the formulation isotonic with the blood of the intended recipient. Pharmaceutical formulations for parenteral administration may also take the form of aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents (R. G. Strickly, Solubilizing Excipients in oral and injectable formulations, Pharmaceutical Research, VoI 21(2) 2004, p 201-230).
A drug molecule that is ionizable can be solubilized to the desired concentration by pH adjustment if the drug's pKa is sufficiently away from the formulation pH value. The acceptable range is pH 2-12 for intravenous and intramuscular administration, but subcutaneously the range is pH 2.7-9.0. The solution pH is controlled by either the salt form of the drug, strong acids/bases such as hydrochloric acid or sodium hydroxide, or by solutions of buffers which include but are not limited to buffering solutions formed from glycine, citrate, acetate, maleate, succinate, histidine, phosphate, tris(hydroxymethyl)- aminomethane (TRIS), or carbonate. The combination of an aqueous solution and a water-soluble organic solvent/surfactant (i.e., a cosolvent) is often used in injectable formulations. The water-soluble organic solvents and surfactants used in injectable formulations include but are not limited to propylene glycol, ethanol, polyethylene glycol 300, polyethylene glycol 400, glycerin, dimethylacetamide (DMA), N-methyl-2-pyrrolidone (NMP; Pharmasolve), dimethylsulphoxide (DMSO), Solutol HS 15, Cremophor EL, Cremophor RH 60, and polysorbate 80. Such formulations can usually be, but are not always, diluted prior to injection.
Propylene glycol, PEG 300, ethanol, Cremophor EL, Cremophor RH 60, and polysorbate 80 are the entirely organic water-miscible solvents and surfactants used in commercially available injectable formulations and can be used in combinations with each other. The resulting organic formulations are usually diluted at least 2-fold prior to IV bolus or IV infusion. Alternatively increased water solubility can be achieved through molecular complexation with cyclodextrins.
Liposomes are closed spherical vesicles composed of outer lipid bilayer membranes and an inner aqueous core and with an overall diameter of <100 μm. Depending on the level of hydrophobicity, moderately hydrophobic drugs can be solubilized by liposomes if the drug becomes encapsulated or intercalated within the liposome. Hydrophobic drugs can also be solubilized by liposomes if the drug molecule becomes an integral part of the lipid bilayer membrane, and in this case, the hydrophobic drug is dissolved in the lipid portion of the lipid bilayer. A typical liposome formulation contains water with phospholipid at 5-20 mg/ml, an isotonicifier, a pH 5-8 buffer, and optionally cholesterol.
The formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. The pharmaceutical formulation can be prepared by lyophilising a compound of Formula (I) or acid addition salt thereof. Lyophilisation refers to the procedure of freeze-drying a composition. Freeze-drying and lyophilisation are therefore used herein as synonyms. A typical process is to solubilise the compound and the resulting formulation is clarified, sterile filtered and aseptically transferred to containers appropriate for lyophilisation (e.g. vials). In the case of vials, they are partially stoppered with lyo-stoppers. The formulation can be cooled to freezing and subjected to lyophilisation under standard conditions and then hermetically capped forming a stable, dry lyophile formulation. The composition will typically have a low residual water content, e.g. less than 5% e.g. less than 1 % by weight based on weight of the lyophile.
The lyophilisation formulation may contain other excipients for example, thickening agents, dispersing agents, buffers, antioxidants, preservatives, and tonicity adjusters. Typical buffers include phosphate, acetate, citrate and glycine. Examples of antioxidants include ascorbic acid, sodium bisulphite, sodium metabisulphite, monothioglycerol, thiourea, butylated hydroxytoluene, butylated hydroxyl anisole, and ethylenediaminetetraacetic acid salts. Preservatives may include benzoic acid and its salts, sorbic acid and its salts, alkyl esters of para-hydroxybenzoic acid, phenol, chlorobutanol, benzyl alcohol, thimerosal, benzalkonium chloride and cetylpyridinium chloride. The buffers mentioned previously, as well as dextrose and sodium chloride, can be used for tonicity adjustment if necessary.
Bulking agents are generally used in lyophilisation technology for facilitating the process and/or providing bulk and/or mechanical integrity to the lyophilized cake. Bulking agent means a freely water soluble, solid particulate diluent that when co-lyophilised with the compound or salt thereof, provides a physically stable lyophilized cake, a more optimal freeze-drying process and rapid and complete reconstitution. The bulking agent may also be utilised to make the solution isotonic.
The water-soluble bulking agent can be any of the pharmaceutically acceptable inert solid materials typically used for lyophilisation. Such bulking agents include, for example, sugars such as glucose, maltose, sucrose, and lactose; polyalcohols such as sorbitol or mannitol; amino acids such as glycine; polymers such as polyvinylpyrrolidine; and polysaccharides such as dextran.
The ratio of the weight of the bulking agent to the weight of active compound is typically within the range from about 1 to about 5, for example of about 1 to about 3, e.g. in the range of about 1 to 2.
Alternatively they can be provided in a solution form which may be concentrated and sealed in a suitable vial. Sterilisation of dosage forms may be via filtration or by autoclaving of the vials and their contents at appropriate stages of the formulation process. The supplied formulation may require further dilution or preparation before delivery for example dilution into suitable sterile infusion packs.
Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets. In one preferred embodiment of the invention, the pharmaceutical composition is in a form suitable for i.v. administration, for example by injection or infusion.
In another preferred embodiment, the pharmaceutical composition is in a form suitable for subcutaneous (s.c.) administration. Pharmaceutical dosage forms suitable for oral administration include tablets, capsules, caplets, pills, lozenges, syrups, solutions, powders, granules, elixirs and suspensions, sublingual tablets, wafers or patches and buccal patches.
Pharmaceutical compositions containing compounds of the formula (I) can be formulated in accordance with known techniques, see for example, Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, PA, USA.
Thus, tablet compositions can contain a unit dosage of active compound together with an inert diluent or carrier such as a sugar or sugar alcohol, eg; lactose, sucrose, sorbitol or mannitol; and/or a non-sugar derived diluent such as sodium carbonate, calcium phosphate, calcium carbonate, or a cellulose or derivative thereof such as methyl cellulose, ethyl cellulose, hydroxypropyl methyl cellulose, and starches such as corn starch. Tablets may also contain such standard ingredients as binding and granulating agents such as polyvinylpyrrolidone, disintegrants (e.g. swellable crosslinked polymers such as crosslinked carboxymethylcellulose), lubricating agents (e.g. stearates), preservatives (e.g. parabens), antioxidants (e.g. BHT), buffering agents (for example phosphate or citrate buffers), and effervescent agents such as citrate/bicarbonate mixtures. Such excipients are well known and do not need to be discussed in detail here.
Capsule formulations may be of the hard gelatin or soft gelatin variety and can contain the active component in solid, semi-solid, or liquid form. Gelatin capsules can be formed from animal gelatin or synthetic or plant derived equivalents thereof.
The solid dosage forms (eg; tablets, capsules etc.) can be coated or un-coated, but typically have a coating, for example a protective film coating (e.g. a wax or varnish) or a release controlling coating. The coating (e.g. a Eudragit ™ type polymer) can be designed to release the active component at a desired location within the gastro-intestinal tract. Thus, the coating can be selected so as to degrade under certain pH conditions within the gastrointestinal tract, thereby selectively release the compound in the stomach or in the ileum or duodenum. Alternatively or additionally, the coating can be used as a taste masking agent to mask unpleasant tastes such as bitter tasting drugs. The coating may contain sugar or other agents that assist in masking unpleasant tastes.
Instead of, or in addition to, a coating, the drug can be presented in a solid matrix comprising a release controlling agent, for example a release delaying agent which may be adapted to selectively release the compound under conditions of varying acidity or alkalinity in the gastrointestinal tract. Alternatively, the matrix material or release retarding coating can take the form of an erodible polymer (e.g. a maleic anhydride polymer) which is substantially continuously eroded as the dosage form passes through the gastrointestinal tract. As a further alternative, the active compound can be formulated in a delivery system that provides osmotic control of the release of the compound. Osmotic release and other delayed release or sustained release formulations may be prepared in accordance with methods well known to those skilled in the art. The pharmaceutical formulations may be presented to a patient in "patient packs" containing an entire course of treatment in a single package, usually a blister pack. Patient packs have an advantage over traditional prescriptions, where a pharmacist divides a patient's supply of a pharmaceutical from a bulk supply, in that the patient always has access to the package insert contained in the patient pack, normally missing in patient prescriptions. The inclusion of a package insert has been shown to improve patient compliance with the physician's instructions.
Compositions for topical use include ointments, creams, sprays, patches, gels, liquid drops and inserts (for example intraocular inserts). Such compositions can be formulated in accordance with known methods.
Compositions for parenteral administration are typically presented as sterile aqueous or oily solutions or fine suspensions, or may be provided in finely divided sterile powder form for making up extemporaneously with sterile water for injection.
Examples of formulations for rectal or intra-vaginal administration include pessaries and suppositories which may be, for example, formed from a shaped moldable or waxy material containing the active compound. Thus, unit-dose suppositories or pessaries may be prepared by admixture of the active ingredient with one or more conventional solid carriers, for example coca butter, and shaping the resulting mixture. Further examples of mouldable waxy materials include polymers such as high molecular weight polyalkylene glycols, e.g. high molecular weight polyethylene glycols.
Alternatively, in the case of vaginal administration, the formulation may be presented as a tampon impregnated with the active ingredients and optionally one or more excipients or diluents. Other formulations suitable for rectal and vaginal administration include creams, gels, foams, pastes and sprays.
Further examples of topical compositions include dressings such as bandages and adhesive plasters impregnated with active ingredients and optionally one or more excipients or diluents. Carriers which may be used include e.g. polyhydric alcohols such as polyethylene glycols, propylene glycol or glycerol. Suitable excipients are those known in the art to be appropriate.
Compositions for administration by inhalation may take the form of inhalable powder compositions or liquid or powder sprays, and can be administrated in standard form using powder inhaler devices or aerosol dispensing devices. Such devices are well known. For administration by inhalation, the powdered formulations typically comprise the active compound together with an inert solid powdered diluent such as lactose. The compounds of the formula (I) will generally be presented in unit dosage form and, as such, will typically contain sufficient compound to provide a desired level of biological activity. For example, a formulation may contain from 1 nanogram to 2 grams of active ingredient, e.g. from 1 nanogram to 2 milligrams of active ingredient. Within this range, particular sub-ranges of compound are 0.1 milligrams to 2 grams of active ingredient (more usually from 10 milligrams to 1 gram, e.g. 50 milligrams to 500 milligrams), or 1 microgram to 20 milligrams (for example 1 microgram to 10 milligrams, e.g. 0.1 milligrams to 2 milligrams of active ingredient).
For oral compositions, a unit dosage form may contain from 1 milligram to 2 grams, more typically 10 milligrams to 1 gram, for example 50 milligrams to 1 gram, e.g. 100 miligrams to 1 gram, of active compound.
The active compound will be administered to a patient in need thereof (for example a human or animal patient) in an amount sufficient to achieve the desired therapeutic effect.
Methods of Treatment
It is envisaged that the compounds of the formula (I) and sub-groups as defined herein will be useful in the prophylaxis or treatment of a range of disease states or conditions mediated by Hsp90 client proteins. Examples of such disease states and conditions are set out above.
The compounds are generally administered to a subject in need of such administration, for example a human or animal patient, preferably a human.
The compounds will typically be administered in amounts that are therapeutically or > prophylactically useful and which generally are non-toxic. However, in certain situations (for example in the case of life threatening diseases), the benefits of administering a compound of the formula (I) may outweigh the disadvantages of any toxic effects or side effects, in which case it may be considered desirable to administer compounds in amounts that are associated with a degree of toxicity. The compounds may be administered over a prolonged term to maintain beneficial therapeutic effects or may be administered for a short period only. Alternatively they may be administered in a continuous manner or in a manner that provides persistent intermittent dosing (e.g. a pulsatile manner).
A typical daily dose of the compound of formula (I) can be in the range from 100 picograms to 100 milligrams per kilogram of body weight, more typically 5 nanograms to 25 milligrams per kilogram of bodyweight, and more usually 10 nanograms to 15 milligrams per kilogram (e.g. 10 nanograms to 10 milligrams, and more typically 1 microgram per kilogram to 20 milligrams per kilogram, for example 1 microgram to 10 milligrams per kilogram) per kilogram of bodyweight although higher or lower doses may be administered where required. The compound can be administered on a daily basis or on a repeat basis every 2, or 3, or 4, or 5, or 6, or 7, or 10 or 14, or 21 , or 28 days for example. In one particular dosing schedule, a patient will be given an infusion of a compound for periods of one hour to 4 hours daily for up to ten days in particular up to two days for one week, every two weeks in three, and the treatment repeated at a desired interval such as three to six weeks, in particular every three weeks. More particularly, a patient may be given an infusion of a compound for periods of one hour daily twice a week for two weeks in three weeks and the treatment repeated every three weeks.
Alternatively, a patient may be given an infusion of a compound for periods of one hour daily twice a week for three weeks in four weeks and the treatment repeated every four weeks.
In another particular dosing schedule, a patient will be given an infusion of a compound for periods of one hour daily for up to ten days in particular up to five days for one week, and the treatment repeated at a desired interval such as two to four weeks, in particular every three weeks.
More particularly, a patient may be given an infusion of a compound for periods of one hour daily for 5 days and the treatment repeated every three weeks.
In another particular dosing schedule, a patient is given an infusion over 30 minutes to 1 hour followed by maintenance infusions of variable duration, for example 1 to 5 hours, e.g. 3 hours.
In a further particular dosing schedule, a patient is given a continuous infusion for a period of 12 hours to 5 days, an in particular a continuous infusion of 24 hours to 72 hours.
Ultimately, however, the quantity of compound administered and the type of composition used will be commensurate with the nature of the disease or physiological condition being treated and will be at the discretion of the physician.
The compounds as defined herein can be administered as the sole therapeutic agent or they can be administered in combination therapy with one of more other compounds for treatment of a particular disease state, for example a neoplastic disease such as a cancer as hereinbefore defined. Examples of other therapeutic agents or treatments that may be administered together (whether concurrently or at different time intervals) with the compounds of the formula (I) include but are not limited to:
• Topoisomerase I inhibitors
• Antimetabolites • Tubulin targeting agents
• DNA binder and topoisomerase II inhibitors
• Alkylating Agents
• Monoclonal Antibodies.
• Anti-Hormones • Signal Transduction Inhibitors
• Proteasome Inhibitors • DNA methyl transferases
• Cytokines and retinoids
• Chromatin targeted therapies, e.g. HDAC or HAT modulators
• Radiotherapy; and • Other therapeutic or prophylactic agents; for example agents that reduce or alleviate some of the side effects associated with chemotherapy. Particular examples of such agents include anti-emetic agents and agents that prevent or decrease the duration of chemotherapy-associated neutropenia and prevent complications that arise from reduced levels of red blood cells or white blood cells, for example erythropoietin (EPO), granulocyte macrophage-colony stimulating factor (GM-CSF), and granulocyte-colony stimulating factor (G-CSF). Also included are agents that inhibit bone resorption such as bisphosphonate agents e.g. zoledronate, pamidronate and ibandronate, agents that suppress inflammatory responses (such as dexamethazone, prednisone, and prednisolone) and agents used to reduce blood levels of growth hormone and IGF-I in acromegaly patients such as synthetic forms of the brain hormone somatostatin, which includes octreotide acetate which is a long-acting octapeptide with pharmacologic properties mimicking those of the natural hormone somatostatin. Further included are agents such as leucovorin, which is used as an antidote to drugs that decrease levels of folic acid, or folinic acid itself and agents such as megestrol acetate which can be used for the treatment of side-effects including oedema and thromboembolic episodes.
For the case of Hsp90 inhibitors combined with other therapies, the two or more treatments may be given in individually varying dose schedules and via different routes.
Where the compound is administered in combination therapy with one, two, three, four or more other therapeutic agents (preferably one or two, more preferably one), the compounds can be administered simultaneously or sequentially. When administered sequentially, they can be administered at closely spaced intervals (for example over a period of 5-10 minutes) or at longer intervals (for example 1 , 2, 3, 4 or more hours apart, or even longer periods apart where required), the precise dosage regimen being commensurate with the properties of the therapeutic agent(s).
The compounds of the invention may also be administered in conjunction with non- chemotherapeutic treatments such as radiotherapy, photodynamic therapy, gene therapy; surgery and controlled diets.
For use in combination therapy with another chemotherapeutic agent, the compound and one, two, three, four or more other therapeutic agents can be, for example, formulated together in a dosage form containing two, three, four or more therapeutic agents. In an alternative, the individual therapeutic agents may be formulated separately and presented together in the form of a kit, optionally with instructions for their use.
A person skilled in the art would know through his or her common general knowledge the dosing regimes and combination therapies to use. Methods of Diagnosis
Prior to administration of a compound, a patient may be screened to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against Hsp90. For example, a biological sample taken from a patient may be analysed to determine whether a condition or disease, such as cancer, that the patient is or may be suffering from is one which is characterised by a genetic abnormality or abnormal protein expression which leads to the mutation or over-activation of an Hsp90 client protein. Examples of such abnormalities that result in activation of Hsp90 client proteins include; Bcr-ABL translocation, Flt-3 internal duplication, and mutation of Braf, or over-expression of ErbB2.
Thus, the patient may be subjected to a diagnostic test to detect a marker characteristic of up- regulation. The term diagnosis includes screening. By marker we include genetic markers including, for example, the measurement of DNA composition to identify mutations of Braf, BCR- abl, and Flt3 or other affected client proteins. The term marker also includes proteins such as ErbB2, including levels or concentrations of the protein or some fragments or degradation product and for enzymes the enzymic activity. The protein (e.g. phosphorylated or not) and mRNA levels of the aforementioned proteins could also be assessed to characterise a change in activity. For example the level of phosphorylated AKT can be an indicator of sensitivity to HSP90 inhibitors
The diagnostic tests are typically conducted on a biological sample selected from for example tumour biopsy samples, blood samples (isolation and enrichment of shed tumour cells), stool biopsies, sputum, chromosome analysis, pleural fluid, peritoneal fluid, buccal spears or biopsy or from urine.
The screening process will typically involve direct sequencing, oligonucleotide or protein microarray analysis, proteomic analysis by mass spectrometry, immunohistochemical techniques or detection using a specific antibody.
Methods of identification and analysis of mutations and up-regulation of proteins are well known to a person skilled in the art. Screening methods could include, but are not limited to, standard methods such as reverse-transcriptase polymerase chain reaction (RT-PCR), in-situ hybridisation or immunoblotting. In screening by RT-PCR, the level of mRNA in the tumour is assessed by creating a cDNA copy of the mRNA followed by amplification of the cDNA by PCR. Methods of PCR amplification, the selection of primers, and conditions for amplification, are known to a person skilled in the art. Nucleic acid manipulations and PCR are carried out by standard methods, as described for example in Ausubel, F. M. et al., eds. Current Protocols in Molecular Biology, 2004, John Wiley & Sons Inc., or Innis, M.A. et-al., eds. PCR Protocols: a guide to methods and applications, 1990, Academic Press, San Diego. Reactions and manipulations involving nucleic acid techniques are also described in Sambrook et al., 2001 , 3rd Ed1 Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press. Alternatively a commercially available kit for RT-PCR (for example Roche Molecular Biochemicals) may be used, or methodology as set forth in United States patents 4,666,828; 4,683,202; 4,801,531; 5,192,659, 5,272,057, 5,882,864, and 6,218,529 and incorporated herein by reference, An example of an in-situ hybridisation technique for assessing mRNA expression would be fluorescence in-situ hybridisation (FISH) (see Angerer, 1987 Meth. Enzymol., 152: 649).
Generally, in situ hybridization comprises the following major steps: (1) fixation of tissue to be analyzed; (2) prehybridization treatment of the sample to increase accessibility of target nucleic acid, and to reduce non-specific binding; (3) hybridization of the mixture of nucleic acids to the nucleic acid in the biological structure or tissue; (4) post-hybridization washes to remove nucleic acid fragments not bound in the hybridization, and (5) detection of the hybridized nucleic acid fragments. The probes used in such applications are typically labelled, for example, with radioisotopes or fluorescent reporters. Preferred probes are sufficiently long, for example, from about 50, 100, or 200 nucleotides to about 1000 or more nucleotides, to enable specific hybridization with the target nucleic acid(s) under stringent conditions. Commercially available FISH probes also exist for cytogenetic detection of chromosome rearrangemrnts, which can be used to detect Flt3 and Bcr-Abl translocations within leukeamia cell populations. Standard methods for carrying out FISH are described in Ausubel, F. M. et al., eds. Current Protocols in Molecular Biology, 2004, John Wiley & Sons lnc and Fluorescence In Situ Hybridization: Technical Overview by John M. S. Bartlett in Molecular Diagnosis of Cancer, Methods and Protocols, 2nd ed.; ISBN: 1-59259-760-2; March 2004, pps. 077-088; Series: Methods in Molecular Medicine.
Methods for gene expression profiling are described by (DePrimo et al., BMC Cancer 2003, 3:3). Briefly, the protocol is as follows: double-stranded cDNA is synthesized from total RNA Using a (dT)24 oligomer for priming first-strand cDNA synthesis, followed by second strand cDNA synthesis with random hexamer primers. The double-stranded cDNA is used as a template for in vitro transcription of cRNA using biotinylated ribonucleotides. cRNA is chemically fragmented according to protocols described by Affymetrix (Santa Clara, CA, USA), and then hybridized overnight on Human Genome Arrays.
Alternatively, the protein products expressed from the mRNAs may be assayed by immunohistochemistry of tumour samples, solid phase immunoassay with microtiter plates,
Western blotting, 2-dimensional SDS-polyacrylamide gel electrophoresis, ELISA, flow cytometry and other methods known in the art for detection of specific proteins. Detection methods would include the use of site specific antibodies. The skilled person will recognize that all such well- known techniques for detection of the "Philadelphia chromosome" indicative of BCR-ABL translocation.
Therefore, all of these techniques could also be used to identify tumours particularly suitable for treatment with the compounds of the invention. EXAMPLES
The invention will now be illustrated, but not limited, by reference to the specific embodiments described in the following examples.
In the examples, the following abbreviations may be used. AcOH acetic acid
BOC te/f-butyloxycarbonyl
Bn benzyl
CDI 1 ,1-carbonyldiimidazole
DMAW90 Solvent mixture: DCM: MeOH, AcOH, H2O (90:18:3:2) DMAW120 Solvent mixture: DCM: MeOH, AcOH, H2O (120:18:3:2)
DMAW240 Solvent mixture: DCM: MeOH, AcOH, H2O (240:20:3:2)
DCM dichloromethane
DMF dimethylformamide
DMSO dimethyl sulphoxide EDC 1-ethyl-3-(3'-dimethylaminopropyl)-carbodiimide
Et3N triethylamine
EtOAc ethyl acetate
Et2O diethyl ether h hour(s) HOAt 1-hydroxyazabenzotriazole
HOBt 1-hydroxybenzotriazole
MeCN acetonitrile
MeOH methanol min. minutes P. E. petroleum ether r.t. room temperature
SiO2 silica
TBTU N,N,N',N'-tetramethyl-0-(benzotriazol-1-yl)uronium tetrafluoroborate
THF tetrahydrofuran Proton magnetic resonance (1H NMR) spectra were recorded on a Bruker AV400 instrument operating at 400.13 MHz, in DMSO-d6 or MeOH-d4 (as indicated) at 27 0C, unless otherwise stated and are reported as follows: chemical shift δ/ppm (number of protons, multiplicity where s=singlet, d=doublet, t=triplet, q=quartet, m=multiplet, br=broad). The residual protic solvent was used as the internal reference. In the examples, the compounds prepared were characterised by liquid chromatography and mass spectroscopy using the system and operating conditions set out below. Where atoms with different isotopes are present and a single mass quoted, the mass quoted for the compound is the monoisotopic mass (i.e. 35CI; 79Br etc.). Different systems were used, as described below, and these were equipped with, and were set up to run under, closely similar operating conditions. The operating conditions used are also described below.
System description:
System 1 (analytical system): HPLC System: Waters 2795
Mass Spec Detector: Micromass Platform LC
PDA Detector: Waters 2996 PDA
System 2 (preparative and analytical system):
HPLC System: Waters Fractionlynx system Mass Spec Detector: Waters ZQ
PDA Detector: Waters 2996 PDA
System 3 (preparative and analytical system):
HPLC System: Agilent 1 100 system
Mass Spec Detector: LC/MSD UV Detector: Agilent MWD
Operating conditions: Acidic analytical conditions:
Eluent A: H2O (0.1% Formic Acid)
Eluent B: CH3CN (0.1 % Formic Acid) Gradient: 5-95% eluent B over 3.5 minutes (over 15 minutes w/ column 2)
Flow: 0.8 ml/min
Column 1 : Phenomenex Synergi 4μ MAX-RP 8OA, 2.0 x 50 mm
Column 2: Phenomenex Synergi 4μ MAX-RP 8OA, 2.0 x 150 mm
Basic analytical conditions:
Eluent A: H2O (1OmM NH4HCO3 buffer adjusted to pH=9.2 with NH4OH)
Eluent B: CH3CN
Gradient: 5-95% eluent B over 3.5 minutes
Flow: 0.8 ml/min
Column: Phenomenex Gemini 5μ 2. O x 50 mm MS conditions (Waters systems):
Capillary voltage: 3.6 kV (3.40 kV on ES negative)
Cone voltage: 25 V
Source Temperature: 12O 0C
Scan Range: 125-800 amu lonisation Mode: ElectroSpray Positive, Negative or Positive & Negative
MS conditions (Agilent systems):
Capillary voltage: 4000 V (3500 V on ES Negative)
Fragmentor/Gain: 150 / 1 Drying gas Temp/flow: 350 0C / 13.0 Lmin"1 Nebuliser pressure: 50 psig
Scan Range: 125-800 amu lonisation Mode: ElectroSpray Positive or Negative
The starting materials for each of the Examples are commercially available unless otherwise specified.
PREPARATION OF PHENOLIC STARTING MATERIALS A. GENERAL SYNTHETIC METHODS
In the following general methods, the volumes stated may vary according to the scale of the reaction, as will be apparent to the skilled person. METHOD A1
Amide coupling (acid chloride method)
A mixture of a carboxylic acid (1 equivalent) and thionyl chloride (1.5 equivalents) in benzene (or toluene) was stirred and held at reflux for 2 hours. Excess amine was added dropwise to the hot solution and the mixture stirred at room temperature for 15 minutes. Alternatively, the acid chloride could be isolated by evaporation and then re-dissolved in a 9:1 mixture of dichloromethane: triethylamine and the amine then added and the mixture stirred under nitrogen at room temperature for 1-18 hours. In either case, the mixture was diluted with ethyl acetate and extracted successively with water, saturated aqueous sodium bicarbonate and 2M hydrochloric acid. The organic layer was reduced to dryness in vacuo and the pure products were obtained either by trituration with ethyl acetate or by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether) or in a few cases by preparative HPLC/MS.
METHOD A2
Amide coupling (EDC, HOBt method)
A stirred solution of the acid (1 equivalent) in dichloromethane (10 ml) was treated successively with N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.2 equivalents), 1- hydroxybenzotriazole (1.2 equivalents) and the amine (1.5 equivalents) and the mixture was stirred at room temperature overnight. The mixture was washed successively with 2M hydrochloric acid and 2M sodium hydroxide, the organic layer was separated and the solvent removed in vacuo to afford the products. The products were either obtained pure or were purified by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether or methanol in ethyl acetate as appropriate).
METHOD A3
Anisole or benzyl ether dealkylation (BBr2 method) A stirred solution of the anisole or benzyl ether (1 equivalent) in dichloromethane at 0 0C was treated dropwise with a 1 M solution of boron tribromide in dichloromethane (1.5 equivalents per group to be deprotected) and the mixture was stirred for 2 hours. The reaction was quenched by the addition of water and saturated aqueous sodium bicarbonate, the organic layer was separated and the solvent was removed in vacuo. The pure products were obtained either by trituration with diethyl ether or ethyl acetate or by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether).
METHOD A4
Amide coupling (EDC, HOAt method)
A stirred solution of the acid (1 equivalent) in dimethylformamide (5 ml) was treated successively with N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.2 equivalents), 1-hydroxy-7- aza-benzotriazole (1.2 equivalents) and the amine (1.5 equivalents) and the mixture was stirred at room temperature overnight. DMF was evaporated and crude dissolved in EtOAc and was washed successively with saturated sodium bicarbonate, the organic layer was separated and the solvent removed in vacuo. The products were either obtained pure or were purified by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether or methanol in ethyl acetate as appropriate).
METHOD A5 Hvdroqenation
A stirred solution of protected derivative (1 equivalent) and a catalytic amount of 10% palladium on carbon (typically 30-50 mg) in ethanol (5-10 ml), methanol (5-10 ml) or methanol/DCM (3ml / 3ml) was stirred at room temperature under an atmosphere of hydrogen for 2-16 hours. The catalyst was removed by filtration, washed with methanol (5 ml) and the solvent removed in vacuo to afford the products. Some required purification by flash chromatography, eluting typically with ether.
METHOD A6 Suzuki coupling
The aryl bromide (1 equivalent, typically 0.5 mmol), boronic acid or potassium vinyl trifluoroborate derivative (1.2 equivalents) and caesium carbonate (3 equivalents) were dissolved in THF (10 ml) under nitrogen. 1 ,1'-Bis(diphenylphosphino)ferrocene] palladium(ll) dichloride (0.1 equivalent) was added and then water (1 ml). The mixture begins to darken until black. The mixture was then heated at reflux under nitrogen until the reaction is complete (8-45 hrs). The mixture was cooled, diluted with DCM and magnesium sulphate added. The mixture was filtered and the solvent evaporated. The resulting residues were purified by flash chromatography in pet. ether / ether mixtures, and generally gave product in good yield (~60-80%).
METHOD A7 Resorcinol mono-O-methylation
Dimethyl sulphate (1 equivalent) was added to a stirred solution of the resorcinol (1 equivalent) and potassium carbonate (2.2 equivalents) in acetonitrile (10 ml per mmol of substrate) and the mixture was stirred at room temperature for 16 hours. The solvent was removed in vacuo, the residue partitioned between dichloromethane and water, the organic layer separated and the solvent removed in vacuo. The pure products were obtained either after column chromatography on silica (eluting with mixtures of petroleum ether and ethyl acetate) or by preparative HPLC/MS.
METHOD A8
Electrophilic aromatic fluorination
1-(Chloromethyl)-4-fluoro-1 ,4-diazoniabicyclo[2.2.2]octane bis(tetrafluoroborate) (1 equivalent) was added to a solution of the substrate (1 equivalent) in acetonitrile (15 ml per mmol of substrate) and the mixture was stirred at room temperature for 16 hours. The solvent was removed in vacuo and the residue was partitioned between ethyl acetate and water. The organic layer was separated and reduced to dryness in vacuo. The pure products were obtained either after column chromatography on silica (eluting with mixtures of petroleum ether and ethyl acetate) or by preparative HPLC/MS.
B. SYNTHESIS OF CARBOXYLIC ACID INTERMEDIATES
PREPARATION B1
4-Hvdroxy-3-isopropylbenzoic acid
Figure imgf000120_0001
Carbon tetrachloride (28 ml, 0.26 mol) and copper powder (1.0 g) were added to a stirred solution of 2-isopropylphenol (27.2 g, 0.2 mol) in 50% aqueous sodium hydroxide (120 ml) and the mixture was held at reflux for 16 hours. Upon cooling the mixture was acidified to pH 2 or below by the addition of concentrated hydrochloric acid and was extracted with ethyl acetate. The organic layer was extracted with a saturated aqueous solution of sodium bicarbonate and the aqueous layer acidified to pH 2 or below by the very careful addition of concentrated hydrochloric acid. The solution was extracted with ethyl acetate, the organic layer was washed with water, separated and the solvent removed in vacuo to afford 4-hydroxy-3-isopropylbenzoic acid (12.5 g, 35%) as a bright red solid that was used without further purification. 1H NMR (DMSOd6) 12.36 (1 H, br s), 10.13 (1 H, br s), 7.73 (1 H, d), 7.63 (1 H, dd), 6.85 (1 H, d), 3.22 (1 H, m), 1.19 (6H, d). MS: [M-H]+ 179.
Alternatively, if required, the crude product may be purified using a three step procedure involving di-benzylation [according to the conditions outlined below in Preparation B5 for the synthesis of methyl 5-acetyl-2,4-bis-benzyloxybenzoate (BnBr, K2CO3, MeCN, reflux)], column chromatography on silica to remove highly coloured impurites (eluting with 3-5% ethyl acetate in petroleum ether) and catalytic hydrogenation [according to Method A5 outlined above (10% Pd/C, EtOH, H2)] to afford 4-hydroxy-3-isopropylbenzoic acid as a colourless solid.
PREPARATION B2 5-Ethyl-2-methoxybenzoic acid
Figure imgf000121_0001
n-Butyl lithium (2.5M in hexanes, 38.5 m , 100.0 mmol) was added dropwise under a nitrogen atomosphere to a stirred solution of 4-ethylanisole (11.7 g, 86.0 mmol) and Λ/,Λ/,Λ/',Λ/'- tetramethylethylenediamine (10 ml, 88.0 mmol) in anhydrous diethyl ether (100 ml) and the mixture was stirred and held at 3O0C for 16 hours. The mixture was cooled and poured slowly in to a mixture of excess solid carbon dioxide in anhydrous diethyl ether. Upon warming to room temperature the mixture was made basic by the addition of 2M sodium hydroxide, the aqueous layer was separated and acidified to pH 2 or below by the addition of concentrated hydrochloric acid. The mixture was extracted with diethyl ether, the organic layer separated and the solvent removed in vacuo to afford 5-ethyl-2-methoxybenzoic acid (5.7 g, 37%) as a pale yellow oil. 1H
NMR (DMSO-d6) 12.50 (1 H, br s), 7.48 (1 H, d), 7.33 (1 H, dd), 7.03 (1 H, d), 2.56 (2H, q), 1.17 (3H, q). MS: [M+H]+ 181.
