EA044393B1 - TETRAHYDROBENZOFURO[2,3-C]PYRIDINE AND BETA-CARBOLINE COMPOUNDS FOR THE TREATMENT, ALLIANCE, OR PREVENTION OF DISORDERS ASSOCIATED WITH TAU PROTEIN AGGREGATES - Google Patents

Description

Field of technology to which the invention relates

The invention relates to new compounds that can be used for the treatment, amelioration or prevention of a group of disorders and disorders associated with tau protein aggregates (associated with tubulin), including, but not limited to, neurofibrillary tangles (NFTs), e.g.

Alzheimer's (AD).

State of the art

Many age-related diseases are based on or associated with extracellular or intracellular deposits of amyloid or amyloid-like proteins, which contribute to the pathogenesis as well as disease progression. The best characterized amyloid protein that forms extracellular aggregates is amyloid beta protein (Ae). Other examples of amyloid proteins that form extracellular aggregates are prion, ATTR (transthyretin) or ADan (ADanPP). Amyloid-like proteins that form primarily intracellular aggregates include, but are not limited to, tau protein, alpha-synuclein, TAR DNA binding protein 43 (TDP-43), and huntingtin (htt). Diseases involving tau protein aggregates are typically referred to as tauopathies, such as AD.

Amyloid or amyloid-like deposits result from the misfolding of proteins followed by aggregation to form multiple beta sheets in which multiple peptides or proteins are held together by intermolecular hydrogen bonds. Although amyloid or amyloid-like proteins have different primary amino acid sequences, their deposits often contain many common molecular components, in particular β-sheet quaternary structures are present. The relationship between amyloid deposits and disease remains largely unclear. A variety of protein aggregates, including both those associated with and not associated with disease pathologies, have been found to be toxic, suggesting that common molecular features of amyloid are involved or responsible for disease (Bucciantini et al., Nature, 2002, 416, 507-11). A variety of β-sheet-aggregated peptide or protein multimers have also been associated with toxicity to a variety of peptides or proteins ranging from dimers to soluble low molecular weight oligomers, protofibrils, or insoluble fibrillar deposits.

Alzheimer's disease (AD) is a neurological disorder thought to be primarily caused by amyloid plaques, an extracellular accumulation of pathological deposits of Ae (amyloid-beta) aggregates in the brain. Other major neuropathological hallmarks of AD are intracellular neurofibrillary tangles (NFTs), which result from the aggregation of hyperphosphorylated tau, misfolded tau, or pathological tau and its conformers. AD shares its etiopathology with many neurodegenerative tauopathies, particularly certain types of frontotemporal dementia (FTD). Tau protein is a freely soluble, naturally unfolded protein that actively binds to microtubules (MTs), promoting their assembly and stability. MTs are of great importance for the integrity of the neuronal cytoskeleton and, therefore, for the proper formation and functioning of neural circuits, and therefore for learning and memory. The binding of tau to MTs is controlled by dynamic phosphorylation and dephosphorylation, as demonstrated primarily in vitro and in nonneuronal cells. In the brains of AD patients, tau protein pathology (tauopathy) develops later than amyloid pathology, but there is still debate as to whether the Ae protein is the causative agent of AD, which is the essence of the so-called amyloid cascade hypothesis (Hardy et al., Science 1992, 256, 184-185; Musiek et al., Nature Neurosciences 2015, 18(6), 800-806). The precise mechanisms that link amyloid to tau pathology remain largely unknown, but neuronal signaling pathways that act on or through GSK3 and cdk5 as major tau kinases have been proposed (Muyllaert et al., Rev. Neurol. (Paris), 2006, 162, 903-7; Muyllaert et al., Genes Brain and Behav. 2008, Suppl 1, 57-66). Even if tauopathy develops later than amyloid, it is not just an innocent side effect, but a major pathological culprit in AD. In experimental mouse models, cognitive defects caused by amyloid pathology are almost completely alleviated by the absence of tau protein (Roberson et al., Science, 2007, 316 (5825), 750-4), and the severity of cognitive dysfunction and dementia correlates with tauopathy rather than with amyloid pathology.

Diseases involving tau protein aggregates are typically referred to as tauopathies, and these include, but are not limited to, Alzheimer's disease (AD), familial AD, primary age-related tauopathy (PART), Creutzfeldt-Jakob disease, Boxer dementia, Down syndrome, Gerstmann-Straussler-Scheinker disease (GSS), inclusion body myositis, cerebral prion protein-induced amyloid angiopathy, traumatic brain injury (TBI), amyotrophic lateral sclerosis (ALS), parkinsonian dementia (Guam syndrome), non-Guam disease motor neurons with neurofibrillary tangles, disease characterized by the appearance of argyrophilic granules, corticobasal degeneration (CBD), diffuse neurofibrillary tangles with calcification, frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), Hallervorden-Spatz disease , multiple system atrophy (MSA), Niemann-Pick disease type C, pallidoponto-nigral degeneration, Pick disease (PiD), progressive subcortical gliosis, progressive supranuclear palsy (PSP), subacute sclerosing panencephalitis, tangle-predominant dementia, postencephalitic parkinsonism , myotonic dystrophy, subacute sclerosing panencephalitis, LRRK2 mutations, chronic traumatic encephalopathy (CTE), familial British dementia, familial Danish dementia, other frontotemporal lobar degenerations, Guadalupean parkinsonism, neurodegeneration with brain iron accumulation, SLC9A6-related mental retardation , white matter tauopathy with globular glial inclusions, epilepsy, dementia with Lewy bodies (LBD), mild cognitive impairment (MCI), multiple sclerosis, Parkinson's disease, HIV-associated dementia, adult-onset diabetes, senile cardiac amyloidosis, glaucoma, ischemic stroke, psychosis in AD and Huntington's disease.

(Williams et al., Intern. Med. J., 2006, 36, 652-60; Kovacs et al., J Neuropathol Exp Neurol. 2008; 67(10): 963975; Higuchi et al., Neuropsychopharmacology - 5th Generation of Progress, 2002, Section 9, Chapter 94: 1339-1354; Hilton et al., Acta Neuropathol. 1995;90(1):101-6; Iqbal et al., Biochimica et Biophysica Acta 1739 (2005) 198-210; McQuaid et al., Neuropathol Appl Neurobiol. 1994 Apr;20(2): 103-10; Vossel et al., Lancet Neurol 2017; 16: 311-22; Stephan et al., Molecular Psychiatry (2012) 17, 1056- 1076; Anderson et al., Brain (2008), 131, 1736-1748; Savica et al., JAMA Neurol. 2013;70(7):859-866; Brown et al. Molecular Neurodegeneration 2014, 9:40; El Khoury et al., Front. Cell. Neurosci., 2014, volume 8, article 22: 1-18; Tanskanen et al., Ann. Med. 2008;40(3):232-9; Gupta et al., CAN J OPHTHALMOL-VOL.43, No. 1, 2008: 53-60; Dickson et al., Int J Clin Exp Pathol 2010;3(1):1-23; Fernandez-Nogales et al., Nature Medicine, 20, 881885 (2014); Bi et al., Nature Communications volume 8, Article number: 473 (2017); Murray et al., Biol Psychiatry. 2014 April 1; 75(7): 542-552).

Of all the drugs in clinical trials for Alzheimer's disease in 2017, those targeting tau protein are very rare, representing only 8% of phase II clinical trials (Cummings et al., Alzheimer's & Dementia: Translational Research & Clinical Interventions 3 (2017) 367-384). Current therapeutic approaches that target tau include primarily antibody-based approaches with the major limitation of targeting only extracellular tau. Among small molecule approaches, several tau kinase inhibitors have been developed, although they are very challenging in terms of toxicity and specificity. However, currently only one kinase inhibitor, Nilotinib, is being tested in clinical trials. Finally, among the tau aggregation inhibitors, only one, LMTX, is currently in clinical trials (Cummings et al., 2017). Although tau-based therapies have received increasing attention in recent years, there is still a great need for additional therapeutics that target pathological tau conformers known or suspected to cause tauopathies.

WO2011/128455 relates to specific compounds that are useful for treating disorders associated with amyloid proteins or amyloid-like proteins.

Description of graphic materials

The figure illustrates the reduction of intracellular tau protein misfolding by immunocytochemistry in differentiated neuroblastoma cells using Example 44. Data are shown as mean + standard deviation.

The essence of the invention

The purpose of this invention was to provide compounds that can be used in the treatment, amelioration or prevention of a group of disorders and disorders associated with tau protein aggregates, including, but not limited to, NFTs, for example, Alzheimer's disease (AD). There is also a need in the art for compounds that can be used as therapeutic agents to: (a) reduce tau/NFT aggregates by recognizing the aggregated tau and disaggregating the tau, for example, by changing the molecular conformation of the tau aggregate -protein, and/or (b) preventing the formation of tau protein aggregates, and/or (c) intracellular interaction with tau protein aggregates. The inventors of the present invention have surprisingly discovered that these objectives can be achieved using compounds of formula (I) as described herein below.

The compounds of formula (I) demonstrate a high ability to reduce tau protein aggregates by recognizing aggregated tau protein and disaggregating tau protein, e.g.

- 2 044393 changes in the molecular conformation of the tau protein aggregate. Certain compounds of formula (I) prevent the formation of tau protein aggregates and/or interact intracellularly with tau protein aggregates. Without being bound by any theory, the compounds of formula (I) are believed to inhibit tau protein aggregation or disaggregate preformed tau protein aggregates, including when present intracellularly. Due to their unique structural features, these compounds exhibit properties such as appropriate lipophilicity and molecular weight, brain uptake and pharmacokinetics, cell permeability, solubility and metabolic stability in order to be a successful drug for the treatment, amelioration or prevention of tauopathies.

Ultrastructural analysis revealed that tau inclusions consist of paired helical filaments (PHF) or straight filaments (SF). High-resolution structural analysis revealed that these filaments consist of a central region containing amino acids 306–378 of tau, which adopts a transverse beta/beta helical structure. The compounds of this invention can recognize aggregated tau protein and disaggregate tau protein, for example, by changing the molecular conformation of the tau protein aggregate, and therefore can be expected to facilitate the clearance of tau protein.

In addition, it has been shown that tau protein is capable of both replicating from cell to cell and that certain forms of tau protein (acting as seeds) are capable of causing a structural change in the native tau protein in a healthy cell to undergo misfolding and aggregation. Aggregated tau protein is thought to be responsible for seeding and therefore spreading tau pathology. The compounds of this invention can interact intracellularly with aggregated tau protein and, therefore, can be expected to reduce the spread of tau protein pathology and ultimately prevent or reduce the associated cognitive deficits in AD.

The present invention discloses new compounds of formula (I) having the ability to reduce tau protein aggregates, recognize aggregated tau protein, and disaggregate tau protein, for example, by changing the molecular conformation of the tau protein aggregate.

The present invention discloses certain new compounds of formula (I) having the ability to prevent the formation of tau protein aggregates and/or interact intracellularly with tau protein aggregates.

The present invention provides methods for treating disorders and disorders associated with tau protein aggregates, including, but not limited to, NFTs, using a compound of formula (I) or a pharmaceutical composition thereof. The present invention further provides a pharmaceutical composition comprising a compound of formula (I) and a pharmaceutically acceptable carrier or excipient.

The present invention is summarized in the following paragraphs. 1. Compound of formula (Ia)

^Rd

R ^a

or its stereoisomers, racemic mixtures, tautomers, pharmaceutically acceptable salts;

- 3 044393 to the N atom in any accessible position and, at the same time, more substituents R ^j ;

B represents NR ^a ;

replaced by one or

E and V are independently selected from the group consisting of N, NR ⁵ , O and S;

G is selected from the group consisting of a benzene ring and a pyridine ring;

J is selected from the group consisting of O, NR ¹ and CH2 or J is selected from the group consisting of CH or C if J is attached to R ² ;

Y, Y ¹ , Y ² and Y ³ represent CZ;

Z is independently selected from the group consisting of H, halogen, O-(C ₁ -C ₆ alkyl), C1-C6 alkyl and CN;

(R ² )n -N O

R is independently selected from the group consisting of and -NR ³ R ⁴ ;

R ^a is selected from the group consisting of H and C1-C6 alkyl;

R ^d , R ^e , R ^f and R ^g are H or one of R ^d and R ^e , and one of R ^f , R ^g can be joined to form a 5-7 membered ring;

where when A represents

R ^j is independently selected (R ² )n ^a /-k ^b from the group consisting of -halogen, -O-(C1-C ₆ alkyl), -NR ³ R ⁴ , -CN,

—N

' wherein a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom and carbon atom c or d, or where a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom b and carbon atom c or d;

and where when A represents

R ^j independently selected

from the group consisting of -F, -O-(C ₁ -C ₆ alkyl), -NR ³ R ⁴ , -CN, where the bridge or bond containing a C1 carbon atom. ₂ may be present between carbon atom and carbon atom c or d, or wherein a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom b and carbon atom c or d;

R ¹ is selected from the group consisting of H and C1-C ₆ alkyl;

R ² is independently selected from the group consisting of C1-C ₆ alkyl or -O-(C ₁ -C ₆ alkyl), and if two R ² are geminal, they can be combined to form a 3-6 membered ring ;

R ³ and R ⁴ are independently selected from the group consisting of H and C1-C ₆ alkyl;

R ⁵ is selected from the group consisting of H and C1-C ₆ alkyl;

n is 0, 1, 2, 3 or 4;

r and s are independently equal to 0,1,2 or 3; and t and u are independently equal to 1, 2 or 3.