PREPARATION B3 2,4-Bis-benzyloxy-5-chloro-benzoic acid
Figure imgf000121_0002
WO 2004/050087
1-(2,4-Bis-benzyloxy-5-chloro-phenyl)-ethanone [prepared as per WO 2004/0500087] (1.10 g, 3.0 mmol) was added to a stirred solution of sodium hydroxide (1.20 g, 30.0 mmol) in water (10 ml) and dioxane (10 ml). Bromine (1.44 g, 9.0 mmol) was added dropwise and the mixture stirred at room temperature for 3 hours. The dioxane was removed by evaporation in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with ethyl acetate, the organic layer separated and the solvent removed in vacuo to afford 2,4-bis- benzyloxy-5-chloro-benzoic acid (900 mg, 81 %) as a pale yellow solid. 1H NMR (DMSO-d6) 12.58 (1H1 br s), 7.77 (1 H1 s), 7.55-7.30 (10H, m), 7.11 (1H, s), 5.31 (2H, s), 5.27 (2H, s). MS: [M+H]+ 369.
PREPARATION B4
3-(1 ,2-Dimethyl-allyl)-4-hydroxy-benzoic acid
Figure imgf000122_0001
Ethyl 4-hydroxybenzoate (1.66 g, 10.0 mmol) and anhydrous potassium carbonate (2.07 g, 15.0 mmol) in acetonitrile (30 ml) was treated with 3-methyl-2-butenyl chloride (1.35 ml, 12.0 mmol) and the mixture was stirred and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between dichloromethane and water. The organics were separtaed and the solvent removed in vacuo to afford ethyl 4-(3-methyl-but-2-enyloxy)-benzoate (2.23 g, 95%) as a pale yellow liquid which was used without further purification. 1H NMR (DMSO- dβ) 7.89 (2H, d), 7.04 (2H, d), 5.44 (1 H, t), 4.62 (2H, d), 4.28 (2H, q), 1.77 (3H, s), 1.73 (3H, s), 1.31 (3H, t). MS: [M+H]+ 235.
Ethyl 4-(3-methyl-but-2-enyloxy)-benzoate (2.23 g, 9.53 mmol) was dissolved in anisole (8 ml) and the mixture stirred and held at reflux for 4 days. The solvent was removed in vacuo and the residue subjected to column chromatography on silica. Elution with 20% ethyl acetate in petroleum ether afforded ethyl 3-(1 ,2-dimethyl-allyl)-4-hydroxy-benzoate (600 mg, 27%) as a colorless solid. 1H NMR (DMSOd6) 10.32 (1 H, br s), 7.67 (1 H, dd), 7.62 (1 H, s), 6.90 (1 H, d), 4.90 (1H, s), 4.85 (1 H, s), 4.25 (2H, q), 3.75 (1 H, q), 1.61 (3H, s), 1.30 (3H, t), 1.26 (3H, d). MS: [M+H]+ 235.
Ethyl 3-(1 ,2-dimethyl-allyl)-4-hydroxy-benzoate (600 mg, 2.56 mmol) was dissolved in methanol (20 ml), a solution of potassium hydroxide (560 mg, 10.0 mmol) in water (10 ml) was added and the mixture was stirred and held at reflux for 16 hours. Upon cooling the methanol was removed in vacuo and the solution acidified to pH 2 or below by the addition of 2M hydrochloric acid. The solution was extracted with dichloromethane, the organic layer was separated and the solvent was removed in vacuo to afford 3-(1 ,2-dimethyl-allyl)-4-hydroxy-benzoic acid (270 mg, 51 %) as a colourless gum. 1H NMR (DMSO-d6) 12.38 (1 H, br s), 10.22 (1 H, br s), 7.63 (2H, m), 6.88 (1 H, d), 4.90 (1 H, s), 4.87 (1 H, s), 3.75 (1H, q), 1.60 (3H, s), 1.28 (3H, d). MS: [M-H]+ 205.
PREPARATION B5 2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid
Figure imgf000123_0001
Acetic anhydride (3.06 g, 30.0 mmol) was added to methyl 2,4-dihydroxybenzoate (5.04 g, 30.0 mmol) in boron trifluoride diethyl etherate (7.6 ml) and the mixture was stirred and held at reflux for 3 hours and then allowed to cool to room temperature. Water (80 ml) was added and the mixture stirred at room temperature for 30 minutes. The resulting yellow solid was removed by filtration and sucked as dry as possible under vacuum. The solid was dissolved in dichloromethane and was washed with water, the organic layer was separated and the solvent removed in vacuo to afford methyl 5-acetyl-2,4-dihydroxybenzoate as a bright yellow solid (2.62 g, 42%) which was used without further purification. 1H NMR (DMSO-d6) 12.58 (1 H, s), 11.22 (1 H, s), 8.33 (1 H, s), 6.45 (1 H, s), 3.90 (3H, s), 2.62 (3H, s). MS: [M+H]+ 211. Methyl 5-acetyl-2,4-dihydroxybenzoate (2.62 g, 12.48 mmol) was dissolved in acetonitrile (40 ml), anhydrous potassium carbonate (4.93 g, 35.7 mmol) was added and the stirred mixture was treated with benzyl bromide (5.09 g, 29.75 mmol) and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between water and dichloromethane. The organic layer was separated and the solvent removed in vacuo to afford methyl 5-acetyl-2,4- bis-benzyloxybenzoate (3.48 g, 71%) as a colourless solid which was dried at 50 0C in a vacuum oven and used without further purification. 1H NMR (DMSO-d6) 8.21 (1 H, s), 7.55 (4H, m), 7.43 (4H, m), 7.37 (2H, m), 7.04 (1 H, s), 5.38 (4H, s), 3.79 (3H, s), 2.48 (3H, s). MS: [M+H]+ 391.
A stirred suspension of methyltriphenylphosphonium bromide (1.96 g, 5.5 mmol) in anhydrous tetrahydrofuran (20 ml) at 0 0C under a nitrogen atmosphere was treated dropwise with n-butyl lithium (1.6 M in hexanes, 3.5 ml, 5.5 mmol) and the resulting bright yellow solution was stirred at 0 0C for 30 minutes. A solution of methyl 5-acetyl-2,4-bis-benzyloxybenzoate (1.95 g, 5.00 mmol) in anhydrous tetrahydrofuran (20 ml) was added dropwise and the resulting mixture was allowed to warm to room temperature and was stirred for 16 hours. Methanol (10 ml) was added and the solvent was removed in vacuo. The residues were partitioned between dichloromethane and water, the organic layer was separated and the solvent removed in vacuo to afford a brown gum that was purified by column chromatography on silica. Elution with 7% ethyl acetate in petroleum ether afforded methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate as a colourless solid (700 mg, 36%). 1H NMR (DMSO-d6) 7.59 (1 H, s), 7.52 (2H, d), 7.64-7.32 (8H, m), 6.97 (1 H, s), 5.28 (2H, s), 5.22 (2H, s), 5.09 (1 H, s), 5.04 (1 H, s), 3.76 (3H, s), 2.02 (3H, s). MS: [M+H]+ 389.
Methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate (700 mg, 1.80 mmol) was dissolved in methanol (20 ml), a solution of potassium hydroxide (286 mg, 5.1 mmol) in water (4 ml) was added and the mixture was stirred and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with dichloromethane, the organic layer was separated and the solvent removed in vacuo to afford 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (600 mg, 89%) as a colourless solid. 1H NMR (DMSO-de) 7.52 (2H, d), 7.47-7.29 (9H, m), 6.82 (1 H, s), 5.20 (2H, S)1 5.17 (2H, s), 5.06 (1 H, s), 5.04 (1 H, s), 2.03 (3H1 s). MS: [M+H]+ 375. PREPARATION B6
2,4-β/s-benzvloxv-5-bromo-benzoic acid
Figure imgf000124_0001
2,4-dihydroxy-5-bromobenzoic acid (5.16 g, 22.15 mmol) was dissolved in DMF (40 ml) and potassium carbonate (12.2 g) and benzyl bromide (8 ml) were sequentially added. The mixture was stirred at room temperature for 18 hours under nitrogen. An aqueous solution of potassium hydroxide (2 g) in water (25 ml) was then added, followed by methanol (50 ml) and the mixture heated to reflux with vigorous stirring for 24 hours. The mixture was then allowed to cool, was poured into 1 N HCI (250ml) and was then extracted with ether and then DCM. The combined organic layers were dried over magnesium sulphate and the solvent evaporated in vacuo. The resulting solid material was washed with P. E. and then Et2O (3 x 50ml) to yield pure product (5.2g, 56%). 1H NMR (MeOH-d4) 8.06 (1 H, s), 7.51-7.30 (10H, m), 6.85 (1 H1 s), 5.22 (2H, s), 5.20 (2H, s). MS: [M+H]+ 413.
PREPARATION B7
Synthesis of (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoic acid
Figure imgf000125_0001
Methyl 3-bromo-4-hydroxybenzoate [prepared as per Tetrahedron, 2003, 59, 9173] (3.47 g, 15.0 mmol) was dissolved in acetonitrile (50 ml), anhydrous potassium carbonate (3.11 g, 22.5 mmol) was added and the stirred mixture was treated with benzyl bromide (3.08 g, 18.0 mmol) and held at reflux for 5 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between water and dichloromethane. The organic layer was separated, the solvent removed in vacuo and the residue subjected to column chromatography on silica. Elution with 10% ethyl acetate in petroleum ether afforded methyl 4-benzyloxy-3-bromobenzoate (3.6 g, 75%) as a colourless solid. 1H NMR (DMSOd6) 8.12 (1 H, d), 7.96 (1 H, dd), 7.51 (2H, m), 7.43 (2H, t), 7.35 (2H, m), 5.32 (2H, s), 3.84 (3H, s).
Methyl 4-benzyloxy-3-bromobenzoate (1.61 g, 5.0 mmol), caesium carbonate (4.89 g, 15.0 mmol), (£)-2-buten-2-yl boronic acid (600 mg, 6.0 mmol) and [1 ,1'-bis(diphenylphosphino)ferrocenyl] palladium (II) chloride (204 mg, 0.25 mmol) were dissolved in anhydrous tetrahydrofuran (100 ml), water (10 ml) was added and the mixture was stirred and held at reflux under an atmosphere of nitrogen for 16 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between dichloromethane and water. The organic layer was separated, the solvent removed in vacuo and the residue subjected to column chromatography on silica. Elution with 5% ethyl acetate in petroleum ether afforded methyl (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoate (600 mg, 41 %) as a colourless solid. 1H NMR (DMSO-d6) 7.88 (1H, dd), 7.59 (1 H, d), 7.40 (4H, m), 7.34 (1 H, m), 7.23 (1 H, d), 5.57 (1 H, q), 5.21 (2H, s), 3.82 (3H, s), 1.94 (3H, s), 1.38 (3H, d).
Methyl (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoate (592 mg, 2.0 mmol) was dissolved in methanol (20 ml), a solution of potassium hydroxide (336 mg, 6.0 mmol) in water (7 ml) was added and the mixture was stirred and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with dichloromethane, the organic layer was separated and the solvent removed in vacuo to afford (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoic acid (460 mg, 82%) as a colourless solid. 1H NMR (DMSOd6) 7.85 (1 H, dd), 7.57 (1 H, d), 7.40 (4H, m), 7.34 (1 H, m), 7.18 (1 H, d), 5.57 (1 H, q), 5.21 (2H, s), 1.96 (3H1 S)1 1.40 (3H, d). MS: [M+H]+ 283.
PREPARATION B8 Synthesis of 2,4-bis-benzyloxy-5-tert-butyl-benzoic acid
Figure imgf000126_0001
1-(2,4-Bis-benzyloxy-5-tert-butyl-phenyl)-ethanone [prepared as per WO 2004/072051] (2.02 g, 5.2 mmol) was dissolved in 1 ,4-dioxane (30 ml), a solution of sodium hydroxide (2.08 g, 52.0 mmol) in water (30 ml) was added and the mixture was stirred and treated dropwise with bromine (0.8 ml, 15.6 mmol). The resulting mixture was stirred at room temperature for 16 hours. The 1 ,4-dioxane was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with ethyl acetate, the organic layer was separated, the solvent removed in vacuo and the residue subjected to column chromatography on silica. Elution with 30% ethyl acetate in petroleum ether afforded 2,4-bis-benzyloxy-5-tert-butyl-benzoic acid (1.6 g, 79%) as a pale yellow oil. 1H NMR (DMSOd6) 12.18 (1 H, br s), 7.69 (1 H, s), 7.52 (4H, t), 7.45-7.33 (6H, m), 6.93 (1 H, s), 5.24 (2H, s), 5.23 (2H, s), 1.32 (9H, s). MS: [M+H]+ 391.
PREPARATION B9
Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (Alternative synthesis)
Figure imgf000126_0002
Stepi : Synthesis of 2.4-bis-benzyloxy-5-bromo-benzoic acid benzyl ester
Figure imgf000126_0003
To a 1OL jacketed vessel, fitted with a flange lid containing stirrer, thermometer and dropping funnel, was charged acetone (2.5 L) followed by 5-bromo-2,4-dihydroxybenzoic acid (100 g, 0.43 mol) and potassium carbonate (356 g, 2.58 mol). To the stirring mixture at ambient was added benzyl bromide (185 mL, 1.55 mol) at a rate of ~20 ml/min. The mixture was heated at 60 0C for 18 h and then taken to 45 0C. Water (1.5 L) was added and the mixture stirred for 30 min. The mixture was extracted with EtOAc (2 x 1 L) and the combined organic portions reduced in vacuo. To the residue was added Et2O (200 mL) and petroleum ether (1 L), the mixture stirred for 30 min and the solid formed collected by filtration and dried in vacuo to give the title compound (197.2 g) as a white solid.
Step 2: Synthesis of potassium isopropenyl trifluoroborate
Figure imgf000127_0001
To a solution of 2-bromopropene (20 mL, 225 mmol) in anhydrous THF (250 mL) stirring under a N2 atmosphere at -78 0C was added over 30 mins n-BuLi (2.5M in hexanes) (100 mL, 250 mmol) and the mixture stirred for 30 mins. To the mixture at -78 °C was slowly added triethyl borate (58 mL, 340 mmol) at a rate to ensure that the temperature of the reaction mixture did not exceed -65 0C. The resulting solution was then stirred at -78 0C for 30 mins, allowed to slowly warm to ambient and stirred for a further 90 mins. Potassium hydrogen fluoride (105 g, 1.35 mol) was added to the mixture followed by water (250 mL). The mixture was stirred at ambient for 14 h and then reduced to dryness. The procedure was repeated as above and following reduction to dryness the two residues were combined for further work-up.
To the combined residues was added acetone (800 mL), the mixture stirred for 1 h and then filtered. The solid collected was washed with acetone (200 mL) and the combined filtrates reduced in vacuo to give a solid. This solid was triturated with Et2O (250 mL) and then dried in vacuo to give the title compound (28.2 g) as a white solid.
Step 3: Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid benzyl ester
Figure imgf000127_0002
To a mixture of 2,4-bis-benzyloxy-5-bromo-benzoic acid benzyl ester (42.9 g, 85.7 mmol), potassium isopropenyl trifluoroborate (14.0 g, 95.2 mmol) and caesium carbonate (83.8 g, 257.1 mmol) in THF (800 mL) was added Pd(PPh3)4 (2.0 g) followed by water (150 mL). The mixture was heated at reflux for 72 h then allowed to cool to ambient. The mixture was reduced in vacuo to remove THF and then partitioned between water (500 mL) and EtOAc (300 mL). The organic portion was washed with brine, dried (MgSO4), filtered and reduced in vacuo to give the title compound (40.9 g) as a brown oil. Step 3A
Alternative Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid benzyl ester
Figure imgf000127_0003
A mixture of 2,4-bis-benzyloxy-5-bromo-benzoic acid benzyl ester (10.0 g, 20 mmol), potassium isopropenyl trifluoroborate (4.0 g, 27.2 mmol) and n-butylamine (6.0 ml_, 60 mmol) in 2- propanol/water (2:1 , 200 m L) was purged with N2 for 5 minutes. To this mixture was added [1 ,1'- bis(diphenyl-phosphino)ferrocene]dichloropalladium(ll) (816 mg, 1.09 mmol) and the mixture was heated at reflux for 20 h. The mixture was allowed to cool to ambient then diluted with water (400 ml_) and extracted with EtOAc (2 x 300 ml_). The combined organic extracts were washed with 1 M aqueous HCI, brine, dried (MgSO4), filtered through a plug of Celite and the filtrate reduced in vacuo to give the title compound (11.1 g) as a brown gum.
Step 4: Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid
Figure imgf000128_0001
To a solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid benzyl ester (40.8 g, 87.9 mmol) in THF-MeOH-water (3:1 :1 , 300 mL total) was added lithium hydroxide (8.42 g, 352 mmol). The mixture was heated at 50 0C for 16 h, allowed to cool to ambient and then diluted with water (300 mL). The mixture was taken to pH~1 using cone. HCI (~30 mL) and then extracted with EtOAc (2 x 200 mL). The combined organic extracts were washed with brine, dried (MgSO4), filtered and reduced in vacuo. The solid residue was taken up in P. E-MeOH (9:1 , 300 mL total), the slurry stirred for 1 h at ambient and the solid collected by filtration. The solid was dried in vacuo to give the title compound (26.8 g) as an off-white solid. 1H NMR (400 MHz, DMSO-c/6) 12.30 (s, 1 H), 7.61 (s, 1 H), 7.53 (d, J = 7.0 Hz, 2H), 7.47 - 7.31 (m, 8H), 6.94 (s, 1 H), 5.23 (d, J = 14.0 Hz, 4H), 5.08 (d, J = 9.0 Hz, 2H), 2.04 (s, 3H).
PREPARATION B10
2,4-Bis-benzyloxy-5-isopropyl-benzoic acid
Step i
Preparation of 1-(2-4-Bis-benzyloxy-phenyl)-ethanone
Figure imgf000128_0002
Material inputs:
S.No. Item Quantity Eq
1. 1 ,3 Dihydroxy acetophenone 5O g 1
2. Benzyl bromide 97 ml 3 3. Acetonitrile 750 ml 15 times
4. Potassium carbonate 115 g 3
1 ,3 Dihydroxy acetophenone (50 g) was placed in a 2 L single neck RB flask equipped with a reflux condenser and a guard tube. Acetonitrile (750 ml), potassium carbonate (115 g) and benzyl bromide (97 ml) were added and the mixture was heated at reflux (90° C) for 16 hours. On completion, the acetonitrile was removed under reduced pressure. Water (200 ml) was added to the reaction mixture which was then extracted with ethyl acetate (500 ml). The organic layer was separated and dried over sodium sulphate. The solvent was removed under reduced pressure to give a residue which was washed n-hexane (600 ml) to give the product.
Quantity of the product obtained : 105.1 g
Yield : 96.24%
Nature : Solid
Colour : Brown
Step 2 Preparation of 2-4-Bis-benzyloxy-1-isopropenylbenzene
Figure imgf000129_0001
Material inputs:
S. No. Item Quantity Eq.
1. Compound of Step 1 2O g 1
2. n-BuLi (1.6 M) 92.6 ml 2.3
3. Methyl-triphenylphosphonium iodide 53.4 g 2.2
4. THF 200 ml 10 times
Methyl-triphenylphosphonium iodide (53.4 g) and THF (100 ml) were introduced into a 1 L 3-neck RB flask equipped with an addition funnel and an inlet for nitrogen atmosphere and the mixture was cooled to 0° C. n-BuLi (92.6 ml) was added dropwise to the reaction mixture over a period of 15 min at 0° C. The reaction mixture was stirred for 10 min at 0° C and further stirred at RT for 30 min. 1-(2-4-Bis-benzyloxy-phenyl)-ethanone (20 g) in THF (100 ml) was added dropwise to the reaction mixture over a period of 10 min at 0° C and the reaction mixture was stirred overnight at RT. The progress of the reaction was monitored by TLC (10% EtOAc/n-hexane, product Rf~0.9). On completion, methanol (-100 ml) was added to the reaction mixture and the solvent was removed under reduced pressure to give a residue. n-Hexane (1 L) was added to the residue which was refluxed (75° C) for 30 min. before filtering the mixture was through a Celite bed and washing the bed with n-hexane (500 ml). The solvent was removed under reduced pressure to give a residue, which was further purified by column chromatography (SiO2 2% EtOAc/n-hexane).
Quantity of the product obtained : 12.5 g
Yield : 63.13 %
Nature : Liquid.
Colour : Colorless
Step 3 4-lsopropyl-benzene-1 ,3-diol
Figure imgf000130_0001
Material inputs: S.No. Item Quantity Eq.
1. 2-4-Bis-benzyloxy-1 -isopropenylbenzene 12.5 g 1
2. Ethanol 125 ml 10 times
3. 20% Palladium hydroxide 2 g
To a mixture of 2-4-bis-benzyloxy-1 -isopropenylbenzene (12.5 g) in ethanol (125 ml) in a 500 ml hydrogenation flask was added 20% palladium hydroxide (2 g). The reaction mixture was hydrogenated at 80 psi for 36 h. The progress of the reaction was monitored by TLC (10% EtOAc/n-hexane, product Rf-0.1). On completion, the reaction mixture was filtered through a bed of Celite and the bed was washed with ethanol (300 ml). The solvent was removed under reduced pressure to give a crude product, which was used as such for the next step.
Quantity of the product obtained : 5.8 g (crude)
Nature : Solid.
Colour : Colourless.
Step 4 1-(2,4-Dihydroxy-5-isopropyl-phenyl)-ethanone
Figure imgf000130_0002
Material inputs: S.No. Item Quantity Eq.
1. 4-lsopropyl-benzene-1 ,3-diol 5.8 g 1
2. Boron trifluoride etherate 28.7 ml 6
3. Acetic acid 4.55 ml 2
4-lsopropyl-benzene-1 ,3-diol (5.8 g) and boron trifluoride etherate (28.7 ml) were introduced into a 250 ml single neck RB flask equipped with a reflux condenser and an inlet for nitrogen atmosphere stirred at room temperature for 10 minutes. Acetic acid (4.55 ml) was added to the reaction mixture and stirred at 90 0C for 16 hours. On completion, 10% sodium acetate (300 ml) was added to the reaction mixture which was stirred at room temperature for 4 hours. The reaction mixture was then extracted with ethyl acetate (300 ml) and washed with saturated sodium bicarbonate (100 ml) and the organic layer was dried over sodium sulphate. The reaction was monitored by TLC (10% EtOAc/n-hexane, product Rf~0.5). The solvent was removed under reduced pressure to give a residue, which was further purified by column chromatography (SiO2, 10% EtOAc/n-hexane).
Quantity of the product obtained : 3.2 g
Yield : 43.24 %
Nature : Solid.
Colour : Colourless
Step 5 1-(2,4-Bis-benzyloxy-5-isopropyl-phenyl)-ethanone
Figure imgf000131_0001
Material inputs:
S.No. Item Quantity Eq.
1. 1-(2,4-Dihydroxy-5-isopropyl-phenyl)-ethanone 3.2 g 1
2. Benzyl bromide 5.89 ml 3
3. Potassium carbonate 6.82 g 3
4. Acetonitrile 60 ml 20 times
To a mixture of 1-(2,4-dihydroxy-5-isopropyl-phenyl)-ethanone (3.2 g), acetonitrile (60 ml) and potassium carbonate (10.6 g) in a a 250 ml single neck RB flask equipped with a reflux condenser and a guard tube was added benzyl bromide (9.1 ml). The reaction mixture was refluxed (900C) for 16 hours. The progress of the reaction was monitored by TLC (10% EtOAc/n-hexane, product Rf~0.5). On completion, acetonitrile was removed under reduced pressure. Water (100 ml) was added to the residue obtained and the resulting mixture was extracted with ethyl acetate (200 ml). The organic layer was dried over sodium sulphate. The solvent was removed under reduced pressure to give a residue to which n-hexane (150 ml) was added to give the product.
Quantity of the product obtained : 5.1 g
Yield : 83.6 %
Nature : Solid.
Color : Colorless
2,4-Bis-benzyloxy-5-isopropyl-benzoic acid
Figure imgf000132_0001
Material inputs: S.No. Item Quantity
1. 1-(2,4-Bis-benzyloxy-5-isopropyl-phenyl)-ethanone 7 g
2. Sodium hypobromide 13 g in water 100 ml
3. Dioxane 100 ml Procedure:
A mixture of a mixture of 1-(2,4-Bis-benzyloxy-5-isopropyl-phenyl)-ethanone (7 g) in dioxane (100 ml) in a 500 ml single neck round bottomed flask equipped with a guard tube was cooled to 10° C and sodium hypobromide [13 g in water (100 ml)] was added. The reaction mixture was stirred overnight at room temperature. The progress of the reaction was monitored by TLC (30% EtOAc/n-hexane, product Rf-0.5). On completion, sodium bisulphite (7 g) was added to the reaction mixture which was cooled to 0°C. The reaction mixture was then acidified with HCI (-10 ml) to pH~2, extracted with ethyl acetate (100 ml) and washed with water (25 ml). The organic layer was dried over sodium sulphate, and the solvent was removed under reduced pressure to give a residue, which was further purified by column chromatography (SiO2, 10% EtOAc/n- hexane).
Quantity of the product obtained : 3.4 g
Yield : 48.3 %
Nature : Solid. Colour : Colourless.
C. SYNTHESIS OF ISOINDOLINE INTERMEDIATES
PREPARATION C1
Synthesis of 4,7-difluoroisoindoline
Figure imgf000133_0001
A mixture of 1 ,4-difluoro-2,3-dimethylbenzene (4.26 g, 30.0 mmol), N-bromosuccinimide (10.68 g, 60.0 mmol) and dibenzoyl peroxide (75 wt % in water, 120 mg) in carbon tetrachloride (50 ml) was stirred and held at reflux for 16 hours. Upon cooling to room temperature the mixture was filtered, the solids washed with carbon tetrachloride (10 ml), the organic extracts combined and the solvent removed in vacuo to afford 2,3-bis-bromomethyl-i ,4-difluorobenzene (9.0 g, 100%) as a pale yellow liquid that solidified upon standing. 1H NMR (DMSO-ds) 7.36 (2H, dd), 4.78 (4H, s).
A solution of 4-toluenesulphonamide (2.44 g, 14.28 mmol) in N,N-dimethylformamide (10 ml) was added dropwise to a vigourously stirred suspension of sodium hydride (1.2 g, 60 wt % in mineral oil, 30.0 mmol) in anhydrous N,N-dimethylformamide (60 ml). The mixture was strirred at room temperature for 1 hour, at 1100C for 1 hour and was then cooled to 6O0C and a solution of 2, 3-bis- bromomethyl-1 ,4-difluorobenzene (4.28 g, 14.28 mmol) in N,N-dimethylformamide (30 ml) was added dropwise. The mixture was stirred at 600C for 1 hour and then at room temperature for 16 hours. The solvent was removed in vacuo and the residue partitioned between dichloromethane and 1 M hydrochloric acid. The organic layer was separated, washed with 5% aqueous potassium carbonate solution, the organics were separated and the solvent removed in vacuo. The residue was rinsed with diethyl ether, filtered and the solids sucked dry under reduced pressure to afford 4,7-difluoro-2-(toluene-4-sulphonyl)isoindoline (2.46 g, 56%) as a pale tan solid. 1H NMR (DMSO- dβ) 7.82 (2H, d), 7.43 (2H, d), 7.15 (2H, dd), 4.66 (4H, s), 2.36 (3H, s). MS: [M+H]+ 310.
A mixture of 4,7-difluoro-2-(toluene-4-sulphonyl)isoindoline (2.36 g, 7.64 mmol), phenol (2.36 g, 25.11 mmol), 48% hydrogen bromide in water (20 ml) and propionic acid (4 ml) was stirred and held at reflux for 6 hours. Upon cooling to room temperature water (50 ml) was added and the mixture extracted with diethyl ether (2 x 100 ml). The aqueous layer was basified by the addition of 2M sodium hydroxide and was extracted with diethyl ether (3 x 100 ml). The combined extracts were evaporated to dryness in vacuo to afford 4,7-difluoroisoindoline (586 mg, 50%) as a brown oil that solidified upon standing. 1H NMR (DMSO-d6) 7.06 (2H, dd), 4.12 (4H, s). MS: [M+H]+ 156. PREPARATION C2
Synthesis of 5-hydroxyisoindoline hvdrobromide
Figure imgf000134_0001
A solution of dimethyl 4-methoxyphthalate (36.75 g, 0.16 mol) in methanol (100 ml) was treated with a solution of potassium hydroxide (28.0 g, 0.5 mol) in water (50 ml) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the methanol was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 5M hydrochloric acid. The solid material was filtered off, washed with water and sucked dry under reduced pressure overnight to afford 4-methoxyphthalic acid (31.8 g, 99%) as an off white solid. 1H NMR (DMSOd6) 12.90 (2H, br s), 7.74 (1 H, d), 7.12-7.05 (2H, m), 3.84 (3H, s). MS: [M+H]+ 197.
Acetic anhydride (40 ml) was added to a mixture of 4-methoxyphthalic acid (30.8 g, 0.16 mol) in anhydrous tetrahydrofuran (150 ml) and the mixture was strirred and held at reflux for 4 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford 4-methoxyphthalic anhydride (27.8 g, 99%) as an off white solid. 1H NMR (DMSO-d6) 8.02 (1 H, d), 7.59 (1 H, d), 7.49 (1 H1 dd), 3.97 (3H1 s). MS: [M+H]+ 179.
A mixture of 4-methoxyphthalic anhydride (27.8 g, 0.16 mol) and formamide (175 ml) was stirred and held at 21 O0C for 5 hours and was then allowed to cool to room temperature overnight. The solid material was filtered off, washed sequentially with water (100 ml), 50% aqueous acetone (50 ml) and diethyl ether (200 ml) and sucked dry under reduced pressure to afford A- methoxyphthalimide (21.3 g, 77%) as a pale yellow solid. 1H NMR (DMSO-d6) 11.15 (1 H, br s), 7.74 (1 H, d), 7.33-7.28 (2H, m), 3.92 (3H, s).
A stirred solution of 4-methoxyphthalimide (21.3 g, 0.12 mol) in anhydrous tetrahydrofuran (425 ml) at O0C was treated dropwise with a solution of borane in tetrahydrofuran (1 M, 340 ml, 0.34 mol) and the resulting mixture was stirred and held at reflux for 16 hours. The mixture was cooled to 00C, methanol (150 ml) was added dropwise followed by 5M hydrochloric acid (150 ml) and the mixture was stirred and held at reflux for 3 hours. Upon cooling to room temperature the organic solvent was removed in vacuo, the mixture was diluted with water (750 ml) and was extracted with dichloromethane (3 x 750 ml). The aqueous layer was basified to pH 12 or above by the addition of 5M sodium hydroxide, extracted with dichloromethane (3 x 750 ml) and the combined extracts were evaporated to dryness in vacuo to afford 5-methoxyisoindoline (8.34 g, 47%) as a brown oil. 1H NMR (DMSOd6) 7.13 (1 H, d), 6.84 (1 H, d), 6.74 (1 H, dd), 4.05 (2H, s), 4.01 (2H, s), 3.73 (3H, s). MS: [M+H]+ 150.
5-Methoxyisoindoline (8.34 g, 55.97 mmol) in 48% aqueous hydrobromic acid (100 ml) was stirred and held at reflux for 16 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford 5-hydroxyisoindoline hydrobromide (11.32 g, 93%) as a tan solid. 1H NMR (DMSOd6) 9.63 (1 H, br s), 9.32 (2H, br s), 7.18 (1 H, d), 6.79 (1 H, d), 6.76 (1 H, dd), 4.42 (2H, t), 4.38 (2H, t). MS: [M+H]+ 136.
PREPARATION C3
Synthesis of 5-chloro-2,3-dihvdro-1 H-isoindole
Figure imgf000135_0001
A mixture of 3,4-dimethylchlorobenzene (1Og, 71.1 mmol), N-bromosuccinimide (25g,142.2mmol), and benzoyl peroxide (0.147g, 0.6mmol), was refluxed in 80 ml of carbon tetrachloride for 18 hours. After cooling, the insoluble material was filtered off and washed with a small amount of carbon tetrachloride. The filtrate and the washings were combined and concentrated under reduced pressure to obtain 2Og of 1 ,2-bis-bromomethyl-4-chloro-benzene as a pale yellow oil product as a major component. To a suspension of 60% sodium hydride (3.0 g, 0.125 mmol) in mineral oil in 80 ml of anhydrous DMF (100 ml) was dropwise added a solution of para-toluene sulphonamide (5.6 g, 32.60 mmol) in 30 ml of DMF over 1 hour with vigorous stirring at room temperature. After the addition, the mixture was stirred for 1 hour at room temperature and another 1 hour heating at 9O0C. To this mixture was added dropwise a solution of 1 ,2-bis-bromomethyl-4-chloro-benzene (4g, 14.18 mmol) in 20 ml of anhydrous DMF at 60 0C and then stirred overnight at room temperature. The resultant mixture was poured onto ice and the resulting precipitate was collected by filtration. The precipitate was washed with 1 N hydrochloric acid, 5% sodium carbonate and brine then dried (MgSO4), filtered and evaporated to give 2.8 g of 5-Chloro-2-(toluene-4-sulphonyl)-2,3-dihydro-1 H-isoindole as a pale yellow solid. MS: [M+ H]+ 308 1.Og of 2-(p-toluensulphonyl)-5-chloroisoindoline and 1.Og of phenol were added to a mixture of 8 ml of 48% hydrobromic acid and 1.4 ml of propionic acid, and then mixture was heated at reflux for 6 hours. The resultant reaction mixture was diluted with 10ml of water and extracted twice with 50ml of ethyl acetate. The water layer was basified with aqueous sodium hydroxide solution and extracted with ethyl acetate three times. The extract was concentrated and the crude product was diluted with 4N HCI/dioxane and stirred for 15minutes before evaporating the HCI and then re- evaporating with toluene three times to give 0.3g of 5-chloro-2,3-dihydro~1H-isoindole hydrochloride as a black solid. MS: [M+H]+ 153-15
PREPARATION C4
Synthesis of 5-chloro-6-methoxy-2,3-dihydro-1 H-isoindole
Figure imgf000136_0001
A mixture of 1-chloro-2-methoxy-4,5-dimethyl-benzene (3 g, 17.6 mmol), N-bromosuccinimide (6.3 g, 35.3 mmol), and benzoyl peroxide (0.100 g, 0.41 mmol) in carbon tetrachloride (40 ml) was heated at reflux for 18 hours. After cooling, the insoluble material was removed by filtration, washed with a small amount of carbon tetrachloride and the filtrate evaporated to give 1 ,2-bis- bromomethyl-4-chloro-5-methoxy-benzene as an oil product as a major component. MS: [M+H]+ 329 A solution of 4-methoxybenzyIamine (2.4 g, 17.6 mmol) in acetone (110 ml) was added dropwise to a mixture of 1 ,2-bis-bromomethyl-4-chloro-5-methoxy-benzene (assumed theoretical, 17.6 mmol) and Na2CO3 (12 g, 114 mmol) in acetone/water (10 ml:12.5 ml) then stirred at room temperature for 2 hours and concentrated in vacuo. The crude material was dissolved in ethyl acetate and extracted with 2N HCI. The aqueous layerwas neutralized with sodium carbonate, extracted with ethyl acetate (x 2), dried (MgSO4) and evaporated under vacuum to give 5-chloro-6- methoxy-2-(4-methoxy-benzyl)-2,3-dihydro-1 H-isoindole (0.8 g, 2.6 mmol) as a brown gum. MS: [M+H]+ 304
A solution of 5-chloro-6-methoxy-2-(4-methoxy-benzyl)-2,3-dihydro-1 H-isoindole (600 mg) and anisole (0.3 ml) in trifluoroacetic acid (6 ml) was heated at 180°C (50W) for 40 minutes in a CEM discover microwave synthesiser. The reaction mixture was evaporated and re-evaporated with toluene. The crude material was partitioned between DCM and water, the aqueous layer washed with DCM (x 3) then evaporated and re-evaporated with toluene to give 5-chloro-6-methoxy-2,3- dihydro-1 H-isoindole (256 mg) as green crystals. MS: [M+H]+ 184
PREPARATION C5 Synthesis of 2,3-dihvdro-1 H-isoindol-5-ylamine trifluoroacetate
Figure imgf000137_0001
A solution of 4-nitro-o-xyIene (15.1 g; 0.1 mol) in carbon tetrachloride (150 ml) was treated with N- bromosuccinimide (36 g; 0.2 mol) followed by benzoyl peroxide (1 g) then heated at reflux overnight. The reaction was allowed to cool to ambient temperature, filtered and the filtrate evaporated to give 32 g of crude 1 ,2-bis-bromomethyl-4-nitro-benzene as a mobile oil. The crude product was dissolved in benzene (200 ml) then treated dropwise over 30 minutes with a solution of 2,4-dimethoxybenzylamine (15 ml) and triethylamine (27.85 ml) in benzene (100 ml) then heated at 800C for 3 hours. The reaction was cooled, washed with water followed by saturated sodium bicarbonate. The organics were extracted with 2M HCI (2 x 150 ml) then combined aquesous basified with 2M NaOH and extracted with EtOAc (x 2). The combined EtOAc layer was dried (MgSO4), evaporated then purified by flash column chromatography eluting with EtOAc / P. E. (1 :3 - 1.2 - 1 : 1 ). Product containing fraction were combined and evaporated to give 10.15 g of 2- (2,4-dimethoxy-benzyl)-5-nitro-2,3-dihydro-1 H-isoindole as a brown solid. 1H NMR (DMSO-d6) 8.12 (2H, m), 7.50 (1 H, d), 7.25 (1 H, d), 6.55 (1 H, d), 6.52 (1 H, dd), 3.93 (4H, s), 3.80 (3H, s), 3.78 (2H, s), 3.75 (3H, s).
2-(2,4-dimethoxy-benzyl)-5-nitro-2,3-dihydro-1 H-isoindole (13 g) in TFA (18 ml) was treated with anisole (6 ml) then heated in a CEM microwave synthesiser at 12O0C (30 Watts) for 20 minutes (carried out batch wise, 6 times). The reaction mixture was evaporated in vacuo and the residue partitioned between DCM and water. The water layer was separated, washed with DCM (x 3) then evaporated and re-evaporated with toluene / MeOH (x 3) to give 9.8 g of 5-nitro-2,3-dihydro-1 H- isoindole trifluoroacetic acid salt as a beige solid. 1H NMR (DMSO-d6) 9.85 (2H, br s), 8.32 (1 H, d), 8.25 (1 H, dd), 7.70 (1 H, d), 4.68 (2H, s), 4.65 (2H, s).
A mixture of 5-nitro-2,3-dihydro-1 H-isoindole trifluoroacetic acid salt (9.8g) and 10 % palladium on carbon (1 g) in methanol (75 ml) was hydrogenated at room temperature and pressure for 16 hours. The reaction was filtered through Celite™, the filtrate evaporated and re-evaporated with toluene to give 8.76 g of 2,3-dihydro-1 H-isoindol-5-ylamine mono trifluoroacetic acid salt as a dark brown solid. 1 H NMR (DMSO-d6) 9.45 (2H, br s), 7.05 (1 H, d), 6.60 (2H, m), 5.35 (2H, br s), 4.40 (2H, s), 4.30 (2H, s). PREPARATION C6
Synthesis of 5-morpholin-4-ylmethyl-2,3-dihvdro-1 H-isoindole ditrifluoroacetate
Figure imgf000138_0001
Steps 1 and 2 were carried out in a manner analogous to that described in Preparation C5 using methyl 3,4-dimethylbenzoate as the starting material.
A mixture of 2-(2,4-dimethoxybenzyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methyl ester (4.65 g; 14.2 mmol) and lithium hydroxide monohydrate (660 mg; 1.1 equiv.) in 4:1 :1 THF-MeOH-H2O (60 ml) was stirred at room temperature overnight. A further 170 mg of base were added and stirring continued for 7 hours. The reaction was evaporated then re-evaporated with MeOH / toluene (x 2). A mixture of the crude 2-(2,4-dimethoxy-benzyl)-2,3-dihydro-1 H-isoindole-5- carboxylic acid lithium salt (1.5 g; 4.7 mmol), morpholine (820 μl; 2 equiv.), EDAC (1.1 g; 1.2 equiv.) and HOBt (760 mg; 1.2 equiv.) in DMF (25 ml) was stirred at room temperature overnight then evaporated in vacuo. The residue was partitioned between EtOAc and saturated NaHCO3, the EtOAc layer was separated, washed with brine, dried (MgSO4) and evaporated. Purification by flash column chromatography (2% then 5% MeOH / DCM as eluant) gave 1.1g of [2-(2,4- dimethoxybenzyl)-2,3-dihydro-1 H-isoindol-5-yl]-morpholin-4-yl-methanone as a red / brown gum. 1H NMR (DMSO-d6) 7.30-7.18 (4H, m), 6.56 (1 H, d), 6.52 (1 H, dd), 3.85 (4H, s), 3.78 (5H, m), 3.73 (3H, s).
A solution of [2-(2,4-dimethoxybenzyl)-2,3-dihydro-1 H-isoindol-5-yl]-morpholin-4-yl-methanone (1.05 g; 2.75 mmol) in dry THF (20 ml) under a nitrogen atmosphere was treated with 1M lithium aluminium hydride solution then stirred at room temperature overnight. The reaction was quenched by the cautious addition of saturated sodium sulphate solution, then diluted with EtOAc (40 ml), filtered through Celite™ and evaporated. Purification by flash column chromatography (2% then 5% MeOH / DCM as eluant) gave 340 mg of 2-(2,4-dimethoxybenzyl)-5-morpholin-4-ylmethyl-2,3- dihydro-1 H-isoindole as a pale brown gum.
A mixture of 2-(2,4-dimethoxybenzyl)-5-morpholin-4-ylmethyl-2,3-dihydro-1 H-isoindole (340 mg) and anisole (350 μl) in trifluoroacetic acid (1.5 ml) was heated at 130C in a CEM microwave synthesiser for 1 hour then evaporated and re-evaporated with toluene. The residue was partitioned between DCM and water. The water layer was separated, washed with DCM (x 3) then evaporated and re-evaporated with toluene / MeOH (x 3) to give 422 mg of 5-morpholin-4- ylmethyl-2,3-dihydro-1 H-isoindole ditrifluoroacetate as a brown gum. 1H NMR (DMSO-d6) 10.30 (1 H, br s), 9.60 (2H, br s), 7.55-7.45 (3H, m), 4.45 (4H, s), 4.45-4.30 (2H, m), 4.20-3.88 (2H, m), 3.70-3.55 (2H, m), 3.30-3.00 (4H, m).
PREPARATION C7
Synthesis of ethyl-2,3-dihydro-1 H-isoindole-5-carboxylate trifluoroacetate
Figure imgf000139_0001
A solution of 2-(2,4-dimethoxy-benzyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methyl ester (215 mg) and anisole (200 μl) in 1 ml of TFA was heated at 1400C for 30 minutes in a CEM discover microwave synthesiser. The reaction was partititioned between water and DCM, the water layer was separated, washed with DCM then evaporated and re-evaporated with toluene / MeOH (x 2) to give 105 mg of the title compound. 1H NMR (DMSO-d6) 9.70 (2H, br s), 8.02 (1H, s), 8.98 (1 H, d), 7.57 (1 H, d), 4.60 (2H, s), 4.56 (2H, s), 3.89 (3H, s).
PREPARATION C8
4-Hvdroxy-2-(4-methoxy-benzvD-isoindole-1 ,3-dione
Figure imgf000139_0002
A mixture of 3-hydroxyphthalic anhydride (543 mg, 3.31 mmol), 4-methoxybenzylamine (0.43 ml_, 3.31 mmol) and acetic acid (3 mL) was heated at 10O0C for 4 hours. The mixture was allowed to cool and diluted with water (20 mL). The white solid was collected by filtration, washed well with water and dried to give the title compound (760 mg, 81 %). 1H NMR (DMSO-d6) 11.03 (1 H, s), 7.61 (1 H, dd), 7.28 (1 H, d), 7.23-7.19 (3H, m), 6.89-6.86 (2H, m), 4.63 (2H, s), 3.71 (3H, s). MS: [M-H+] 282.
PREPARATION C9
4-Hvdroxy-2-(2,4-dimethoxy-benzyl)-isoindole-1 ,3-dione
Figure imgf000139_0003
A mixture of 3-hydroxyphthalic anhydride (1.24 g, 7.6 mmol), 2,4-dimethoxybenzylamine (1.14 mL, 7.6 mmol) and acetic acid (5 mL) was heated at 800C for 24 hours. The mixture was allowed to cool and diluted with water (20 mL). The white solid was collected by filtration, washed well with water and dried to give the title compound (1.73 g, 73%). 1H NMR (DMSO-d6) 11.00 (1 H, s), 7.62 (1 H, dd), 7.29 (1 H, d), 7.21 (1 H, d), 6.90 (1 H, d), 6.56 (1 H, d), 6.43 (1 H, dd), 4.59 (2H, s), 3.79 (3H, s), 3.72 (3H, s). MS: [M-H+] 314.
PREPARATION C10
2-(4-Methoxy-ben2yl)-4-r2-(2-methoxy-ethoxy)-ethoxy1-isoindole-1 ,3-dione
Figure imgf000140_0001
1-(2-Bromo-ethoxy)-2-methoxy-ethane (107 mg, 0.58 mmol) was added to a suspension of 4- hydroxy-2-(4-methoxy-benzyl)-isoindole-1 ,3-dione (150 mg, 0.53 mmol) and potassium carbonate (200 mg, 1.4 mmol) in DMF (2 ml_). After 3.5 hours, a catalytic amount of potassium iodide was added. After a further 17 hours, the mixture was warmed to 60°C. After 3 hours, an additional amount of 1-(2-bromo-ethoxy)-2-methoxy-ethane (20 mg, 0.11 mmol) was added and the mixture maintained at 6O0C for a further 20 hours. The mixture was concentrated in vacuo then the residue was taken up in ethyl acetate and washed with potassium carbonate solution and brine. The organic phase was dried (MgSO4) and concentrated to give the title compound as a yellow oil (149 mg, 73%). 1H NMR (methanol-d4) 7.71 (1 H, t), 7.43-7.40 (2H, m), 7.31-7.27 (2H, m), 6.87-6.83 (2H, m), 4.71 (2H, s), 4.37-4.34 (2H, m), 3.92-3.89 (2H, m), 3.77-3.74 (5H, m), 3.55-3.53 (2H, m), 3.33 (3H, s). MS: [M+H]+ 386.
PREPARATION C11 2-(2,4-Dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-isoindole-1 ,3-dione
Figure imgf000140_0002
A mixture of 2-(2,4-dimethoxy-benzyl)-4-hydroxy-isoindole-1 ,3-dione (317 mg, 1.01 mmol), 2- dimethylaminoethyl chloride hydrochloride (160 mg, 1.11 mmol) and potassium carbonate (350 mg, 2.5 mmol) in DMF (4 ml_) was heated at 6O0C for 18 hours. The mixture was concentrated in vacuo, taken up in ethyl acetate and extracted twice with 1 N hydrochloric acid. The aqueous extracts were made basic with solid potassium carbonate and extracted with ethyl acetate (*2). The combined organic extracts were washed with brine, dried (MgSO4) and concentrated to give the title compound (236 mg, 61%) as an off-white solid. 1H NMR (methanol-d4) 7.73 (1 H, t), 7.44- 7.40 (2H, m), 7.02 (1 H, d), 6.51 (1 H, d), 6.42 (1 H, dd), 4.72 (2H, s), 4.33 (2H, t), 3.80 (3H, S)1 3.76 (3H, s), 2.87 (2H, t), 2.40 (6H, s). MS: [M+H]+ 385.
PREPARATION C12 2-(2,4-Dimethoxy-benzyl)-4-(3-morpholin-4-yl-propoxy)-isoindole-1 ,3-dione
Figure imgf000141_0001
A mixture of 2-(2,4-dimethoxy-benzyl)-4-hydroxy-isoindole-1 ,3-dione (313 mg, 1.00 mmol), 4-(3- chloropropyl)morpholine (160 mg, 1.11 mmol) and potassium carbonate (350 mg, 2.5 mmol) in DMF (5 mL) was heated at 6O0C for 18 hours. The mixture was diluted with ethyl acetate and extracted twice with 1 N hydrochloric acid. The aqueous extracts were made basic with solid potassium carbonate and extracted with ethyl acetate. The organic extract was washed with brine, dried (MgSO4) and concentrated to give a yellow solid which was recrystallised from methanol/petrol then ethyl acetate/chloroform/petrol to give the title compound (298 mg, 68%) as an off-white solid. 1H NMR (methanol-d4) 7.72 (1 H, t), 7.41 (1 H, d), 7.39 (1 H, d), 7.02 (1 H, d), 6.51 (1 H, d), 6.43 (1 H, dd), 4.72 (2H, s), 4.27 (2H, t), 3.81 (3H1 s), 3.76 (3H, s), 3.68 (4H, t), 2.61 (2H, t), 2.50 (4H, m), 2.05 (2H, qn). MS: [M+H]+ 441.
PREPARATION C13
2-(4-Methoxy-benzyl)-4-r2-(2-methoxy-ethoxy)-ethoxy1-2,3-dihydro-1 H-isoindole
Figure imgf000141_0002
2-(4-Methoxy-benzyl)-4-[2-(2-methoxy-ethoxy)-ethoxy]-isoindole-1 ,3-dione (149 mg, 0.38 mmol) was treated with a 1 M solution of lithium aluminium hydride in THF (5 mL, 5 mmol). The mixture was maintained at r.t. for 4 hours, 60 0C for 1 hour, then r.t. for a further 18 hours. The mixture was then cooled in ice and quenched by the dropwise addition of water (0.2 mL), 2N sodium hydroxide solution (0.4 mL) and water (0.4 mL). Magnesium sulphate was added, followed by ethyl acetate and then the mixture was stirred at r.t. for 15 minutes. The solids were removed by filtration, being well washed with ethyl acetate. Concentration of the filtrate gave a residue which was absorbed onto an SCX cartridge and washed with 5% methanol/dichloromethane then eluted with 10% 1 M ammonia in methanol/dichloromethane to afford the title compound (134 mg, 97%). 1H NMR (methanol-d4) 7.43-7.39 (2H, m), 7.27 (1 H, t), 6.99-6.96 (2H, m), 6.90 (1 H, d), 6.88 (1 H, d), 4.33 (2H, s), 4.28 (2H, s), 4.23 (2H, s), 4.18-4.15 (2H, m), 3.85-3.79 (5H, m), 3.67-3.64 (2H, m), 3.54- 3.51 (2H, m), 3.33 (3H, s). MS: [M+H]+ 358.
PREPARATION C14 2-(2,4-Dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-2,3-dihvdro-1 H-isoindole
Figure imgf000142_0001
2-(2,4-Dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-isoindole-1 ,3-dione (201 mg, 0.52 mmol) was treated with a 1 M solution of lithium aluminium hydride in THF (5 ml_, 5 mmol). After 7.5 hours at r.t. a further portion of lithium aluminium hydride solution (5 ml_, 5 mmol) was added and the mixture maintained for further 18 hours. The mixture was then cooled in ice and quenched by the dropwise addition of water (0.4 ml_), 2N sodium hydroxide solution (0.8 mL) and water (0.8 mL).
Magnesium sulphate was added, followed by ethyl acetate and then the mixture was stirred at r.t. for 1 hour. The solids were removed by filtration, being well washed with ethyl acetate.
Concentration of the filtrate gave the title compound (192 mg, 103%) as a brown oil which was carried forward without further purification. 1H NMR (methanol-d4) 7.24 (1H, d), 7.16 (1H, t), 6.82-
6.78 (2H, m), 6.55 (1 H, d), 6.51 (1 H, dd), 4.12 (2H, t), 3.92 (4H, s), 3.86 (2H, s), 3.82 (3H, s), 3.80
(3H, s), 2.76 (2H, t), 2.33 (6H, s). MS: [M+H]+ 357.
PREPARATION C15 2-(2,4-Dimethoxy-benzyl)-4-(3-morpholin-4-yl-propoxy)-2,3-dihvdro-1 H-isoindole
Figure imgf000142_0002
2-(2,4-Dimethoxy-benzyl)-4-(3-morpholin-4-yl-propoxy)-isoindole-1 ,3-dione (298 mg, 0.68 mmol) was treated with a 1 M solution of lithium aluminium hydride in THF (5 mL, 5 mmol) and maintained at room temperature for 21 hours. The mixture was heated to 75 °C for 1 hour then cooled in ice and quenched by the dropwise addition of water (0.2 mL), 2N sodium hydroxide solution (0.4 mL) and water (0.4 mL). Magnesium sulphate was added, followed by ethyl acetate and then the mixture was stirred at room temperature for 1 hour. The solids were removed by filtration, being well washed with ethyl acetate. Concentration of the filtrate gave a crude product which was purified by flash chromatography on silica, eluting with 5% methanol in DCM. This afforded the title compound (233 mg, 83%) as a red oil. 1H NMR (methanol-d4) 7.24 (1 H, d), 7.15 (1 H, t), 6.80 (1 H, d), 6.78 (1 H, d), 6.56 (1 H, d), 6.52 (1 H, dd), 4.05 (2H, t), 3.94 (2H, s), 3.88 (2H, s), 3.87 (2H, S)1 3.83 (3H, s), 3.80 (3H, s), 3.70-3.68 (4H, m), 2.54-2.50 (2H, m), 2.49-2.47 (4H, m), 2.00-1.93 (2H, m). MS: [M+H]+ 413.
PREPARATION C16 4-f2-(2-Methoxy-ethoxy)-ethoxyl-2,3-dihvdro-1 H-isoindole
Figure imgf000143_0001
A solution of 2-(4-methoxy-benzyl)-4-[2-(2-methoxy-ethoxy)-ethoxy]-2,3-dihydro-1 H-isoindole (45 mg, 0.13 mmol) in 1 ,2-dichloroethane (2 mL) was treated with α-chloroethyl chloroformate (0.1 ml_, 0.93 mmol). After 17 hours, methanol (5 mL) was added and the mixture stirred for 3 hours. The solvents were removed in vacuo to afford the title compound as a greenish-black solid, which was used without further purification. 1H NMR (methanol-d4) 7.36 (1 H, t), 6.98 (2H, d), 4.60 (2H, s), 4.57 (2H, s), 4.23-4.21 (2H, m), 3.85-3.83 (2H, m), 3.69-3.67 (2H, m), 3.57-3.54 (2H, m), 3.36 (3H, s). MS: [M+H]+ 238.
PREPARATION C17 [2-(2,3-Dihvdro-1 H-isoindol-4-yloxy)-ethyll-dimethvl-amine
Figure imgf000143_0002
A solution of 2-(2,4-dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-2,3-dihydro-1 H-isoindole (170 mg, 0.48 mmol) in trifluoroacetic acid (0.5 mL) and anisole (0.5 mL) was heated at 150 0C under microwave irradiation for 10 minutes. The mixture was diluted with ethyl acetate and extracted twice with water. The combined aqueous extracts were concentrated to give the title compound as a purple oil (240 mg, including residual TFA and/or water). 1H NMR (methanol-d4) 7.42 (1 H, t), 7.07 (1 H, d), 7.04 (1 H, d), 4.64 (4H, br.s), 4.47-4.44 (2H, m), 3.65-3.63 (2H, m), 3.01 (6H, s). MS: [M+H]+ 207.
PREPARATION C18 4-(3-Morpholin-4-yl-propoxy)-2,3-dihvdro-1 H-isoindole
Figure imgf000144_0001
A solution of 2-(2,4-dimethoxy-benzyI)-4-(3-morpholin-4-yl-propoxy)-2,3-dihydro-1 H-isoindole (233 mg, 0.56 mmol) in trifluoroacetic acid (1.0 mL) and anisole (0.5 mL) was heated at 150 0C under microwave irradiation for 10 minutes. The mixture was diluted with diethyl ether and extracted twice with water. The combined aqueous extracts were concentrated to give an oil which was dissolved in methanol and concentrated in vacuo to afford the title compound as a brown oil (348 mg, including residual TFA and/or water). 1H NMR (methanol-d4) 7.40 (1 H, t), 7.03 (1 H, d), 6.99 (1 H, d), 4.63 (2H, s), 4.59 (2H, s), 4.21 (2H, t), 4.14-4.04 (2H, m), 3.85-3.73 (2H, m), 3.61-3.52 (2H, m), 3.41-3.36 (2H, m), 3.25-3.13 (2H, m), 2.32-2.25 (2H, m). MS: [M+H]+ 263.
PREPARATION C19
Synthesis of 4-bromo-2,3-dihvdro-1 H-isoindole trifluoroacetate
Figure imgf000144_0002
Prepared in a manner analogous to 5-nitro-2,3-dihydro-1 H-isoindole (described in preparation C5). 1H NMR (DMSO-d6) 9.73 (2H, br s), 7.60 (1H, d), 7.45 (1 H, d), 7.35 (1 H, t), 4.65 (2H, s), 4.55 (2H, s).
PREPARATION C20
Synthesis of 5-bromo-2,3-dihydro-1 H-isoindole
Figure imgf000144_0003
A mixture of 4-bromophthalic anhydride (25 g) in formamide (75 ml) was heated at 2000C for 16 hours then allowed to cool to room temperature. The reaction mixture was diluted with water (200 ml), filtered, the filter cake was washed with water then diethyl ether and sucked dry to give 20.85 g of light mustard solid.
280 ml of 1M Borane-THF complex was added dropwise to a stirred solution of 4- bromophthalimide (20.85 g; 92.2 mmol) in anhydrous THF (200 ml) at O0C then heated at reflux overnight. The reaction was cooled to 00C then treated cautiously with methanol (100 ml) followed by 2M HCI (100 ml) then heated at reflux for 3 hours. The reaction mixture was cooled and the organics evaporated. The aqueous was diluted with water (100 ml) the extracted with DCM (x3). The aqueous was basified with 2M NaOH then extracted with DCM (x3). The combined DCM extracts were dried (MgSCU), filtered and evaporated to give 6.99 g of 5-bromo-2,3-dihydro-1 H- isoindole as a dark brown gummy solid. 1H NMR (DMSOd6) 7.45 (1 H, s), 7.36 (1 H, d), 7.20 (1 H, d), 4.05 (4H, s).
PREPARATION C21
Synthesis of 2.3-dihvdro-1 H-isoindole-5-carboxylic acid methyl ester trifluoroacetate
Figure imgf000145_0001
2-(2,4-Dimethoxybenzyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methyl ester (preparation C6, step 2 product) was deprotected in a manner analogous to 5-nitro-2,3-dihydro-1 H-isoindole
(described in preparation C5) to give the title compound. 1H NMR (DMSO-d6) 9.70 (2H, br s), 8.00 (1 H, s), 7.95 (1 H, d), 7.57 (1 H, d), 4.60 (4H, s), 2.88 (3H, s).
D. SYNTHESIS OF BENZYLATED RESORCINOL INTERMEDIATES PREPARATION D1 Synthesis of (2,4-bis-benzyloxy-5-isoproDyl-phenyl)-r5-(2-methoxy-ethoxy)-1 ,3-dihydro-isoindol-2- yli-methanone
Figure imgf000145_0002
(2,4-Bis-benzyloxy-5-isopropyl-phenyl)-(5-hydroxy-1 ,3-dihydro-isoindol-2-yl)-methanone (A2 from 2,4-bis-benzyloxy-5-isopropyl-benzoic acid (Preparation B10) and 5-hydroxyisoindoline) (100 mg, 0.2 mmol), i-chloro-2-methoxy-ethane (23.6 mg, 0.25 mmol) and K2CO3 (34.5 mg, 0.25 mmol) in DMF (4 ml) were combined and stirred for 2 hours at room temperature. A further 0.25 mmol of 1- chloro-2-methoxy-ethane and K2CO3 was added then heated at 90 0C for 16 hours. Reaction cooled to room temperature and diluted with EtOAc then filtered. The filtrate was reduced in vacuo then purified by flash column chromatography, eluting with 100% petroleum ether to 100 % ethyl acetate to afford 115 mg of the title compound as a colourless gel. MS: [M+H]+ 552
PREPARATION D2
Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-f5-(3-morpholin-4-yl-propoxy)-1 ,3-dihydro- isoindol-2-vπ-methanone
Figure imgf000146_0001
A mixture of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-hydroxy-1 ,3-dihydro-isoindol-2-yl)- methanone (100 mg, 0.2 mmol), 4-(3-chloropropyl)morpholine (82 mg, 0.5 mmol) and K2CO3 (104 mg, 0.75 mmol) in DMF (5ml) was heated at 900C for 16 hours. The reaction mixture was diluted with EtOAc and filtered. The filtrate was reduced in vacuo and purified by flash column chromatography, eluting with 0-100% P.E./EtOAc then 0-10% MeOH/EtOAc to give the title compound as a colourless gel (90.1 mg). MS: [M+H]+ 621.
PREPARATION D3
Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-ethoxy)-1 ,3-dihvdro- isoindol-2-yll-methanone
Figure imgf000146_0002
A mixture of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-hydroxy-1 ,3-dihydro-isoindol-2-yl)- methanone (100 mg, 0.2 mmol), 2-dimethylaminoethylchloride.HCI (72 mg, 0.5 mmol) and K2CO3 (173 mg, 1.25 mmol) in DMF (5ml) was heated at 90 0C for 16 hours. Dilute reaction mixture with EtOAc and filtered. The reaction mixture was diluted with EtOAc and filtered. The filtrate was reduced in vacuo and purified by flash column chromatography, eluting 100% DCM then 90% DMAW 90 to give the title compound as an off white gel (79 mg). MS: [M+H]+ 565
PREPARATION D4
Synthesis of 2.4-bis-benzyloxy-5-isopropyl-benzoyl chloride
Figure imgf000146_0003
2,4-Bis-benzyloxy-5-isopropyl-benzoic acid (Preparation B10) (0.2 g, 0.53 mmol) was dissolved in
DCM (10ml) and treated with oxalyl chloride (1.5g, 12 mmol) and a catalytic amount of DMF. The reaction mixture was stirred at room temperature for 14 hours and the solvent was then removed in vacuo. The crude material was dissolved in toluene and evaporated. Crude 2,4-bis-benzyloxy-5- isopropyl-benzoyl chloride was obtained as an oil (200 mg).
PREPARATION D5
Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenvπ-(5-morpholin-4-yl-1 ,3-dihydro-isoindol-2-yl)- methanone
A solution of 2,4-bis-benzyloxy-5-isopropyl-benzoic acid (505 mg; 1.3 mmol) (Preparation B5), 5- nitroisoindoline, trifluoroacetate (360 mg; 1 equiv.), EDAC (300 mg; 1.2 equiv.), HOBt (210 mg; 1.2 equiv.) and NEt3 (270 μl; 1.5 equiv.) in DMF (10 ml) was stirred at room temperature overnight then evaporated in vacuo. The residue was partitioned between EtOAc and 2M HCI, the EtOAc layer was separated, washed with saturated NaHCO3, dried (MgSO4) and evaporated. Purification by flash column chromatography (1 :4 then 1 :2 then 1 :1 EtOAc/P.E. as eluant) gave 460 mg of (2,4-bis-benzyloxy-5-isopropyl-phenyI)-(5-nitro-1 ,3-dihydro-isoindol-2-yl) methanone. MS: [M+H]+ 523. A solution of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-nitro-1 ,3-dihydro-isoindol-2-yl) methanone (460 mg; 0.88 mmol) in ethanol (25 ml) was treated with tin (II) chloride dihydrate (1 g; 5 equiv.) then heated at reflux overnight then evaporated in vacuo. The residue was partitioned between EtOAc and saturated NaHCO3, the EtOAc layer was separated, dried (MgSO4) and evaporated to give 380 mg of (5-amino-1 ,3-dihydro-isoindol-2-yl)-(2,4-bis-benzyloxy-5-isopropyl-phenyl)- methanone.
A mixture of (5-amino-1 ,3-dihydro-isoindol-2-yl)-(2,4-bis-benzyloxy-5-isopropyl-phenyl)-methanone (100 mg; 0.2 mmol), bis(2-chloroethyl)ether (30 μl; 1.1 equiv.), Hunigs base (125 μl; 3.5 equiv.) and tetrabutylammonium iodide (10 mg) in NMP (1 ml) was heated in a CEM microwave synthesiser at 1500C for 30 minutes. A further 30 μl of Hunig's base and 125 μl of bis(2- chloroethyl)ether were added and heating repeated for the same time. The reaction mixture was partitioned between EtOAc and saturated NH4CI solution, the EtOAc layer was separated, washed with more saturated NH4CI solution, then brine, dried (MgSO4) and evaporated. Purification by flash column chromatography (1 :2 then 1 :1 then 2:1 EtOAc/P.E. as eluant) gave 60 mg of (2,4-bis- benzyloxy-5-isopropyl-phenyi)-(5-morpholin-4-yl-1 ,3-dihydro-isoindol-2-yl)-methanone. MS: [M+H]+ 563.
PREPARATION D6
Synthesis of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihvdro-1 H-isoindole-5-carboxylic acid
Figure imgf000148_0001
A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methyl ester (390 mg) in methanol (10 ml) and 2M NaOH (10 ml) was heated at 5O0C for 48 hours then evaporated. The residue was acidified with 2M HCI, the solid collected by filtration, washed with water and sucked dry to give 255 mg of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3- dihydro-1 H-isoindole-5-carboxylic acid as a white solid. [M+H]+ 520.
EXAMPLES
By following the methods described above, the compounds set out in the Table below were prepared.
Figure imgf000148_0002
Figure imgf000149_0001
Figure imgf000150_0001
03860
150
Figure imgf000151_0001
Figure imgf000152_0002
EXAMPLE 17
Synthesis of (5-chloro-1 ,3-dihvdro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone
Figure imgf000152_0001
A solution of 2, 4-bis-benzyloxy-5-isopropyl-benzoic acid (Preparation B10) (0.451 g, 1.2 mmol ), EDC (0.276 mg, 1.44 mmol), HOAt (0.196 mg, 1.44 mmol), triethylamine (0.5 ml, 3.6 mmol) and 5- chloro-2,3-dihydro-1 H-isoindole ( 0.187 g, 1.2 mmol) (Preparation C3) in DMF (5 ml) was stirred at room temperature for 16 hours, then evaporated under vacuum. The crude material was dissolved in ethyl acetate and extracted twice with saturated NaHCO3, organics washed with water three times, then evaporated under vacuum to give 0.5 g of 2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5- chloro-1 ,3-dihydro-isoindol-2-yl)-methanone. MS: [M+H]+ 512
Boron trichloride (1 M in DCM) was added dropwise to a solution of 2,4-bis-benzyloxy-5-isopropyl- phenyl)-(5-chloro-1 ,3-dihydro-isoindol-2-yl)-methanone (0.5 g, 0.97mmol) in dry DCM (10 ml) at O0C under nitrogen, then stirred for at 0 0C for 1 hour, warmed to room temperature and stirred for a further 3 hours. The reaction was quenched with ice, partitioned between DCM and water. The DCM layer was dried (MgSO4), evaporated under vacuum, then purified by flash silica column chromatography eluting with 80% P. E.: EtOAc to give 0.1 g of (5-chloro-1 ,3-dihydro-isoindol-2-yl)- (2,4-dihydroxy-5-isopropyl-phenyl)-methanone as a white solid. MS: [M+H]+ 332. 1H NMR (DMSO- d6) 10.0 (1 H, s) 9.60 (1 H, s), 7.45 (1 H, br s), 7.33 (2H, br s), 7.0 (1 H, s), 6.4 (1 H, s), 4.80 (4H, br s), 3.10 (1H, m), 1.15 (6H, d).
EXAMPLE 18
Synthesis of f5-(3-amino-propoxy)-1.3-dihvdro-isoindol-2-vπ-(2.4-dihvdroxy-5-isopropyl-phenyl)- methanone hydrochloride
Figure imgf000153_0001
A solution of {3-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindol-5-yloxy]-propyl}- carbamic acid tert-butyl ester (Example 46) (1 g) in EtOAc (10 ml) was treated with a saturated solution of HCI in EtOAC (20 ml) then stirred at room temperature for 2 hours,. The reaction mixture was evaporated and re-evaporated with ethanol (x 3). The title compound was isolated as a cream foam (840 mg). 1 H NMR (DMSO-d6) 10.05 (1 H, br s), 9.60 (1 H, s), 7.88 (3H1 br s), 7.30- 7.18 (1 H, m), 7.05(1 H, s), 7.00-6.85 (2H, m), 6.42 (1 H, s), 4.75 (2H, br s) 4.70 (2H, br s), 4.05 (2H, t), 3.10 (1 H, m), 3.00-2.95 (2H, m), 2.00 (2H, tt), 1.15 (6H, d). MS: [M+H]+ 371.