2. The compound according to claim 1, characterized in that A is

and

can be attached to the N atom in any accessible position, with substituents R ^j , and R ^j is as defined in paragraph 1.

O ₆ ¹⁴ is replaced by one or more

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3. A compound according to any one of claims 1 and 2, which is a compound of formula (Ib)

. (ib) where R ^a , R ^j and Z are as defined in paragraph 1, and p is 1 or 2.

4. A connection selected from the group consisting of

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/

I o

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5. A pharmaceutical composition containing a therapeutically effective amount of a compound according to any one of claims. 1-4 and a pharmaceutically acceptable carrier or excipient.

6. Use of a compound according to any one of claims 1 to 4 for the treatment, amelioration or prevention of a disorder or disorder associated with tau protein aggregates.

7. A method of treating, preventing, or alleviating a disorder associated with tau protein aggregates, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof.

8. Use of a compound according to any one of claims 1 to 4 in the manufacture of a medicament for the treatment of a disorder or disorder associated with tau protein aggregates.

9. Use of a compound according to any one of claims 1 to 4 in the manufacture of a medicinal product for the treatment of Alzheimer's disease.

10. Use of a compound according to any one of claims 1 to 4 for the manufacture of a medicament for the treatment of progressive supranuclear palsy (PSP).

11. A method of treating, preventing or alleviating Alzheimer's disease, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof.

12. A method of treating, preventing or alleviating PSP, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof.

13. A method of reducing tau protein aggregation, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof.

14. A method for preventing the formation of tau protein aggregates and/or inhibiting tau protein aggregation, comprising administering an effective amount of a compound according to any one of claims. 1 -4 to the subject who needs it.

15. A method of intracellular interaction with tau protein aggregates, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof.

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16. Method according to any one of claims 11 to 15, characterized in that the subject is an animal or a human.

17. A combination pharmaceutical composition containing a therapeutically effective amount of a compound according to any one of claims 1 to 4 and a therapeutically effective amount of at least one additional biologically active compound selected from a therapeutic agent other than a compound according to any one of claims 1 to 4, pharmaceutically acceptable carrier, diluent and excipient.

18. The composition according to claim 17, characterized in that the additional biologically active compound is a compound used in the treatment of amyloidosis.

19. The composition according to claim 17 or claim 18, characterized in that the additional biologically active compound is selected from the group consisting of compounds against oxidative stress, anti-apoptotic compounds, metal chelators, DNA repair inhibitors, 3-amino-1-propanesulfonic acid ( 3APS), 1,3-propane disulfonate (1,3PDS), α-secretase activators, β- and γ-secretase inhibitors, tau proteins, neurotransmitters, β-sheet disruptors, cellular component attractants, beta-amyloid scavengers/depleters, truncated inhibitors at the N-terminus of amyloid beta, including amyloid beta pyroglutamate 3-42, anti-inflammatory compounds or cholinesterase inhibitors (ChEl), M1 agonists, amyloid or tau-modifying drugs and dietary supplements, antibodies, including any functionally equivalent antibody or functional ones thereof parts or vaccine.

20. Composition according to any one of claims 17 to 19, characterized in that the compound and/or additional biologically active compound is/are present in a therapeutically effective amount.

21. Use according to claim 6 or 8, characterized in that the disorder is selected from Alzheimer's disease (AD), familial AD, primary age-related tauopathy (PART), Creutzfeldt-Jakob disease, Boxer dementia, Down syndrome, Gerstmann-Straussler-Scheinker disease (GSS), inclusion body myositis, prion cerebral amyloid angiopathy, traumatic brain injury (TBI), amyotrophic lateral sclerosis (ALS), parkinsonian dementia (Guam syndrome), non-Guam motor neuron disease with neurofibrillary tangles, a disease characterized by the appearance of argyrophilic grains , corticobasal degeneration (CBD), diffuse neurofibrillary tangles with calcification, frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), Hallerwarden-Spatz disease, multiple system atrophy (MSA), Niemann-Picatype C disease, pallidoponto -nigral degeneration, Pick's disease (PiD), progressive subcortical gliosis, progressive supranuclear palsy (PSP), subacute sclerosing panencephalitis, tangle-predominant dementia, postencephalitic parkinsonism, myotonic dystrophy, subacute sclerosing panencephalitis, mutations in LRRK2, chronic traumatic encephalopathy (CTE) , familial British dementia, familial Danish dementia, other frontotemporal lobar degenerations, Guadalupean parkinsonism, brain iron accumulation neurodegeneration associated with SLC9A6 mental retardation, white matter tauopathy with globular glial inclusions, epilepsy, Lewy body dementia (LBD), mild cognitive disorder (MCI), multiple sclerosis, Parkinson's disease, HIV-associated dementia, adult-onset diabetes, age-related cardiac amyloidosis, glaucoma, ischemic stroke, AD psychosis and Huntington's disease.

22. The method according to claim 7, characterized in that the disorder is selected from Alzheimer's disease (AD), familial AD, primary age-related tauopathy (PART), Creutzfeldt-Jakob disease, Boxer dementia, Down syndrome, Gerstmann-Straussler-Scheinker disease (GSS ), inclusion body myositis, prion cerebral amyloid angiopathy, traumatic brain injury (TBI), amyotrophic lateral sclerosis (ALS), parkinsonian dementia (Guam syndrome), non-Guam motor neuron disease with neurofibrillary tangles, a disease characterized by the appearance of argyrophilic grains, corticobasal degeneration (CBD), diffuse neurofibrillary tangles with calcification, frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), Hallerwarden-Spatz disease, multiple system atrophy (MSA), Niemann-Pick disease type C, pallidoponto -nigral degeneration, Pick's disease (PiD), progressive subcortical gliosis, progressive supranuclear palsy (PSP), subacute sclerosing panencephalitis, tangle-predominant dementia, postencephalitic parkinsonism, myotonic dystrophy, subacute sclerosing panencephalitis, mutations in LRRK2, chronic traumatic encephalopathy (CTE) , familial British dementia, familial Danish dementia, other frontotemporal lobar degenerations, Guadalupean parkinsonism, neurodegeneration with brain iron accumulation associated with SLC9A6 mental retardation, white matter tauopathies with globular glial inclusions, epilepsy, dementia with Lewy bodies (LBD), mild cognitive impairment (MCI), multiple sclerosis, Parkinson's disease, HIV-associated dementia, midlife diabetes, senile cardiac amyloidosis, glaucoma, ischemic stroke, AD psychosis and Huntington's disease.

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23. Use of a compound according to any one of paragraphs. 1-4 as an analytical standard or in vitro screening tool to characterize tissue with Tau pathology or to test compounds targeting Tau pathology in such tissue.

24. Compound formula

or a pharmaceutically acceptable salt thereof.

Definitions.

In the context of this invention, the following definitions apply.

The term alkyl refers to a saturated straight or branched chain organic moiety consisting of carbon and hydrogen atoms. Examples of suitable alkyl groups have from 1 to 6 carbon atoms, preferably from 1 to 4 carbon atoms, and include methyl, ethyl, propyl, isopropyl, n-butyl, t-butyl and isobutyl.

The term Hal or halogen refers to F, Cl, Br and I, preferably F.

The term 3-8 membered ring refers to a three-, four-, five-, six-, seven-, or eight-membered ring in which none, one, or more of the carbon atoms in the ring are replaced by 1 or 2 (for a three-membered ring), 1 , 2 or 3 (for a four-membered ring), 1, 2, 3 or 4 (for a five-membered ring) or 1, 2, 3, 4 or 5 (for a six-membered ring), 1, 2, 3, 4, 5 or 6 ( for a seven-membered ring) or 1, 2, 3, 4, 5, 6 or 7 (for an eight-membered ring) with the same or different heteroatoms, the heteroatoms being selected from O, N and S.

The compounds of this invention having one or more optically active carbon atoms may exist as racemates and racemic mixtures (including mixtures in all proportions), stereoisomers (including diastereomeric mixtures and individual diastereomers, enantiomeric mixtures and individual enantiomers, mixtures of conformers and individual conformers ), tautomers, atropoisomers and rotamers. All isomeric forms are included in this invention. The compounds described in this invention containing olefinic double bonds include the geometric isomers E and Z. Also included in this invention are all pharmaceutically acceptable salts, prodrugs, polymorphs, hydrates and solvates.

The term polymorphs refers to the different crystal structures of the compounds of this invention. It may include, but is not limited to, crystal (and amorphous materials) morphology and all crystal lattice shapes. The salts of this invention may be crystalline and may exist as more than one polymorph.

Solvates, hydrates, as well as anhydrous salt forms are also covered by this invention. The solvent included in the solvates is not particularly limited and may be any pharmaceutically acceptable solvent. Examples include water and C1.4 alcohols (such as methanol or ethanol).

Pharmaceutically acceptable salts are defined as derivatives of the disclosed compounds in which the parent compound is modified by producing acidic or basic salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic moieties, such as amines, alkali metal salts or organic salts of acidic moieties, for example, carboxylic acids, and similar salts. Pharmaceutically acceptable salts include conventional non-toxic salts or quaternary ammonium salts of the parent compound derived, for example, from non-toxic inorganic or organic acids. For example, such conventional non-toxic salts include those derived from inorganic acids, such as, but not limited to, hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric and similar acids; and salts derived from organic acids, such as, but not limited to, acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamic, maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic, sulfanilic , 2-acetoxybenzoic, fumaric, toluenesulfonic, methanesulfonic, ethanedisulfonic, oxalic, isethionic acid and similar acids. The pharmaceutically acceptable salts of this invention can be synthesized from a parent compound that contains a basic or acidic moiety by conventional chemical methods. Typically, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of a suitable base or acid in water or an organic solvent, or a mixture of water and an organic solvent. Organic solvents include, but are not limited to, non-aqueous media such as ethers, ethyl acetate, ethanol, isopropanol or acetonitrile. Lists of suitable salts can also be found in Remington's Pharmaceutical Sciences, ^18th ed., Mack Publishing Company, Easton, PA, 1990, p. 1445, the description of which is incorporated herein by reference.

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The compounds of this invention can also be obtained in the form of a prodrug, that is, a compound that is metabolized in vivo to an active metabolite. As used hereinafter in the specification and claims, the term prodrug means any covalently linked compound that releases the active parent drug through biotransformation in vivo. Reference to Goodman and Gilman (The Pharmacological Basis of Therapeutics, 8 ed, McGraw-Hill, Int. Ed. 1992, Biotransformation of Drugs, pp. 13-15) describing prodrugs is generally incorporated herein by reference.

As used herein, the phrase pharmaceutically acceptable refers to compounds, materials, compositions and/or dosage forms that, based on careful medical examination, are suitable for use in contact with human and animal tissue without causing excessive toxicity, irritation, allergic response or other problem or complications commensurate with an acceptable benefit-to-risk ratio.

Patients or subjects of the present invention are typically animals, in particular mammals, and more particularly humans.

The term tau protein, as used herein, refers to a highly soluble microtubule-binding protein primarily found in neurons and includes the major 6 isoforms, cleaved or processed forms, and other modified forms, such as those resulting from phosphorylation, glycosylation, glycation, prolyl isomerization, nitration, acetylation, polyamination, ubiquitination, sumoylation and oxidation.

The term aggregated tau protein refers to aggregated monomers of tau peptides or proteins that are folded into oligomeric or polymeric structures.

The term neurofibrillary tangles (NFTs), as used herein, refers to insoluble aggregates of hyperphosphorylated tau protein containing paired helical filaments (PHFs) and straight filaments. Their presence is a characteristic feature of AD and other diseases known as tauopathies.

The definitions and preferred definitions given in the Definitions section apply to all embodiments described below unless otherwise noted.

Detailed description of the invention

The compounds of this invention will be described below. It should be understood that all possible combinations of the following definitions are also provided.

In one embodiment, this invention relates to a compound of formula (I)

^Rd

(I) or its stereoisomers, racemic mixtures, tautomers, pharmaceutically acceptable salts, prodrugs, hydrates, solvates and polymorphs.

A preferred embodiment of the compound of formula (I) is

^Rd

(la)

R ^a

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A further preferred embodiment of a compound of formula (I) is

The following definitions A apply to compounds of formula (I) and preferred embodiments thereof.

In a preferred embodiment of the invention

selected from

R

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In another preferred embodiment of the invention

yourself

represents embodiments of the invention A represents

In one preferred embodiment

In a more preferred embodiment of the invention, A is selected from

In a further preferred embodiment of the invention, A is selected from

In an even more preferred embodiment of the invention, A is ^N

In the above definitions of A and its preferred embodiments

can be attached to the N atom in any available position.

In the above definitions of A and its preferred embodiments, A is substituted with one or more R ^j substituents, for example, R ^j may be present once or twice. If A is a phenyl ring, then preferably 1 or 2 substituents may be present.

If A is a pyridine ring, then preferably 1 substituent may be present.

The following definitions and their preferred embodiments apply to formula (I), as appropriate.

B is selected from the group consisting of O and NR ^a . More preferably, B is NR ^a .

E and V are independently selected from the group consisting of N, NR ⁵ , O and S, thus

selected from

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R ⁵

J is selected from the group consisting of O, NR ¹ and CH2 or J is selected from the group consisting of CH or C if J is attached to R ² . In one embodiment, J is O, in another embodiment, J is NR ¹ (preferably N-Me), and in a further embodiment, J is CH2, CH, or C.

Y, Y ¹ , Y ² and Y ³ are CZ, more preferably Y, Y ¹ , Y ² and Y ³ are CH.

Z is independently selected from the group consisting of H, halogen (preferably F), O-alkyl, alkyl and CN, preferably H, halogen (preferably F) and O-alkyl. In one preferred embodiment, one Z is halogen (preferably F) or O-alkyl and the other Zs are H. In a more preferred embodiment, one Z is halogen (preferably F) and the other Zs are H.

(R ² )n -ν ^ΑΛ ο

R is independently selected from the group consisting of / and -NR ³ R ⁴ , preferably R is -S is /

R ^a is selected from the group consisting of H and alkyl, more preferably H and Me.

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R ^d , R ^e , R ^f and R ^g are independently selected from the group consisting of H and alkyl, or any two of R ^d , R ^e , R ^f and R ^g can be joined to form a 3-8 membered ring. Preferably, ^Rd , ^Re , ^Rf , ^Rg , independently representing H or ^Rd and ^Rf , can be joined together to form a bridge containing C _1-2 carbon atoms. More preferably, ^Rd , ^Re , ^Rf , and ^Rg are H.

Rj is independently selected from the group consisting of -halogen, -O-alkyl, -NR ³ R ⁴ , -CN,

wherein a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom and carbon atom c or d, or where a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom b and carbon atom c or d. More preferably, Rj is selected from the a, -B -\ group consisting of -halogen (preferably -F), -O-alkyl (preferably -O-Me) and cd -r/ S - νΎ^Ο - J\-Me (preferably or \-7), even more preferably Rj represents -1\/ S is /.

R ¹ is selected from the group consisting of H and alkyl, preferably alkyl, more preferably CH ₃ .

R ² is independently selected from the group consisting of alkyl or -O-alkyl, and if two R ² are geminal, they can be joined to form a 3-6 membered ring. In one embodiment, R ² is alkyl, in another embodiment, R ² is -O-alkyl, in a further embodiment, two R ² that are geminal can be joined to form a 3-6 membered ring.

R ³ and R ⁴ are independently selected from the group consisting of H and alkyl. In one embodiment, R ³ or R ⁴ is alkyl and the other is H. In another embodiment, R ³ is alkyl and R ⁴ is alkyl. In a further embodiment of the invention, R ³ and R ⁴ are H.

R ⁵ is selected from the group consisting of H and alkyl. In one embodiment, R ⁵ is H. In another embodiment, R ⁵ is alkyl.

n is 0, 1, 2, 3 or 4, more preferably 0, 1 or 2, even more preferably n is 0.

p is 1 or 2, more preferably 1.

r and s are independently equal to 0,1,2 or 3.

t and u are independently equal to 1, 2 or 3.

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Preferred compounds of formula (I) are

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Preferred compounds are also illustrated in the examples.

Any combination of embodiments, preferred embodiments, and more preferred embodiments disclosed herein are also contemplated by this invention.

Pharmaceutical compositions.

Although the compounds of this invention can be administered separately, in accordance with standard pharmaceutical practice, it is preferable to formulate them into a pharmaceutical composition. Thus, the present invention also provides a pharmaceutical composition that contains a therapeutically effective amount of a compound of formula (I), optionally in admixture with a pharmaceutically acceptable carrier, diluent, adjuvant or excipient.

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Pharmaceutically acceptable excipients are well known in the pharmaceutical art and are described, for example, in Remington's Pharmaceutical Sciences, ^15th Ed., Mack Publishing Co., New Jersey (1975). The pharmaceutical excipient may be selected based on the intended route of administration and standard pharmaceutical practice. The excipient must be acceptable in the sense that it is not harmful to the recipient.

Pharmaceutically applicable excipients that can be used in the pharmaceutical composition of this invention may include, for example, carriers, carrier vehicles, diluents, solvents, for example, monohydric alcohols, for example, ethanol, isopropanol, and polyhydric alcohols, for example, glycols as well as edible oils, e.g. soybean oil, coconut oil, olive oil, safflower oil, cottonseed oil, fatty esters, e.g. ethyl oleate, isopropyl myristate, binders, adjuvants, solubilizers, thickeners, stabilizers, disintegrants, glidants, lubricants , buffering agents, emulsifiers, wetting agents, suspending agents, sweeteners, colors, flavors, coating agents, preservatives, antioxidants, emulsifiers, drug delivery modifiers and enhancing agents, e.g. calcium phosphate, magnesium stearate, talc, monosaccharides, disaccharides, starch , gelatin, cellulose, methylcellulose, sodium carboxymethylcellulose, dextrose, hydroxypropyl-β-cyclodextrin, polyvinylpyrrolidone, low-melting waxes and ion exchange resins.

Routes of administration (delivery) of the compounds of this invention include, but are not limited to, one or more of: oral (e.g., as a tablet, capsule, or ingestible solution), topical, transmucosal (e.g., as a nasal inhalation spray or aerosol), nasal, parenteral (eg, injectable), gastrointestinal, intraspinal, intraperitoneal, intramuscular, intravenous, intrauterine, intraocular, intradermal, intracranial, intratracheal, intravaginal, intracerebroventricular, intracerebral, subcutaneous, ophthalmic (including intravitreal or intracameral), transdermal, rectal, buccal, epidural and sublingual.

For example, the compounds may be administered orally in the form of tablets, capsules, ovules, tinctures, solutions or suspensions, which may contain flavorings or colors, for immediate, sustained, modified, sustained, pulsed or controlled release.

Tablets may contain excipients, for example, microcrystalline cellulose, lactose, sodium citrate, calcium carbonate, dibasic calcium phosphate and glycine, disintegrants, for example, starch (preferably corn, potato or tapioca starch), sodium starch glycolate, croscarmellose sodium and some complex silicates, and also granulation binders such as polyvinylpyrrolidone, hydroxypropyl methylcellulose (HPMC), hydroxypropylcellulose (HPC), sucrose, gelatin and gum acacia. In addition, lubricants such as magnesium stearate, stearic acid, glyceryl behenate and talc may be included. Solid compositions of this type can also be used as fillers in gelatin capsules. Preferred excipients in this regard include lactose, starch, cellulose, milk sugar or high molecular weight polyethylene glycols. For aqueous suspensions and/or tinctures, the agent may be combined with a variety of sweeteners or flavoring agents, coloring agents or dyes, emulsifying and/or suspending agents, and diluents such as water, ethanol, propylene glycol and glycerin, and combinations of the above.

When the compounds of this invention are administered parenterally, examples of such administration include one or more of: intravenous, intraarterial, intraperitoneal, intrathecal, intraventricular, intraurethral, intrasternal, intracranial, intramuscular, or subcutaneous administration of the compounds; and/or using infusion methods. For parenteral administration, the compounds are best used in the form of a sterile aqueous solution, which may contain other substances, for example, sufficient salts or glucose to render the solution isotonic with blood. Aqueous solutions should be suitably buffered (preferably to a pH of 3 to 9) if necessary. The preparation of suitable parenteral compositions under sterile conditions is readily accomplished by standard pharmaceutical techniques well known to those skilled in the art.

As indicated, the compounds of the present invention can be administered intranasally or by inhalation and are conveniently delivered in the form of a dry powder by an inhaler or in the form of an aerosol spray from a pressurized container, pump, atomizer or nebulizer using a suitable propellant, for example, dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane , hydrofluoroalkane, for example 1,1,1,2-tetrafluoroethane (HFA134AT) or 1,1,1,2,3,3,3-heptafluoropropane (HFA 227EA), carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be defined by providing a valve for delivering the metered amount. The pressurized container, pump, atomizer or nebulizer may contain a solution or suspension of the active compound, for example using a mixture of ethanol and a propellant as a solvent, which may further contain a lubricant,

- 19 044393 for example, sorbitan trioleate. Capsules and cartridges (made, for example, from gelatin) for use in an inhaler or insufflator can be formulated to contain a powder mixture of the compound and a suitable powder base, for example lactose or starch.

Alternatively, the compounds of this invention may be administered in the form of a suppository or pessary, or they may be applied topically in the form of a gel, hydrogel, lotion, solution, cream, ointment, or powder. The compounds of this invention can also be administered transdermally or transdermally, for example, using a skin patch.

They can also be administered by the pulmonary or rectal route. They can also be administered through the eye. For ophthalmic use, the compounds may be formulated as micronized suspensions in isotonic, pH-adjusted, sterile saline or, preferably, as solutions in isotonic, pH-adjusted, sterile saline, optionally in combination with a preservative, e.g. benzylalkonium chloride. Alternatively, they may be formulated as an ointment such as Vaseline.

For topical application to the skin, the compounds of this invention may be formulated as a suitable ointment containing the active compound suspended or dissolved, for example, in a mixture with one or more of the following: mineral oil, liquid petrolatum, white petrolatum, propylene glycol, emulsifying wax and water. Alternatively, they may be formulated as a suitable lotion or cream, suspended or dissolved, for example, in a mixture of one or more of the following: mineral oil, sorbitan monostearate, polyethylene glycol, liquid paraffin, polysorbate 60, cetyl ester wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.

Typically, the physician will determine the actual dosage that will be most appropriate for the individual subject. The specific dosage level and frequency of dosage for any given individual may vary and will depend on a variety of factors, including the potency of the particular compound used, the metabolic stability and duration of action of said compound, age, body weight, general health, sex, diet, route and time of administration. , elimination rate, drug combination, severity of specific condition, and individual treatment.

The suggested dose of the compounds of this invention for administration to humans (approximately 70 kg body weight) is from 0.1 mg to 1 g, preferably from 1 mg to 500 mg of active ingredient per dosage unit. A unit dose may be administered, for example, 1 to 4 times a day. The dose will depend on the route of administration. It is understood that regular dosage adjustments may be required depending on the age and weight of the patient and the severity of the condition being treated. The exact dose and route of administration will ultimately be left to the discretion of the treating physician or veterinarian.

The compounds of this invention can also be used in combination with other therapeutic agents. If a compound of this invention is used in combination with a second therapeutic agent active against the same disease, the dose of each compound may be different from the dose when the compound is used alone.

The combinations mentioned above may, for convenience, be presented for use in the form of a pharmaceutical composition. The individual components of such combinations may be administered sequentially or simultaneously in separate or combined pharmaceutical compositions in any convenient manner. If administration is sequential, both the compound of this invention and the second therapeutic agent may be administered first. If administration is simultaneous, the combination may be administered in the same or in different pharmaceutical compositions. When combined in the same composition, it is understood that the two compounds must be stable and compatible with each other and the other components of the composition. When formulated separately, they may be provided in any convenient composition, for convenience, in a manner known for such compounds in the art.

The pharmaceutical compositions of this invention can be prepared in a manner well known to one skilled in the art, as described, for example, in Remington's Pharmaceutical Sciences, ^15th Ed., Mack Publishing Co., New Jersey (1975).

Diseases or pathological conditions that can be treated, alleviated or prevented using the compounds of this invention are disorders or disorders associated with tau protein aggregates, for example, neurodegenerative disorders. Examples of diseases and conditions that can be treated, ameliorated, or prevented are caused by or are associated with the formation of neurofibrillary lesions. This is the predominant brain pathology in tauopathy. The diseases and conditions include a heterogeneous group of neurodegenerative diseases or conditions, including diseases or conditions that exhibit the coexistence of tau protein and amyloid pathologies.

Examples of diseases and conditions that may be treated, ameliorated, or prevented include, but are not limited to, Alzheimer's disease (AD), familial AD, PART (primary age-related tauopathy), Creutzfeldt-Jakob disease, Boxer dementia, Down syndrome,

- 20 044393 Gerstmann-Streissler-Scheinker disease (GSS), inclusion body myositis, prion cerebral amyloid angiopathy, traumatic brain injury (TBI), amyotrophic lateral sclerosis (ALS), parkinsonian dementia (Guam syndrome), non-Guam motor neuron disease with neurofibrillary tangles, argyrophilic grain disease, corticobasal degeneration (CBD), diffuse neurofibrillary tangles with calcification, frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), Hallervorden-Spatz disease, multiple system atrophy (MSA) , Niemann-Pick disease type C, pallidopontonigral degeneration, Pick disease (PiD), progressive subcortical gliosis, progressive supranuclear palsy (PSP), subacute sclerosing panencephalitis, tangle-predominant dementia, postencephalitic parkinsonism, myotonic dystrophy, subacute sclerosing panencephalitis, mutations in LRRK2 , chronic traumatic encephalopathy (CTE), familial British dementia, familial Danish dementia, other frontotemporal lobar degenerations, Guadalupean parkinsonism, neurodegeneration with brain iron accumulation, SLC9A6-related mental retardation, white matter tauopathy with globular-glial inclusions, epilepsy, diffuse Lewy body disease (LBD), mild cognitive impairment (MCI), multiple sclerosis, Parkinson's disease, HIV-associated dementia, adult-onset diabetes, frailty, glaucoma, ischemic stroke, AD psychosis, and Huntington's disease. Preferably, diseases and pathological conditions that can be treated, ameliorated or prevented include Alzheimer's disease (AD), as well as other neurodegenerative tauopathies, for example, Creutzfeldt-Jakob disease, boxer dementia, amyotrophic lateral sclerosis (ALS), a disease characterized by appearance of argyrophilic granules, corticobasal degeneration (CBD), frontotemporal dementia with parkinsonism associated with chromosome 17 (FTDP-17), Pick's disease (PiD), progressive supranuclear palsy (PSP), tangle-predominant dementia, parkinsonian dementia (Guam syndrome), Hallervorden-Spatz disease, chronic traumatic encephalopathy (CTE), traumatic brain injury (TBI), and other frontotemporal lobar degeneration. More preferably, Alzheimer's disease (AD), corticobasal degeneration (CBD), Pick's disease (PiD) and progressive supranuclear palsy (PSP).