EXAMPLE 19 (5-Bromo-2,4-dihvdroxy-phenylH1 ,3-dihvdro-isoindol-2-yl)-methanone
Figure imgf000153_0002
A solution of 5-bromo-2,4-dihydroxy-benzoic acid (520 mg, 2.33 mmol) in DMF (5 mL) was treated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (471 mg, 2.45 mmol) then HOBt (362 mg, 2.68 mmol). After 25 min, 2,3-dihydro-1 H-isoindoIe (0.5 mL, 2.63 mmol) was added then the mixture was stirred at r.t. for 18 h. The solvent was removed in vacuo then the residue was taken up in ethyl acetate and washed with 1 N hydrochloric acid, saturated sodium bicarbonate solution and brine then dried (MgSO4) and concentrated. The residue was triturated with methanol to afford the title compound as a grey solid (328 mg, 44%). 1H NMR (DMSO-d6) 10.45 (1 H, s), 10.32 (1 H, s), 7.36 (1 H, br.s), 7.35 (1 H, s), 7.28 (3H, br.s), 6.59 (1 H, s), 4.77 (2H, br.s), 4.71 (2H, br.s). MS: [M+H]+ 332/334.
EXAMPLE 20
(1 ,3-Dihvdro-isoindol-2-vπ-(2,4-dihvdroxy-5-trifluoromethyl-phenyl)-methanone 2OA. (2,4-Bis-benzyloxy-5-bromo-phenyl)-(1.3-dihydro-isoindol-2-yl)-methanone
Figure imgf000154_0001
According to general method A2, 2,4-bis-benzyloxy-5-bromo-benzoic acid (1.02 g, 2.47 mmol) gave a residue which was purified by flash chromatography on silica (ethyl acetate/petrol gradient, O - 20%) to afford the title compound as a white crystalline solid (501 mg, 39%). 1H NMR (methanol-d4) 7.52 (1 H, s), 7.49-7.46 (2H, m), 7.42-7.37 (2H, m), 7.34 (t, 2H), 7.30-7.24 (4H1 m), 7.23-7.20 (3H, m), 7.16 (1 H, d), 6.94 (1 H, s), 5.24 (2H, s), 5.16 (2H, s), 4.86 (2H, s), 4.60 (2H, s). MS: [M+H]+ 514/516.
2OB. (2,4-Bis-benzyloxy-5-trifluoromethyl-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone
Figure imgf000154_0002
A mixture of (2,4-bis-benzyloxy-5-bromo-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone (491 mg, 0.95 mmol), sodium trifluoroacetate (649 mg, 4.8 mmol) and copper (I) iodide (364 mg, 1.91 mmol) were dried under vacuum (0.04 mbar) for 6 hours. The flask was flushed with nitrogen, DMF (5 mL) was added and the mixture heated at 150 0C for 17 hours. After cooling to r.t., the mixture was diluted with DCM (100 mL) and filtered through Celite, rinsing with DCM. The filtrate was concentrated to dryness and the residue was partially purified by flash chromatography on silica (ethyl acetate/petrol gradient, 0 - 20%). The purest fraction was recrystallised from methanol to afford the title compound as a white solid (140 mg, 29%). 1H NMR (methanol-d4) 7.60 (1 H, s), 7.48-7.44 (2H1 m), 7.40 (2H, t), 7.37-7.21 (m, 9H), 7.17 (1 H, d), 7.02 (1 H, s), 5.29 (2H, s), 5.24 (2H, s), 4.88 (2H, s), 4.62 (2H, s). MS: [M+H]+ 504. 2OC. (1 ,3-Dihvdro-isoindol-2-yl)-(2,4-dihvdroxy-5-trifluoromethyl-phenyl)-methanone
Figure imgf000154_0003
A solution of (2,4-bis-benzyloxy-5-trifluoromethyl-phenyl)-(1 ,3-dihydro-isoindol-2-yl)-methanone (140 mg, 0.28 mmol) in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (34 mg) for 4 hours. A further portion of catalyst was added (31 mg) and hydrogenation continued for a further 1.5 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to afford the title compound as a white solid (91 mg, quant.). 1H NMR (DMSO-d6) 10.79 (1 H, s), 10.70 (1 H, s), 7.40-7.35 (2H, m), 7.31- 7.35 (3H, m), 6.61 (1 H, s), 4.79 (2H, br.s), 4.68 (2H, br.s). MS: [M+H]+ 324.
EXAMPLE 21
(2,4-Dihvdroxy-5-isopropyl-phenvπ-{4-[2-(2-methoxy-ethoxy)-ethoxy1-1 ,3-dihydro-isoindol-2- vDmethanone
Figure imgf000155_0001
A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (96 mg, 0.26 mmol) and DMF (1 drop, cat.) in DCM (3 mL) was cooled in ice then treated with oxalyl chloride (112 μl_, 1.28 mmol). After 2 hours the mixture was concentrated in vacuo then azeotroped with toluene. The resulting acid chloride was dissolved in DCM (4 mL) and added to a solution of 4-[2-(2-methoxy-ethoxy)-ethoxy]- 2,3-dihydro-1 H-isoindole (0.26 mmol, assuming a quantitative yield from the preceding step (debenzylation procedure C16)) and triethylamine (0.20 mL, 1.4 mmol) in DCM (1 mL). After 2 hours the mixture was diluted with ethyl acetate and washed with 1 N hydrochloric acid, brine, sodium bicarbonate solution and brine. The organic phase was dried (MgSO4) and concentrated to give a black residue. This was partially purified by flash chromatography on silica (ethyl acetate/petrol gradient, 20 - 33%) to afford an impure sample of the intermediate (2,4-bis- benzyloxy-5-isopropenyl-phenyl)-{4-[2-(2-methoxy-ethoxy)-ethoxy]-1 ,3-dihydro-isoindol-2-yl}- methanone.
A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-{4-[2-(2-methoxy-ethoxy)-ethoxy]-1 ,3- dihydro-isoindol-2-yl}-methanone in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (12 mg) for 3 hours. A further portion of catalyst (12 mg) was added and hydrogenation continued for a further 7 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to give a residue which was purified by preparative HPLC (basic method). This afforded the title compound as a white solid (17 mg, 16% over two steps). 1H NMR (methanol-d4) 7.25 (1 H, t), 7.17 (1 H, s), 6.95-6.82 (2H, m), 6.37 (1H, s), 4.89 (2H, br.s), 4.83 (overlaps with H2O, br.s), 4.16 (2H, br.s), 3.82 (2H, br.s), 3.66 (2H, br.s), 3.52 (2H, br.s), 3.39-3.28 (overlaps with MeOH, m), 3.20 (1 H, sept), 1.21 (6H, d). MS: [M+H]+ 416.
EXAMPLE 22
(2,4-Dihvdroxy-5-isopropyl-phenvπ-f4-(2-dimethylamino-ethoxy)-1 ,3-dihvdro-isoindol-2-yl1- methanone
DCM
Figure imgf000156_0001
Figure imgf000156_0002
A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (189 mg, 0.50 mmol) and DMF (1 drop, cat.) in DCM (5 ml_) was cooled in ice then treated with oxalyl chloride (112 μL, 1.28 mmol). After 2 hours the mixture was concentrated in vacuo then azeotroped with toluene. The resulting acid chloride was dissolved in DCM (5 mL) and added to a solution of [2-(2,3-dihydro-1 H-isoindol-4- yloxy)-ethyl]-dimethyl-amine (0.48 mmol, assuming a quantitative yield from the preceding step (C17)) and triethylamine (0.50 mL, 3.6 mmol) in DCM (3 mL). After 16 hours the mixture was diluted with ethyl acetate and washed with saturated potassium carbonate solution and brine. The organic phase was dried (MgSO4) and concentrated to give a residue which was partially purified by flash chromatography on silica (methanol/DCM gradient, 5 - 10% followed by 10% 2M methanolic ammonia/DCM) to afford an impure sample of the intermediate (2,4-bis-benzyloxy-5- isopropenyl-phenyl)-[4-(2-dimethylamino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]-methanone. A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(2-dimethylamino-ethoxy)-1 ,3-dihydro- isoindol-2-yl]-methanone in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (40 mg) for 22 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to give a residue which was purified by preparative HPLC (acidic method). This afforded the formate salt of the title compound as a white solid (9 mg, 5% over two steps). 1H NMR (methanol-d4) 8.52 (0.7H, s), 7.29 (1 H, t), 7.17 (1 H, s), 6.98-6.86 (2H, m including 6.90 (1 H, d)), 6.37 (1 H, s), 4.89 (2H, br.s), 4.87 (2H, br.s), 4.28 (2H, br.s), 3.29-3.5 (3H, m including 3.20 (1 H, sept)), 2.81-2.51 (6H, br.d), 1.21 (6H, d). MS: [M+H]+ 385.
EXAMPLE 23 (2.4-Dihvdroxy-5-isopropyl-phenyl)-f4-(3-morpholin-4-yl-propoxy)-1 ,3-dihvdro-isoindol-2- ylimethanone
Figure imgf000157_0001
A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (210 mg, 0.56 mmol) and diisopropylethylamine (0.25 ml_, 1.4 mmol) in DCM (5 ml.) was treated with bromo-tris-pyrrolidino- phosphonium hexafluorophosphate (PyBrOP) (287 mg, 0.62 mmol). After 1 hour a solution of 4-(3- morpholin-4-yl-propoxy)-2,3-dihydro-1 H-isoindole (0.56 mmol, assuming a quantitative yield from the preceding step(C18)) in DCM (5 mL) was added. After 4 hours the mixture was diluted with ethyl acetate and washed with water, 1 N sodium hydroxide solution and brine. The organic phase was dried (MgSO4) and concentrated to give a residue which was absorbed onto an SCX column. This was washed with 10% methanol/DCM then the product was eluted with 25% 2M methanolic ammonia / DCM) to afford an impure sample of the intermediate (2,4-bis-benzyloxy-5-isopropenyl- phenyl)-[4-(3-morpholin-4-yl-propoxy)-1 ,3-dihydro-isoindol-2-yl]-methanone.
A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1 ,3-dihydro- isoindol-2-yl]-methanone in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (45 mg) for 4 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to give a residue which was purified by preparative HPLC (basic method). This afforded the title compound as a white solid (16 mg, 6% over two steps). 1H NMR (methanol-d4) 7.24 (1 H, t), 7.18 (1 H, s), 6.89 (1 H, d), 6.84 (1 H, d), 6.37 (1 H, s), 4.87 (2H, br.s), 4.78 (2H, br.s), 4.11-4.04 (2H, m), 3.72-3.66 (4H, m), 3.21 (1 H, sept), 2.60- 2.42 (6H, m), 2.05-1.92 (2H, m), 1.21 (6H1 d). MS: [M+H]+ 441.
EXAMPLES 24 TO 47
By following the methods described above, the compounds of Examples 24 to 47 were prepared.
Figure imgf000157_0002
Figure imgf000158_0001
Figure imgf000159_0001
Figure imgf000160_0001
Figure imgf000161_0001
Figure imgf000162_0001
Figure imgf000163_0001
Figure imgf000164_0002
EXAMPLE 48
Synthesis of (Σ^-dihvdroxy-δ-isopropyl-phenvD-Fδ-fΣ-isopropylamino-ethoxyVI .S-dihvdro-isoindol-
2-vH-methanone
Figure imgf000164_0001
To a suspension of [5-(3-amino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]-(2,4-dihydroxy-5-isopropyl- phenyl)-methanone hydrochloride (Example 57) (250 mg, 0.702 mmoles) in 1 ,2-dichloroethane (10ml) was added acetone (62 μl, 0.842 mmoles), sodium triacetoxyborohydride (178 mg, 0.842 mmoles) and acetic acid (48 μl, 0.842 mmoles) and then heated at 6O0C for 24 hours. To the reaction mixture was added further acetone (52 μl, 0.702 mmoles), sodium triacetoxyborohydride (149 mg, 0,702 mmoles) and acetic acid (40 μl, 0.702 mmoles) and heated at 6O0C for a further 2 hours. The reaction mixture was then filtered and the mother liquor purified by flash chromatography [Biotage SP4: 25M, flow rate 25ml/min, gradient 20% to 100% DMAW 90 in DCM) to give (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(2-isopropylamino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]- methanone as a light brown viscous oil (140mg, 50%). 1H NMR (DMSO-d6) 10.05 (1H, br s); 9.60 (1 H, br s); 7.23 (1 H, br s); 7.05 (1 H, s); 6.93 (1 H, br s); 6.85 (1 H, br d); 6.40 (1 H, s); 4.70 (4H, br m); 4.00 (2H, t); 3.10 (1 H, m); 2.90 (2H, t); 2.80 (1 H, m); 1.15 (6H, d); 1.00 (6H, d). MS: [M+H]+ 399.
EXAMPLE 49 Synthesis of N-{2-[2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihvdro-1 H-isoindol-5-yloxy1-ethyl)-2- morpholin-4-yl-acetamide
Figure imgf000165_0001
To a solution of [5-(3-amino-ethoxy)-1 ,3-dihydro-isoindol-2-yl]-(2,4-dihydroxy-5-isopropyl-phenyI)- methanone hydrochloride (100 mg, 0.255 mmoles) in DMF (10 ml) was added EDC (59 mg, 0.306 mmoles), HOBt (41 mg, 0.306 mmoles), morpholin-4-yl-acetic acid (37 mg, 0.255mmoles) and triethylamine (43μl, 0.306m moles) and stirred at ambient temperature for one hour. To the reaction mixture was added further EDC (20 mg, 0.104 mmoles), HOBt (14 mg, 0.104 mmoles), morpholin- 4-yl-acetic acid (12 mg, 0.083 mmoles) and triethylamine (14 μl, 0.100 mmoles) and stirred at ambient temperature for a further 2 hours. Solvent removed in vacuo. The residue was purified by flash chromatography [Biotage SP4: 25S, flow rate 25 ml/min, gradient 20% DMAW 90 in DCM to 100% DMAW 90] and then by preparative HPLC to give N-{2-[2,4-dihydroxy-5-isopropyl-benzoyl)- 2,3-dihydro-1 H-isoindol-5-yloxy]-ethyl}-2-morpholin-4-yl-acetamide as a colourless viscous oil (40 mg, 33%). 1H NMR (Me-d3-OD) 7.20 (1 H, br s); 7.18 (1 H, s); 6.90 (2H, br m); 6.40 (1 H, s); 4.10 (2H, t); 3.73 (4H, m); 3.63 (2H, t); 3.20 (1 H, m); 3.18 (2H, s); 2.60 (4H, m); 1.25 (6H1 d). MS: [M+H]+ 484.
EXAMPLE 50
Synthesis of (2,4-dihvdroxy-5-isopropyl-phenyl)-[5-(4-hvdroxy-1-methyl-piperidin-4-yl)-1 ,3-dihydro- isoindol-2-yli-methanone
Figure imgf000166_0001
5OA: Synthesis of 5-bromo-1 ,3-dihvdro-isoindole-2-carboxylic acid tert-butyl ester
A mixture of 5-bromo-2,3-dihydro-1 H-isoindole (1.26 g; 6.4 mmol), di-tert-butyl dicarbonate (1.53 g; 1.1 equiv.) and 4-dimethylaminopyridine (catalytic amount) in DMF (20 ml) was stirred at room temperature overnight then evaporated. The residue was partitioned between EtOAc and brine, the EtOAc layer was separated, dried (MgSO4) and evaporated. Purification by flash column chromatography using a Biotage SP4 (4OS, 40 ml/min) eluting with 0% to 5% MeOH / DCM gave 695 mg of 5-bromo-1 ,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester as a brown gum. 1HNMR (DMSO-d6) 7.55 (1 H, d), 7.48 (1 H, d), 7.30 (1 H, dd), 4.63-4.51 (4H, m), 1.46 (9H, s). 5OB. Synthesis of 5-(4-hvdroxy-1-methyl-piperidin-4-yl)-1 ,3-dihydro-isoindole-2-carboxylic acid tert- butyl ester
0.69 ml of n-Butyl lithium (2.5M solution in hexane) was added dropwise to a stirred solution of 5- bromo-1 ,3-dihydro-isoindole-2-carboxy!ic acid tert-butyl ester (429 mg; 1.44 mmol) in anhydrous THF (10 ml) at -780C under an atmosphere of nitrogen. The reaction was stirred for 50 minutes then 1-methyl-4-piperidone (212 μl; 1.2 equiv.) was added and stirred at -780C for a further 60 minutes then warmed to room temperature. The reaction was quenched with saturated ammonium chloride solution then extracted with EtOAc. The EtOAc layer was washed with saturated NaHCO3, brine, dried (MgSO4) and evaporated. Purification by flash column chromatography on SiO2, gradient elution from 0% to 10% 2M methanolic ammonia / DCM gave 111 mg of 5-(4-hydroxy-1- methyl-piperidin-4-yl)-1 ,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester as a colourless oil.
5OC. Synthesis of 4-(2,3-dihvdro-1 H-isoindol-5-vO-1-methyl-piperidin-4-ol
A solution of 5-(4-hydroxy-1-methyl-piperidin-4-yl)-1.S-dihydro-isoindole^-carboxylic acid tert-butyl ester (107 mg; 0.32 mmol) in THF (4 ml) was treated with concentrated hydrochloric acid (1.5 ml) then heated at reflux for 4 hours, then evaporated and re-evaporated with toluene to give 4-(2,3- dihydro-1 H-isoindol-5-yl)-1-methyl-piperidin-4-ol dihydrochloride as a brown gum.
5OD. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenylH5-(4-hvdroxy-1-methyl-piperidin-4-yl)- 1 ,3-dihvdro-isoindol-2-yl]-methanone
A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (145 mg; 1.2 equiv.) in DCM (5 ml) was treated with EDC (80 mg; 1.3 equiv.) and HOAt (66 mg; 1.5 equiv.) then stirred at room temperature for 30 minutes. This solution was then added to a mixture of 4-(2,3-dihydro-1 H- isoindol-5-yl)-1-methyl-piperidin-4-ol dihydrochloride (112 mg; 0.32 mmol) and triethylamine (90 μl; 2 equiv.) in THF (5 ml) and DMF (2 ml), the reaction was then stirred at room temperature overnight. The reaction mixture was diluted with EtOAc, washed with water, 1N NaOH and brine, the EtOAc layer was separated, dried (MgSO4) and evaporated. Purification by flash column chromatography on SiO2, gradient elution from 0% to 5% 2M methanolic ammonia / DCM gave 104 mg of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1 ,3- dihydro-isoindol-2-yl]-methanone as a yellow glass.
5OE. Synthesis of (2,4-dihvdroxy-5-isopropyl-phenvπ-r5-(4-hvdroxy-1-methyl-piperidin-4-yl)-1 ,3- dihvdro-isoindol-2-vπ-methanone Hydrogenation of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)- 1 ,3-dihydro-isoindol-2-yl]-methanone (as described in method A5) afforded 72 mg of the title compound as a cream solid. 1H NMR (Me-d3-OD) 7.35 (2H, m), 7.18 (1H, br m) 7.08 (1H, s), 6.25 (1 H, s), 4.78 (4H, m), 3.10 (1 H, m), 2.65 (2H, m), 2.45 (2H, m), 2.25 (3H, s), 2.00 (2H, m), 1.65 (2H, m), 1.10 (6H, d). MS: [IvHH]+ 411. EXAMPLE 51
Synthesis of (2,4-dihvdroxy-5-isopropyl-phenvπ-{5-r4-(4-methyl-piperazin-1-yl)-piperidin-1-vπ-1 ,3- dihvdro-isoindol-2-vD-methanone
Figure imgf000167_0001
51 A. Synthesis of (2.4-bis-benzyloxy-5-isopropenyl-phenvD-(5-bromo-1 ,3-dihvdro-isoindol-2-vD- methanone
A solution of benzyloxy-5-isopropenyl-benzoic acid (2.85 g; 7.6 mmol), 5-bromo-2,3-dihydro-1 H- isoindole (1.5 g; 1 equiv.) , EDC (1.75g; 1.2 equiv.) and HOBt (1.25 g; 1.2 equiv.) in DMF (25 ml) was stirred at room temperature overnight then evaporated. The residue was dissolved in EtOAc, washed with 2M HCI then saturated NaHCO3, dried (MgSO4) and evaporated. Purification using a Biotage SP4 (4OS, 40 ml/min) eluting with 1:4-1 :3-1 :2 EtOAc / P. E. gave 2.45 g of (2,4-bis- benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1 ,3-dihydro-isoindol-2-yl)-methanone as a light brown solid.
51 B. (2,4-bis-benzyloxy-5-isopropenyl-phenylH5-r4-(4-methyl-piperazin-1 -yl)-piperidin-1 -yli-1 ,3- dihvdro-isoindol-2-vD-methanone A solution of (2,4-bis-benzyIoxy-5-isopropenyl-phenyl)-(5-bromo-1 ,3-dihydro-isoindol-2-yl)- methanone (200 mg; 0.36 mmol) and 1-methyl-4-(piperidin-4-yl)piperazine (80 mg; 1.2 equiv.) in toluene (5 ml) was treated with (2-biphenyl)-di-tert-butylphosphine (6 mg; 5 mol%), tris(dibenzylidene)palladium(O) (10 mg; 2.5 mol%) and sodium tert-butoxide (50 mg; 1.4 equiv.) then heated at 1200C for 30 minutes in a CEM explorer microwave synthesiser. The reaction mixture was diluted with DCM, washed with brine, dried (MgSO4) and evaporated. Purification by flash column chromatography (Biotage SP4 - 25S, 25 ml/min) eluting with DMAW 240-120-90 followed by evaporation of product containing fractions gave 105 mg of (2,4-bis-benzyloxy-5- isopropenyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1 ,3-dihydro-isoindol-2-yl}- methanone as the acetic acid salt.
51 C. (2,4-dihvdroxy-5-isopropyl-phenyl)-(5-r4-(4-methyl-piperazin-1-yl)-piperidin-1-vπ-1 ,3-dihvdro- isoindol-2-vD-methanone hydrochloride
A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1- yl]-1 ,3-dihydro-isoindol-2-yl}-methanone acetic acid salt in methanol (10 ml) was treated with 10% palladium on carbon (wet), hydrogenated at room temperature and pressure overnight then filtered and evaporated. The crude compound was purified by flash column chromatograph (Biotage SP4 - 25S, 25 ml/min) eluting with DMAW 240-120-90-60. Product containing fractions were evaporated, treated with saturated HCI / EtOAc then evaporated and re-evaporated with methanol and dried under high vacuum at 6O0C overnight. (2,4-dihydroxy-5-isopropyl-phenyl)-{5-[4-(4-methyl- piperazin-1-yl)-piperidin-1-yl]-1 ,3-dihydro-isoindol-2-yl}-methanone hydrochloride was isolated as a cream solid (62 mg). 1H NMR (DMSO-d6) 12.40-12.00 (2H, br m), 9.75-9.55 (1 H, br m), 7.45-7.05 (3H, m), 7.03 (1 H, s), 6.45 (1 H, s), 4.70-4.55 (4H, m), 3.85-3.65 (6H, m), 3.60-3.40 (5H, m), 3.15- 3.05 (1 H, m), 3.0-2.78 (5H, m), 2.30-2.20 (2H, m), 2.05-1.90 (2H, m), 1.15 (6H, d). MS: [M+H]+ 479. EXAMPLE 52
Synthesis of (2,4-dihvdroxy-5-isopropyl-phenyl)-[5-(4-piperazin-1 -yl-phenyl)-1 ,3-dihvdro-isoindol-2- yli-methanone
Figure imgf000168_0001
52A. Synthesis of 4-f4-[2-(2,4-bis-benzyloxy-5-isopropenyl-benzoyl)-2,3-dihvdro-1 H-isoindol-5-yil- phenvDpiperazine-1-carboxylic acid tert-butyl ester A mixture of (2,4-bis-beπzyloxy-5-isopropenyI-phenyl)-(5-bromo-1 ,3-dihydro-isoindol-2-yl)- methanone (240 mg, 0.43 mmol), t-butyl-4-[4-(4,4,5,5-tetramethyl-1 ,3,2-dioxaborolan-2- yl)phenyl]piperazine carboxylate (210 mg, 1.25 equiv.), bis(tri-t-butylphosphine)palladium(0) (12.5 mg, 2.5 mol%) and potassium carbonate (350 mg, 6 equiv.) in toluene/water/ethanol (1 ml: 1 ml: 4 ml) was heated at 1350C for 30 minutes in a CEM explorer microwave synthesiser. The reaction mixture was diluted with EtOAc, washed with saturated NaHCO3, dried (MgSO4) and evaporated. Purification by flash column chromatography (Biotage SP4 - 25S, 25 ml/min) eluting with 1 :3 then 1 :1 EtOAc / P. E.. Evaporation of product containing fractions gave 85 mg of 4-{4-[2-(2,4-bis- benzyloxy-S-isopropenyl-benzoyO^.S-dihydro-I H-isoindol-δ-yll-phenylJpiperazine-i-carboxylic acid tert-butyl ester. MS: [M+H]+ 736.
52B. Synthesis of (2,4-dihvdroxy-5-isopropyl-phenyl)-F5-(4-piperazin-1-yl-phenyl)-1 ,3-dihvdro- isoindol-2-yli-methanone
Hydrogenation of 4-{4-[2-(2,4-bis-benzyloxy-5-isopropenyI-benzoyl)-2,3-dihydro-1 H-isoindol-5-yl]- phenyl}piperazine-1-carboxylic acid tert-butyl ester (as described in method A5), followed by BOC deprotection (as described in example 70) afforded 10 mg of the title compound as the hydrochloride salt after flash column chromatography (Biotage SP4, 25S) eluting with DMAW 240- 120-90 and evaporation from saturated HCI / EtOAc. 1H NMR (Me-d3-OD) 7.63 (2H, d), 7.55 (2H, m) 7.45-7.30 (1 H, m), 7.25 (1 H, s), 7.20 (2H, d), 5.03 (4H, m), 3.55 (4H, m), 3.47 (4H, m), 3.23 (1 H, m), 1.25 (6H1 d). MS: [M+H]+ 458. EXAMPLE 53
Synthesis of 2,4-dihvdroxy-5-isopropyl-phenyl)-r5-(1-dimethylamino-2-hvdroxy-ethyl)-1 ,3-dihydro- isoindol-2-vH-methanone, and dihvdroxy-5-isopropyl-phenylH5-(2-dimethylamino-1-hvdroxy-ethyl)- 1 ,3-dihvdro-isoindol-2-yl1-methanone
Figure imgf000169_0001
53A. Synthesis of 5-bromo-2,4-dimethoxybenzoic acid methyl ester A solution of 5-bromo-2,4-dihydroxybenzoic acid (24.9 g, 107 mmol) in acetone (355 ml), was treated with methyl iodide (39.9 ml, 640mmol) and K2CO3 (88 g, 640 mmol) then heated at reflux overnight. The salts were filtered off and washed with acetone. The filtrate was evaporated to dryness and the product was purified by flash column chromatography (100% DCM) to yield 5- bromo-2,4-dimethoxybenzoic acid methyl ester as a colourless solid (28 g). 1H NMR (Me-Cf3-OD) 7.98 (1 H, s), 6.74 (1 H, s), 3.99 (3H, s), 3.94 (3H, s), 3.85 (3H, s). MS: [M+H]+ 275/277.
53B. Synthesis of -isopropenyl-2,4-dimethoxy-benzoic acid methyl ester
To potassium isopropylidene trifluoroborate (4.87 g, 32.7 mmol) and 5-bromo-2,4- dimethoxybenzoic acid methyl ester (7.5 g, 27.3 mmol) in THF (195 ml) was added Cs2CO3 (26.6 g, 81.8 mmol) in water (39 ml). The reaction was degassed and Pd(PPh3)4 (1.58 g, 1.36 mmol) added. The reaction was heated at reflux for three days then quenched by adding water and extracted with EtOAc (x2). The combined organic layers were washed with brine, dried (MgSO4), filtered and evaporated to leave an orange solid. The product was taken up in EtOAc again and the precipitate filtered. The filtrate was evaporated to dryness to yield 5-isopropenyl-2,4- dimethoxy-benzoic acid methyl ester (6.2 g). 1H NMR (Me-Cf3-OD) 7.68 (1 H, s), 6.66 (1 H, s), 5.10- 5.08 (1 H, m), 5.02-5.00 (1 H, m), 3.93 (3H, s), 3.92 (3H, s), 3.84 (3H, s), 2.08-2.06 (3H, m). MS: [M+Hf 237.
53C. Synthesis of 5-isopropyl-2,4-dimethoxy-benzoic acid methyl ester
A solution of 5-isopropenyl-2,4-dimethoxy-benzoic acid methyl ester (6.0 g, 25.4 mmol) in MeOH (85 ml) was shaken with 10 % Pd/C under an atmosphere of H2 at room temperature for 3 hours. The catalyst was filtered through GF/A paper but a little fine powder passed through. The filtrate was passed through a small pad of silica and evaporated to dryness to yield a colourless solid. The product was purified by flash column chromatography (DCM:Petrol gradient elution) to yield 5- isopropyl-2,4-dimethoxy-benzoic acid methyl ester a colourless solid (5.5 g). 1H NMR (Me-Cl3-OD) 7.68 (1 H, s), 6.64 (1 H, s), 3.94 (3H, s), 3.91 (3H, s), 3.84 (3H, s), 3.23 (1 H, sept), 1.20 (6H, d). MS: [M+H]+ 239.
53D. Synthesis of 5-isopropyl-2,4-dimethoxy-benzoic acid
5-lsopropyl-2,4-dimethoxy-benzoic acid methyl ester (5.5 g, 23.1 mmol) and NaOH (1.38 g, 34.6 mmol) in THF (46 ml) and water (46 ml) was warmed to 50 0C overnight. The reaction was cooled and diluted with water and EtOAc. The aqueous layer was neutralised with HCI (1 N, aq.). The product was extracted with EtOAc (x3) and the combined organic layers were washed with brine and dried over MgSO4. The product was filtered and evaporated to dryness to yield 5-isopropyl- 2,4-dimethoxy-benzoic acid as a pale peach solid (4.7 g). 1H NMR (DMSO-Cf6) 12.1 (1 H, br s), 7.62 (1 H, s), 6.71 (1 H, s), 3.95 (3H, s), 3.91 (3H, s), 3.19 (1 H, sept), 1.18 (6H, d). MS: [M+H]+ 225.
Figure imgf000171_0001
53E Synthesis of (5-bromo-1 ,3-dihvdro-isoindol-2-yl)-(5-isopropyl-2,4- dimethoxyphenvDmethanone
To a mixture of 5-isopropyl-2,4-dimethoxybenzoic acid (2.45 g, 10.9 mmol), HOBt (1.61 g, 11.9 mmol) and EDC (1.85 g, 11.9 mmol) in anhydrous DMF (33 ml) under N2 was added 5-bromo-2,3- dihydro-1 H-isoindole (1.97 g, 9.95 mmol) and stirred at room temperature overnight. The reaction was quenched by diluting with NaOH (1 M, aq.) and extracting the product with EtOAc (x2). The combined organic layers were washed with brine and dried over MgSO4. The product was filtered and evaporated to dryness to leave a brown oil. The product was purified by flash column chromatography using gradient elution (ether/petrol) to yield (5-bromo-1 ,3-dihydro-isoindol-2-yl)-(5- ιsopropyl-2,4-dimethoxyphenyl)-methanone as a beige solid (3 g). 1H NMR (Me-Cf3-OD) 7.60-7.13 (3H, m), 7.14 (1 H, s), 6.71 (1 H, s), 4.89 (2H, d), 4.64 (2H, d), 3.93 (3H, s), 3.90 (3H, s), 3.27 (1 H, sept), 1.20 (6H, d). MS: [M+H]+ 404/406.
53F. Synthesis of 5-isopropyl-2,4-dimethoxy-phenyl)-(5-vinyl-1 ,3-dihvdro-isoindol-2-yl)-methanone To (5-bromo-1 ,3-dihydro-isoindol-2-yl)-(5-isopropyl-2,4-dimethoxyphenyl)methanone (2.2 g, 5.44 mmol), and 2-vinyl-4,4,5,5-tetramethyl-1 ,3,2-dioxaboralane (1.2 ml, 6 53 mmol) in MeOH (25 ml) and toluene (25 ml) was added Na2CO3 in water (25 ml). The reaction was degassed, Pd(PPh3)4 (0.38 g, 0.05 mmol) added then heated at 800C overnight. The reaction was worked up by adding water and extracting with EtOAc (x3). The combined organic layers were washed with brine and dried over MgSO4. The product was filtered and evaporated to dryness then purified by flash column chromatography, gradient elution (ether: petrol) to yield 5-isopropyl-2,4-dimethoxy-phenyl)- (5-vinyl-1 ,3-dihydro-isoindol-2-yl)-methanone as a yellow oil (1.6 g). 1H NMR (Me-d3-OD) 7.47- 7.15 (3H, m), 7.15 (1 H, s), 6.82-6.72 (1 H, m), 6.71 (1 H, s), 5.79 (1 H, dd), 5.24 (1 H, dd), 4.90 (2H, d), 4.64 (2H, d), 3.93 (3H, s), 3.91 (3H, s), 3.27 (1 H, sept), 1.23 (6H, d). MS: [M+H]+ 352. 53G. Synthesis of (5-isopropyl-2,4-dimethoxy-phenylH5-oxiranyl-1 ,3-dihvdro-isoindol-2-yl)- methaπone
To (5-isopropyl-2,4-dimethoxy-phenyl)-(5-vinyl-1 ,3-dihydro-isoindol-2-yl)-methanone (0.80 g, 2.28 mmol) in DCM (22 ml) was added mCPBA (0.61 g, 2.73 mmol) at 0 0C. The reaction was stirred at room temperature for an hour. The reaction was diluted with NaOH (1M, aq.) and extracted the product with EtOAc. The EtOAc layer was washed with NaOH again. The organic layer was washed with brine and dried over MgSO4. The product was filtered and evaporated to dryness to yield crude (5-isopropyl-2,4-dimethoxy-phenyl)-(5-oxiranyl-1 ,3-dihydro-isoindol-2-yl)-methanone as a very pale yellow oil. MS: [M+H]+ 368.