The compounds of this invention can also be used to reduce protein aggregation, in particular tau protein aggregation. The ability of a compound to reduce tau protein aggregation can, for example, be determined by the ThT assay (Hudson et al., FEBS J., 2009, 5960-72).

The compounds of this invention can be used to treat a wide range of disorders in which the process of neuroinflammation is associated with misfolding and/or pathological aggregation of tau protein.

The compounds of this invention can be used as an analytical standard or in vitro screening tool to characterize tissue with tau pathology and to test compounds targeting tau pathology in such tissue.

The compounds of this invention may also be provided in the form of a mixture with at least one additional biologically active compound and/or a pharmaceutically acceptable carrier and/or diluent and/or excipient. The compound and/or additional biologically active compound is preferably present in a therapeutically effective amount.

The nature of the additional biologically active compound will depend on the intended use of the mixture. The additional biologically active substance or compound may exert its biological action by the same or similar mechanism as the compound of the invention, or by an unrelated mechanism of action, or by multiple related and/or unrelated mechanisms of action.

Typically, the additional biologically active compound may include neutron transmission enhancers, psychotherapeutic drugs, acetylcholinesterase inhibitors, calcium channel blockers, biogenic amines, benzodiazepine tranquilizers, acetylcholine synthesis, release or release enhancers, acetylcholine postsynaptic receptor agonists, monoamine oxidase-A inhibitors, or - B, N-methyl-O-aspartate glutamate receptor antagonists, non-steroidal anti-inflammatory drugs, antioxidants and serotonergic receptor antagonists. In particular, the additional biologically active compound may be selected from the group consisting of a compound used in the treatment of amyloidosis, anti-oxidative stress compounds, anti-apoptotic compounds, metal chelates, DNA repair inhibitors, for example, pirenzepine and metabolites, 3-amino-1-propanesulfonic acid (3APS), 1,3-propane disulfonate (1,3PDS), α-secretase activators, β- and γ-secretase inhibitors, tau proteins, neurotransmitter, β-sheet disruptors, attractants for cellular components, beta-amyloid scavenging/depleting , N-terminally truncated beta-amyloid inhibitors, including pyroglutamate beta-amyloid 3-42, anti-inflammatory molecules or cholinesterase (ChEl) inhibitors, for example, tacrine, rivastigmine, donepezil and/or galantamine, M1 agonists, other drugs, including any amyloid or tau-modifying drug and nutritional supplements, antibodies, including any functionally equivalent antibody or functional parts thereof, or a vaccine.

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In a further embodiment, mixtures of this invention may contain niacin or memantine together with a compound of this invention and, optionally, a pharmaceutically acceptable carrier and/or diluent and/or excipient.

In yet another embodiment, the invention provides mixtures that contain, as an additional biologically active compound, an atypical antipsychotic agent, for example, clozapine, ziprasidone, risperidone, aripiprazole or olanzapine, for the treatment of positive and negative psychotic symptoms, including hallucinations, delusions, thought disorders (manifested by in marked incoherence, slippage of the thought process, tangentiality of thought) and strange or disorganized behavior, as well as anhedonia, affective flattening, apathy and social withdrawal, together with a compound of this invention and, optionally, a pharmaceutically acceptable carrier and/or diluent and/or excipient.

Other compounds which may suitably be used in mixtures in combination with a compound of this invention are described, for example, in WO 2004/058258 (in particular see pages 16 and 17), including therapeutic drug targets (pages 36-39) , alkanesulfonic acids and alkanesulfuric acids (pages 39-51), cholinesterase inhibitors (pages 5156), NMDA receptor antagonists (pages 56-58), estrogens (pages 58-59), non-steroidal anti-inflammatory drugs (pages 60 and 61), antioxidants ( pages 61 and 62), peroxisome proliferator-activated receptor (PPAR) agonists (pages 63-67), cholesterol lowering agents (pages 68-75), amyloid inhibitors (pages 75-77), amyloid inhibitors (pages 77- 78), metal chelates (pages 78 and 79), antipsychotics and antidepressants (pages 80-82), dietary supplements (pages 83-89) and compounds that increase the availability of bioactive substances in the brain (see pages 89-93) and prodrugs (pages 93 and 94), incorporated herein by reference.

The present invention also includes all suitable isotopic variants of the compounds of the invention. An isotopic variant of a compound of this invention is defined as one in which at least one atom is replaced by an atom having the same atomic number but an atomic mass different from the atomic mass typically found in nature. Examples of isotopes that may be included in the compounds of this invention include isotopes of hydrogen, carbon, nitrogen, oxygen, sulfur, fluorine and chlorine, for example, ^{2H , 3H} , 13C, ^14C , ^15N , ^17O , ¹⁸ O, ³⁵ S, ¹⁸ F and ³⁶ Cl, respectively. Certain isotopic embodiments of the invention, for example those that include a radioactive isotope, such as ³ H or ¹⁴ C, are useful in drug and/or substrate tissue distribution studies. Tritiated isotopes, i.e. ³ H and carbon-14, i.e. ¹⁴ C are particularly preferred due to their ease of preparation and detectability. 1T-labeled compounds are particularly suitable for imaging applications such as PET. In addition, substitution with isotopes such as deuterium, i.e. ^2H may provide certain therapeutic benefits due to greater metabolic stability, such as increased in vivo half-life or reduced dosage requirements, and may therefore be preferred in some cases. Isotopic variations of the compounds of this invention can generally be prepared by conventional techniques, for example, the exemplary methods or the synthetic methods described below in the Examples and Preparation Methods sections using suitable isotopic variations of suitable reagents.

The compounds of this invention can be synthesized by one of the general methods illustrated in the schemes below. These methods are for illustrative purposes only and should not be construed as limiting.

General synthesis schemes for obtaining building blocks according to the invention

Scheme 1

Heating commercially available phenylhydrazine derivatives (Z=H, F or OMe) with commercially available tert-butyl-3-oxopiperidine-1-carboxylate in a suitable solvent under acidic conditions (Fischer indole synthesis) resulted in tricyclic derivatives (possibly formation of a regiomer for Z =F or OMe) after cleaning. If regioisomers were formed, they were separated by supercritical fluid chromatography (SFC) to obtain the desired tetrahydro7H-pyrido[3,4-b]indole derivatives. The aliphatic secondary NH moiety of the tricyclic building blocks was further protected with a Boc protecting group using a suitable solvent and base to obtain the desired Boc protected building blocks after purification.

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Scheme 2

The NH moiety of the tricyclic building blocks was treated with methyl iodide or tosyl chloride in a suitable solvent using a suitable base to obtain, after purification, the N-methyl or N-tosyl derivatives. The Boc protecting group was cleaved by acid treatment in a suitable solvent to obtain, after purification, the desired N-methyl or N-tosyl tricyclic building blocks. In the absence of base treatment after cleavage of the Boc protecting group, the corresponding salts were obtained.

Scheme 3

Z = F

Heating commercially available phenylhydrazine derivatives (Z=F) with commercially available tert-butyl-2-oxo-8-azabicyclo[3.2.1]octane-8-carboxylate in a suitable solvent under acidic conditions (Fischer indole synthesis) gave tricyclic derivatives. In the case of 2- or 3-substituted phenylhydrazine derivatives, regioisomers were separated by supercritical fluid chromatography (SFC). The aliphatic secondary amine moiety was then protected with a Boc group using suitable solvents and base to obtain, after purification, the desired building block. The NH moiety of the indole moiety was then treated with tosyl chloride in a suitable solvent using a suitable base to obtain, after purification, the N-tosyl derivatives. The Boc-protecting group was cleaved by acid treatment in an appropriate solvent to obtain, after purification, tricyclic building blocks containing a secondary amine. In the absence of base treatment after cleavage of the Boc protecting group, the corresponding salts were obtained.

Scheme 4 /—\

ΗΝ O HN Η,Ν-alk -k alk

R ^x , x = Hal (halogen) EhV=N,O,S

G = Ph (phenyl) Ru (pyridine) solvent, base or microwave radiation, solvent

R = morpholine, -N(alk) ₂ , -NH(alk)

R ^x = Hal (halogen)

EhV=N,O,S

G = Ph (phenyl)

Roux (pyridine)

Commercially available benzo[d]thiazole (G=Ph) or benzo[d]oxazole (G=Ph) derivatives containing two halogen atoms (Br or Cl) were treated with primary or secondary amines in a suitable solvent and an additional base. Leaving group X was replaced by nucleophilic substitution with primary or secondary amines to obtain, after purification, the corresponding amino-substituted benzo[d]thiazole or benzo[d]oxazole derivatives. For less reactive amines, the desired benzo[d]thiazole or benzo[d]oxazole derivatives were prepared by performing a nucleophilic substitution reaction under microwave reactor conditions. The corresponding thiazolo[5,4-b]pyridine (G=Py) and thiazolo[4,5-b]pyridine (G=Py) derivatives containing two halogen atoms (Br or Cl) were treated with morpholine in a suitable solvent and an additional base , and after purification the corresponding products of nucleophilic substitution (derivatives of morpholinothiazolo[5,4-b]pyridine (G=Py) and morpholinothiazolo[4,5-b]pyridine (G=Py)) were obtained.

General synthetic schemes for preparing the compounds of the invention

Scheme 5

Tricyclic building blocks with B=NCH ₃ , O were combined with substituted benzo[d]thiazole or benzo[d]oxazole derivatives or substituted phenyl or pyridine derivatives using palladium chemistry with a suitable palladium source, for example palladium(P) acetate (Pd( OAc)2), suitable ligand tris-(dibenzylideneacetone)dipalladium(0) (Pd2(dba) ₃ ), for example, 2dicyclohexylphosphino-2',6'-diisopropoxybiphenyl (RuPhos), 4,5-bis-(diphenylphosphino)-9 ,9dimethylxanthene (xanthos), 2-dicyclohexylphosphino-2',4', _6' -triisopropylbiphenyl (Xphos) and a suitable base, e.g. cesium carbonate ( _Cs2CO3 ) and sodium tert-butoxide (NaOtBu) in a suitable solvent, e.g. , dioxane, to obtain, after purification, the desired compounds of formula (I).

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Scheme 6

pyrazine

Tricyclic building blocks containing an N-tosyl group on the indole moiety were coupled to substituted benzo[d]thiazole or benzo[d]oxazole derivatives or substituted phenyl or pyridine derivatives using palladium chemistry with a suitable palladium source, e.g. palladium(P) acetate. (Pd(OAc)2), a suitable ligand for tris-(dibenzylideneacetone)dipalladium(0) (Pd2(dba) ₃ ), for example, 2-dicyclohexylphosphino-2',6'-diisopropoxybiphenyl (RuPhos),4,5-bis- (diphenylphosphino)-9,9-dimethylxanthene (xanthos), 2-dicyclohexylphosphino-2',4',6'triisopropylbiphenyl (Xphos) and a suitable base such as cesium carbonate (Cs ₂ CO ₃ ) and sodium tert-butoxide (NaOtBu ) in a suitable solvent, for example dioxane, to obtain, after purification, the desired compounds of formula (I). In the event that the tosyl group is not removed during the palladium coupling reaction, the tosyl-protected compounds are typically treated with a suitable base, such as sodium tert-butoxide (NaOtBu), in an appropriate mixture of solvents, such as dioxane and methanol, to obtain, after purifying the desired compounds of formula (I).

Scheme 7

base, solvent

Pd catalyst, ligand

Q = NCH ₃ NTs

R ^x = Hal

Pd catalyst, ligand base, solvent morpholine

Tricyclic building blocks containing an N-tosyl or NCH ₃ group on the indole moiety were coupled to halogenated benzo[d]thiazole or benzo[d]oxazole derivatives using palladium chemistry with a suitable palladium source, such as palladium acetate (P) (Pd(OAc)2), suitable ligand tris-(dibenzylideneacetone)dipalladium(0) (Pd ₂ (dba) ₃ ), for example 2dicyclohexylphosphino-2',6'-diisopropoxybiphenyl (RuPhos), 4,5-bis -(diphenylphosphino)-9,9dimethylxanthene (xanthos), 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (Xphos) and a suitable base such as cesium carbonate (Cs ₂ CO ₃ ) and sodium tert-butoxide (NaOtBu ) in a suitable solvent, such as dioxane, to obtain the desired intermediates. Subsequent palladium coupling of the monohalogenated intermediates with morpholine using similar conditions described for the first step afforded, after purification, the desired compounds of formula (I). In the event that the tosyl group is not eliminated during the palladium coupling reaction, the tosyl-protected compounds are typically treated with a suitable base, such as sodium tert-butoxide (NaOtBu), in an appropriate mixture of solvents, such as dioxane and methanol, to obtain, after purification, the desired compounds of formula (I).

Examples

All reagents and solvents were obtained from commercial sources and used without further purification. 1H-NMR spectra were recorded on Bruker AV 300 and 400 MHz spectrometers in deuterated solvents. Chemical shifts (δ) are presented in parts per million and coupling constants (J values) are presented in hertz. Spin multiplicities are denoted by the following symbols: s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), sh.s (wide singlet). Mass spectra were acquired on an Agilent 1290 Infinity II spectrometer with Chemstation 6130 and on an Agilent 1200 Infinity II spectrometer with Chemstation 6130. GC-MS data were collected using an Agilent 7890B gas chromatograph and 5977B mass spectrometer. Infrared spectra were obtained on a PerkinElmer spectrometer. Chromatography was carried out using silica gel (Fluka: Silica gel 0I 60, 0.0630.2 mm) and suitable solvents as indicated in the specific examples. Flash purification was performed using Biotage Isolera with HP-Sil or KP-NH SNAP cartridges (Biotage), and the solvent gradient was specified in specific examples. Thin layer chromatography (TLC) was performed on silica gel plates with UV detection.

Preparative example 1.

Stage A.

To a solution of 3-(fluorophenyl)hydrazine (1 g, 6.1 mmol) and tert-butyl-4-oxopiperidine-1-carboxylate (1.2 g, 6.1 mmol) in 1,4-dioxane (10 ml) added conc. H ₂ SO ₄ (1 ml) at 0°C. The reaction mixture was then heated at 110°C for 3 hours. The reaction mixture was cooled to 25°C, and the precipitate was filtered off. The solid was dissolved in water, made basic with NaOH solution and extracted with dichloromethane. The organic phase was separated, dried over Na ₂ SO ₄ , filtered and the solvent was removed under reduced pressure to give the cyclized title compound as a pale yellow solid (0.6 g, 54%).

1H-NMR (400 MHz, DMSOA δ=10.73 (b.s., 1H), 7.21-7.22 (m, 1H), 7.06-7.07 (m, 1H), 6.78 -6.79 (m, 1H), 3.83 (s, 2H), 2.94-2.95 (m, 2H), 2.49-2.50 (m, 2H).

MS: 191 (M+H)+.

Stage B.

To a solution of the title compound obtained in Step A above (0.6 g, 3.15 mmol) in THF (10 ml) was added triethylamine (1.3 ml, 9.47 mmol) and di-tert-butyl dicarbonate (0.757 g, 3.46 mmol) and the reaction mixture was stirred for 12 hours. After completion of the reaction, as indicated by TLC, the solvent was removed under reduced pressure and the crude reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (80:20) to give the title compound as a pale yellow solid (0.55 g, 60%).

1H-NMR (400 MHz, DMSOA δ=10.94 (b.s., 1H), 7.26-7.28 (m, 1H), 7.12-7.13 (m, 1H), 6.83 -6.84 (m, 1H), 4.55 (s, 2H), 3.64-3.66 (m, 2H), 2.65 (s, 2H), 1.47 (s, 9H). MS: 191 (M-Vos) ⁺ .

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Stage C.

To a solution of the title compound obtained in Step B above (0.55 g, 1.89 mmol) in THF (5 ml) was added sodium hydride (0.136 g, 5.6 mmol), followed by methyl iodide (0.13 ml, 2.07 mmol). The reaction mixture was stirred at room temperature for 2 hours. The mixture was dissolved in ethyl acetate (20 ml) and washed with water and brine. The organic phase was separated, dried over Na ₂ SO ₄ , filtered and the solvent was removed under reduced pressure. The crude reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (70:30) to give the methylated title compound (0.42 g, 73%).

1H-NMR (400 MHz, DMSO-d6) δ=7.40-7.42 (m, 1H), 7.18-7.19 (m, 1H), 6.92-6.93 (m, 1H ), 4.61 (s, 2H), 3.64-3.65 (m, 5H), 2.67 (br.s, 2H), 1.47 (s, 9H). MS: 305.0 (M+H)+.

Stage D

To a solution of the title compound obtained in Step C above (0.42 g, 1.38 mmol) in dichloromethane (10 ml) was added 2N. HCl (5 ml) in 1,4-dioxane.

The reaction mixture was stirred overnight. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the title compound as an off-white solid (0.25 g, 80%).

1H-NMR (400 MHz, DMSO-06) δ=9.83 (b.s., 1H), 7.47-7.50 (m, 1H), 7.30 (d, J=9.60 Hz, 1H), 7.02 (b.s., 1H), 4.42 (s.s, 2H), 3.67 (s.s, 3H), 3.40 (b.s., 2H), 2.92 (b.s. , 2H).

MS: 205.2 (M+H) ⁺ .

Preparative example 2.

Stage A.

To a solution of the title compound obtained in Step B of Preparation Example 1 (0.55 g, 1.89 mmol) in THF (5 ml) was added sodium hydride (0.136 g, 5.6 mmol), followed by ptoluenesulfonyl chloride (0.396 g, 2.07 mmol). The reaction mixture was stirred for 30 minutes at room temperature. The mixture was dissolved in ethyl acetate (20 ml) and washed with water and brine. The organic phase was separated, dried over Na2SO4, filtered and the solvent was removed under reduced pressure. The crude reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (70:30) to give the title compound (0.4 g, 47%). 1H-NMR (400 MHz, DMSO-06) δ=8.00 (s, 1H), 7.77 (s, 2H), 7.30-7.32 (m, 3H), 7.167.18 (m, 1H), 4.87 (s, 2H), 3.64 (s, 2H), 2.62 (br.s, 2H), 2.32 (s, 3H), 1.39 (s, 9H). MS: 445.2 (M+H) ⁺ .

Stage B.

To a solution of the title compound obtained in step A above (0.4 g, 0.9 mmol) in dichloromethane (10 ml) was added 2N. HCl (5 ml) in 1,4-dioxane. The reaction mixture was stirred at room temperature for 12 hours. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the title compound as an off-white solid (0.25 g, 80%). The crude product was taken without further purification for the next stage. MS: 345.0 (M+H) ⁺ .

Preparative example 3.

Stage A.

Conc. . H ₂ SO ₄ (10 ml) at 0°C. The reaction mixture was then heated at 110°C for 3 hours. The reaction mixture was cooled to 25°C, and the precipitate was filtered off. The solid was dissolved in water, made basic with NaOH solution and extracted with dichloromethane. The organic phase was separated, dried over Na ₂ SO ₄ , filtered and the solvent was removed under reduced pressure to give the cyclized title compound as a brown gum solid (10 g, crude). The product was taken without further purification for the next step. MS: 203.2 (M+H) ⁺ .

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Stage B.

To a solution of the title compound obtained in Step A above (10 g, 49.5 mmol) in THF (10 ml) was added triethylamine (20 ml, 148.5 mmol) and di-tert-butyl dicarbonate (11.8 g, 54 mmol) and the reaction mixture was stirred for 12 hours. After completion of the reaction, as indicated by TLC, the solvent was removed under reduced pressure and the crude reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (80:20) to give the above title compound as a white solid (4 g, 26.7%).

1H-NMR (400 MHz, DMSO-d6) δ=10.67 (br.s, 1H), 7.19 (d, J=8.40 Hz, 1H), 6.89 (d, J=2, 00 Hz, 1H), 6.666.67 (m, 1H), 4.54 (s, 2H), 3.75 (s, 3H), 3.65-3.66 (m, 2H), 2.64- 2.66 (m, 2H), 1.44 (s, 9H).

MS: 303.2 (M+H)+.

Stage C.

To a solution of the title compound obtained in Step B above (2 g, 6.6 mmol) in THF (10 ml) was added sodium hydride (0.317 g, 13.2 mmol), followed by methyl iodide (0.4 ml, 7.26 mmol). The reaction mixture was stirred at room temperature for 2 hours. The mixture was dissolved in ethyl acetate (200 ml) and washed with water and brine. The organic phase was separated, dried over Na ₂ SO ₄ , filtered and the solvent was removed under reduced pressure. The crude reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (80:20) to give the methylated title compound (0.65 g, 30%).

1H-NMR (400 MHz, DMSO-d6) δ=7.30 (d, J=8.80 Hz, 1H), 6.92 (s, 1H), 6.73-6.73 (m, 1H) , 4.59 (s, 2H), 3.76 (s, 3H), 3.60-3.64 (m, 5H), 2.66-2.68 (m, 2H), 1.45 (s , 9H). MS: 317.2 (M+H) ⁺ .

Stage D

To a solution of the title compound obtained in Step C above (0.65 g, 2.05 mmol) in dichloromethane (10 ml) was added 4 N. HCl (5 ml) in 1,4-dioxane. The reaction mixture was stirred overnight. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the title compound as an off-white solid (0.5 g, 98%).

1H-NMR (400 MHz, DMSO-d6) δ=9.80 (s, 2H), 7.35 (d, J=8.80 Hz, 1H), 6.99 (d, J=2.00 Hz , 1H), 6.796.80 (m, 1H), 4.39 (s, 2H), 3.77 (s, 3H), 3.63 (s, 3H), 3.38-3.40 (m, 2H), 2.90-2.92 (m, 2H).

MS: 217.3 (M+H) ⁺ .

Preparative example 4.

Stage A.

To a solution of the title compound obtained in Step B of Preparative Example 3 (2 g, 6.6 mmol) in THF (10 ml) was added sodium hydride (0.317 g, 13.2 mmol), followed by ptoluenesulfonyl chloride (1.5 g, 7.9 mmol). The reaction mixture was stirred for 2 hours at room temperature. The mixture was dissolved in ethyl acetate (200 ml) and washed with water and brine. The organic phase was separated, dried over Na2SO4, filtered and the solvent was removed under reduced pressure to give the title compound (1.5 g, 50%). The crude product was taken without further purification for the next step. MS: 357.2 (M-Vos) ⁺ .

Stage B.

To a solution of the title compound obtained in step A above (1.5 g, 3.28 mmol) in dichloromethane (15 ml) was added 2N. HCl (10 ml) in 1,4-dioxane. The reaction mixture was stirred at room temperature for 12 hours. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the title compound as an off-white solid (1 g, 77%).

1H-NMR (400 MHz, DMSO-d6) δ=9.91 (b.s., 2H), 7.83 (d, J=8.40 Hz, 3H), 7.35 (d, J=8, 40 Hz, 2H), 7.06 (d, J=2.00 Hz, 1H), 6.93-6.94 (m, 1H), 4.61 (s, 2H), 3.76 (s, 3H), 3.57 (s, 4H), 3.44 (s, 4H), 2.89 (br.s, 2H), 2.32 (s, 3H).

MS: 357.2 (M+H) ⁺ .

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Preparative example 5.

Stage A.

A solution of 5-fluoro-2-hydrazineylpyridine (5 g) and tert-butyl-3-oxopiperidine-1-carboxylate (5 g) in methanol (50 ml) was stirred at 25°C for 12 hours. After completion of the reaction by TLC, the reaction mixture was concentrated and the dark brown crude product (10 g) was sent to the next step without further purification. MS: 309.1 (M+H)+.

Stage B.

A solution of the crude title compound obtained in Step A above (10 g) in diethylene glycol (20 ml) was heated at 180° C. using a microwave reactor for 90 minutes (4 loads). After completion of the reaction by LCMS, the reaction mixture was poured into water and then extracted with dichloromethane. The organic layer was separated, concentrated to give the crude product, which was washed with diethyl ether and dried in vacuo to give the title compound as a pale brown solid (1 g, 16%). MS: 192.2 (M+H)+.

Stage C.

To a solution of the title compound obtained in Step B above (1 g, 5.23 mmol) in THF (10 ml) was added triethylamine (2.2 ml, 15.39 mmol), and then di-tert-butyl dicarbonate ( 1.36 g, 6.28 mmol). The reaction mixture was then stirred at room temperature for 12 hours. After completion of the reaction as indicated by TLC, the solvent was removed and the crude reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (60:40) to give the title compound as a yellow solid (1.1 g, 72%).

1H-NMR (400 MHz, DMSO-d6) δ=11.54 (s, 1H), 8.08 (s, 1H), 7.65 (d, J=12.40 Hz, 1H), 3.65 -3.66 (m, 2H), 2.82 (t, J=8.80 Hz, 2H), 1.95 (t, J=6.40 Hz, 2H), 1.46 (s, 9H) . MS: 292.2 (M+H)+.

Stage D

To a solution of the title compound obtained in Step C above (1.1 g, 3.78 mmol) in K,R'-dimethylformamide (10 ml) was added sodium hydride (0.226 g, 5.67 mmol) at 0°C . The reaction mixture was stirred at 0°C for 30 min, and then methyl iodide (0.25 ml, 4.15 mmol) was added at 0°C. The reaction mixture was then stirred at 0°C for 30 minutes. After completion of the reaction by TLC, the reaction mixture was quenched with ice water and then extracted with ethyl acetate (20 ml). The organic layer was concentrated to give the crude product as a yellow solid (1 g), which was carried to the next step without further purification. MS: 306.1 (M+H)+.

Stage E.