Figure imgf000172_0001
53H. Synthesis of (2,4-dihvdroxy-5-isopropyl-phenyl)-[5-(1-dimethylamino-2-hvdroxy-ethyl)-1 ,3- dihvdro-isoindol-2-yl]-methanone (Compound 121 H-i) and (2,4-dihydroxy-5-isopropyl-phenyl)-f5- (2-dimethylamino-1-hvdroxy-ethvπ-1 ,3-dihvdro-isoindol-2-vn-methanone(Compound 121 H-H)
(5-lsopropyl-2,4-dimethoxy-phenyl)-(5-oxiranyl-1 ,3-dihydro-isoindol-2-yl)-methanone (~120 mg, crude) was dissolved in dimethylamine in EtOH (20 ml, -33%, 5.6 M) and heated at 60 0C overnight. The reaction was evaporated to dryness and the product crudely purified by flash column chromatography MeOH:DCM (1 :5) to yield impure material which was used without further purification. To a mixture of [5-(1-dimethylamino-2-hydroxy-ethyl)-1 ,3-dihydro-isoindol-2-yl]-(5- isopropyl-2,4-dimethoxy-phenyl)-methanone and [5-(2-dimethylamino-1-hydroxy-ethyl)-1 ,3- dihydro-isoindol-2-yl]-(5-isopropyl-2,4-dimethoxy-phenyl)-methanone (-100 mg) was added DCM (5 ml) and then boron tribromide (3 eq.) under N2. The reaction was left to stir at room temperature until completion. The reaction was quenched with ice and diluted with water and EtOAc. The aqueous layer was extracted with EtOAc (x2). The combined organic layers were washed with brine, dried over MgSO4 then filtered and evaporated to dryness, to leave a yellow residue which was purified by preparative HPLC to yield the two resorcinol isomers. (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(1-dimethylamino-2-hydroxy-ethyl)-1 ,3-dihydro-isoindol-2-yl]- methanone, (Compound 121 H-i) 1H NMR (Me-Cf3-OD) 7.42-7.30 (3H, m), 7.19 (1 H, s), 6.39 (1 H, s), 4.98-4.87 (4H, m), 4.03-3.97 (1 H, m), 3.94-3.86 (1 H, m), 3.68 (1 H, br s), 3.22 (1 H, sept), 2.40 (6H, s), 1.23 (6H, d). MS: [M+H]+ 384. (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-1-hydroxy-ethyl)-1 ,3-dihydro-isoindol-2-yl]- methanone, (Compound 121 H-ii) 1H NMR (Me-Cf3-OD) 7.39-7.25 (3H, m), 7.18 (1 H, s), 6.38 (1 H, s), 6.94-6.88 (5H, m), 3.22 (1 H, sept), 2.77-2.68 (1 H, m), 2.61-2.51 (1 H, m), 2.42 (6H, s), 1.23 (6H, d). MS: [M+H]+ 384.
EXAMPLE 54 Synthesis of (2,4-dihvdroxy-5-isoi3ropyl-phenyl)-r5-(piperazine-1-carbonyl)-1 ,3-dihvdro-isoindol-2- yli-methanone hydrochloride
Figure imgf000173_0001
54A. Synthesis of 4-[2-(2,4-dihvdroxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5-carbonvn- piperazine-1-carboxylic acid tert-butyl ester
A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid (Preparation D6) (0.5 g, 0.96 mmol ), EDC (0.22 g, 1.15 mmol), HOBT (0.196 g, 1.15 mmol) and BOC piperazine ( 0.117 ml, 1.06 mmol) in DMF (10 ml) was stirred at room temperature for 48 hours, then evaporated under vacuum. The crude material was dissolved in ethyl acetate and extracted twice with saturated NaHCO3, organics washed with brine, dried (MgSO4), filtered then evaporated under vacuum and purified by flash column chromatography (80% EtOAc- P. E. as eluant) to give 0.5 g of 4-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5- carbonyl]-piperazine-1-carboxylic acid tert-butyl ester. MS: [M+H] + 688.
54B. Synthesis of (2.4-dihvdroxy-5-isopropyl-phenvπ-[5-(piperazine-1-carbonyl)-1.3-dihydro- isoindol-2-yli-methanone hydrochloride
Hydrogenation as Method A5 to give (0.2 g, 0.30 mmol) 4-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)- 2,3-dihydro-1 H-isoindole-5-carbonyl]-piperazine-1-carboxylic acid tert-butyl ester [used crude] dissolved in EtOAc then treated with saturated EtOAc/HCI, stirred at ambient for 3 hours, reaction diluted with ether, solid filtered to give 0.19 g of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(piperazine- 1-carbonyl)-1 ,3-dihydro-isoindol-2-yl]-methanone hydrochloride. 1H NMR (Me-d3-OD) 7.50-7.42 (3H, m), 7.18 (1 H, s), 6.39 (1 H, s), 5.00-4.95 (4H, br s), 3.92-3.79 (4H, br s), 3.35-3.28 (4H, br s), 3.26-3.15 (1 H, m), 1.23 (6H, d). MS: [M+H] + 410. EXAMPLE 55
Synthesis of (2,4-dihvdroxy-5-isopropyl-phenvn-f5-(4-methy]-piperazin-1-ylmethyl)-1 ,3-dihvdro- isoindol-2-yli-methanone
Figure imgf000174_0001
55A. Synthesis of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methoxy-methyl-amide
A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid (Preparation D6) (1.76 g, 3.39 mmol ), EDC (0.78 g, 4.06 mmol), HOBT (0.55 g, 4.06 mmol), Et3N ( 1 ml, 6.78mmol) and N,O-dimethylhydroxylamine hydrochloride (0.36 g, 3.72 mmol) in DMF (20 ml) was stirred at room temperature for 48 hours, then evaporated under vacuum. The crude material was dissolved in ethyl acetate and extracted twice with saturated NaHCO3, organics washed with brine, dried (MgSO4), filtered then evaporated to give 1.84 g of 2-(2,4-bis-benzyloxy- 5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methoxy-methyl-amide. MS: [M+H] + 563. 55B. Synthesis of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihvdro-1 H-isoindole-5- carbaldehyde
A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5-carboxylic acid methoxy-methyl-amide (0.226g, 0.4mmol) in THF (5ml) cooled to 00C , treated with 1M LiAIH/THF (0.3ml, 0.3mmol), stirred 1 hour, further LiAIH4 (0.05ml) added then stirred for 30 minutes. The reaction was quenched with saturated KHSO4 solution, extracted with EtOAc, dried (MgSO4), filtered and evaporated to give 0.2g of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H- isoindole-5-carbaldehyde. MS: [M+H] + 504.
55C. Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3- dihvdro-isoindol-2-yli-methanone To a solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1 H-isoindole-5- carbaldehyde (0.316 g, 0.63 mmol) and n-methyl piperazine (63 mg, 0.63 mmol) in CH2CI2 (10 ml) was added AcOH (38 mgs 0.63 mmol) and NaBH(OAc)3 (0.28 g, 1.33 mmol), then stirred at ambient for 5 hours. The reaction was quenched with water, layers separated and aqueous washed CH2CI2. The organics were combined, washed with brine, dried (MgSO4), filtered and evaporated to give 0.32 g of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1- ylmethyl)-1 ,3-dihydro-isoindol-2-yl]-methanone. MS: [M+H]+ 588. 55D, Synthesis of (2,4-dihvdroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3- dihydro-isoindol-2-vπ-methanone
Hydrogenation was carried out using Method A5 but with the addition of K2CO3 (2 equiv.) in a MeOH/H2O [9.1]. After evaporation of methanol the reaction was diluted with water, neutralised using 1M HCI and extracted with CH2CI2 (x2). Organics dried (MgSO4), filtered and evaporated under vacuum then purified by preparative HPLC to give 21 mg of (2,4-dihydroxy-5-isopropyl- phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihydro-isoindol-2-yl]-methanone. MS: [M+H] + 410. 1H NMR (Me-ds-OD) 7.37-7.23 (3H, br s), 7.19 (1 H, s), 6.39 (1 H, s), 4.94-4.87 (4H, br s), 3.57 (2H, s), 3.27-3.16 (1 H, m), 2.67-2.39 (8H1 m), 2.31 (3H, s), 1.23 (6H, d). EXAMPLE 56
Synthesis of (2.4-dihvdroxy-5-isopropyl-phenyl)-f4-(3-morpholin-4-yl-propoxy)-1 ,3-dihvdro-isoindol- 2-yl1methanone
56A. Synthesis of 4-hydroxyisoindoline hvdrobromide
J. Chem
Figure imgf000175_0001
A suspension of dimethyl 3-methoxyphthalate (69.45 g, 0.31 mol) [prepared as per J. Chem. Soc, Perkin Trans. 1, 1989, 391] in water (300 ml) was treated with potassium hydroxide (43.7 g, 0.78 mol) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the methanol liberated during the course of the reaction was removed in vacuo, the mixture acidified to pH 2 or below by the addition of 5M hydrochloric acid and evaporated gently in vacuo to induce crystallization. The solid material was filtered off, washed with a little ice cooled water, sucked dry under reduced pressure and dried in a vacuum oven at 5O0C overnight to afford 3- methoxyphthalic acid (51.0 g, 84%) as a colourless solid. 1H NMR (DMSO-d6) 13.05 (2H, br s), 7.48 (2H, m), 7.33 (1 H, m), 3.82 (3H, s). MS: [M+H]+ 197.
Acetic anhydride (70 ml) was added to a mixture of 3-methoxyphthalic acid (51.0 g, 0.26 mol) in anhydrous tetrahydrofuran (250 ml) and the mixture was strirred and held at reflux for 4 hours. Upon cooling to room temperature the solvent was removed in vacuo and the resulting solid material was dried in a vacuum oven at 5O0C overnight to afford 3-methoxyphthalic anhydride (45.9 g, 99%) as a colourless solid. 1H NMR (DMSO-d6) 7.97 (1 H, dd), 7.63 (1 H, d), 7.60 (1 H, d), 4.02 (3H, s). MS: [M+H]+ 179. A mixture of 3-methoxyphthalic anhydride (24.0 g, 134.8 mmol) and formamide (120 ml) was stirred and held at 21O0C for 5 hours and was then allowed to cool to room temperature overnight. Water (100 ml) was added and the solid material filtered off under reduced pressure. The crude product was washed sequentially with 50% aqueous acetone (50 ml) and diethyl ether (200 ml) and sucked dry under reduced pressure to afford 3-methoxyphthalimide (8.95 g, 37%) as an off- white solid. 1H NMR (DMSO-d6) 11.08 (1 H, br s), 7.78 (1 H, dd), 7.45 (1 H, d), 7.36 (1 H1 d), 3.93 (3H, s). MS: [M+H]+ 178.
A stirred solution of 3-methoxyphthalimide (8.95 g, 50.56 mmol) in anhydrous tetrahydrofuran (200 ml) at 00C was treated dropwise with a solution of borane in tetrahydrofuran (1 M, 150 ml, 0.15 mol) and the resulting mixture was stirred and held at reflux for 16 hours. The mixture was cooled to O0C, methanol (60 ml) was added dropwise followed by 5M hydrochloric acid (60 ml) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the organic solvent was removed in vacuo and the mixture diluted with water (250 ml) and extracted with dichloromethane (3 x 250 ml). The aqueous layer was basified to pH 12 or above by the addition of 5M sodium hydroxide, extracted with dichloromethane (3 x 250 ml) and the combined extracts were evaporated to dryness in vacuo to afford 4-methoxyisoindoline (4.44 g, 59%) as a green oil which was used without further purification. 1H NMR (DMSOd6) 7.18 (1 H, t), 6.83 (1 H1 d), 6.78 (1 H, d), 4.07 (2H1 s), 4.02 (2H, s), 3.78 (3H, s). MS: [M+H]+ 150.
4-Methoxyisoindoline (4.4 g, 29.53 mmol) in 48% aqueous hydrobromic acid (50 ml) was stirred and held at reflux for 16 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford 4-hydroxyisoindoline hydrobromide (5.0 g, 78%) as a pale orange solid. 1H NMR (DMSOd6) 9.95 (1 H, br s), 9.37 (2H, br s), 7.19 (1 H, t), 6.84 (1H1 d), 6.80 (1 H, d), 4.48 (2H1 1), 4.40 (2H1 1). MS: [MH-H]+ 136.
56B. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenylH4-hvdroxy-1 ,3-dihvdro-isoindol-2-vD- methanone
Figure imgf000176_0001
A mixture of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (8.1 g, 21.65 mmol), A- hydroxyisoindoline hydrobromide (4.91 g, 22.73 mmol), N-ethyl-N'-(3- dimethylaminopropyl)carbodiimide hydrochloride (5.0 g, 25.98 mmol), 1 -hydroxy benzotriazole (3.5 g, 25.98 mmol) and triethylamine (6 ml, 43.3 mmol) in N,N-dimethylformamide (50 ml) was stirred at room temperature for 16 hours. The solvent was removed in vacuo and the residue was treated with a saturated aqueous solution of sodium hydrogen carbonate (200 ml). The mixture was filtered, the solid material was washed copiously with water, sucked dry under reduced pressure and dried in a vacuum oven at 5O0C overnight to afford (2,4-bis-benzyloxy-5-isopropenyl-phenyl)- (4-hydroxy-1 ,3-dihydro-isoindol-2-yl)-methanone (10.25 g, 96%) as a pale tan solid. 1H NMR (DMSOd6) (mixture of amide rotamers) 9.68 and 9.60 (1 H, 2 x br s), 7.45-7.25 (1OH, m), 7.20-7.00 (3H, m), 6.82 and 6.72 (1 H, 2 x d), 6.68 (1 H, m), 5.23 and 5.22 (2H, 2 x s), 5.18 (2H, s), 5.11 (1 H, s), 5.09 (1 H, s), 4.77 and 6.67 (2H, 2 x s), 4.53 and 4.44 (2H, 2 x s), 2.04 (3H, s). MS: [M+H]+ 492.
56C. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenylH4-(3-morpholin-4-yl-propoxy)-1 ,3- dihvdro-isoindol-2-vπ-methanone
A mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(4-hydroxy-1 ,3-dihydro-isoindol-2-yl)- methanone (2g; 4.07 mmol), 4-(3-chloropropyl)morpholine (1.66 g; 2.5 equiv.) and caesium carbonate (8.3 g; 6.25 equiv) in DMF was heated at 900C overnight then evaporated. The residue was dissolved in EtOAc, washed with brine, dried (MgSO4) and evaporated. Purification of the crude material using a Biotage SP4 (4OS, 40 ml/min), using gradient elution form 0% to 10% MeOH / EtOAc gavelδ g of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl- propoxy)-1 ,3-dihydro-isoindol-2-yl]-methanone as a pale yellow gum. MS: [M+H]+ 619. 56D. Synthesis of (2,4-dihvdroxy-5-isopropyl-phenylH4-(3-morpholin-4-yl-propoxy)-1 ,3-dihvdro- isoindol-2-vπmethanone
Figure imgf000177_0001
Hydrogenation of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1 ,3- dihydro-isoindol-2-yl]-methanone (as described in method A5) followed by treatment with saturated HCI/EtOAc and trituration with hot acetone afforded 890 mg of the title compound (hydrochloride salt) as a cream solid. 1H NMR (DMSO-d6) 10.78 (1 H, br s), 10.05 (1 H, br s), 9.55 (1 H, br s), 7.30 (1 H, t), 7.08 (1 H, s) 6.98-6.90 (2H, m), 6.45 (1 H, s), 4.80 (2H, S), 4.75 (2H, s), 4.15 (2H, t), 3.95 (2H, br m), 2.80 (2H, br m), 3.50-3.35 (2H, br m), 3.25 (2H, br m,), 3.18-3.02 (3H, br m), 2.20 (2H, br m), 1.15 (6H, d). MS: [M+H]+ 441. EXAMPLES 57 TO 74
By following the methods described above, the following compounds were prepared.
Figure imgf000178_0001
Figure imgf000179_0001
Figure imgf000180_0001
Figure imgf000181_0001
Figure imgf000182_0001
Figure imgf000182_0002
Figure imgf000183_0002
EXAMPLE 75 (5-Chloro-2,4-dihvdroxy-phenyl)-r5-(4-methyl-piperazin-1-vπ-1 ,3-dihydro-isoindol-2-yll-methanone
75A. 5-(4-Methyl-piperazin-1-yl)-1 ,3-dihvdro-isoindole-2-carboxylic acid tert-butyl ester
Figure imgf000183_0001
5-Bromo-1 ,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester (2.97 g, 10 mmol) was azeotropically dried by evaporation from toluene. Tris(dibenzylideneacetone)dipalladium (0) (228 mg, 0.25 mmol), 2-(di-tert-butylphosphino)biphenyl (149 mg, 0.50 mmol) and sodium tert-butoxide (1.34 g, 13.9 mmol) were added and the flask was purged with nitrogen. Toluene (25 mL) then N-methylpiperazine (1.33 mL, 12 mmol) were added and the mixture was heated to 80°C for 2 hours. After allowing to cool to r.t. the mixture was diluted with ether, filtered through Celite and concentrated to give a residue that was purified by flash chromatography on silica (2M methanolic ammonia/dichloromethane, 1 % to 3% gradient). This afforded the title compound as a brown solid (1.45g, 46%). 1H NMR (MeOH-Cl4) 7.15 (1 H, m), 6.94-6.88 (2H, m), 4.60-4.54 (4H, m), 3.20-3.17 (4H, m), 2.63-2.60 (4H, m), 2.34 (3H, s), 1.52 (9H, s). MS: [M+H]+ 318.
75B. 5-(4-Methyl-piperazin-1-yl)-2,3-dihvdro-1H-isoindole dihvdrochloride
Figure imgf000184_0001
5-(4-Methyl-piperazin-1-yl)-1 ,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester (247 mg, 0.78 mmol) was treated with 4M HCI in dioxane (4 mL, 4 mmol) for 24 hours. Concentration in vacuo afforded the title compound quantitatively, which was used directly in the coupling reaction. 1H NMR (DMSO-d6) 11.13 (1 H, br.s), 9.99 (2H, br.s), 7.27 (1H, d), 7.02-7.00 (2H, m), 4.43-4.37 (4H, m), 3.82-3.75 (2H, m), 3.49-3.43 (2H, m), 3.15-3.10 (4H, m), 2.79-2.78 (3H, s), 1.52 (9H, s). MS: [M+H]+ 218.
75C. (5-Chloro-2,4-dihvdroxy-phenylH5-(4-methyl-piperazin-1-yl)-1 ,3-dihvdro- isoindol-2-vπ-methanone
Figure imgf000184_0002
A solution of 5-chloro-2,4-dihydroxy-benzoic acid (176 mg, 0.93 mmol) in DMF (5 mL) was treated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (179 mg, 0.93 mmol) then HOBt (126 mg, 0.93 mmol). After 45 min, the solution of the activated acid was added to a mixture 5-(4- methyl-piperazin-1-yl)-2,3-dihydro-1 H-isoindole dhihydrochloride (290 mg, 0.78 mmol) and triethylamine (0.28 mL, 2 mmol) then the mixture was stirred at r.t. for 3 h. The solvent was removed in vacuo then the residue was partitioned between ethyl acetate and water (*3). Each extract was washed with saturated sodium bicarbonate solution and brine then dried (MgSO4), combined and concentrated. Some insoluble material remained and this was dissolved in 1 N hydrochloric acid and methanol then combined with the organic extracts. The pH was adjusted to 14 with solid sodium hydroxide and the mixture allowed to stand overnight. The pH was adjusted to 7 with 1 N hydrochloric acid and the resulting precipitate was filtered off then subjected to purification by preparative HPLC to afford the title compound as a red solid. This was converted to its hydrochloride salt by treatment with 4M HCI in dioxane, concentration in vacuo and trituration with ether which gave a brown solid (91 mg, 27%). 1H NMR (DMSO-d6) 1 1.10 (1 H, br.s), 10.50 (1 H, br.s), 7.26-7.15 (2H, m), 7.02-6.93 (2H, m), 6.69 (1 H, s), 4.72-4.61 (4H, m), 3.78-3.72 (2H, m), 3.45 (2H, br.s), 3.12 (4H, br.s), 2.78 (3H, s). MS: [M+H]+ 386/388.
EXAMPLE 76
(2,4-dihvdroxy-5-isopropyl-phenyl)-f5-(4-methyl-piperazin-1-yl)-1 ,3-di-hvdro-isoindol-2-vn- methanone
76A. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenvπ-(5-bromo-1 ,3-dihydro-isoindol-2-yl)- methanone
Figure imgf000185_0001
Coupling of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (5.0 g, 13.4 mmol) (Preparation B9) and 5-bromo-2,3-dihydro-1 H-isoindole (Preparation C20) was completed according to method A4, using CH2CI2 as the reaction solvent to give the title compound (8.34 g) as a beige solid.
76B. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-r5-(4-methyl-piperazin-1-vO-1 ,3- dihydro-isoindol-2-yll-methanoπe
Figure imgf000185_0002
To a mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1 ,3-dihydro-isoindol-2-yl)- methanone (8.30 g, 15.0 mmol), 2-(di-t-butylphosphino)biphenyl (223 mg, 0.75 mmol), tris(dibenzylideneacetone)dipalladium (344 mg, 0.38 mmol), sodium tert-butoxide (2.17 g, 22.5 mmol) and 1-methyl-piperazine (2.16 ml_, 19.5 mmol) under a N2 atmosphere was added anhydrous toluene (100 ml_). The mixture was taken to 80 0C and heated at this temperature for 16 h. The mixture was allowed to cool to ambient temperature, diluted with ether (150 mL) and filtered through a plug of Celite, washing with ether. The filtrate was reduced in vacuo and the residue purified by column chromatography using an eluant of CH2CI2-DMAWI 20 (1 :0 - 0:1) to give the title compound (9.39 g) as a red gum.
76C. (2,4-dihvdroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-vπ-1 ,3-di-hvdro-isoindol-2-yl1- methanone
Figure imgf000186_0001
A mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-methyl-piperazin-1 -yl)-1 ,3-dihydro- isoindol-2-yl]-methanone (8.61 g, 15.0 mmol) and 10% Pd/C (1.0 g) in methanol (200 mL) was stirred vigorously under a hydrogen atmosphere (~1 atm) for 18 h at ambient temperature. The mixture was filtered through a plug of Celite and reduced in vacuo to give a purple oil. This residue was purified by column chromatography using an eluant of DMAW120 to give the title compound as its acetate salt. This salt was taken up in MeOH (30 mL) and to the solution was added saturated HCI in EtOAc (20 mL). This mixture was stirred at ambient for 2 h and the solid formed collected by filtration and dried in vacuo to give the title compound as its hydrochloride salt (2.64 g) as a white solid.
EXAMPLE 77 (5-Chloro-2,4-dihvdroxy-phenyl)-[5-(4-methyl-piperazin-1-vn-1 ,3-dihydro-isoindol-2-vn-methanone
Figure imgf000186_0002
A solution of 5-chloro-2,4-dihydroxy-benzoic acid (176 mg, 0.93 mmol) in DMF (5 mL) was treated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (179 mg, 0.93 mmol) then HOBt (126 mg, 0.93 mmol). After 45 minutes, the solution of the activated acid was added to a mixture 5- (4-methyl-piperazin-1-yl)-2,3-dihydro-1 H-isoindole dhihydrochloride (290 mg, 0.78 mmol) and triethylamine (0.28 mL, 2 mmol) then the mixture was stirred at r.t. for 3 h. The solvent was removed in vacuo then the residue was partitioned between ethyl acetate and water (*3). Each extract was washed with saturated sodium bicarbonate solution and brine then dried (MgSO4), combined and concentrated. Some insoluble material remained and this was dissolved in 1 N hydrochloric acid and methanol then combined with the organic extracts. The pH was adjusted to 14 with solid sodium hydroxide and the mixture allowed to stand overnight. The pH was adjusted to 7 with 1 N hydrochloric acid and the resulting precipitate was filtered off then subjected to purification by preparative HPLC to afford the title compound as a red solid. This was converted to its hydrochloride salt by treatment with 4M HCI in dioxane, concentration in vacuo and trituration with ether which gave a brown solid (91 mg, 27%). 1H NMR (DMSO-Ci6) 11.10 (1 H, br.s), 10.50 (1 H, br.s), 7.26-7.15 (2H, m), 7.02-6.93 (2H, m), 6.69 (1 H, s), 4.72-4.61 (4H, m), 3.78-3.72 (2H, m), 3.45 (2H, br.s), 3.12 (4H, br.s), 2.78 (3H, s). MS: [M+H]+ 386/388.
EXAMPLE 78
Alternative Synthesis of (2,4-Dihvdroxy-5-isopropyl-phenylH5-(4-hydroxy-1 -methyl-piperidin-4-vD- 1 ,3-dihvdro-isoindol-2-vπ-methanone
78A. 5-bromo-2-trityl-2,3-dihvdro-1 H-isoindole
Figure imgf000187_0001
Trityl chloride (2.30 g, 8.23 mmol) was added to a solution of 5-bromo-2,3-dihydro-1 H-isoindole (1.64 g, 8.23 mmol) and triethylamine (1.4 mL, 9.9 mmol) in dichloromethane (20 mL). After 18 hours the solvent was removed in vacuo, the residue taken up in ethyl acetate and washed with water (*2) and brine, dried (MgSO4) and concentrated. The crude material was purified by flash chromatography on silica eluting with 1 % triethylamine/10% ethyl acetate/petrol to give 5-bromo-2- trityl-2,3-dihydro-1 H-isoindole as a reddish-brown solid (3.10 g, 85%). 1H NMR (CDCI3) 7.91-7.84 (1 H, m), 7.57 (6H, d), 7.45-7.41 (1 H, m), 7.33-7.14 (9H, m), 6.95 (1 H, d), 3.90 (2H, s), 3.86 (2H, s). MS: Ph3C+ 243.
78B. 1 -methyl-4-(2-trityl-2,3-dihydro-1 H-isoindol-5-yl)-piperidin-4-ol
Figure imgf000187_0002
Under nitrogen, a solution of 5-bromo-2-trityl-2,3-dihydro-1 H-isoindole (2.03 g, 4.6 mmol) in THF (20 mL) was cooled to -780C. n-Butyllithium solution (2.5M in hexanes, 2.0 mL, 5 mmol) was added over 5 minutes, then after 10 minutes, 1-methyl-4-piperidone was added dropwise. After a further hour, the cooling bath was removed and the reaction quenched with sodium bicarbonate solution. The mixture was extracted with ethyl acetate then the organic phase was washed with brine, dried (MgSO4) and concentrated. The residue was purified by flash chromatography on silica (gradient elution with 2M methanolic ammonia/dichloromethane, 0% to 5%) to afford 1- methyl-4-(2-trityl-2,3-dihydro-1 H-isoindol-5-yl)-piperidin-4-ol as a pink foam (1.25 g, 57%). 1H NMR (MeOH-d4) 7.56 (6H, dd), 7.28 (6H, t), 7.25-7.21 (2H, m), 7.15 (3H, t), 7.03 (1 H, d), 3.92 (2H, s), 3.91 (2H, s), 2.70 (2H, d), 2.53 (2H, td), 2.33 (3H, s), 2.06 (2H, td), 1.70 (2H, d). MS: [M+H]+ 475.
78C. 4-(2,3-Dihvdro-1 H-isoindol-5-vD-1 -methyl-piperidin-4-ol dihydrochloride
Figure imgf000188_0001
A mixture of 1-methyl-4-(2-trityl-2,3-dihydro-1 H-isoindol-5-yl)-piperidin-4-ol (1.42 g, 3.0 mmol), 5N hydrochloric acid (5 ml_) and methanol (10 ml_) was placed under nitrogen then heated to reflux for 80 minutes. After cooling, the mixture was concentrated in vacuo to remove methanol, diluted with water and washed with ethyl acetate (χ2). The aqueous phase was concentrated to dryness to afford the title compound in quantitative yield as a black solid. 1H NMR (MeOH-d4) 7.62 (1 H, s), 7.57 (1 H, d), 7.45 (1 H, d), 4.64 (2H, s), 4.63 (2H, s), 3.49-3.46 (4H, m), 2.95 (3H, s), 2.40-2.32 (2H, m), 1.97 (2H, dd).
78D. (2,4-Bis-ben2yloxy-5-isopropenyl-phenvπ-r5-(4-hvdroxy-1-methyl-piperidin-4-ylV1.3-dihvdro- isoindol-2-vπ-methanone
Figure imgf000188_0002
2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid (1.65 g, 4.4 mmol), 1-[3-(dimethylamino)propyl)]-3- ethylcarbodiimide hydrochloride (843 mg, 4.4 mmol) and 1-hydroxybenztriazole (595 mg, 4.4 mmol) were dissolved in DMF (20 mi_). After 35 minutes, the solution was added to a suspension of 4-(2,3-dihydro-1 H-isoindol-5-yl)-1-methyl-piperidin-4-ol dihydrochloride (1.22 g, 4.0 mmol) in DMF (5 mL) and triethylamine (1.4 mL, 10 mmol). The mixture was stirred for 3 hours then concentrated in vacuo. The residue was taken up in ethyl acetate and washed with a mixture of water (adjusted to pH 14 with 2N sodium hydroxide solution) and brine. The aqueous phase was extracted twice further with ethyl acetate then the combined organic extracts were washed with sodium bicarbonate solution and brine, dried (MgSCU) and concentrated. The crude product was purified by flash chromatography (gradient elution with 2M methanolic ammonia/dichloromethane, 2% to 10%) to afford the title compound as a brown foam (1.62 g, 69%). 1H NMR (methanol-d4) 7.51-7.14 (14H, m), 6.85 (0.5H, s), 6.84 (0.5H, s), 5.16 (2H, s), 5.15 (2H, s), 5.10-5.08 (1 H, m), 5.07-5.05 (1 H, m), 4.87 (1 H, s), 4.86 (1 H, s), 4.61 (2H, br.s), 2.78-2.70 (2H, m), 2.57 (1 H, td), 2.54 (1 H, td), 2.36 (1.5H, s), 2.34 (1.5H, s), 2.16-2.05 (5H, m including 2.09 (3H, s)), 1.78-1.70 (2H, m). MS: [M+Hf 589.
78E. (2,4-Dihvdroxy-5'-isopropyl-phenyl)-f5-(4-hvdroxy-1-methyl-piperidin-4-yl)-1 ,3-dihvdro- isoindol-2-vπ-methanone
Figure imgf000189_0001
(2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-f5-(4-hydroxy-1-methyI-piperidin-4-yl)-1 ,3-dihydro- isoindol-2-yl]-methanone (Example 50F) (1.62 g, 2.75 mmol) was dissolved in methanol (50 mL) and hydrogenated at 500C over 10% palladium on charcoal using an H-cube hydrogenation apparatus, under free hydrogen conditions. Concentration afforded the title compound (1.14 g, 100%) as a yellow solid, the NMR and mass spectrometric data of which were as set out in Example 5OE.
EXAMPLE 79
(2.4-Dihvdroxy-5-isopropyl-phenvπ-[5-(2-dimethylamino-ethoxy)-7-methyl-1 ,3-dihvdro-isoindol-2- yll-methanone
79A. 7-Methyl-2,3-dihvdro-1 H-isoindol-5-ol hydrobromide
Figure imgf000189_0002
Using the method of preparation C2, 5-methoxy-3-methyl-phthalic acid dimethyl ester (prepared according to Tarn and Coles, Synthesis 1988, 383) was hydrolysed to 5-methoxy-3-methyl-phthalic acid. 1H NMR (DMSOd6) 12.95 (2H, br.s), 7.15 (1 H, d), 7.04 (1 H, d), 3.80 (3H, s), 2.29 (3H, s). MS: [M-H]+ 209.
5-Methoxy-3-methyl-phthalic acid was converted to 5-methoxy-3-methyl-phthalic anhydride. 1H NMR (DMSOd6) 7.40 (1H, d), 7.34-7.33 (1H, m), 3.94 (3H, s), 2.58 (3H, s).
5-Methoxy-3-methyl-phthalic anhydride was used to prepare 6-methoxy-4-methyl-isoindole-1 ,3- dione. 1H NMR (DMSOd6) 11.05 (1H, br.s), 7.13 (1H, d), 7.10 (1H, d), 3.88 (3H, s), 2.55 (3H, s).
Reduction of 6-methoxy-4-methyl-isoindole-1,3-dione according to the method of preparation C2 afforded 6-methoxy-4-methyl-isoindole. 1H NMR (DMSOd6) 6.64 (1 H, s), 6.57 (1 H, s), 4.05 (2H, s), 3.96 (2H, s), 3.70 (3H, s), 2.16 (3H, s). MS: [M+H]+ 164.