To a solution of the title compound obtained in Step D above (1 g) in dichloromethane (10 ml) was added 2N. HCl (5 ml) in 1,4-dioxane. The reaction mixture was stirred at room temperature for 3 hours. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the title compound as a pale yellow solid (0.25 g, 37%).

1H-NMR (400 MHz, DMSO-d6) δ=11.32 (t, J=6.92 Hz, 2H), 8.32 (s, 1H), 7.87 (d, J=9.12 Hz , 1H), 3.73 (s, 3H), 3.45 (s, 2H), 2.94 (t, J=5.88 Hz, 2H), 2.17 (t, J=4.80 Hz , 2H).

MS: 206.1 (M+H)+.

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Preparative example 6.

Stage A.

To a solution of the title compound obtained in Step C of Preparation Example 5 (1.1 g, 3.78 mmol) in N,N'-dimethylformamide (10 mL) was added sodium hydride (0.226 g, 5.67 mmol) at 0 °C. The reaction mixture was stirred at 0°C for 30 min and then ptoluenesulfonyl chloride (0.788 g, 4.15 mmol) was added. The reaction mixture was stirred for 30 min at 0°C. The mixture was quenched with ice water and extracted with ethyl acetate (20 ml). The organic phase was separated, dried over Na2SO4, filtered and the solvent was removed under reduced pressure to give the crude title compound as a pale brown solid (1 g). The crude product was taken without further purification for the next step. MS: 446.1 (M+H)+.

Stage B.

To a solution of the title compound obtained in step A above (1 g) in dichloromethane (10 ml) was added 2N. HCl (5 ml) in 1,4-dioxane. The reaction mixture was stirred at room temperature for 3 hours. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the title compound as a pale yellow solid (0.75 g, 65%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ=8.40 (d, J=1.24 Hz, 1H), 7.97 (d, J=8.24 Hz, 2H), 7.88 ( kv, J=2.64 Hz, 1H), 7.40 (d, J=8.24 Hz, 2H), 3.37 (kv, J=5.20 Hz, 2H), 3.22 (t, J=6.12 Hz, 2H), 2.33 (d, J=12.52 Hz, 3H), 2.09 (t, J=5.00 Hz, 2H).

MS: 346.1 (M+H) ⁺ .

Preparative example 7.

.HCI

NH ₂

N N

N Vos

H ₂ S0 ₄ /100°C

Stage A

HCI

Ts

Stage E hci/DXM

„n minor

major

„ H minor (Boc) ₂ O

THF

Stage B

NaH/TsCl THF

major

Stage C separation by SFC method

Stage D ^m major

Stage A.

To a solution of 3-(fluorophenyl)hydrazine HCl salt (25 g, 153.7 mmol) and tert-butyl-3-oxopiperidine-1-carboxylate (30.59 g, 153.7 mmol) in 1,4-dioxane (250 ml ) added conc. H ₂ SO ₄ (25 ml) at 0°C. The reaction mixture was then heated at 110°C for 3 hours. The reaction mixture was cooled to 25°C, and the precipitate was filtered off. The solid was dissolved in water, made basic with NaOH solution and extracted with dichloromethane. The organic phase was separated, dried over Na ₂ SO ₄ , filtered and the solvent was removed under reduced pressure to give the cyclized title compounds as a pale brown solid (23 g). The crude mixture of regioisomers was taken without further purification for the next step. MS: 191 (M+H) ⁺ .

Stage B.

To a solution of the title compound obtained in Step A above (23 g, 121.05 mmol) in THF (250 ml) was added triethylamine (33.7 ml, 242.1 mmol) and di-tert-butyl dicarbonate (32 g, 145.2 mmol) and the reaction mixture was stirred for 12 hours. After completion of the reaction, as indicated by TLC, the solvent was removed under reduced pressure and the crude reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (80:20) to give of the title compounds as a pale yellow solid (7 g, 20%). MS: 191.2 (M-Vos) ⁺ .

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Stage C.

The mixture of regioisomers obtained in step B above (7.0 g, ratio: 95/5) was separated on a chiral SFC column (YMC Amylose-SA) to obtain the desired major regioisomer as a pale yellow solid with 100% purity of the regioisomer (3 g, 43%).

1H-NMR (400 MHz, DMSO-Y6) δ=10.95 (b.s., 1H), 7.35-7.37 (m, 1H), 7.09-7.12 (m, 1H), 6.80-6.85 (m, 1H), 4.54 (s, 2H), 3.66 (t, J=5.64 Hz, 2H), 2.67 (t, J=5.48 Hz , 2H), 1.44 (s, 9H).

MS: 191.2 (M-Vos) ⁺ .

A minor regioisomer was also isolated (0.3 g, 4.3%).

1H-NMR (400 MHz, DMSO-Y6) δ=11.16 (s, 1H), 7.13 (d, J=8.12 Hz, 1H), 6.96-7.01 (m, 1H) , 6.68-6.70 (m, 1H), 4.56 (s, 2H), 3.67 (t, J=5.64 Hz, 2H), 2.78 (t, J=5.24 Hz, 2H), 1.44 (s, 9H).

MS: 191.2 (M-Vos) ⁺ .

Stage D

To a solution of the major regioisomer obtained in step C above (1.5 g, 5.16 mmol) in N,N'dimethylformamide (15 ml) was added sodium hydride (0.3 g, 7.74 mmol), and then ptoluenesulfonyl chloride was added (1.17 g, 6.19 mmol). The reaction mixture was stirred for 30 minutes at room temperature. The mixture was dissolved in ethyl acetate (50 ml) and washed with water and brine. The organic phase was separated, dried over Na ₂ SO ₄ , filtered and the solvent was removed under reduced pressure. The crude material was purified by flash column chromatography using hexane/ethyl acetate (80:20) to give the title compound (2 g, 87%).

1H-NMR (400 MHz, DMSO-66) δ=7.75-7.78 (m, 3H), 7.47-7.48 (m, 1H), 7.38-7.40 (m, 2H ), 7.16-7.17 (m, 1H), 4.84 (s, 2H), 3.61-3.62 (m, 2H), 2.63-2.66 (m, 2H), 2.32 (s, 3H), 1.44 (s, 9H). MS: 345.2 (M-Vos) ⁺ .

Stage E.

To a solution of the title compound obtained in step D above (2 g, 4.5 mmol) in dichloromethane (120 ml) was added 2N. HCl (10 ml) in 1,4-dioxane. The reaction mixture was stirred at room temperature for 12 hours. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and the residue was washed with diethyl ether to obtain the title compound as an off-white solid (1.6 g, 94%).

1H-NMR (400 MHz, DMSO-Y6) δ=9.65 (b.s., 2H), 7.93 (d, J=7.92 Hz, 2H), 7.72-7.72 (m, 1H), 7.58-7.59 (m, 1H), 7.40 (d, J=8.24 Hz, 2H), 7.19-7.24 (m, 1H), 4.65 (s , 2H), 3.36 (br.s, 2H), 2.91 (br.s, 2H), 2.35 (s, 3H). MS: 345.2 (M+H) ⁺ .

Preparative example 8.

Stage A.

To a solution of the minor regioisomer (0.3 g, 1.034 mmol) obtained from Step C of Preparative Example 7 in THF (10 ml) was added sodium hydride (0.082 g, 2.06 mmol), followed by ptoluenesulfonyl chloride (0.294 g, 1.06 mmol). 55 mmol). The reaction mixture was stirred for 30 minutes at room temperature. The mixture was dissolved in ethyl acetate (30 ml) and washed with water and brine. The organic phase was separated, dried over Na ₂ SO ₄ , filtered and the solvent was removed under reduced pressure to give the crude title compound (0.25 g, 55%). The crude product was taken without further purification for the next step.

MS: 345.1 (M+H) ⁺ .

Stage B.

To a solution of the title compound obtained in step A above (0.25 g, 0.56 mmol) in dichloromethane (10 ml) was added 2N. HCl (2 ml) in 1,4-dioxane. The reaction mixture was stirred at room temperature for 12 hours. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the crude title compound as an off-white solid (0.2 g). MS: 345.1 (M+H) ⁺ .

Preparative example 9.

Stage A.

To a solution of the major regioisomer (1.0 mg, 3.44 mmol) obtained from step C of Preparative Example 7 in THF (15 ml) was added sodium hydride (0.275 g, 6.89 mmol), followed by methyl iodide (10.3 ml , 4.14 mmol). The reaction mixture was stirred at room temperature for 30 minutes. The mixture was dissolved in ethyl acetate (40 ml) and washed with water, brine and dried over Na ₂ SO ₄ . The non-purified reaction mixture was purified by flash column chromatography using hexane/ethyl acetate (70:30) to give the title compound (1.0 g, 96%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ=7.38-7.40 (m, 1H), 7.28-7.29 (m, 1H), 6.87 (d, J=12, 80 Hz, 1H), 4.59 (s,

2H), 3.61-3.63 (m, 5H), 2.68 (s, 2H), 1.45 (s, 9H). MS: 305.1 (M+H)+.

Stage B.

To a solution of the title compound obtained in step A above (1.0 g, 3.28 mmol) in dichloromethane (10 ml) was added 2N. HCl (5 ml) in 1,4-dioxane.

The reaction mixture was stirred at room temperature for 12 hours. After completion of the reaction, the reaction mixture was evaporated to remove the solvent and washed with diethyl ether to obtain the title compound as an off-white solid (0.75 g, 96%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ=9.86 (b.s., 2H), 7.46-7.47 (m, 1H), 7.34-7.37 (m, 1H) , 6.89-6.90 (m, 1H), 4.39 (s, 2H), 3.63 (s, 3H), 3.37 (d, J=6.04 Hz, 2H), 2, 91-2.93 (m, 2H).

MS: 205.1 (M+H) ⁺ .

Preparative example 10.

Stage A.

To a solution of (4-fluorophenyl)hydrazine HCl salt (2 g, 12.3 mmol) and tert-butyl-2-oxo-8azabicyclo[3.2.1]ocgan-8-carboxylate (2.7 g, 12.3 mmol ) in 1,4-dioxane (30 ml) was added conc. H2SO4 (2 ml) at 0°C. The reaction mixture was then heated at 100°C for 12 hours. After completion of the reaction, the reaction mixture was cooled to room temperature and the solvent was removed under high vacuum to obtain the crude material. The crude material was made alkaline with 30% sodium hydroxide solution and then extracted with ethyl acetate (50 ml). The ethyl acetate layer was washed with water (20 ml) and saline (10 ml). The organic layer was dried over Na ₂ SO ₄ , filtered and evaporated to give the crude title compound as a pale brown gum (1.6 g, 83%). The crude product was taken without further purification for the next step. MS: 217.2 (M+H) ⁺ .

Stage B.

To a solution of the title compound obtained in Step A above (2 g, 5.1 mmol) in THF (20 ml) was added triethylamine (2.56 ml, 18 mmol) and di-tert-butyl dicarbonate (2.21 g, 10 mmol), and the reaction mixture was stirred for 12 hours. After completion of the reaction, as indicated by TLC, the solvent was removed under reduced pressure and the crude reaction mixture was purified by flash column chromatography using 40%-50% ethyl acetate in petroleum ether to give the title compound as a pale yellow solid (0.9 g, 31%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ=10.94 (b.s., 1H), 7.22-7.23 (m, 2H), 6.80-6.81 (m, 1H) , 5.09 (d, J=5.20 Hz, 2H), 4.46 (sh.s, 1H), 3.22 (sh.s, 1H), 2.50-2.51 (m, 1H ), 2.22 (b.s., 1H), 2.06 (b.s., 1H), 1.79 (b.s., 1H), 1.581.60 (m, 1H), 1.42 (s. 9H). MS: 261.2 (M-tert-butyl) ⁺ .

Stage C.

To a solution of the title compound obtained in Step B above (1 g, 3.16 mmol) in THF (15 ml) was added sodium hydride (60% in mineral oil; 0.25 g, 6 .32 mmol). After addition was complete, the reaction mixture was left stirring at room temperature for 30 minutes, and then the reaction mixture was cooled again to 0°C. A solution of p-toluenesulfonyl chloride (0.72 g, 3.79 mmol) in THF (5 ml) was then added dropwise to the reaction mixture at 0° C. After addition was complete, the reaction mixture was left stirring at room temperature for 2 hours. After completion of the reaction by TLC, the reaction mixture was cooled to 0°C and quenched with ice water, and then extracted using ethyl acetate (50 ml). The ethyl acetate layer was washed with water (10 ml) and saline (10 ml). The organic layer was dried over

Na2SO4, filtered and evaporated to give the crude product, which was purified on a silica gel column using 15% to 25% ethyl acetate in petroleum ether to give the title compound as an off-white solid (0.9 g, 61%).

1H-NMR (400 MHz, DMSO-de) δ=8.01-8.02 (m, 1H), 7.58-7.60 (m, 2H), 7.50-7.51 (m, 1H ), 7.50-7.51 (m, 2H), 7.15-7.16 (m, 1H), 5.06 (b.s., 1H), 4.46 (b.s., 1H), 3.35 (b.s., 1H), 2.91 (b.s., 1H), 2.44 (b.s., 3H), 1.77-1.79 (m, 3H), 1.56- 1.58 (m, 1H), 1.27-1.33 (m, 9H). MS: 371.1 (M-Vos) ⁺ .