6-Methoxy-4-methyl-isoindole was demethylated to give the title compound as its hydrobromide salt. 1H NMR (DMSOd6) 9.52 (1H, br.s), 9.29 (2H, br.s), 6.59 (1 H, S)1 6.56 (1 H, s), 4.41 (2H, t), 4.34 (2H1 1), 2.17 (3H1 s). 7913, (2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-(5-hvdroxy-7-methyl-1 ,3-dihydro-isoindol-2-yl)- methanone
Figure imgf000190_0001
2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid (248 mg, 0.66 mmol), 1-[3-(dimethylamino)propyl)]- 3-ethylcarbodiimide hydrochloride (127 mg, 0.66 mmol) and 1-hydroxybenztriazole (89 mg, 0.66 mmol) were dissolved in DMF (5 mL). After 20 minutes, 7-methyl-2,3-dihydro-1 H-isoindol-5-ol hydrobromide (152 mg, 0.66 mmol) and triethylamine (0.14 mL, 0.99 mmol) were added. After a further 3.5 hours the mixture was concentrated in vacuo and the residue was treated with 1 N hydrochloric acid and ethyl acetate. The aqueous phase was removed, brine was added and the title compound was collected by filtration as a grey solid (168 mg, 57%). 1H NMR (DMSO-d6) 9.30 (0.47H, s), 9.24 (0.53H, s), 7.48-7.25 (10H, m), 7.09 (0.47H, s), 7.08 (0.53H, s), 6.99 (0.47H, s), 6.98 (0.53H, S), 6.56 (0.47H, s), 6.50 (0.53H, s), 6.48 (0.47H, S)1 6.44 (0.53H, s), 5.24 (0.47H, s), 5.22 (0.53, s), 5.18 (2H, s), 5.10-5.07 (2H, m), 4.70 (0.47H, s), 4.61 (0.53H, s), 4.46 (0.47H, s), 4.36 (0.53H, s), 2.17 (1.41 H1 s), 2.04 (3H, s), 1.99 (1.59H, s). MS: [M+H]+ 506. 79B. (2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-r5-(2-dimethylamino-ethoxy)-7-methyl-1 ,3-dihydro- isoindol-2-vπ-methanone
Figure imgf000190_0002
A mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-hydroxy-7-methyl-1 ,3-dihydro-isoindol-2- yl)-methanone (164 mg, 0.32 mmol), potassium carbonate (112 mg, 0.81 mmol) and 2- (dimethylamino)ethyl chloride hydrochloride (93 mg, 0.64 mmol) in DMF (5 mL) was heated at 6O0C for 17 hours then 9O0C for 6 hours. Further portions of potassium carbonate (112 mg, 0.81 mmol) and 2-(dimethylamino)ethyl chloride hydrochloride (93 mg, 0.64 mmol) were added and the mixture maintained at 6O0C for 72 hours and finally, a further 24 hours at 900C. The mixture was concentrated in vacuo then the residue was partitioned between ethyl acetate and 0.5N aqueous sodium hydroxide. The organic phase was washed with brine (*2), dried (MgSO4) and concentrated to give a residue which was purified by preparative HPLC (acidic method) to afford the title compound as a formate salt (37 mg, 20%). 1H NMR (MeOH-d4) 8.51 (1 H, br.s), 7.43-7.27 (7H, m), 7.24-7.20 (3H, m), 7.17 (0.5H, s), 7.16 (0.5H, s), 6.85 (0.5H, s), 6.84 (0.5H, s), 6.81 (0.5H, s), 6.77 (0.5H, s), 6.74 (0.5H, s), 6.62 (0.5H, s), 5.16 (1 H, s), 5.14 (3H, s), 5.09 (1 H, m), 5.06 (1 H, m), 4.83 (1H, s), 4.74 (1 H, s), 4.60 (1 H, s), 4.48 (1 H, s), 4.28 (1 H, t), 4.23 (1 H, t), 3.41 (1 H, t), 3.37 (1 H, t), 2.84 (3H, s), 2.81 (3H, s), 2.27 (1.5H, s), 2.09 (3H, s), 2.07 (1.5H, s). MS: [M+H]+ 577.
79C. (2,4-Dihvdroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-ethoxy)-7-methyl-1 ,3-dihydro- isoindol-2-yll-nnethanone
Figure imgf000191_0001
(2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-[5-(2-dimethylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]- methanone (37 mg, 0.06 mmol) was hydrogenated in methanol at 50°C over 10% palladium on charcoal using an H-cube hydrogenation apparatus, under free hydrogen conditions. The product was purified by preparative HPLC (basic method) to give the title compound as an off-white solid (9 mg, 35%). 1H NMR (MeOH-d4) 7.18 (1H, s), 6.77-6.65 (2H, br.m), 6.37 (1H, s), 4.85 (water obscuring CH2), 4.77 (2H, s), 4.08 (2H, t), 3.20 (1 H, sept), 2.81 (2H, t), 2.39 (6H, s), 2.22 (3H, br.s), 1.21 (6H, d). MS: [M+Hf 399.
EXAMPLE 80
(2,4-Dihvdroxy-5-isopropyl-phenyl)-r5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihydro-isoindol-2-vn- methanone
Step 1
4-Acetoxy-2-hvdroxy-benzoic acid methyl ester
Figure imgf000191_0002
Resorcinol methyl ester (50 g, 0.298 mol) and N,N-dimethyl-4-aminopyridine (0.27 g, 0.0022 mol, 0.74 mol%) were added to toluene 0.2 L followed by acetic anhydride (30 mL, 0.318 mol). The solution was heated to 50 0C for 2h. The solvent was removed by evaporation at 50 0C to a small volume and the residue was azeotroped once with toluene. To the residual oil was immediately added toluene (100 mL) whilst still warm and the solution used for Step 2 without further purification.
Step 2
5-Acetyl-2,4-dihydroxy-benzoic acid methyl ester
Figure imgf000192_0001
The toluene solution from Step 1 was cooled in an ice bath under N2 and triflic acid (26 mL) added slowly over 30 min. On stirring a fine white solid was formed which dissolved on stirring for 16 h at RT to give a yellow solution. To the solution was added acetyl chloride (2 mL) and the solution stirred at RT for a further 1 h. This solution was cannulated into a stirred cooled (O0C) solution of EtOAc (600 mL) and NaOAc.3H2O (40 g) dissolved in water (400 mL). The organic phase was washed with water (twice, 200 mL), saturated brine and was evaporated to a small volume without drying. The residue was azeotroped with heptane (twice, 100 mL) and heptane (100 mL) was added and the crystalline solid removed by filtration, washed well on sinter with heptane and dried to give 49.5 g (79%). Final purification of combined batches
The combined batches of solid (96.3g ) was heated to boiling with 10% IPA/heptane (250 mL) then cooled to RT and finally to O0C, filtered and the residue dried 72h (oil pump) to give (88.04 g, 91.5%), pure by hplc, tic and NMR.
1H NMR (DMSO-de) 12.58 (1H, s), 11.22 (1H, s), 8.33 (1H, s), 6.45 (1H, s), 3.90 (3H, s), 2.62 (3H, s).
Step 3
5-Acetyl-2,4-dihvdroxy-benzoic acid methyl ester (Alternative procedure)
Figure imgf000192_0002
Resorcinol methyl ester (50 g, 0.298 mol) and Amberlyst 15 resin (40 g) were suspended in toluene 150 mL (under a nitrogen atmosphere) and the solution was heated in an oil bath at 7O0C (internal temp 560C). Acetyl chloride (22 mL, 308 mmol) was added in 5mL portions over 30 mins giving evolution of gaseous HCI (which was scrubbed by passing the nitrogen stream through aqueous NaOH). The solution was stirred at 7O0C for 4.5 h then heated in an oil bath temp (internal temperature 960C) for 3.5 h. The solution was cooled to 5O0C and EtOAc (100 mL) was added and the solution filtered whilst at this temperature. The residual resin was washed with EtOAc (50 mL) and the combined filtrates were concentrated to slurry of crystalline solid (total weight of 128g for solid plus solvent). To the slurry was added heptane (100 mL) and after 10 mins at RT the solid was removed by filtration. The residue was washed with heptane : toluene ( 2 : 1 , 60 mL) then with petroleum ether bp 40- 6O0C and dried in vacuo to give crop 1 29 g (46.4%) (NMR showed 3% of material resulting from saponification of the methyl ester).
The filtrate was evaporated to a small volume and 20% EtOAc in heptane (100 mL) was added. After standing at RT 16 h a second crop of 4.75 g (7.6%) was obtained (NMR identical to crop 1). Step 4
5-Acetyl-2,4-bis-benzyloxy-benzoic acid methyl ester
Figure imgf000193_0001
Benzyl bromide (70 ml, 0.59 mol) was added to a stirred mixture of methyl 5-acetyl-2, 4- dihydroxybenzoate (60.7 g, 0.29 mol) and anhydrous potassium carbonate (87.8 g, 0.64 mol) in acetonitrile (800 ml) and the mixture was stirred and held at reflux for 16 hours. Upon cooling to room temperature the mixture was poured onto water (3 L) and stirred vigorously for 2 hours. The solids were collected by filtration, rinsed with water (2 L), sucked dry under reduced pressure and dried to constant mass in a vacuum oven at 6O0C overnight to afford methyl 5-acetyl-2,4-bis- benzyloxybenzoate (112.1 g, 99%) as a cream solid. 1H NMR (DMSO-d6) 8.21 (1 H, s), 7.55 (4H, m), 7.43 (4H, m), 7.37 (2H, m), 7.04 (1 H, s), 5.38 (4H, s), 3.79 (3H, s), 2.48 (3H, s). MS: [M+H]+ 391.
Step 5
2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid methyl ester
Figure imgf000193_0002
Potassium terf-butoxide (29.1 g, 0.26 mol) was added to a stirred suspension of methyltriphenylphosphonium bromide (92.8 g, 0.26 mol) in anhydrous tetrahydrofuran (1 L) and the mixture was stirred at room temperature for 10 minutes whereupon methyl 5-acetyl-2,4-bis- benzyloxybenzoate (78.0 g, 0.2 mol) was added and the mixture stirred at room temperature for a further 30 minutes. Methanol (100 ml) was added to quench excess phosphorus ylide and the solvent was removed in vacuo to afford an orange oil that crystallized on standing. The residue was recrystallized from methanol (330 ml). The solids were collected by suction filtration, washed with methanol (50 ml) and sucked dry under reduced pressure to afford methyl 2,4-bis-benzyloxy- 5-isopropenyl-benzoate as pale yellow needles. The mother liquor deposited a second crop of material upon standing overnight (combined yield : 56.55 g, 73%) 1H NMR (DMSO-d6) 7.59 (1H, s), 7.52 (2H1 d), 7.64-7.32 (8H, m), 6.97 (1H, s), 5.28 (2H, s), 5.22 (2H, S)1 5.09 (1H, s), 5.04 (1 H, s), 3.76 (3H, s), 2.02 (3H, s). MS: [M+HJ* 389.
A further crop of the ester could be obtained as follows. The crystallization residues were evaporated to dryness in vacuo and the oily solid was treated with 5% ethyl acetate in heptane
(250 ml). Ethyl acetate was added in small portions to the vigourously stirred mixture until the residue deposited a large quantity of solid triphenylphosphine oxide. The solids were removed by filtration and the filtrate evaporated to dryness in vacuo to afford an orange oil. Recrystallization from methanol (as described above) afforded further methyl 2,4-bis-benzyloxy-5-isopropenyl- benzoate as a pale yellow crystalline solid (total yield 85-90%).
Step 6 2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid
Figure imgf000194_0001
Potassium hydroxide (10.96 g, 0.19 mmol) was added to a stirred suspension of methyl 2,4-bis- benzyloxy-5-isopropenyl-benzoate (61.0 g, 0.16 mol) in methanol (750 ml) and water (250 ml) and the mixture was stirred and held at reflux for 16 hours. Upon cooling the organic solvent was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid (200 ml). The mixture was diluted with water (2 L) and extracted with ethyl acetate (2 L), the organic layer was separated and the solvent removed in vacuo to afford 2,4-bis-benzyloxy-5- isopropenyl-benzoic acid (58.8 g, 100%) as a colourless solid. 1H NMR (DMSO-d6) 7.52 (2H, d),
7.47-7.29 (9H1 m), 6.82 (1H, s), 5.20 (2H, s), 5.17 (2H, s), 5.06 (1H, s), 5.04 (1H, s), 2.03 (3H1 s).
MS: [M+H]+ 375.
Step 7
Di-prop-2-ynyl-carbamic acid benzyl ester
Figure imgf000195_0001
To a cooled (O0C) solution of dipropargylamine (46.7 g, 502 mmol) in EtOAc (200 mL) and 10% aqueous K2CO3 (700 mL, 507 mmol) was slowly added a solution of N- (benzyloxycarbonyloxy)succinimide (125 g, 502 mmol) in EtOAc (500 mL) over 20 mins. The solution was stirred at 0 0C for 2 h then at RT 16h. The phases were separated and the organic phase was washed with 10% aqueous K2CO3 (700 mL, 507 mmol) and then with saturated brine (500 mL) and was diluted to 1000 mL with EtOAc to give a 0.5M solution.
Step 8
5-Hvdroxymethyl-1 ,3-dihvdro-isoindole-2-carboxylic acid benzyl ester
Figure imgf000195_0002
A solution of propargyl alcohol (26.4 mL, 424 mmol) in toluene (120 mL) was degassed. The 0.5M- diyne solution above (440 mL, 220 mmol) was evaporated and the residue dissolved in toluene (80 mL). This protected diyne solution and Wilkinson's catalyst (2.26 g, 2.44 mmol, 1.11% were added in 14 equal portions over a 2 h period with constant monitoring of the internal temperature such that the temperature remained 50- 1000C. The solution was allowed to cool to 50 0C over 30 min when the solution was evaporated (to remove excess propargyl alcohol). The residue was heated with toluene (500 mL) and charcoal (Darco 4-12 mesh, 20 g) at 100 0C for 30 min and then filtered hot through a bed of Celite and the brown solution was evaporated. The residue was dissolve in EtOAc (400 mL) at 80 0C when silica gel (chromatography grade 65 g) was added and heating continued for 20 mins. The solution was filtered whilst hot and then evaporated (with seeding) to give a pale brown solid. 10 % EtOAc/heptane (v/v, 100 mL) was added and the solid removed by filtration. The solid was washed on the sinter with heptane (100 mL) and the dried (50 0C, oil pump, 16 h) to give the title compound 59.0 g (95%). 1 H NMR (400 MHz, Me-d3-OD): 7.51-7.16 (m, 8H), 5.21 (s, 2H), 4.74 (s, 2H)1 4.70 (s, 2H), 4.61 (s, 2H). Step 9 δ-Methanesulfonyloxymethyl-I .S-dihvdro-isoindole^-carboxylic acid benzyl ester
Figure imgf000195_0003
To a solution of 5-hydroxymethyl-1 ,3-dihydro-isoindole-2-carboxylic acid benzyl ester (65.75 g, 0.232 mol) in THF (470 mL) and EtOAc (770 mL) was added Et3N (39 ml_, 0.28 mol). The solution was cooled in an ice-bath and a solution of methanesulphonyl chloride (19 mL,0.245 mol) dissolved in EtOAc (50 mL) was added (so that the internal temp <12 0C). After stirring for 2h in the ice-bath further additions of methanesulphonyl chloride (1.9 mL and 0.95 mL) and Et3N (3.9 mL) were made (so that by tic there was no remaining starting material after a further 1 h of stirring). NaHCO3 (550 mL) was added and the solution stirred for 20 mins then saturated brine (200 mL) was added and the phases were separated. The organic phase was dried (MgSO4) and evaporated with seeding to give a damp solid which was used in the next step without thorough drying.
5-(4-Methyl-piperazin-1-ylmethyl)-1 ,3-dihydro-isoindole-2-carboxylic acid benzyl ester dihvdrochloride salt
Figure imgf000196_0001
The solid from Step 9 (assume 0.232 mol) was dissolved in acetone (700 mL) and this solution was added over 45 mins to a cooled (internal temp 15-17 0C) suspension of K2CO3 (48 g) and N- methylpiperazine (50 mL, 0.45 mol) in acetone (330 mL). The suspension was stirred at 15 0C for 3h (complete removal of starting material by tic) when the solution was evaporated to a small volume and the residue partition between EtOAc (1000 mL) and a mixture of water (500 mL) and saturated brine (50 mL). The organic phase was washed with a mixture of water (500 mL) and saturated brine (150 mL) and finally washed with saturated brine (300 mL). The solution was dried (MgSO4) and filtered and to this solution was added 1 M-HCI in MeOH (430 mL, 0.43 mol). The suspension was cooled (O0C for 30 mins) and the solid removed by filtration which was washed with EtOAc and then heptane on the sinter and the solid dried (oil-pump, RT 72 h) to give crop 1 of the title compound 66.34 g (65%) as a colourless solid. 1 H NMR (400 MHz, Me-d3-OD): 7.64-7.51 (m, 2H), 7.51-7.29 (m, 6H), 5.23 (s, 2H), 4.79 (dd, J = 16.2, 6.1 Hz, 4H), 4.49 (s, 2H), 3.66 (s, 8H), 3.03 (s, 3H).
Alternative Step 10A
5-(4-Methyl-piperazin-1-ylmethvπ-1.3-dihvdro-isoindole-2-carboxylic acid benzyl ester dihvdrochloride
Figure imgf000196_0002
Step 10A can be used as an alternative route to replace steps 9 and 10 above.
To a suspension of manganese dioxide (15.5 g, 178 mmol) in DCM (100 mL) was added 5- hydroxymethyl-1 ,3-dihydro-isoindole-2-carboxylic acid benzyl ester (3.35g, 11.8 mmol) and after 6 h stirring at RT a further addition of manganese dioxide (5g, 57 mmol) was made. After a further 1h stirring at RT Celite (7g) was added and the solution was filtered through a bed of Celite™ giving a clear pale yellow solution. The Celite™ was washed with DCM and the volume of the combined organic solution adjusted to 100 mL by evaporation. N-Methylpiperazine (1.31 mL, 11.8 mmol) and acetic acid (0.68 mL) were added followed by sodium triacetoxyborohydride (4.98 g, 23.5 mmol). The yellow solution was stirred 16 h giving a colourless solution. To the solution was added 2M-HCI (10 mL, 20 mmol) giving an effervescence. After 30 min water (1 OmL) and K2CO3 (5.5g, 39.8 mmol) were added and the organic phase was dried (Na2SO4). After filtration 4M-HCI in dioxan (6 mL) was added with stirring and the suspension was evaporated to dryness. The residue was dissolved in MeOH with warming and after evaporation the solid was washed on a sinter with EtOAc then petrol (bp 40-600C) followed by drying in vacuo at 5O0C to give the title compound 3.61 g (70%). 1 H NMR (400 MHz, Me-d3-OD): 7.65-7.51 (2H, m), 7.51-7.27 (6H, m), 5.23 (2H, s), 4.83- 4.69 (4H, m), 4.49 (2H, s), 3.66 (8H, d), 3.03 (3H, s)
5-(4-Methvt-piperazin-1-ylmethyl)-2,3-dihvdro-1 H-isoindole
Figure imgf000197_0001
To 5-(4-Methyl-piperazin-1-ylmethyl)-1 ,3-dihydro-isoindole-2-carboxylic acid benzyl ester dihydrochloride salt (Step 10, 59.8 g, 136.7 mmol) was added EtOAc (400 mL) and 10% aqueous K2CO3 (400 mL). The organic phase was washed with saturated brine (200 mL) and then dried (MgSO4). The solution was filtered and was evaporated to an oil (which crystallised on standing with petroleum ether (bp 40-600C)). The solid was dried in vacuo to give a colourless solid: 48.8g (133.5 mmol).
A portion of the solid (24.4 g, 66.8 mmol) was dissolved in MeOH (170 mL) and after degassing the solution and purging with nitrogen 10% Pd/C (1.22 g) was added and the mixture hydrogenated at latmosphere for 2.5 h. The solution was filtered and the solution evaporated and the residue was azeotroped twice with toluene at 30-400C. The residue was dissolved in DMF (92 mL) and the solution was immediately degassed and purged with N2.
(NB The product at this stage is sensitive to air and darkens on contact with oxygen. The DMF solution was used immediately but can be stored by degassing and storing under an atmosphere Of N2) (2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-f5-(4-methyl-piperazin-1-ylrnethvπ-1 ,3-dihydro-isoinclol- 2-vπ-methanone
Figure imgf000198_0001
A solution of the resorcinol acid (Step 6, 23.7 g, 63.4 mmol) and 1-hydroxybenzotriazole (10.21 g,66.7 mmol) were dissolved in DMF (92 mL) and to this solution was added N-ethyl-N'-(3- dimethylaminopropyl)carbodiimide hydrochloride (12.8 g, 66.8 mmol). The solution was stirred at RT for 40 mins and this solution was added to the solution of the amine from Step 11 (66.8 mmol) together with DMF (5 mL) washings. The solution was degassed and the solution stirred at RT for 16 h. To the solution was added 10% K2CO3 (500 mL) and EtOAc (500 mL) and the organic phase was washed sequentially with 10% K2CO3 (500 mL), water (4 x 100 mL) and saturated brine (200 mL). The solution was evaporated to a small volume and 20% EtOAc in heptane (250 mL) was added and stored at O0C. The solid which had formed was removed by filtration, washed with heptane twice and was dried in vacuo to give the title compound 35.05 g (94.4 %). 1 H NMR (400 MHz, Me-d3-OD): 7.49-7.10 (m, 14H), 6.86 (d, J = 2.5 Hz, 1 H), 5.17 (d, J = 2.5 Hz, 4H), 5.09 (d, J = 11.3 Hz, 2H), 4.88 (s, 2H), 4.63 (s, 2H), 3.54 (d, J = 16.0 Hz, 2H), 2.50 (s, 7H), 2.28 (d, J = 7.6 Hz, 3H), 2.11 (s, 3H).
(2,4-Dihvdroxy-5-isopropyl-phenvπ-f5-(4-methyl-piperazin-1-ylmethyl')-1 ,3-dihvdro-isoindol-2-vn- methanone
Figure imgf000198_0002
The product from Step 12 (4.7 g) was dissolved in 1 :1 MeOH/water (98 mL) and after purging with N2 10% Pd/C and K2CO3 (2.38g, 17.2 mmol) were, added and the suspension was hydrogenated for 16 h under an atmosphere of H2. The solution was filtered and the solvent evaporated. To the residue was added aqueous 2M-HCI (40 mL) and the solution was washed with 1 :1 EtOAc/petrol (40 mL x 2) and then the pH adjusted to pH 8.5 by addition of NaOH and EtOAc (50 mL) added. The solution was heated to 60 0C and the aqueous phase removed. The hot organic phase was washed with water (30 mL) and then evaporated to a small volume (ca. 5 mL) and allowed to stand at RT 16h with seeding. To the crystalline material was added 1 :1 EtOAc/ petrol (10 mL) and the mixture was filtered and dried to give the title compound as the free base 1.76 g . 1 H NMR (400 MHz, Me-d3-OD): 7.29 (s, 3H), 7.19 (s, 1H), 6.39 (s, 1 H), 4.91 (s, 4H), 3.56 (s, 2H), 3.28-3.15 (m, 1H), 2.53 (S, 8H), 2.31 (s, 3H)1 1.23 (d, J = 6.9 Hz, 7H).
Optional Step 14
Purification of (2,4-Dihvdroxy-5-isopropyl-phenvπ-f5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihvdro- isoindol-2-yll-methanone
In some batches of product, the title compound (X = H in the formula) can contain small amounts of the impurity 2,4-Dihydroxy-5-(2-hydroxyprop-2-yl)-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)- 1,3-dihydro-isoindol-2-yl]-methanone (X = OH in the formula). The impurities can be removed by the following method.
Figure imgf000199_0001
Acetic anhydride (1.04 ml, 11.0 mmol) was added to a stirred suspension of impure 2-(2,4- dihydroxy-5-isopropylbenzoyl)-5-(4-methylpiperazin-1-ylmethyl)-1 ,3-dihydroisoindole (2.05 g, 5.0 mmol) in toluene (20 ml) and the resulting mixture was stirred and held at 100 0C for 16 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford a brown oil which was dissolved in methanol (20 ml). Concentrated hydrochloric acid (1 ml) was added and the mixture was stirred and held at reflux for 5 hours. Upon cooling to room temperature, the organic solvent and volatile material were removed in vacuo and the aqueous residue was diluted with water (25 ml) and basified to pH 8 with vigourous stirring by the careful addition of 10% aqueous potassium carbonate solution. 50% Ethyl acetate in heptane (50 ml) was added and the mixture was stirred vigourously at room temperature for 16 hours. The solid material was collected by suction filtration, rinsed with 50% ethyl acetate in heptane (50 ml), sucked dry under reduced pressure and dried overnight in a vacuum oven at 50 0C to afford 2-(2,4-dihydroxy-5- isopropylbenzoyl)-5-(4-methylpiperazin-1-ylmethyl)-1 ,3-dihydroisoindole (1.85 g, 90%) as an off- white solid. 1H NMR (DMSO-d6) 10.07 (1H, br s), 9.60 (1 H, br s), 7.24 (3H1 m), 7.06 (1H, s), 6.40 (1 H, s), 4.76 (4H, br s), 3.44 (2H, s), 3.10 (1H, m), 2.32 (8H, m), 2.14 (3H, s), 1.15 (6H, d). MS: [M+H]+ 410. EXAMPLE 81
(2,4-Dihvdroxy-5-isopropyl-phenyl)-r5-(4-methyl-piperazin-1-ylmethvπ-1,3-dihydro-isoindol-2-vπ- methanone
Example 81 describes a synthetic route containing essentially the same process steps as the route described in Example 80 but wherein the process conditions are more suited to larger scale reactions.
Step 1
4-Acetoxy-2-hvdroxy-benzoic acid methyl ester
To a heated solution (500C) of resorcinol methyl ester (16.5 Kg, 98.1 mol) and N,N-dimethyl-4- aminopyridine (89.1 g, 0.73 mol, 7.4 mol%) in toluene (66 L) was slowly added (over 2 h) acetic anhydride (9.9 L, 104.9 mol). The solution was heated to 50 0C for a further 1.5h and then the solvent was removed by evaporation at 50 0C to a small volume and the residue was azeotroped once with toluene. To the residual oil was immediately added toluene (33 L) whilst still warm and the solution used for Step 2 without further purification. Step 2
5-Acetyl-2,4-dihvdroxy-benzoic acid methyl ester
The toluene solution from Step 1 was cooled in an ice bath under N2 and triflic acid (9.44 L) added slowly over 3 h. On stirring a fine white solid was formed which dissolved on warming to RT over 20 h and then stirring at RT for 37 h to give a yellow solution. To the solution was added acetyl chloride (726 mL) and the solution stirred at RT for a further 1 h. This solution was cannulated into a stirred cooled (O0C) solution of EtOAc (217.8 L) and NaOAc.3H2O (14.52 Kg) dissolved in water (145 L). The organic phase was washed with saturated brine (twice, 72.6 L), and was evaporated to 5.5 Kg. Toluene : lsopropanol (2 : 3) was added and the crystalline solid removed by filtration and dried to give 12.6 Kg (61% over 2 steps), mp 124-126 0C. Step 3
5-Acetyl~2,4-bis-benzyloxy-benzoic acid methyl ester
To a stirred solution of benzyl bromide (16.14 L, 136 mol) and anhydrous potassium carbonate (20.25 Kg, 147.6 mol) in acetonitrile (184.5 L) was added methyl 5-acetyl-2,4-dihydroxybenzoate (14 Kg, 66.6 mol, step 2) in 6 portions over 5 h. The mixture was stirred and held at reflux for 20 hours, cooled to room temperature the mixture was poured onto water (682 L) and stirred vigorously for 2 hours. The solids were collected by centrifugation and dried under reduced pressure to constant mass in a vacuum oven at 600C overnight to afford methyl 5-acetyl-2,4-bis- benzyloxybenzoate (23.5 Kg, 97.3%) as a cream solid mp 114-115 0C.
Step 4 2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid methyl ester A solution of potassium terf-butoxide (6.72 Kg, 60.1 mol) in anhydrous THF (60 L) was added over 3 h to a stirred suspension of methyltriphenylphosphonium bromide (21 ,43 Kg, 60.1 mol) and methyl 5-acetyl-2,4-bis-benzyloxybenzoate (21.3 Kg, 54.6 mol, step 3) in anhydrous tetrahydrofuran (213 L) at 15 0C. The mixture was stirred at 15 0C for 70 mins and the warmed to 20 0C over 60 mins. Methanol (27.3 L) was added to quench excess phosphorus ylide and the solvent was concentrated in vacuo followed by addition of EtOAc and water. The organic phase was treated with activated charcoal, filtered and evaporated to a small volume. The residue was crystallised from boiling MeOH and the solids were collected by suction filtration, washed with methanol and dried under reduced pressure to afford methyl 2,4-bis-benzyloxy-5-isopropenyl- benzoate 18.1 Kg (85%) as pale yellow needles mp 92-94 0C (99.6% pure by hplc).
2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid
Potassium hydroxide (0.527 Kg, 9.4 mol) was added to a stirred suspension of methyl 2,4-bis- benzyloxy-5-isopropenyl-benzoate (3.1 Kg, 8 mol, step 4) in methanol (18.6 L) and water (12.4 L) and the mixture was stirred and held at reflux for 3 hours. The methanol was removed under partial vacuum from the vessel, and to the remaining solution was added toluene (62 L). The solution was heated to 40 0C and to the mixture was added cone HCI (1.36L). The biphasic mixture is heated to 50 0C and the phases separated. The organic phase was washed with water (31 L) at 50 0C and the organic phase was evaporated under redeuced pressure to give 2,4-bis-benzyloxy- 5-isopropenyl-benzoic acid 2.851 Kg (95% yield) as a colourless solid.
Di-prop-2-vnyl-carbamic acid benzyl ester
To a cooled (5 0C) solution of K2CO3 (4 Kg, 29.0 mol) in water (17.5 L) and toluene (12.5 L) was added dipropargylamine (2.50 Kg, 26.88 mol). Benzyloxychloroformate (4.8 Kg, 28.14 mol) was added at a rate such that T<10 0C. The solution was stirred at 5 0C for 10 mins and then allowed to warm to RT. The aqueous phase was separated and the organic phase was washed with 0.2M HCI (12.5 L), sat NaHCO3 (13.5 L) and brine (17 L) and the resultant solution used in step 7 (assayed to contain 6.23 Kg, 102 % based on an evaporated portion).
Step 7 5-Hvdroxymethyl-1 ,3-dihvdro-isoindole-2-carboxylic acid benzyl ester
A solution of propargyl alcohol (2.11 Kg, 37.7 mol) in toluene (32.48L) was degassed and heated to 55 0C. The solution of di-prop-2-ynyl-carbamic acid benzyl ester (4.06 Kg, 17.86 mol, step 6) in toluene and Wilkinsons catalyst (0.162 Kg) were added in 10 equal portions such that temperature <65 0C (the exotherm was allowed to subside before the next addition was made). The solution was then stirred at 55 0C for 1 h and then cooled to 20 0C. DCM (8.12 L) was added and the mixture was concentrated to a small volume. Toluene (8L) was added and the solution evaporated to constant weight giving the title compound 5.72 Kg (113%).
Step 8 δ-Methanesulfonyloxymethyl-I .S-dihvdro-isoindole^-carboxylic acid benzyl ester To a cooled solution (5 0C) of 5-hydroxymethyl-1 ,3-dihydro-isoindole-2-carboxylic acid benzyl ester (11 Kg, 38.8 mol, step 7) and Et3N (7.04 L, 50.6 mol) in DCM (55 L) was added methanesulphonyl chloride (2.97 L, 38.4 mol) so that the internal temp <10 0C. After stirring for 0.5 h at 5 0C the solution was used below in step 9.
Step 9 5-(4-Methyl-piperazin-1-ylmethyl)-1 ,3-dihvdro-isoindole-2-carboxylic acid benzyl ester dihvdrochloride salt
Figure imgf000202_0001
2HCI
The solid from Step 8 (assume 0.232 mol) was dissolved in acetone (700 mL) and this solution was added over 45 mins to a cooled (internal temp 15-17 0C) suspension of K2CO3 (48 g) and N- methylpiperazine (50 mL, 0.45 mol) in acetone (330 mL). The suspension was stirred at 15 0C for 3h (complete removal of starting material by tic) when the solution was evaporated to a small volume and the residue partition between EtOAc (1000 mL) and a mixture of water (500 mL) and saturated brine (50 mL). The organic phase was washed with a mixture of water (500 mL) and saturated brine (150 mL) and finally washed with saturated brine (300 mL). The solution was dried (MgSO4) and filtered and to this solution was added 1M-HCI in MeOH (430 mL, 0.43 mol). The suspension was cooled (O0C for 30 mins) and the solid removed by filtration which was washed with EtOAc and then heptane on the sinter and the solid dried (oil-pump, RT 72 h) to give crop 1 of the title compound 66.34 g (65%) as a colourless solid. 1H NMR (400 MHz, Me-d3-OD): 7.64-7.51 (m, 2H), 7.51-7.29 (m, 6H), 5.23 (s, 2H), 4.79 (dd, J = 16.2, 6.1 Hz, 4H), 4.49 (s, 2H), 3.66 (s, 8H), 3.03 (s, 3H).
Step 9
5-(4-Methyl-piperazin-1-ylmethyl)-1 ,3-dihvdro-isoindole-2-carboxylic acid benzyl ester
DCM (33L) and N-methylpiperazine (21.45 L, 193.4 mol) were stirred at 25 0C and the solution from step 8 added over a minimum of 30 mins such that temperature 20 - 30 0C. After stirring the solution for a further 30 mins water (55 L) was added and the organic phase was washed with water (2 x 55 L). The product was extracted into 0.8M HCI (66 L) and the layers separated. The aqueous phase was washed with DCM (55 L) and then basified with 2M NaOH to pH 10-11 and the product was extracted into EtOAc (2 x 55 L). The combined organic phase were filtered to remove solids and the evaporated followed by azeotroping with toluene and drying to constant weight to give the title compound, 6.63kg (47% yield, 98% pure by hplc).
Step 10 5-(4-Methyl-piperazin-1-ylmethyl)-2,3-dihvdro-1 H-isoindole To a degassed solution of 5-(4-Methyl-piperazin-1-ylmethyl)-1 ,3-dihydro-isoindole-2-carboxylic acid benzyl ester (Step 9, 1.3 Kg, 3.55 mol) dissolved in EtOH (13 L) was added 10% Pd/C (0.065 Kg). Hydrogen was passed through the mixture at 30 0C for 4 h or until complete by NMR. The solution was then stirred for 1 h under an atmosphere of N2 and then filtered to remove the catalyst through a GF/F filter followed by filtration through a Cuno filter. The filtrate was evaporated to a small volume, azeotroped with toluene (3.9 L) and dried to constant weight yielding the title compound as a red/ black oily solid (0.78 Kg) which was stored under nitrogen until required.
Step 11
(2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethvπ-1 ,3-dihvdro-isoindol- 2-vπ-methanone 1 ,1'-Carbonyldiimidazole (4.82 Kg, 29.8 mol) was added to a solution of 2,4-bis-benzyloxy-5- isopropenyl-benzoic acid (10.58 Kg, 28.3 mol, step 5) in DMF (21.2 L) at 25 0C. After 20 mins at 25 0C a solution of 5-(4-Methyl-piperazin-1-ylmethyl)-2,3-dihydro-1 H-isoindole (7.2 Kg, 31.1 mol, step 10) in DMF (7.2 L) maintaining a temperature below 35 0C and the solution stirred at 25 0C for a minimum of 12 h. The solid which had formed was removed by filtration, washed with isopropyl acetate (2 x 21.6 L) and dried at 35 0C to constant weight to give the title compound 8.7 Kg (77% yield, purity by hplc 97.5%).
(2,4-Dihvdroxy-5-isopropyl-phenyl)-f5-(4-methyl-piperazin-1-ylmethvπ-1 ,3-dihvdro-isoindol-2-vn- methanone The product from Step 11 (0.9 Kg, 1.53 mol) was dissolved in isopropanol ( 6.8L) and water (1.04 L) and after purging with N2 10% Pd/C (9Og) and K2CO3 (0.212 Kg, 1.53 mol) were added and the suspension was hydrogenated for 60 to 70 mins under an 3 Barr pressure of H2. The solution was diluted with water (0.5 L) and filtered. To the filtrate was added aqueous HCI (30% hydrochloric acid, 0.85 Kg diluted with water 5.42 Kg) and the solution was concentrated at 60 0C under vacuum (removing 10 L isopropanol). Water (0.45 L) was added to the solution and concentration continued (until a further 10 L isopropanol had been removed). The aqueous phase was washed with EtOAc (4.61 L), diluted with acetonitrile (4.06L) and netralised to pH 7.5- 8.5 by addition of cone ammonia solution (0.35 Kg). The suspension was stirred for 2.5 h and then the solid was removed by filtration. The residue was washed with acetonitrile (2 x 0.8 L) and dried at 40 0C to constant weight to give the title compound 588 g (94 % yield).
EXAMPLE 82 Alternative synthesis of 5-(4-Methyl-piperazin-1-ylmethvπ-2,3-dihvdro-1 H-isoindole 82A, Synthesis of 1-methyl-4-prop-2-vnyl-piperazine
Figure imgf000204_0001
To 1-methylpiperazine (37.7 ml, 337 mmol) and K2CO3 (46.6 g, 337 mmol) in acetone (380 ml) was added propargyl bromide (25 ml, 225 mmol, 80% in toluene) in acetone (70 ml) dropwise at 0 0C under N2. The internal temperature of the reaction was kept <10 0C. The reaction was stirred at room temperature for 3 hours. The reaction was filtered, and the salts were washed with small portions of acetone (x2). The filtrates were combined evaporated to concentration (gently). To the residue was added water and the product was extracted with DCM (x3). The combined organic layers were washed with brine and dried over MgSO4. The product was filtered and evaporated to dryness to yield 1-methyl-4-prop-2-ynyl-piperazine as a yellow oil.
82B. Synthesis of 5-(4-Methyl-piperazin-1-ylmethyl)-1 ,3-dihvdro-isoindole-2-carboxylic acid tert- butyl ester
Figure imgf000204_0002
A solution of N-boc-dipropargylamine (36.3 ml, 226 mmol, 86% pure) in EtOAc (30 ml) was made up and degassed by bubbling through N2, in a separating funnel. Tris(triphenylphosphine)rhodium(l) chloride (1.39 g, 1.50 mmol, 1mol%) was added to pre- degassed EtOAc (15 ml) in a second separating funnel. (NB CpRu(COD)CI) can also be used as an alternative catalyst).
In the main reactor flask, 1-propargyl-4-methylpiperazine (32.3 ml, 150 mmol, 90% pure) was diluted with EtOAc (75 ml) and was degassed by bubbling N2 through the mixture The mixture was cooled in a ice-water bath and then the tris(triphenylphosphine)rhodium(l) chloride (1.39 g, 1 mol%) in EtOAc was added. Slow addition of N-boc-dipropargylamine /EtOAc was undertaken to yield a mild exotherm. The internal temperature rose to 25 0C and remained at this temeprature. After addition was approximately one third complete (-45 minutes), the exotherm tailed off (despite the continual slow addition of N-boc-dipropargylamine/EtOAc). Another portion of tris(triphenylphosphine) rhodium(l) chloride catalyst (1.39 g, 1 mol%) in EtOAc (15 ml, pre- degassed) was made up and added very slowly to the reaction. After a couple of minutes a new exotherm started and grew to 30 0C. The reaction temperature was cooled gently by the addition of a small amount of ice to the water bath. Once the exotherm began to subside, slow addition of N-boc-dipropargylamine /EtOAc was continued. The entire addition was carried out over a 2 hour period. The reaction mixture was then left at room temperature overnight before diluting with EtOAc and washing with NH4CI (x2) (aqueous, saturated) to remove excess 1-propargyl-4- methylpiperazine. The mixture was diluted with a small amount of water to dissolve the salts. The organic layer was washed with water, brine and dried over MgSO4. The product was filtered and evaporated to dryness to leave a brown oil.
To the oil residue obtained was added n-heptane. The oil/heptane was left to stand (-10 minutes) until a red precipitate formed. The precipitate was filtered and washed with fresh /7-heptane (x2). The filtrates were dried to yield the product as a red oil.
The desired product was further purified by forming the toluenesulphonic acid (TsOH) salt. Thus, the crude product was taken up in MeOH (20 ml) and the TsOH. H2O (1 eq to estimated purity by NMR) was added. The solution was evaporated to dryness, and then dissolved in toluene (x1) and re-evaporated. The resulting product was taken up in ether. After a few minutes, a precipitate and solution formed. The precipitate was filtered and washed with more ether (x2) until the filtrate was colourless. The yellow solid was dried to yield the product as the TsOH salt. MS: [M+H]+ 332. 82C. Synthesis of 5-(4-Methyl-piperazin-1-ylmethvO-2,3-dihvdro-1H-isoindole
Figure imgf000205_0001
The isoindoline tosylate salt was taken up in DCM (0.3 M) and TFA (12 eq.) added slowly at 0 °C. The reaction was stirred overnight at room temperature. The reaction was evaporated to dryness and then with toluene/MeOH (X3) to yield the product as a mixture of acid addition salts. MS: [M+H] + 232.
The compound of Example 82C can be used in the method of Example 80 Step 12.
EXAMPLE 83
Alternative synthesis of 5-hydroxymethyl-1 ,3-dihvdro-isoindole-2-carboxylic acid benzyl ester
83A. Methyl 2-benzyl-2,3-dihvdro-1H-isoindole-5-carboxylate
Figure imgf000205_0002
Benzylamine (3.21 g, 30.0 mmol) in anhydrous tetrahydrofuran (25 ml) was added to a stirred mixture of methyl 3,4-bis-(bromomethyl)benzoate (9.66 g, 30.0 mmol) (obtained from Fluorochem) and triethylamiπe (9 ml, 64.7 mmol) in anhydrous tetrahydrofuran (50 ml) and the resulting mixture was stirred at room temperature for 3 hours. The solvent was removed in vacuo at 40 °C and the residue partitioned between ethyl acetate (100 ml) and water (100 ml). The organic layer was washed with a further portion of water (100 ml), separated and the solvent removed in vacuo at 40 0C to afford methyl 2-benzyl-2,3-dihydro-1H-isoindole-5-carboxylate as a pale orange solid that was used immediately without further purification as described below. 1H NMR (DMSOd6) 7.82 (2H, m), 7.40-7.25 (6H, m), 3.90 (3H, s), 3.88 (2H, s), 3.84 (4H, s). MS: [M+H]+ 268.
83B. (2-Benzyl-2,3-dihydro-1 H-isoindol-5-vO-methanol
Figure imgf000206_0001
Methyl 2-benzyl-2,3-dihydro-1 H-isoindole-5-carboxylate (from above) was dissolved in anhydrous tetrahydrofuran (75 ml) and added dropwise over 15 minutes to a rapidly stirred suspension of lithium aluminium hydride (1.71 g, 45.0 mmol) in anhydrous tetrahydrofuran (75 ml). The mixture was stirred at room temperature for 2 hours whereupon excess lithium aluminium hydride was destroyed by the slow dropwise addition of 1M sodium sulphate solution (12 ml). The solids were removed by filtration, rinsed with ethyl acetate (2 x 50 ml) and sucked dry. The solvent was removed in vacuo to afford (2-benzyl-2,3-dihydro-1 H-isoindol-5-yl)-methanol (7.15 g, 99%) as a tan solid. 1H NMR (DMSOd6) 7.40-7.30 (4H, m), 7.28 (1 H1 m), 7.17-7.10 (3H, m), 5.10 (1 H, t), 4.47 (2H, Cl)1 3.85 (2H, s), 3.82 (2H, s), 3.80 (2H, s). MS: [MH-H]+ 240.
83C. (2,3-Dihvdro-1 H-isoindol-5-ylVmethanol
Figure imgf000206_0002
10% Palladium on activated carbon (200 mg) was added to a solution of (2-benzyl-2,3-dihydro-1 H- isoindol-5-yl)-methanol (2.39 g, 10.0 mmol) in ethanol (60 ml) and the resulting mixture was placed in a Parr apparatus, heated to 50 0C and shaken under a hydrogen atmosphere at 60 psi for 30 hours. Upon cooling to room temperature the mixture was filtered under gravity, the solids were rinsed with ethanol (2 x 10 ml) and the solvent removed in vacuo to afford (2,3-dihydro-1 H- isoindol-5-yl)-methanol (1.49 g, 100%) as an off-white solid. 1H NMR (DMSOd6) 7.20 (1H, s), 7.18 (1 H, d), 7.12 (1 H, d), 5.10 (1 H, br s), 4.46 (2H, s), 4.05 (4H, s). MS: [M+H]+ 150.
83D. δ-Hvdroxymethyl-I .S-dihvdro-isoindole-Σ-carboxylic acid benzyl ester
Figure imgf000207_0001
A mixture of (2,3-dihydro-1H-isoindol-5-yl)-methanol (1.34 g, 9.0 mmol) in anhydrous tetrahydrofuran (50 ml) was warmed gently to aid dissolution and allowed to cool to room temperature. Triethylamine (1.5 ml, 10.8 mmol) was added and the stirred mixture was treated dropwise with benzyl chloroformate (1.35 ml, 9.5 mmol) and stirred at room temperature for 3 hours. The solvent was removed in vacuo and the residue partitioned between ethyl acetate (30 ml) and 2M hydrochloric acid (30 ml). The organic layer was washed with water (30 ml), separated and the solvent removed in vacuo to afford a pink oil that solidified upon standing. The solids were triturated with 10% ethyl acetate in hexane (10 ml), filtered, rinsed with heptane (10 ml) and sucked dry to afford the title compound (2.5 g, 98%) as a pale pink solid. 1H NMR (DMSO-d6) 7.45-7.21 (8H, m), 5.20 (1 H, t), 5.17 (2H, s), 4.71 (2H, br s), 4.64 (2H, br s), 4.50 (2H, d). MS: [M+H]+ 284.
The title compound can be used in Step 9 of Example 80.
PREPARATION OF PRO-DRUG COMPOUNDS
The pro-drug compounds of the invention can be prepared from the phenolic compounds of Examples 1 to 79 by means of methods well known to the skilled person, for example the general methods described above in the section headed "Methods for the Preparation of Compounds of the Formula (I)", and the methods described in Examples 80 to 85 below.
EXAMPLE 84
Synthesis of acetic acid δ-acetoxy^-isopropyl^δ-H-methyl-piperazin-i-vO-I .S-dihvdro-isoindole- 2-carbonyl1-phenyl ester
Figure imgf000207_0002
A mixture of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2- yl]-methanone hydrochloride salt (100 mg) and acetic anhydride (350 μl) in pyridine (1 ml) was heated at 12O0C for 20 minutes in a CEM discover microwave synthesiser then evaporated and re- evaporated with toluene (x3). The crude material was purified by flash column chromatography (Biotage SP4), eluting with DCM then DMAW240 then DMAW120. Product containing fractions were evaporated then partitioned between DCM and saturated NaHCO3 solution, the DCM layer was separated, dried (MgSCU) filtered and evaporated to give 75 mg of the title compound as a light tan foam. 1H NMR (400 MHz, DMSO-d6): 7.54 (1 H, d), 7.25-7.02 (2H, m), 7.01-6.83 (2H, m), 4.71 (2H, d), 4.53 (2H, d), 3.10 (4H, dt), 3.06-2.96 (1 H, m), 2.48-2.42 (4H, m), 2.34 (3H, s), 2.22 (3H, d), 2.16 (3H, s), 1.19 (6H, d). MS: [M+H]+ 480.
EXAMPLE 85
Synthesis of 2,2-dimethyl-propionic acid 5-(2,2-dimethyl-propionyloxy)-4-isopropyl-2-[5-(4-methyl- piperazin-1-yl)-1 ,3-dihvdro-isoindole-2-carbonyl1-phenyl ester
Figure imgf000208_0001
The title compound was prepared by the method described in Example 84, but using 0-20% MeOH / EtOAc as eluant during column chromatography. 54 mg of product isolated as a white solid. 1H NMR (400 MHz, DMSO-d6): 7.52 (1 H, d), 7.23-7.10 (1 H, m), 7.05 (1 H, s), 6.98-6.85 (2H, m), 4.68 (2H, d), 4.49 (2H, d), 3.15-3.05 (4H, m), 3.03-2.94 (1 H, m), 2.50-2.40 (4H, m), 2.24 (3H, s), 1.35 (9H, s), 1.20 (6H, d), 1.17 (9H, s). MS: [M+H]+ 564.
Example 86
Synthesis of dimethyl-carbamic acid 5-dimethylcarbamoyloxy-2-isopropyl-4-r5-(4-methyl-piperazin- 1-yl)-1 ,3-dihydro-isoindole-2-carbonvπ-phenvl ester
Figure imgf000209_0001
A mixture of (2,4-dihydroxy-5-isopropyI-phenyl)-E5-(4-methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2- yl]-methanone hydrochloride salt (100 mg) and dimethylcarbamyl chloride (420 μl; 20 equivalents) in pyridine (1 ml) was heated at 1400C for 20 minutes in a CEM discover microwave synthesiser then evaporated and re-evaporated with toluene (x3). The crude material was purified by preparative LC/MS and 55 mg of the title compound were isolated as a brown solid (formate salt). 1H NMR (400 MHz, DMSO-d6): 8.15 (1 H, s), 7.42 (1 H, d), 7.25-7.10 (1 H, m), 7.05-6.85 (3H, m), 4.70 (2H, d), 4.54 (2H, d), 3.20-3.10 (4H, m), 3.09 (3H, s), 3.08-3.00 (1 H, m), 2.95 (3H, s), 2.88 (3H, s), 2.79 (3H, s)2.65-2.47 (4H, m), 2.35 (3H, d), 1.20 (6H, d). MS: [M+H]+ 538.
EXAMPLE 87
Synthesis of acetic acid 5-acetoxy-4-isopropyl-2-f5-(4-fnethyl-piperazin-1-ylmethy])-1 ,3-dihvdro- isoindole-2-carbonvπ-phenyl ester
Figure imgf000209_0002
A mixture of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihydro- isoindol-2-yl]-methanone (100 mg) and acetic anhydride (350 μl) in pyridine (1 ml) was heated at 120°C for 20 minutes in a CEM discover microwave synthesiser then evaporated and re- evaporated with toluene (x3). The crude material was purified by flash column chromatography (Biotage SP4), eluting with DCM then DMAW240 then DMAW120. Product containing fractions were evaporated then partitioned between DCM and saturated NaHCO3 solution, the DCM layer was separated, dried (MgSO4) filtered and evaporated to give the title compound. EXAMPLE 88
Preparation of Glucuronide analogues of (2,4-dihvdroxy-5-isopropyl-phenylH5-(4-methyl- piperazin-1-ylmethyl)-1 ,3-dihvdro-isoindol-2-yll-rriethanone
(2S,3S,4S,5R)-3,4,5-trihvdroxy-6-(5-hvdroxy-4-isopropyl-2-f5-(4-methyl-piperazin-1-ylmethyl)-1 ,3- dihvdro-isoindole-Σ-carbonvn-phenoxyl-tetrahvdro-pyran-Σ-carboxylic acid
(2S.3S,4S,5R)-3,4,5-trihvdroxy-6-(5-hvdroxy-2-isopropyl-4-r5-(4-methyl-piperazin-1-ylmethyl)-1 ,3- dihydro-isoindole^-carbonvπ-phenoxyi-tetrahydro-pyran^-carboxylic acid
Figure imgf000210_0001
Using rat microsomes:
A solution of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihydro- isoindol-2-yl]-methanone dihydrochloride in DMSO (14 μl; 150 mM), was added to 806 μl potassium phosphate buffer (100 mM containing 5 mM magnesium chloride at pH 7.4), followed by UDP Glucuronic acid tri-sodium salt (60 μl_; 100 mM) and alamethicin (20 μl; 5 mg / ml) then incubated at 40C. After 3 minutes rat microsomes were added (100 μl; 20 mg / ml) then incubated at 37 0C for 2-4 hours. The reaction was treated with acetonitrile (1 ml), centrifuged (13000 rpm) and the supernatant removed and retained. This procedure was repeated eleven times, the supernatants were pooled, split into 3 equal batches and dried under a stream of nitrogen at 40 0C to approximately 2 ml. The concentrate was extracted with methyl tert-butyl ether (5 x 5 ml) and the aqueous layer evaporated to dryness under a heated stream of nitrogen (40 0C). Confirmation of the production of glucuronides was carried out by LC-MS/MS.
Using dog microsomes:
A solution of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1 -ylmethyl)-1 ,3-dihydro- isoindol-2-yl]-methanone dihydrochloride in DMSO (14 μl; 150 mM), was added to 806 μl potassium phosphate buffer (100 mM containing 5 mM magnesium chloride at pH 7.4), followed by UDP Glucuronic acid tri-sodium salt (60 μL; 100 mM) and alamethicin (20 μl; 5 mg / ml) then incubated at 40C. After 3 minutes dog microsomes were added (100 μl; 20 mg / ml) then incubated at 37 0C for 2-4 hours. The reaction was treated with acetonitrile (1 ml), centrifuged (13000 rpm) and the supernatant removed and retained. This procedure was repeated seven times, the supernatants were pooled, split into two equal batches and dried under a stream of nitrogen at 40 0C to approximately 2 ml. The concentrate was extracted with methyl tert-butyl ether (5 x 5 ml) and the aqueous layer evaporated to dryness under a heated stream of nitrogen (40 0C). Confirmation of the production of glucuronides was carried out by LC-MS/MS.
Compound A (from dog microsomes). LC/MS: Rt = 13.4-14.1 ; [M+Hf 586. (Mix of monoglucuronides)
Compound B (from rat microsomes). LC/MS: Rt = 20.6; [M+H]+ 586. (Single monoglucoronide). EXAMPLE 89
Preparation of sulphate analogues of (2,4-dihvdroxy-5-isopropyl-phenylH5-(4-methyl-piperazin-1- ylmethyl)-1.3-dihydro-isoindol-2-vn-methanone
A. Sulphuric acid mono-(5-hvdroxy-2-isopropyl-4-[5-(4-methyl-piperazin-1-ylmethyl)-1.3-dihydro- isoindole-2-carbonvn-phenyl) ester
B. Sulphuric acid mono-(5-hvdroxy-4-isopropyl-2-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihvdro- isoindole-2-carbonyl1-phenyl) ester
C. Sulphuric acid mono-{4-isopropyl-2-r5-(4-methyl-piperazin-1-ylmethyl)-1.3-dihvdro-isoindole-2- carbonvπ-5-sulphoxy-phenyl) ester
Figure imgf000211_0001
A solution of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihydro- isoindol-2-yl]-methanone (104 mg; 0.25 mmol) in N,N-dimethy!aniline (2 ml) was cooled in an ice bath then treated dropwise with chlorosulphonic acid (0.1 ml; 1.5 mmol) and stirred for 2 hours. The reaction mixture was basified with 50% KOH (0.3 ml) then partitioned between water and Et2O. The aqueous layer was separated washed with Et2O, then pH adjusted to 10 with the addition of 1M HCI. The resulting solution was evaporated to dryness. Purification by preparative LC/MS gave the three title compounds with the data shown below.
Compound A. LC/MS: Rt = 2.13; [IvHH]+ 490.
Compound B. LC/MS: Rt = 2.24; [M+H]+ 490 (compound appears to rearrange to give compound A or decompose).
Compound C. LC/MS: Rt = 1.96; [M+H]+ 570 (disulphate). BIOLOGICAL ACTIVITY EXAMPLE 90 Isothermal titration calorimetry
The ability of the parent phenolic compounds of the pro-drug compounds of the invention to bind to human Hsp90 proteins was determined using isothermal titration calorimetry.
Isothermal titration calorimetry (ITC) experiments were performed with a VP-ITC titration calorimeter (Microcal Inc., Northampton, MA, USA). Cloning, expression, and purification of the Human Hsp90α N-terminal domain were performed according to published methods (Jez, J. M. et a/, Chem Biol. 2003 Apr;10(4):361-8.) Solutions of the human Hsp90α N-terminal domain and compound were prepared in a buffer comprising 25 mM Tris, 100 mM NaCI, 1 mM MgCI2, 1 mM TCEP, 5% DMSO, pH 7.4. All solutions were filtered and degassed prior to a titration being carried out. The enthalpy change resulting from each injection of ligand was obtained through integration of the calorimetric signal. Data were analysed using Origin 7.0 (Microcal Software Inc., Northampton, MA). Heats of dilution were estimated using the final injections of each individual titration and subtracted before data fitting. Different ITC experimental formats were employed in order to obtain compound dissociation constants (Kd's) over a wide range of affinities. For weakly binding compounds a low c-value ITC method was used (Turnbull W.B. & Daranas A.H. J. Am. Chem. Soc. 2003 Dec 3; 125(48): 14859-66) in which the protein was present at 10-20 μM in the calorimetric cell and the compound concentration was 1-20 mM in the injection syringe. In this type of experiment the stoichiometry parameter (N) was locked at 1 for data fitting. For Kd's in the 20- 0.004 μM range the experiment was configured such that the binding site concentration divided by the Kd (c-value) was between 5 and 1000. For the majority of these experiments the protein concentration in the calorimetric cell was in the range 4-100 μM and the ligand concentration in the injection syringe ranged from 50-1500 μM. In rare cases where compound solubility was limiting, the compound solution was placed in the calorimetric cell and titrated with protein from the injection syringe, maintaining a c-value between 5 and 1000. Competition ITC experiments were used to access Kd's < 4 nM by performing the titration in the presence of a weaker binding competitor according to the method described in Sigurskjold B.W. Anal Biochem. 2000 Jan 15; 277(2):260-6. The compounds of examples 5, 10, 1 1 , 12, 13, 14, 16, 17, 18, 19, 21 , 22, 23, 25, 26, 27, 28, 29, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 48, 49, 50, 51 , 52, 53, 54, 55, 59, 60, 61 , 62, 63, 65, 66, 67, 68, 69, 70, 71 , 72, 73, 74 and 75 were tested and were found to have Kd values of less than 1 micromolar.
The compounds of examples 5, 10, 12, 13, 14, 16, 17, 18, 19, 21 , 22, 23, 25, 26, 27, 28, 29, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 48, 49, 50, 51 , 53, 54, 55, 59, 60, 61 , 62, 63, 65, 66, 67, 68, 69, 70, 71 , 72, 73, 74 and 75 have Kd values of less than 0.1 micromolar and most of these compounds have Kd values of less than 0.01 micromolar. EXAMPLE 91 Anti-proliferative Activity
The anti-proliferative activities of compounds of the invention can be determined by measuring the ability of the compounds to inhibition of cell growth in a number of cell lines such as the human colon cancer cell line HCT116. Inhibition of cell growth is measured using the Alamar Blue assay (Nociari, M. M, Shalev, A., Benias, P., Russo, C. Journal of Immunological Methods 1998, 213, 157-167). The method is based on the ability of viable cells to reduce resazurin to its fluorescent product resorufin. For each proliferation assay cells are plated onto 96 well plates and allowed to recover for 16 hours prior to the addition of inhibitor compounds for a further 72 hours. At the end of the incubation period 10% (v/v) Alamar Blue is added and incubated for a further 6 hours prior to determination of fluorescent product at 535nM ex / 59OnM em. In the case of the non-proliferating cell assay cells are maintained at confluence for 96 hour prior to the addition of inhibitor compounds for a further 72 hours. The number of viable cells is determined by Alamar Blue assay as before. Cell lines can be obtained from the ECACC (European Collection of cell Cultures). The phenolic compounds of examples 5, 12, 13, 14, 17, 18, 19, 21 , 22, 23, 25, 28, 29, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 45, 48, 49, 50, 51 , 52, 53, 54, 55, 59, 60, 61 , 62, 63, 65, 66, 67, 68, 69, 70, 71 , 72, 74 and 75 were tested and were found to have IC50 values of less than 1 micromolar against the HCT116 cell line.
The parent compound of the pro-drugs of Examples 87, 88 and 89, i.e. (2,4-dihydroxy-5-isopropyl- phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1 ,3-dihydro-isoindol-2-yl]-methanone, in the form of its L-lactate salt, was was tested in anti-proliferative assays against one hundred cell lines by Oncodesign (Dijon, France). The IC50 values against each cell line are set out in the table below and the figures in the table refer to nanomolar concentrations. The compounds were tested up to a concentration of 10,000 nanomolar.
Figure imgf000213_0001
Figure imgf000214_0001
Figure imgf000215_0001
Figure imgf000216_0001
The results demonstrate that (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1- ylmethyl)-1 ,3-dihydro-isoindol-2-yl]-methanone has potent anti-proliferative activity against a wide range of different cell lines.
PHARMACEUTICAL FORMULATIONS EXAMPLE 92 (i) Tablet Formulation
A tablet composition containing a compound of the formula (I) is prepared by mixing 50 mg of the compound with 197 mg of lactose (BP) as diluent, and 3 mg magnesium stearate as a lubricant and compressing to form a tablet in known manner. (ii) Capsule Formulation
A capsule formulation is prepared by mixing 100 mg of a compound of the formula (I) with 100 mg lactose and filling the resulting mixture into standard opaque hard gelatin capsules.
(iii) Injectable Formulation I
A parenteral composition for administration by injection can be prepared by dissolving a compound of the formula (I) (e.g. in a salt form) in water containing 10% propylene glycol to give a concentration of active compound of 1.5 % by weight. The solution is then sterilised by filtration, filled into an ampoule and sealed.
(iv) Injectable Formulation Il
A parenteral composition for injection is prepared by dissolving in water a compound of the formula (I) (e.g. in salt form) (2 mg/ml) and mannitol (50 mg/ml), sterile filtering the solution and filling into sealable 1 ml vials or ampoules. v) Injectable formulation III
A formulation for i.v. delivery by injection or infusion can be prepared by dissolving the compound of formula (I) (e.g. in a salt form) in water at 20 mg/ml. The vial is then sealed and sterilised by autoclaving. vi) Injectable formulation IV
A formulation for i.v. delivery by injection or infusion can be prepared by dissolving the compound of formula (I) (e.g. in a salt form) in water containing a buffer (e.g. 0.2 M acetate pH 4.6) at 20mg/ml. The vial is then sealed and sterilised by autoclaving. (vii) Subcutaneous Injection Formulation
A composition for sub-cutaneous administration is prepared by mixing a compound of the formula (I) with pharmaceutical grade corn oil to give a concentration of 5 mg/ml. The composition is sterilised and filled into a suitable container. viii) Lvophilised formulation Aliquots of formulated compound of formula (I) are put into 50 ml vials and lyophilized. During lyophilisation, the compositions are frozen using a one-step freezing protocol at (-45 0C). The temperature is raised to -10 0C for annealing, then lowered to freezing at -45 0C, followed by primary drying at +25 0C for approximately 3400 minutes, followed by a secondary drying with increased steps if temperature to 50 0C. The pressure during primary and secondary drying is set at 80 millitor.
(ix) 2% Topical Gel Formulation
% w/w
Compound 2.00
Hydroxypropyl Methyl cellulose (Methocel F4M) 2.50
Polyethyleneoxide (Polyox WSR -205) 0.25
Propylene glycol 10.00
Methylparaben 0.15
Propylparaben 0.05
Purified Water to 100.00
EXAMPLE 93
Plasma Stability Studies
The ease with which pro-drug compounds of the invention can be converted to the active moiety can be assessed by measuring the in vitro plasma stability of the compounds. The method is based on the ability of plasma enzymes to metabolise compounds to the active moiety. Compounds are added to plasma and incubated at 370C for approximately 1 hour. Aliquots of plasma are removed at timed intervals and the compound extracted by the addition of acetonitrile containing an internal standard. All extracts are then analysed for parent compound, the active moiety and the internal standard using a compound specific LCMS-MS assay. Stability is measured by comparison of the peak area ratio of the parent compound and internal standard at time zero with the peak area ratio of the parent compound and internal standard in the incubated samples. Generation of the active moiety in samples is also assessed.
The compounds of examples 84, 85 and 86 were tested in mouse plasma and were found to have a range of stabilities and all formed the active moiety, (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4- methyl-piperazin-1-yl)-1 ,3-dihydro-isoindol-2-yl]-methanone.
EXAMLPE 94 Determination of Antifungal Activity
The antifungal activity of the compounds of the formula (I) is determined using the following protocol.
The compounds are tested against a panel of fungi including Candida parapsilosis, Candida tropicalis, Candida a/ό/cans-ATCC 36082 and Cryptococcus neoformans. The test organisms are maintained on Sabourahd Dextrose Agar slants at 4 0C. Singlet suspensions of each organism are prepared by growing the yeast overnight at 27 0C on a rotating drum in yeast-nitrogen base broth (YNB) with amino acids (Difco, Detroit, Mich.), pH 7.0 with 0.05 morpholine propanesulphonic acid (MOPS). The suspension is then centrifuged and washed twice with 0.85% NaCI before sonicating the washed cell suspension for 4 seconds (Branson Sonifier, model 350, Danbury, Conn.). The singlet blastospores are counted in a haemocytometer and adjusted to the desired concentration in 0.85% NaCI.
The activity of the test compounds is determined using a modification of a broth microdilution technique. Test compounds are diluted in DMSO to a 1.0 mg/ml ratio then diluted to 64 μg/ml in YNB broth, pH 7.0 with MOPS (Fluconazole is used as the control) to provide a working solution of each compound. Using a 96-well plate, wells 1 and 3 through 12 are prepared with YNB broth, ten fold dilutions of the compound solution are made in wells 2 to 11 (concentration ranges are 64 to 0.125 μg/ml). Well 1 serves as a sterility control and blank for the spectrophotometric assays. Well 12 serves as a growth control. The microtitre plates are inoculated with 10 μl in each of well 2 to 11 (final inoculum size is 104 organisms/ml). Inoculated plates are incubated for 48 hours at 35 0C. The MIC values are determined spectrophotometrically by measuring the absorbance at 420 nm (Automatic Microplate Reader, DuPont Instruments, Wilmington, Del.) after agitation of the plates for 2 minutes with a vortex-mixer (Vorte-Genie 2 Mixer, Scientific Industries, Inc., Bolemia, N.Y.). The MlC endpoint is defined as the lowest drug concentration exhibiting approximately 50% (or more) reduction of the growth compared with the control well. With the turbidity assay this is defined as the lowest drug concentration at which turbidity in the well is <50%of the control (IC50). Minimal Cytolytic Concentrations (MCC) are determined by sub-culturing all wells from the 96-well plate onto a Sabourahd Dextrose Agar (SDA) plate, incubating for 1 to 2 days at 35 0C and then checking viability.
Methods Of Testing For Pain Reducing Or Pain Preventing Activity (I) Inflammatory hyperalgesia test
Mechanical hyperalgesia can be examined in a rat model of inflammatory pain. Paw withdrawal thresholds to an increasing pressure stimulus are measured by the Randal-Sellito technique using an analgesymeter (Ugo Basile, Milan), in naϊve animals prior to an intraplantar injection of complete Freund's complete adjuvant (FCA) into the left hind paw. 24 h later paw withdrawal thresholds are measured again prior to (predose) and then from 10 min to 6 h following drug or vehicle administration. Reversal of hyperalgesia in the ipsilateral paw is calculated according to the formula:
Figure imgf000219_0001
(ii) Neuropathic hyperalgesia test Mechanical hyperalgesia can be examined in a rat model of neuropathic pain induced by partial ligation of the left sciatic nerve. Approximately 14 days following surgery mechanical withdrawal thresholds of both the ligated (ipsilateral) and non-ligated (contralateral) paw are measured prior to (predose) and then from 10 min to 6 h following drug or vehicle administration. Reversal of hyperalgesia at each time point is calculated according to the formula:
Figure imgf000220_0001
All experiments are carried out using groups of 6 animals. Stock concentrations of drugs are dissolved in distilled water and subsequent dilutions were made in 0.9% saline for subcutaneous administration in a volume of 4 rnlkg"1. All drugs are made up in plastic vials and kept in the dark.
Statistical analysis are carried out on withdrawal threshold readings (g) using ANOVA with repeated measures followed by Tukey's HSD test. Efficacy refers to the maximal reversal of hyperalgesia observed at the doses used.
(iii) Testing the effects of compounds of formula (0) a Rat Model of Bone Cancer Pain
Adult female rats are given intra-tibial injections of MRMZ-1 rat mammary gland carcinoma cells (3μl, 107 cells/ml). The animals typically gradually develop mechanical hyperalgesia, mechanical allodynia (skin sensitivity to non-noxious stimuli) and hind limb sparing, beginning on day 12-14 following cell injection. A compound of formula (0) (e.g. at a dose of 10 and 30 μg/kg s.c.) is administered 3 times a week from the day of cell injection, and the extent of inhibition of hind limb sparing and mechanical allodynia is determined in comparison to vehicle-treated controls.
Equivalents
The foregoing examples are presented for the purpose of illustrating the invention and should not be construed as imposing any limitation on the scope of the invention. It will readily be apparent that numerous modifications and alterations may be made to the specific embodiments of the invention described above and illustrated in the examples without departing from the principles underlying the invention. All such modifications and alterations are intended to be embraced by this application.