Stage D

To a solution of the title compound obtained in Step C above (0.9 g, 1.91 mmol) in dichloromethane (5 ml) was added 4 N. HCl (10 ml) in 1,4-dioxane. The reaction mixture was left stirring at ambient temperature for 12 hours. After completion of the reaction, the reaction mixture was evaporated to remove solvent and washed with diethyl ether (10 ml) to obtain the title compound as an off-white solid (0.65 g, 93% ). MS: 371.1 (M+H) ⁺ .

Preparative example 11.

Stage A.

Triethylamine (243 ml, 1.74 mol) and di-tert-butyl dicarbonate (228 g, 1.04 mol), and the reaction mixture was stirred at 25°C for 12 hours. After completion of the reaction (under TLC monitoring), water was added to the reaction mixture at ice-cooling and extracted with ethyl acetate (2x500 ml). The combined organic extracts were washed with brine (1x250 ml), dried over anhydrous Na ₂ SO ₄ , filtered and evaporated under reduced pressure to obtain the crude product. The crude material was stirred with diethyl ether (200 ml) and the solid thus obtained was filtered, washed with diethyl ether (2x100 ml) and dried to give the title compound as a brown solid (200 g, 84%).

1H-NMR (400 MHz, CDCl3) δ=7.94 (b.s., 1H), 7.51 (d, J=7.60 Hz, 1H), 7.33-7.34 (m, 1H) , 7.11-7.13 (m, 2H), 4.67 (s, 2H), 3.79 (sh.s, 2H), 2.83 (sh.s, 2H), 1.53 (s , 9H). MS: 273.2 (M+H) ⁺ .

Stage B.

To a stirred suspension of sodium hydride (15.86 g, 60% mineral oil, 1.10 mol) in dry THF (400 ml) at 0°C was slowly added a solution of the title compound obtained in step A above (100 g, 0.367 mol) in dry THF (1 L) and stirred at the same temperature for 30 min. A solution of p-toluenesulfonyl chloride (105 g, 0.55 mol) in dry THF (100 ml) was then added dropwise at 0°C and the reaction mixture was left to stir at 0°C for 1 hour. After completion of the reaction (under TLC monitoring) the reaction mixture was cooled to 0°C and quenched with ice water (500 ml) and then extracted using ethyl acetate (3x500 ml). The combined organic extracts were washed with water (2x500 ml), brine (1x250 ml) and dried over Na ₂ SO ₄ , filtered and evaporated under reduced pressure to obtain the crude product, which was triturated with hexane (250 ml). The solid thus obtained was filtered, washed with hexane (2x100 ml) and dried to give the title compound as a pale brown solid (130 g, 83%).

1H-NMR (400 MHz, DMSO-de) δ=8.01 (d, J=10.40 Hz, 1H), 7.77 (b.s., 2H), 7.45-7.48 (m, 1H), 7.13-7.24 (m, 4H), 4.88 (s, 2H), 3.63-3.65 (m, 2H), 2.65 (sh.s, 2H), 2 .31 (s, 3H), 1.46 (s, 9H).

MS: 327.1 (M ⁺ -Boc).

Stage C.

To a solution of the title compound obtained in Step B above (130 g, 0.30 mol) in 1,4-dioxane (1.3 L) was added 4 M HCl in 1,4-dioxane (500 ml) at 0° WITH. The reaction mixture was stirred at 25°C for 12 hours. After completion of the reaction, the reaction mixture was evaporated under reduced pressure and the residue was washed with diethyl ether to obtain the title compound as a pale brown solid (95 g, 94%).

1H-NMR (400 MHz, DMSO-de) δ=9.64 (b.s., 2H), 7.86-7.89 (m, 3H), 7.55 (d, J=10.40 Hz, 1H), 7.27-7.30 (m, 4H), 4.67 (s, 2H), 3.47 (b.s., 2H), 2.92 (b.s., 2H), 2.33 (s, 3H). MS: 327.1 (M+H) ⁺ .

- 33 044393

Preparative example 12.

Stage A.

To a stirred suspension of sodium hydride (15.86 g, 60% mineral oil, 1.10 mol) in dry THF (400 ml) at 0° C. was slowly added a solution of the title compound obtained in step A of Preparation Example 11 (100 g , 0.367 mol) in dry THF (1 L) and stirred at the same temperature for 30 minutes. A solution of methyl iodide (10 ml, 0.55 mol) in dry THF (100 ml) was then added dropwise at 0°C, and the reaction mixture was left stirring at 0°C for 1 hour. After completion of the reaction (under TLC monitoring), the reaction the mixture was cooled to 0°C and quenched with ice water (500 ml) and then extracted using ethyl acetate (3x500 ml). The combined organic extracts were washed with water (2x300 ml), brine (1x250 ml) and dried over Na ₂ SO ₄ , filtered and evaporated under reduced pressure to obtain the crude product, which was triturated with hexane (250 ml). The solid thus obtained was filtered, washed with hexane (2x100 ml) and dried to give the title compound as a brown solid (93 g, 88%).

1H-NMR (400 MHz, DMSO-d6) δ=7.39-7.41 (m, 2H), 7.09-7.11 (m, 1H), 6.98-7.00 (m, 1H ), 4.58 (s, 2H), 3.64-3.66 (m, 5H), 2.69 (s, 2H), 1.45 (s, 9H). MS: 287.2 (M+H)+.

Stage B.

To a solution of the title compound obtained in Step A above (93 g, 0.32 mol) in 1,4-dioxane (1.0 L) was added 4 M HCl in 1,4-dioxane (400 ml) at 0° WITH. The reaction mixture was stirred at 25°C for 12 hours. After completion of the reaction, the reaction mixture was evaporated under reduced pressure and the residue was washed with diethyl ether to obtain the title compound as a pale brown solid (58 g, 80%).

1H-NMR (400 MHz, DMSO-d6) δ=9.81 (b.s., 1H), 7.44-7.47 (m, 2H), 7.15-7.18 (m, 1H), 7.03-7.06 (m, 1H), 4.42 (s, 1H), 3.66 (s, 3H), 3.41-3.45 (m, 2H), 2.95 (w. s, 2H). MS: 187.1 (M+H) ⁺ .

Preparative example 13.

_Ν η ₂ ν

L R ^ ^ci ----Br - 'S ΤΓΦ/150 C

Microwave radiation/ZOmin Stage A

Stage A.

To a solution of 6-bromo-2-chlorobenzo[d]thiazole (1 g, 4.0 mmol) in ethanol (6 ml) was added a 4 M solution of methylamine (1.5 ml) and the reaction mixture was heated at 150°C for 45 min using a Biotage microwave reactor. The reaction mixture was cooled to room temperature. The solvent was removed under reduced pressure, the crude product was dissolved in dichloromethane (150 ml) and washed with 1 M NaOH, water and brine and dried over Na ₂ SO ₄ . The solvent was removed under reduced pressure to obtain a crude reaction mixture, which was purified by flash column chromatography using hexane:ethyl acetate (50:50) to obtain the title compound as a solid (0.35 g, 36%).

1H-NMR (400 MHz, CDCla) δ=7.72 (s, 1H), 7.41 (s, 1H), 7.29 (d, J=3.3 Hz, 1H), 5.40 (s , 1H), 3.13 (s, 3H).

Preparative example 14.

h ₂ ni T ^ci _______

Arn

ΤΓΦ/150 °C

Microwave radiation/ZOmin Stage A

Stage A.

To a solution of 5-bromo-2-chlorobenzo[d]thiazole (0.9 g, 3.62 mmol) in ethanol (12 ml) was added a 4 M solution of methylamine (1 ml) and the reaction mixture was heated at 150°C for 45 min using a Biotage microwave reactor. The reaction mixture was cooled to room temperature. The solvent was removed under reduced pressure and the crude reaction mixture was purified by flash column chromatography using hexane:ethyl acetate (50:50) to obtain the title compound as a solid (0.57 g, 65%).

- 34 044393

1H-NMR (400 MHz, CDCl ₃ ) δ=7.68 (t, J=2.0 Hz, 1H), 7.46 (d, J=8.3 Hz, 1H), 7.29 (d, J=0.7 Hz, 1H), 7.22 (dd, J=8.4, 1.9 Hz, 1H), 5.90 (s, 1H), 3.13 (s, 3H).

Preparative example 15.

N ^CI _Y VN ^L '

IXI G '^'S Et ₃ N, DXM, «.t., 4h'

Stage A

Stage A.

To a solution of 2,5-dichlorobenzo[d]thiazole (5 g, 24.5 mmol) in dry dichloromethane (50 ml) was added a 2 M solution of dimethylamine in THF (18.37 ml, 36.65 mmol) and the reaction mixture was cooled to 0°C. Triethylamine (6.8 mL, 49 mmol) was added dropwise to this cold reaction mixture. Once the addition was complete, the reaction mixture was left stirring at room temperature for 4 hours. After completion of the reaction, the reaction mixture was treated with water (2x20 ml) and extracted with dichloromethane. The organic layer was separated, dried over Na ₂ SO ₄ , filtered and evaporated to give a white solid, which was triturated with diethyl ether to give the title compound (4.5 g, 88%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ=7.77 (d, J=11.20 Hz, 1H), 7.46 (d, J=2.40 Hz, 1H), 7.05- 7.05 (m, 1H), 3.14 (s, 6H). MS: 213.4 (M+H)+.

Preparative example 16.

Microwave radiation/ZOmin Stage A

Stage A.

To a solution of 6-bromo-2-chlorobenzo[d]thiazole (0.45 g, 1.81 mmol) in ethanol (12 ml) was added a 2 M solution of dimethylamine in THF (3 ml) and the reaction mixture was heated at 150°C in for 45 minutes using a Biotage microwave reactor. The reaction mixture was cooled to room temperature. The solvent was removed under reduced pressure and the crude reaction mixture was purified by flash column chromatography using hexane:ethyl acetate (50:50) to obtain the title compound (0.441 g, 95%) as a solid.

1H-NMR (400 MHz, DMSO-d6) δ=7.70 (d, J=1.9 Hz, 1H), 7.43-7.35 (m, 2H), 3.20 (s, 6H) .

Preparative example 17.

HN O ⁴ - ^/ \ |G yci ------------*

EtjN, DXM, ct., 4 h CI

Stage A

Stage A.

To a solution of 2,6-dichlorobenzo[d]oxazole (5 g, 26.8 mmol) in dry dichloromethane (50 ml) was added morpholine (3.50 g, 40.3 mmol) and the reaction mixture was cooled to 0°C. Triethylamine (4.0 g, 39.6 mmol) was added dropwise to this cold reaction mixture. Once the addition was complete, the reaction mixture was left stirring at room temperature for 4 hours. After completion of the reaction, the reaction mixture was treated with water (2x20 ml) and extracted with dichloromethane. The organic layer was separated, dried over Na ₂ SO ₄ , filtered and evaporated to give a white solid, which was triturated with diethyl ether to give the title compound (5 g, 78%).

1H-NMR (400 MHz, DMSO-d6) δ=7.59 (d, J=2.80 Hz, 1H), 7.30 (d, J=11.20 Hz, 1H), 7.21 (dd , J=2.80, 11.20 Hz, 1H), 3.71-3.74 (m, 4H), 3.57-3.60 (m, 4H). MS: 239.2 (M+H) ⁺ .

Preparative example 18.

HN O

IC I ы С । Ν / \

YrVa ---- 04°

ChPO Et ₃ N, DXM, k.t., 4h ν ο

Stage A

- 35 044393

Stage A.

To a solution of 2,5-dichlorobenzo[(1]oxazole (5 g, 26.8 mmol) in dry dichloromethane (50 ml) was added morpholine (3.50 g, 40.3 mmol) and the reaction mixture was cooled to 0°C. Triethylamine (4.0 g, 39.6 mmol) was added dropwise to this cold reaction mixture. Once addition was complete, the reaction mixture was left stirring at room temperature for 4 hours. After completion of the reaction, the reaction mixture was treated with water (2x20 ml) and extracted with dichloromethane The organic layer was separated, dried over Na ₂ SO ₄ , filtered and evaporated to give a white solid, which was triturated with diethyl ether to give the title compound (5.2 g, 81%).

'H-NMR (400 MHz, DMSO-a ₆ ) 6=7.44 (d, >8.40 Hz, 1H), 7.36 (d, >2.40 Hz, 1H), 7.06 (dd , >2.00, 8.40 Hz, 1H), 3.71-3.73 (m, 4H), 3.59-3.61 (m, 4H).

MS: 239.2 (M+H) ⁺ .

Preparative example 19.

HN^O

N ^ci ___________► J Г ⁴ >- ^{N 0} s cr^ s

Et ₃ N, CH ₂ CI ₂

Stage A

Stage A.

To a stirred solution of commercially available 2,6-2,6-dichlorobenzo[(1]thiazole (500 g, 2.45 mol) in dichloromethane (4 L) was added triethylamine (1031 ml, 7.35 mol) and morpholine (290 ml , 3.67 mol) at 0° C. The reaction mixture was stirred at 25° C. for 48 hours. After completion of the reaction (under TLC monitoring), water was added to the reaction mixture and then extracted with dichloromethane (2x2.5 L).Organic layer dried over Na ₂ SO ₄ , filtered and evaporated under reduced pressure to obtain the crude product. Methyl tert-butyl ether (1 L) was added to the crude material and the mixture was stirred for 2 hours. The solid was collected by filtration and dried under linear vacuum in for 6 hours to obtain the title compound as a pale brown solid (530 g, 85%).