Claims

1. A compound for use in medicine, the compound being a compound of the formula (Vl0):
Figure imgf000221_0001
or a salt, solvate, tautomer or N-oxide thereof; wherein the bicyclic group:
Figure imgf000221_0002
is selected from the structures C1 , C5 and C6:
Figure imgf000221_0003
wherein n is 0, 1 , 2 or 3;
R1 is hydrogen, hydroxy, or O-Rz;
R2a is hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2a is O-Rz;
Rz is Lp-Rp1; SO3H; a glucuronide residue; a mono-, di- or tripeptide residue; or
Figure imgf000221_0004
Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or S(O)xNRp1; x is 1 or 2;
Rp1 is hydrogen or a C1-2S hydrocarbyl group containing O, 1 or 2 carbocyclic rings and 0, 1 , 2, 3, 4, 5 or 6 carbon-carbon multiple bonds, the C1-2S hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-C-i-6 alkylamino, C1-6 acylamino, halogen or C1^ alkoxy or C1-4 alkyl groups wherein the C1-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 3 non-terminal carbon atoms in the C1-2S hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(O)O or 0(C=O); provided that Rp1 is not hydrogen when Lp is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an 0-0 moiety; and excluding compounds wherein R1 is hydroxy and R2a is methoxy; Rp2 and Rp3 are the same or different and each is a group Rp1;
R3 is selected from hydrogen; halogen; cyano; C1-5 hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and C1-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C1-2 alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members;
R4a is selected from hydrogen, fluorine, chlorine and methoxy;
R8 is selected from hydrogen and fluorine; and
R10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or di-C1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group Ra-Rb; wherein:
Ra is a bond, O, CO, X1C(X2), C(X2)X1, X1C(X2)X1, S, SO, SO2, NR0, SO2NR0 or NR0SO2; and
Rb is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C1-12 hydrocarbyl (such as Ci--I0 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino (e.g. mono- or di-C1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C1-12 hydrocarbyl group (or C1--I0 hydrocarbyl group) may optionally be replaced by O, S, SO, SO2, NR0, X1C(X2), C(X2)X1 or
X1C(X2)X1;
R° is selected from Rb, hydrogen and C1-4 hydrocarbyl; A compound of the formula (Vl):
Figure imgf000223_0001
or a salt, solvate, tautomer or N-oxide thereof; wherein the bicyclic group:
Figure imgf000223_0002
is selected from the structures C1 , C5 and C6:
Figure imgf000223_0003
wherein n is 0, 1, 2 or 3;
R1 is hydrogen, hydroxy, or O-Rz;
R2ais hydroxy, methoxy or O-Rz; provided that at least one of R1 and R2a is O-Rz;
Rz is Lp-Rp1; SO3H; a glucuronide residue; a mono-, di- or tripeptide residue; or
Figure imgf000223_0004
.
Lp is a bond, C=O, (C=O)O, (C=O)NRp1 or S(O)xNRp1; x is 1 or 2;
Rp1 is hydrogen or a Ci-25 hydrocarbyl group containing 0, 1 or 2 carbocyclic rings and 0, 1 ,
2, 3, 4, 5 or 6 carbon-carbon multiple bonds, the Ci-2s hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-C1-6 alkylamino, Ci-6 acylamino, halogen or Ci-4 alkoxy or C1-4 alkyl groups wherein the Ci-4 alkoxy or C1-4 alky! groups are each optionally substituted by two or more fluorine atoms; and wherein up to 3 non-terminal carbon atoms in the C1-2S hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O); provided that Rp1 is not hydrogen when Lp is a bond, C=O or (C=O)O; and provided also that O-Rz does not contain an 0-0 moiety; and excluding compounds wherein R1 is hydroxy and R2a is methoxy;
Rp2 and Rp3 are the same or different and each is a group Rp1;
R3 is selected from hydrogen; halogen; cyano; C1-5 hydrocarbyl and C1-5 hydrocarbyloxy; wherein the C1-5 hydrocarbyl and C1-5 hydrocarbyloxy moieties are each optionally substituted by one or more substituents selected from hydroxy, halogen, C1-2 alkoxy, amino, mono- and di-C^ alkylamino, and aryl and heteroaryl groups of 5 to 12 ring members;
R4a is selected from hydrogen, fluorine, chlorine and methoxy; R8 is selected from hydrogen and fluorine; and
R10 is selected from: halogen; hydroxy; trifluoromethyl; cyano; nitro; carboxy; amino; mono- or Cu-G1-4 hydrocarbylamino; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and a group Ra-Rb; wherein:
Ra is a bond, O, CO, X1C(X2), C(X2)X1, X1C(X2)X1, S, SO, SO2, NRC, SO2NR0 or NR0SO2; and
Rb is selected from hydrogen; carbocyclic and heterocyclic groups having from 3 to 12 ring members; and C1-12 hydrocarbyl (such as C1-10 hydrocarbyl) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino (e.g. mono- or di-C1-4 hydrocarbylamino), and carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the C1-12 hydrocarbyl group (or C1-10 hydrocarbyl group) may optionally be replaced by O, S, SO, SO2, NR0, X1C(X2), C(X2)X1 or X1C(X2)X1; R° is selected from Rb, hydrogen and C1-4 hydrocarbyl; and X1 is O, S or NR0 and X2 is =0, =S or =NR°.
3. A compound according to claim 1 or claim 2 wherein R1 is hydroxy or O-Rz and R2a is hydroxy or O-R1p; provided that at least one of R1 and R2a is O-Rz.
4. A compound according to claim 3 wherein R1 and R2a are both O-Rz.
5. A compound according to any one of the preceding claims wherein Rp1 is hydrogen or a C-ι-25 hydrocarbyl group containing 0 or 1 carbocyclic rings and 0, 1 , 2 or 3 carbon-carbon multiple bonds, the C1-2S hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-C1-6 alkylamino, C1-6 acylamino, halogen or C14 alkoxy or C14 alkyl groups wherein the C1-4 alkoxy or Ci-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 2 non-adjacent non-terminal carbon atoms in the C1-2S hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O).
6. A compound according to claim 5 wherein Rp1 is hydrogen or a C1-8 hydrocarbyl group containing 0 or 1 carbocyclic rings and 0, 1 , 2 or 3 carbon-carbon multiple bonds, the C1-8 hydrocarbyl group being optionally substituted by hydroxy, amino, mono- or di-C1-6 alkylamino, C1-6 acylamino, halogen or C1-4 alkoxy or Ci-4 alkyl groups wherein the C1-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms; and wherein up to 2 non-adjacent non-terminal carbon atoms in the C1-25 hydrocarbyl group may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms may optionally be replaced by C(=O)O or 0(C=O).
7. A compound according to claim 6 wherein Rp1 is hydrogen, a C1-6 alkyl group, a C3-6 cycloalkyl group or a benzyl group wherein the phenyl and benzyl groups are optionally substituted by halogen or C1-4 alkoxy or Ci-4 alkyl groups wherein the C1-4 alkoxy or C1-4 alkyl groups are each optionally substituted by two or more fluorine atoms.
8. A compound according to any one of claims 1 to 5 wherein Rp1 is a C1-10 alkyl group wherein up to 2 non-adjacent non-terminal carbon atoms thereof may optionally be replaced by oxygen, and up to one pair of adjacent non-terminal carbon atoms thereof may optionally be replaced by C(=O)O or 0(C=O).
9. A compound according to any one of claims 1 to 8 wherein Lp is a bond.
10. A compound according to claim 9 wherein Rp1 is a C1-4 alkyl group or a C1-6 alkanoyloxy- C1-2-alkyl group.
11. A compound according to claim 9 wherein Rp1 is a C2-4 alkyl group or a C1-6 alkanoyloxy- C1-2-alkyl group.
12. A compound according to claim 11 wherein Rp1 is a C1-6 alkanoyloxymethyl group or a C1-6 alkanoyloxyethyl group.
13. A compound according to claim 11 or claim 12 wherein the C1-6 alkanoyloxy moiety of the Ci-6 alkanoyloxy-C1-2-alkyl group is acetoxy, propanoyloxy, π-butanoyloxy, 2- methylpropanoyloxy, 3-methylbutanoyloxy or pivaloyloxy.
14. A compound according to any one of claims 1 to 8 wherein Lp is C=O.
15. A compound according to claim 14 wherein Rp1 is C1-6 alkyl optionally substituted by mono- or di-C1-6 alkylamino or a C1-6 acylamino group.
16. A compound according to claim 15 wherein Rp1 is C1.4 alkyl (e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyi or ferf-butyl group) optionally substituted by mono- or CIi-C1-2 alkylamino or a C1-4 acylamino group.
17. A compound according to any one of claims 1 to 8 wherein Lp is (C=O)O.
18. A compound according to claim 17 wherein Rp1 is a C1-6 alkyl group, e.g. a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyi or te/f-butyl group.
19. A compound according to any one of claims 1 to 8 wherein Lp is (C=O)NRp1 and hence the moiety Lp-NRp1 is a group (C=O)N(Rp1)2.
20. A compound according to claim 19 wherein the two groups Rp1 are the same or different and are each hydrogen or a C1-6 alkyl group (e.g. a methyl, ethyl, n-propyl, isopropyl, n- butyl, isobutyi or fe/f-butyl group) or a C3-6 cyclic group such as cyclopropyl, cyclobutyl, cyclopentyl, cylcohexyl or phenyl group.
21. A compound according to claim 20 wherein both Rp1 groups are C1-4 alkyl groups.
22. A compound according to claim 21 wherein both Rp1 groups are methyl groups.
23. A compound according to any one of claims 1 to 8 wherein l_p is SOmNRp1 and hence the moiety Lp-NRp1 is a group SOmN(Rp1)2.
24. A compound according to claim 23 wherein the two groups Rp1 are the same or different and each is hydrogen or a C1-6 alkyl group (e.g. a methyl, ethyl, n-propyl, isopropyl, n- butyl, isobutyi or fe/t-butyl group) or a C3.6 cyclic group such as cyclopropyl, cyclobutyl, cyclopentyl, cylcohexyl or phenyl group.
25. A compound according to claim 24 wherein both Rp1 groups are C1-4 alkyl groups.
26. A compound according to claim 25 wherein both Rp1 groups are methyl groups.
27. A compound according to any one of claims 23 to 26 wherein m is 1.
28. A compound according to any one of claims 23 to 26 wherein m is 2.
29. A compound according to any one of claims 1 to 8 wherein Rz is:
Figure imgf000227_0001
wherein Rp2 and Rp3 are the same or different and each is a group Rp1 as defined in any one of claims 1 to 8.
30. A compound according to claim 29 wherein Rp1 is hydrogen or a Ci-4 alkyl group such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or ferf-butyl.
31. A compound according to claim 30 wherein Rp2 and Rp3 are both hydrogen.
32. A compound according to claim 30 wherein Rp2 is hydrogen and Rp3 is methyl, ethyl, n- propyl, isopropyl, n-butyl, isobutyl or ferf-butyl.
33. A compound according to claim 32 wherein Rp2 and Rp3 are both independently selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl and ferf-butyl groups.
34. A compound according to claim 33 wherein Rp2 and Rp3 are the same or different.
35. A compound according to any one of claims 1 to 8 wherein at least one of R1 and R2a is a group O-Rz where R2 is a glucuronide residue.
36. A compound according to claim 35 wherein R1 is hydroxy and R2a is a group O-Rz where Rz is a glucuronide residue.
37. A compound according to claim 35 wherein R2a is hydroxy and R1 is a group O-Rz where Rz is a glucuronide residue.
38. A compound according to any one of claims 1 to 8 wherein Rz is a mono-, di- or tripeptide residue.
39. A compound according to claim 1 or claim 2 wherein Rz is SO3H.
40. A compound according to claim 39 wherein R1 is OSO3H and R2a is hydroxy.
41. A compound according to claim 39 wherein R2a is OSO3H and R1 is hydroxy.
42. A compound according to claim 39 wherein R1 and R2a are both OSO3H.
43. A compound according to any one of the preceding claims wherein R3 is selected from hydrogen, chlorine, C1-5 hydrocarbyl and C1-5 hydrocarbyloxy.
44. A compound according to claim 43 wherein R3 is selected from hydrogen, C1-5 alkyl, C2.5 alkenyl and C3.4 cycloalkyl groups, preferably secondary alkyl and alkenyl groups such as isopropyl, sec-butyl, te/t-butyl, 1 ,2-dimethylallyl and 1 ,2-dimethylpropyl, or cycloalkyl groups such as cyclopropyl.
45. A compound according to claim 44 wherein R3 is selected from hydrogen, isopropyl and terf-butyl.
46. A compound according to any one of the preceding claims wherein R4 is hydrogen.
47. A compound according to any one of the preceding claims wherein R8 is hydrogen.
48. A compound according to any one of the preceding claims wherein R10 is selected from a group R1Oa consisting of halogen, hydroxy, amino and a group Ra-Rb where Ra is selected from a bond, O, CO, C(O)O, C(O)NR0, NR0C(O), NR0C(O)O, NR0, SO, SO2, SONR0, and SO2NR0; and Rb is selected from hydrogen; carbocyclic and heterocyclic groups having 5 or 6 ring members; and Ci-10 hydrocarbyl (e.g. Ci-8 hydrocarbyl such as C1-8 alkyl or C3-7 cycloalkyl) optionally substituted by one or more substituents selected from hydroxy, oxo, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino (e.g. mono- or Oi-C1-4 hydrocarbylamino), carboxy, and carbocyclic and heterocyclic groups having from 3 to 7 ring members, and wherein one or more of the carbon atoms of the C1-8 hydrocarbyl group may optionally be replaced by O, S, C(O)O, C(O)NR0 or NR0.
49. A compound according to claim 48 wherein wherein R10 is selected from a group R1Oaa consisting of halogen, hydroxy, amino and a group Ra-Rb where Ra is selected from a bond, O, CO, C(O)O, C(O)NR0, NR0C(O), NR0C(O)O, NR0, SO, SO2, SONR0, and SO2NR0; and Rb is selected from hydrogen; carbocyclic and heterocyclic groups having 5 or 6 ring members; and non-aromatic C1-10 hydrocarbyl optionally substituted by one or more substituents selected from hydroxy, oxo, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino, carboxy, and carbocyclic and heterocyclic groups having from 3 to 7 ring members, and wherein one or more of the carbon atoms of the C1-8 hydrocarbyl group may optionally be replaced by O, S, C(O)O, NR0C(O), C(O)NR0 or NR0
50. A compound according to claim 49 wherein n is O, 1 or 2 and R1Oaa is selected from fluorine, chlorine, hydroxy, amino and a group Ra-Rb;
Ra is selected from a bond, O, CO, C(O)NR0, NR0C(O), NR0C(O)O and NR0; Rb is selected from: ■ hydrogen;
■ carbocyclic and heterocyclic groups having 5 or 6 ring members and containing O, 1 or 2 heteroatoms selected from O, N and S; and
■ non-aromatic C1-I0 hydrocarbyl optionally substituted by one or more substituents selected from hydroxy, oxo, amino, mono- or di-C1-8 non-aromatic hydrocarbylamino, carboxy, and carbocyclic and heterocyclic groups having from 3 to 7 ring members and containing 0, 1 or 2 heteroatoms selected from O, N and S; and wherein one or more of the carbon atoms of the C1-8 hydrocarbyl group may optionally be replaced by O, S, C(O)O, NR0C(O), C(O)NR0 or NRC.
51 . A compound according to claim 50 wherein Rb is an optionally substituted non-aromatic carbocyclic and heterocyclic group having 5 or 6 ring members and containing O, 1 or 2 heteroatoms selected from O, N and S.
52. A compound according to any one of claims 1 to 47 wherein R10 is selected from a group R1Ob consisting of halogen, OH, NH2, CH2OH, CH2NH2, 0-C|.6-alkyl, NH-C1-6 alkyl, aryl, heteroaryl, C3-7 cycloalkyl, heterocyclyl, O-heteroaryl, O- C3.7 cycloalkyl, O- heterocycloalkyl, C(=O)C1-6 alkyl, C(=O)OC1.6 alkyl, C(=O)NH2, C(=O)NHC1-6 alkyl,
C(O)N(C1-6 alkyl)2, NH(C1-6 alkyl), N(C1-6 alkyl)2, NC(=O)C1-6 alkyl, C6 aryl, OC6 aryl, C(=O)C6aryl, C(=O)OC6aryl, C(=O)NH2, C(=O)NHC6aryl, C(=O)N(C6 aryl)2, NH(C6 aryl), N(C6 aryl)2, NC(=O)C6 aryl, C5.5 heterocyclyl, OC5.6 heterocyclyl, C(=O)C5.6 heterocyclyl, C(=O)OC5.6 heterocyclyl, C(=O)NHC5.6 heterocyclyl, C(=O)N(C5-6 heterocyclyl)2, NH(C5-6 heterocyclyl), N(C6-6 heterocyclyl)2, NC(=O)C5-6 heterocyclyl, C(=0)NHC,.6 alkyl, C5-6 aryl,
S(O)C1-6 alkyl, S(=O)N-C1-6 alkyl and SO2N-C1-6 alkyl; and a group [sol], CH2[sol] or OCH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000229_0001
and wherein (i) R10b is optionally further selected from a group OCH2CH2CH2[SoI] and/or
(ii) [sol] is further selected from NHR11 wherein R11 is COR12 or R12 and R12 is C1-4 alkyl, aryl or aryl-^.4 alkyl.
53. A compound according to any one of claims 1 to 47 wherein R10 is selected from a group
RRm100 wwhheerree RR11OOcc i is a group [sol], CH2[SoI] or OCH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000230_0001
54. A compound according to any one of claims 1 to 47 wherein R10 is selected from a group
R1Occ consisting of: halogen;
CO2R14 wherein R14 is hydrogen or C1-6 alkyl; C-I-4 alkyl optionally substituted by hydroxy or C1-2 alkoxy; C1-4 alkoxy optionally substituted by hydroxy or C1-2 alkoxy; or a group [sol], CH2[SoI], C(O)[sol], OCH2CH2[SoI] or OCH2CH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000230_0002
11 13
-NHR11 — O-(CH2)-NHR — O-(CH2)-OR wherein X4 is NH or O, m is 0 or 1 , n is 1 , 2 or 3, R11 is hydrogen, COR12, C(O)OR12 or R12; R12 is C1-6 alkyl, C3-6 cycloalkyl, aryl, aryl-C^ alkyl or CH2R15; and R15 is selected from hydrogen, C1-6 alkyl, C3-6 cycloalkyl, hydroxy-C1-s alkyl, piperidine, N-C1-6 alkylpiperazine, piperazine, morpholine, COR13 or C(O)OR13; and R13 is C1-6 alkyl.
55. A compound according to any one of the preceding claims wherein R3 is selected from a group R3d consisting of ethyl and secondary and tertiary alkyl groups of 3 to 6 carbon atoms; provided that when R1 and R2 are both hydroxy, then R3d can additionally be selected from hydrogen.
56. A compound according to claim 55 wherein the secondary and tertiary alkyl groups are selected from isopropyl and tert-butyl.
57. A compound according to any one of the preceding claims having the formula (VII):
Figure imgf000231_0001
wherein R1, R2a, R3 R4a, R8and R10b are as defined in any one of the preceding claims and n is O, 1 2 or 3 (more preferably 0, 1 or 2, e.g. O or 1), and provided that at least one of R1 and R2a is O-Rz.
58. A compound according to 57 wherein both R1 and R2a are O-Rz.
59. A compound according to claim 1 of the formula (Vila):
Figure imgf000231_0002
(Vila) wherein R3 is selected from hydrogen, halogen, C1-5 alkyl, C2-5 alkenyl and C3.4 cycloalkyl groups; R4a is selected from hydrogen, fluorine, chlorine and methoxy; R8 is hydrogen or fluorine; n is 0, 1 2 or 3;
R1 and R2a are as defined in any one of the preceding claims; R10 is selected from halogen, hydroxy, trifluoromethyl, cyano, nitro, carboxy, amino, mono- or di-C-,.4 hydrocarbylamino, carbocyclic and heterocyclic groups having from 3 to 12 ring members; a group Ra-Rb wherein Ra is a bond, O, CO, X1C(X2), C(X2)X1, X1C(X2)X1, S, SO, SO2, NRC, SO2NR0 or NR0SO2; and Rb is selected from hydrogen, carbocyclic and heterocyclic groups having from 3 to 12 ring members, and a C1-12 hydrocarbyl group (such as a C1-10 hydrocarbyl group) optionally substituted by one or more substituents selected from hydroxy, oxo, halogen, cyano, nitro, carboxy, amino, mono- or di-C1-8 non- aromatic hydrocarbylamino (e.g. mono- or di-C1-4 hydrocarbylamino), carbocyclic and heterocyclic groups having from 3 to 12 ring members and wherein one or more carbon atoms of the Ci-12 hydrocarbyl group (or C1-10 hydrocarbyl group) may optionally be replaced by O, S, SO, SO2, NR0, X1C(X2), C(X2)X1 or X1C(X2)X1;
R° is selected from Rb, hydrogen and C1-4 hydrocarbyl; and X1 is O, S or NR0 and X2 is =0, =S or =NR°.
60. A compound according to claim 59 wherein R10 is a group R1Oa or or R1Oaa or R10b or R10oor R1Occ as defined in any one of the preceding claims.
61. A compound according to claim 60 which is represented by the formula (VIIb):
Figure imgf000232_0001
wherein R1 and R2a are as defined in any one of the preceding claims;
R3 is selected from hydrogen, halogen, C1-5 alkyl, C2-5 alkenyl and C3-4 cycloalkyl groups;
R4a is selected from hydrogen, fluorine, chlorine and methoxy; R8 is hydrogen or fluorine; n is 0, 1 2 or 3; and R1Occ is selected from: halogen;
CO2R14 wherein R14 is hydrogen or C1-6 alkyl;
Ci-4 alkyl optionally substituted by hydroxy or Ci-2 alkoxy;
Ci-4 alkoxy optionally substituted by hydroxy or C1-2 alkoxy; or a group [sol], CH2[SOl], C(O)[SoI], OCH2CH2[SoI] or OCH2CH2CH2[SoI] where [sol] is selected from the following groups
Figure imgf000233_0001
wherein X4 is NH or O, m is O or 1 , n is 1 , 2 or 3, R11 is hydrogen, COR12, C(O)OR12 or R12; R12 is C1-6 alkyl, C3-5 cycloalkyl, aryl, aryl-C1-6 alkyl or CH2R15; and R15 is selected from hydrogen, C1-6 alkyl, C3-6 cycloalkyl, hydroxy-C^ alkyl, piperidine, N-C1-6 alkylpiperazine, piperazine, morpholine, COR13 or C(O)OR13; and R13 is C1-6 alkyl.
62. A compound according to any one of claims 57 to 61 wherein R8 is hydrogen.
63. A compound according to any one of claims 57 to 62 wherein R4a is hydrogen.
64. A compound according to any one of claims 57 to 63 wherein R3 is selected from halogen, C1-5 alkyl, C2s alkenyl and C3-4 cycloalkyl groups.
65. A compound according to claim 64 wherein R3 is selected from chlorine, bromine, cyclopropyl and branched C3-5 alkyl.
66. A compound according to claim 65 wherein R3 is isopropyl or fe/t-butyl.
67. A compound according to claim 66 wherein R3 is isopropyl.
68. A compound according to any one of the preceding claims wherein n is 1 or 2 and R10 is selected from a group R1OCOC consisting of: a group [sol] or CH2[SoI] where [sol] is selected from the following groups:
Figure imgf000234_0001
wherein X4 is NH or O, m is 0 or 1 , n is 1 , 2 or 3, R11 is hydrogen, COR12, C(O)OR12 or R12; R12 is C1-6 alky), C3-6 cycloalkyl, aryl, aryl-C,.6 alkyl or CH2R15; and R15 is selected from hydrogen, Ci-6 alkyl, C3-6 cycloalkyl, hydroxy-C1-6 alkyl, piperidine, N-Ci-6 alkylpiperazine, piperazine, morpholine, COR13 or C(O)OR13; and R13 is C1-6 alkyl.
69. A compound according to any one of the preceding claims wherein n is 1 , 2 or 3.
70. A compound according to claim 68 wherein n is 1 or 2.
71. A compound according to claim 70 wherein n is 1.
72. A compound according to any one of claims 57 to 71 wherein the moieties:
Figure imgf000234_0002
can be any one of groups B8, B35, B36, B37, B38, B39, B40, B41 , B42, B43, B45, B46,
B48, B53, B54, B55, B56, B57, B58, B59, B60, B61 , B62, B71, B72, B74, B75, B76, B77, B78, B79, B80, B81 , B82, B83, B85, B86, B87, B93, B94, B95, B97, B98, B99, B100 and B101 as set forth in Table 2 herein.
73. A compound according to claim 1 having the formula (VIII):
Figure imgf000235_0001
wherein Q1 is a bond or a group CH2 and R1 and R2a are as defined in any one of the preceding claims.
74. A compound according to claim 73 wherein one of R1 and R2a is hydroxy, O-C(=O)-R20 or O-C(=O)-N(R20)2, and the other of R1 and R2a is O-C(=O)-R20 or O-C(=O)-N(R20)2 wherein
R20 is C1-4 alkyl.
75. A compound according to any one of claims 1 to 74 in the form of a salt, solvate or N- oxide.
76. A compound according to any one of claims 1 to 74 in the form of a salt or solvate.
77. A compound according to any one of claims 2 to 76 for use in medicine, for example for use as an inhibitor of Hsp90.
78. A compound according to any one of claims 1 to 76 for use in the prophylaxis or treatment of a disease state or condition mediated by Hsp90.
79. A method for the prophylaxis or treatment of a disease state or condition mediated by Hsp90, which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76.
80. A compound according to any one of claims 1 to 76 for use in alleviating or reducing the incidence of a disease state or condition mediated by Hsp90
81. A method for alleviating or reducing the incidence of a disease state or condition mediated by Hsp90, which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76.
82. A compound according to any one of claims 1 to 76 for use in treating a disease or condition comprising or arising from abnormal cell growth in a mammal.
83. A method for treating a disease or condition comprising or arising from abnormal cell growth in a mammal, which method comprises administering to the mammal a compound according to any one of claims 1 to 76 in an amount effective in inhibiting abnormal cell growth.
84. A compound according to any one of claims 1 to 76 for use in alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal.
85. A method for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal, which method comprises administering to the mammal a compound according to any one of claims 1 to 76 in an amount effective in inhibiting abnormal cell growth.
86. A method for treating a disease or condition comprising or arising from abnormal cell growth in a mammal, the method comprising administering to the mammal a compound according to any one of claims 1 to 76 in an amount effective to inhibit Hsp90 activity.
87. A method for alleviating or reducing the incidence of a disease or condition comprising or arising from abnormal cell growth in a mammal, the method comprising administering to the mammal a compound according to any one of claims 1 to 76 in an amount effective to inhibit Hsp90 activity.
88. A method of inhibiting Hsp90, which method comprises contacting the Hsp90 with an HspΘO-inhibiting compound according to any one of claims 1 to 76.
89. A method of modulating a cellular process (for example cell division) by inhibiting the activity of Hsp90 using a compound according to any one of claims 1 to 76.
90. A compound according to any one of claims 1 to 76 for use in the prophylaxis or treatment of a disease state as described herein.
91. The use of a compound according to any one of claims 1 to 76 for the manufacture of a medicament, wherein the medicament is for any one or more of the uses defined herein.
92. A pharmaceutical composition comprising a compound according to any one of claims 1 to 76 and a pharmaceutically acceptable carrier.
93. A compound of the formula (I0), (I), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein
94. A pharmaceutical composition comprising a compound of the formula (I0), (I), (II), (III), (IV), (V), (Vl), (Via), (VII), (Vila), (VIIb) or (VIII) or any sub-groups or examples thereof as defined herein and a pharmaceutically acceptable carrier in a form suitable for oral administration.
95. A pharmaceutical composition comprising a compound according to any one of claims 1 to 76 and a pharmaceutically acceptable carrier in a form suitable for parenteral administration, for example by intravenous (i.v.) administration.
96. A pharmaceutical composition comprising a compound according to any one of claims 1 to 76 and a pharmaceutically acceptable carrier in a form suitable for intravenous (i.v.) administration by injection or infusion.
97. The use of a according to any one of claims 1 to 76 for the manufacture of a medicament for the prophylaxis or treatment of a fungal, protozoal or parasitic disease state or condition (other than a disease state or condition due to Plasmodium falciparum), for example a disease state or condition characterised by an antibody response to Hsp90.
98. A method for the prophylaxis or treatment of a fungal, protozoal or parasitic disease state or condition (other than a disease state or condition due to Plasmodium falciparum), for example a disease state or condition characterised by an antibody response to Hsp90, which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76.
99. The use of a compound according to any one of claims 1 to 76 for the manufacture of a medicament for coadministration with an anti-fungal agent, anti-protozoal agent or antiparasitic agent (preferably an anti-fungal agent) to prevent, reduce or reverse the development of resistance to the anti-fungal agent, anti-protozoal agent or anti-parasitic agent.
100. A method of preventing or reducing development of resistance to an anti-fungal agent in a patient (e.g. a human patient), which method comprises administering to the patient a combination of an anti-fungal agent, anti-protozoal agent or anti-parasitic agent (preferably an anti-fungal agent) and a compound according to any one of claims 1 to 76.
101. The use of a compound according to any one of claims 1 to 76 for the manufacture of a medicament for use in the reduction or elimination of pain in a patient (e.g. a mammal such as a human) suffering from pain.
102. The use of a compound according to any one of claims 1 to 76 for the manufacture of a medicament for the treatment of any one or more of nociception, somatic pain, visceral pain, acute pain, chronic pain, hyperalgesia, allodynia, post operative pain, pain due to hypersensivity, headache, inflammatory pain (rheumatic, dental, dysmenorrhoea or infection), neurological pain, musculoskeletal pain, cancer related pain or vascular pain.
103. A compound according to any one of claims 1 to 76 for use in treating any one or more of nociception, somatic pain, visceral pain, acute pain, chronic pain, hyperalgesia, allodynia, post operative pain, pain due to hypersensivity, headache, inflammatory pain (rheumatic, dental, dysmenorrhoea or infection), neurological pain, musculoskeletal pain, cancer related pain or vascular pain.
104. A method of treating pain in a patient such as a mammal (e.g. human), which method comprises administering to the patient a therapeutically effective amount of a compound according to any one of claims 1 to 76.
105. A method for the reduction or elimination of pain in a patient (e.g. a mammal such as a human) suffering from pain, which method comprises administering to the patient an effective pain-reducing or pain-eliminating amount of a compound according to any one of claims 1 to 76.
106. A method for the treatment of any one or more of nociception, somatic pain, visceral pain, acute pain, chronic pain, hyperalgesia, allodynia, post operative pain, pain due to hypersensivity, headache, inflammatory pain (rheumatic, dental, dysmenorrhoea or infection), neurological pain, musculoskeletal pain, cancer related pain or vascular pain, which method comprises administering to the patient a therapeutically effective amount of a compound according to any one of claims 1 to 76.
107. A method of preventing or reducing neuronal damage in a patient suffering from stroke, which method comprises administering to the patient an effective neuroprotective amount of a compound according to any one of claims 1 to 76.
108. The use of a compound according to any one of claims 1 to 76 for the manufacture of a medicament for the prevention or reduction of risk of stroke in patients at risk for stroke, for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation.
109. A compound according to any one of claims 1 to 76 for use in the prevention or reduction of risk of stroke in patients at risk for stroke, for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation.
110. A method for the prevention or reduction of risk of stroke in patients at risk for stroke, for example a patient exhibiting any one or more risk factors selected from vascular inflammation, atherosclerosis, arterial hypertension, diabetes, hyperlipidemia and atrial fibrillation, which method comprises administering to the patient an effective therapeutic amount of a compound according to any one of claims 1 to 76.
111. A compound according to any one of claims 1 to 76 for use in the prophylaxis or treatment of a viral infection (or viral disease).
112. The use of a compound according to any one of claims 1 to 76 for the manufacture of a medicament for the prophylaxis or treatment of a viral infection (or viral disease).
113. A method for the prophylaxis or treatment of a viral infection (or viral disease), which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76.
114. A compound according to any one of claims 1 to 76 for use in blocking or inhibiting viral replication in a host organism (e.g. an animal such as a mammal (e.g. human)).
115. The use of a compound according to any one of claims 1 to 76 for the manufacture of a medicament for use in blocking or inhibiting viral replication in a host organism (e.g. an animal such as a mammal (e.g. human)).
116. A method of blocking or inhibiting viral replication in a host organism (e.g. an animal such as a mammal (e.g. human)), which method comprises administering to the host organism a compound according to any one of claims 1 to 76.
117. A method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90, which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76, wherein the disease state or condition mediated by Hsp90 is the development of resistance to a cancer drug.
118. A method for: (i) sensitizing malignant cells to an anticancer drug; (ii) alleviating or reducing the incidence of resistance to an anticancer drug; (iii) reversing resistance to an anticancer drug; (iv) potentiating the activity of an anticancer drug; (v) delaying or preventing the onset of resistance to an anticancer drug, which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76.
119. A method for the treatment of a cancer which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76, which method is characterized by the absence of drug resistance.
120. A method for the prophylaxis or treatment (or alleviation or reduction of the incidence) of a disease state or condition mediated by Hsp90 in a subject undergoing treatment with a therapeutic agent (such as an anti-cancer agent), which method comprises administering to the subject a compound according to any one of claims 1 to 76, wherein the disease state or condition mediated by Hsp90 is the development of resistance to the said therapeutic agent.
121. A method for: (i) sensitizing malignant cells to an anti-cancer agent; (ii) alleviating or reducing the incidence of resistance to an anti-cancer agent; (iii) reversing resistance to an anti-cancer agent; (iv) potentiating the activity of an anti-cancer agent; (v) delaying or preventing the onset of resistance to an anti-cancer agent, which method comprises administering to a subject undergoing treatment with said anti-cancer agent a compound according to any one of claims 1 to 76.
122. A method for the treatment of a cancer in a subject undergoing treatment with an anticancer agent, which method comprises administering to a subject in need thereof a compound according to any one of claims 1 to 76, which method is characterized by the absence of drug resistance to the anti-cancer agent.
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