'H-NMR (400 MHz, DMSO-a ₆ ) 6 = 7.93-7.94 (m, 1H), 7.43-7.44 (m, 1H), 7.28-7.29 (m , 1H), 3.72-3.74 (m, 4H), 3.54-3.55 (m, 4H). MS: 255.1 (M+H) ⁺ .

Preparative example 20.

Et ₃ N, DCM, rt, 4 h

Stage A

Stage A.

To a solution of 2,5-dichlorobenzo[(1]thiazole (5 g, 24.5 mmol) in dry dichloromethane (50 ml) was added morpholine (3.19 g, 36.6 mmol) and the reaction mixture was cooled to 0°C. Triethylamine (3.71 mL, 36.7 mmol) was added dropwise to this cold reaction mixture and the reaction mixture was left stirring at room temperature for 4 hours. After completion of the reaction, the reaction mixture was treated with water (2x20 mL) and extracted with dichloromethane.Organic layer separated, dried over Na ₂ SO ₄ , filtered and evaporated under reduced pressure to give a white solid, which was triturated with diethyl ether to give the title compound (4.5 g, 86%).

'H-NMR (400 MHz, DMSO-c1 ₆ ) 6=7.82 (d, >8.00 Hz, 1H), 7.50 (d, >2.00 Hz, 1H), 7.11-7 .11 (m, 1H), 3.72-3.73 (m, 4H), 3.55-3.56 (m, 4H). MS: 255.4 (M+H) ⁺ .

Preparative example 21.

NO, /— _h HN N no ₂ ίι 10% Pd/C nh ₂ a o ^N o^ Τ' ^F K ₂ CO ₃ , CH ₃ CN N _H2 , EtOH y F N. _CuBr2 , _CH3CN F Stage A u Stage B u Stage C

Vg

Stage A.

To a solution of 3,4-difluoronitrobenzene (5 g, 31.4 mmol) in acetonitrile (50 ml) was added Nmethylpiperazine (3.7 g, 37.7 mmol) and potassium carbonate (12.8 g, 94.3 mmol). Then the reaction mixture was refluxed for 3 hours. The reaction mixture was filtered, concentrated

-36044393 under vacuum and the resulting solid was washed with ether to give the title compound as a yellow solid (5.2 g, 69%).

1H-NMR (300 MHz, CDCl3) δ=7.89-8.03 (m, 2H), 6.90-6.91 (m, 1H), 3.33-3.34 (m, 4H), 2.60-2.61 (m, 4H),

2.38 (s, 3H).

MS: 240.1 (M+H)+.

Stage B.

To a solution of the title compound obtained in Step A above (5.2 g, 21.7 mmol) in ethanol (100 ml) was added 10% Pd/C (0.5 g) and the reaction mixture was hydrogenated for 16 hours. The reaction mixture was filtered through a pad of celite and concentrated in vacuo to give the title compound as a brown solid (4.0 g, 88%).

1H-NMR (400 MHz, DMSOL) δ=6.75 (t, J=9.20 Hz, 1H), 6.31 (t, J=9.20 Hz, 2H), 4.97 (s, 2H ), 2.82 (b.s., 4H), 2.43 (b.s., 4H), 2.21 (s., 3H).

MS: 210.2 (M+H) ⁺ .

Stage C.

To a suspension of the title compound obtained in Step B above (3.0 g, 14.33 mmol) in acetonitrile (30 ml) at 0° C., tert-butyl nitrite (2.5 ml, 21.5 mmol) was added via syringe. within 10 min. Copper(II) bromide (3.8 g, 17.2 mmol) was then added in portions at 0°C and stirred for 30 min. The reaction mixture was allowed to warm to 25°C for 1 hour and was heated to 60°C for 4 hours. After completion of the reaction (under TLC monitoring), the reaction mixture was filtered through a pad of celite and washed with ethyl acetate to obtain the crude product. The crude material was purified by silica gel column chromatography (60-120) using dichloromethane/methanol (99:1) to give the title compound as a brown solid (1 g, 25%).

1H-NMR (400 MHz, DMSO-b ₆ ) δ=7.40-7.41 (m, 1H), 7.28-7.31 (m, 1H), 6.96-6.98 (m, 1H), 2.98-2.99 (m, 4H), 2.46-2.47 (m, 4H), 2.22 (s, 3H). MS: 275.1 (M+H) ⁺ .

Preparative example 22.

Stage A.

To a solution of 3,4-difluoronitrobenzene (0.5 g, 3.14 mmol) in ethyl acetate (10 ml) was added 81.3 ml (9.42 mmol) of trimethylamine, followed by 8-oxa-3-azabicyclo[3.2. 1]octane (0.56 g, 3.77 mmol). The reaction mixture was then stirred at room temperature for 16 hours. The reaction mixture was filtered, concentrated in vacuo, and the resulting solid was washed with ether to give the title compound as a yellow solid (0.7 g, 88%).

1H-NMR (400 MHz, DMSO-b ₆ ) δ=7.99-8.02 (m, 2H), 7.10 (t, J=8.88 Hz, 1H), 4.41 (b.s. , 2H), 3.40 (d, J=11.80 Hz, 2H), 3.13 (d, J=10.64 Hz, 2H), 1.89 (b.s., 4H). MS: 253.1 (M+H) ⁺ .

Stage B.

To a solution of the title compound obtained in Step A above (0.7 g, 2.77 mmol) in ethanol (20 ml) was added 10% Pd/C (0.1 g) and the reaction mixture was hydrogenated for 16 hours. The reaction mixture was filtered through a pad of celite and concentrated in vacuo to give the title compound as a pale brown tarry solid (0.6 g, 96%).

1H-NMR (400 MHz, DMSO-b ₆ ) δ=6.71 (t, J=9.64 Hz, 1H), 6.27-6.28 (m, 2H), 4.96 (s, 2H ), 4.29 (b.s., 2H), 2.80 (b.s., 4H), 1.92-1.93 (m, 2H), 1.77-1.78 (m, 2H).

MS: 223.1 (M+H) ⁺ .

Stage C.

To a suspension of the title compound obtained in Step B above (0.50 g, 2.24 mmol) in acetonitrile (10 ml) at 0° C., tert-butyl nitrite (0.4 ml, 3.35 mmol) was added via syringe. within 10 min. Copper(II) bromide (0.6 g, 2.68 mmol) was then added in portions at 0°C and stirred for 30 min. The reaction mixture was allowed to warm to 25°C for 1 hour and was heated to 60°C for 4 hours. After completion of the reaction (under TLC monitoring), the reaction mixture was filtered through a pad of celite and washed with ethyl acetate to obtain the crude product. The crude material was purified by silica gel column chromatography (60-120) using dichloromethane/methanol (99:1) to obtain the title compound as a pale brown solid (0.27 g, 42%).

1H-NMR (300 MHz, CDCl3) δ=7.17-7.19 (m, 2H), 6.75 (t, J=8.68 Hz, 1H), 4.43 (br.s, 2H) , 3.03-3.06 (m, 4H), 2.09 (sh.s, 2H), 1.98 (sh.s, 2H). MS: 288.0 (M+H) ⁺ .

- 37 044393

Preparative example 23.

l l KI, NajCOj L / \

IB, ζ%ΝΗ, _{+ Br/} vg -------► _Br -p_N_O

F DMF F

Stage A

Stage A.

To a solution of 4-bromo-3-fluoroaniline (1 g, 5.26 mmol) in N,N'-dimethylformamide (10 ml) was added potassium iodide (2.18 g, 13.1 mmol) and sodium carbonate (1.95 g, 18.4 mmol) and the reaction mixture was heated to 150°C. 1-Bromo-2-(2-bromoethoxy)ethane (1.34 g, 5.77 mmol) was then added and heating continued for 16 hours. After completion of the reaction (under TLC monitoring), the reaction mixture was diluted with water (25 ml) and extracted with ethyl acetate (2x15 ml). The combined organic extracts were dried over Na ₂ SO ₄ , filtered and evaporated under reduced pressure to obtain the crude product. The crude material was purified by flash column chromatography using hexane/ethyl acetate (80:20) to give the title compound (0.6 g, 43%).

1H-NMR (400 MHz, DMSO-d6) δ=7.43-7.44 (m, 1H), 6.93-6.94 (m, 1H), 6.73-6.74 (m, 1H ), 3.70-3.72 (m, 4H), 3.13-3.14 (m, 4H). MS: 261.9 (M+H) ⁺ .

Preparative example 24.

N m DG

Г Г >С1 ---------*- [ Г > ^Ν

Cl^ ⁰ Et ₃ N, Cl'^ ⁴ ^' ^{0 4}

DXM, c.t., 4 hours

Stage A

Stage A.

To a solution of 2,6-dichlorobenzo[d]oxazole (5 g, 26.6 mmol) in dry dichloromethane (50 ml) at 0°C was added a 2 M solution of dimethylamine in THF (26.6 ml, 53.2 mmol) and triethylamine (5.6 ml, 39.9 mmol). The reaction mixture was then stirred at 25°C for 4 hours. After completion of the reaction (under TLC monitoring), the reaction mixture was diluted with water (20 ml) and extracted with dichloromethane (2x20 ml). The combined organic extracts were dried over Na ₂ SO ₄ , filtered and evaporated under reduced pressure to obtain the crude product. The crude material was triturated with diethyl ether, filtered, washed with diethyl ether and dried to give the title compound as a solid (5 g, 96%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ=7.56 (s, 1H), 7.23-7.24 (m, 1H), 7.16-7.16 (m, 1H), 3 ,13 (s, 6H).

MS: 197.2 (M+H) ⁺ .

Preparative example 25.

Stage A.

To a solution of 5-bromo-2-fluoroaniline (1 g, 5.26 mmol) in N,N'-dimethylformamide (10 ml) was added potassium iodide (2.18 g, 13.1 mmol) and sodium carbonate (1.95 g, 18.4 mmol) and the reaction mixture was heated to 150°C. 1-Bromo-2-(2-bromoethoxy)ethane (1.34 g, 5.77 mmol) was then added and heating continued for 16 hours. After completion of the reaction (under TLC monitoring), the reaction mixture was diluted with water (25 ml) and extracted with ethyl acetate (2x15 ml). The combined organic extracts were dried over Na ₂ SO ₄ , filtered and evaporated under reduced pressure to obtain the crude product. The crude material was purified by flash column chromatography using hexane/ethyl acetate (80:20) to give the title compound (0.8 g, 58%). 1H-NMR (400 MHz, CDCl ₃ ) δ=7.03-7.04 (m, 2H), 6.90-6.92 (m, 1H), 3.87-3.88 (m, 4H) , 3.08-3.10 (m, 4H).

- 38 044393

Preparative example 26.

Acetone, bo°s, 12 h

L-proline, Cul, to ₂ with ₃ 1,4-dioxane, 80*C, 16 hours.

Stage B

Stage A

tert-BuONO, CuCi ₂ , ACN, from 0°С to room temperature, 1 h 65°С, 4 h

Stage D

70%H^o ₄ reflux, 2 hours

Stage C

Stage A.

A solution of 5-bromo-3-iodopyridin-2-amine (5 g, 16.73 mmol) and benzoyl isothiocyanate (3.29, 20.16 mmol) in acetone (10 ml) was stirred at 60 °C for 12 h. When the reaction was complete (under TLC monitoring), the reaction mixture was evaporated under reduced pressure and the solid was filtered, washed with hexane (200 mL) and dried to give the title compound as an off-white solid (4 g, 52%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ=12.35 (s, 1H), 11.86 (s, 1H), 8.64-8.65 (m, 2H), 7.99-7 .99 (m, 2H), 7.67 (s, 1H), 7.56 (d, J=9.40 Hz, 2H).

MS: 461.5 (M+H)+.

Stage B.

To a solution of the title compound obtained in Step A above (4 g, 8.67 mmol) in 1,4-dioxane (60 ml) at 25° C. was added potassium carbonate (2.5 g, 18.1 mmol), L-proline (0.28 g, 2.43 mmol) and copper (I) iodide (0.462 g, 2.42 mmol) and the reaction mixture was stirred at 80°C for 16 hours. After completion of the reaction (monitored by TLC) the reaction mixture was poured into water (100 ml) and saturated aqueous NH ₄ Cl (100 ml) and stirred at 25°C for 1 hour. The solid thus obtained was filtered, washed with saturated aqueous NH ₄ Cl (2x25 ml), water (2x25 ml) and dried to obtain the crude title compound as an off-white solid (2.5 g). MS: 336.0 (M+H) ⁺ .

Stage C.

A suspension of the crude title compound obtained in Step B above (2 g, 5.98 mmol) in H2SO4 (70%, 6 ml) was heated at 120°C for 2 hours. After completion of the reaction (under TLC monitoring), the reaction mixture cooled to 25°C and slowly poured into 100 ml of cold water. The reaction mixture was then made basic with aqueous NaOH (50%) and extracted with ethyl acetate (6x25 ml). The combined organic layers were dried over Na ₂ SO ₄ , filtered and the solvent was concentrated under reduced pressure to give the title compound as a light yellow solid (0.3 g, 23%). 1H-NMR (400 MHz, DMSO-d ₆ ) δ=8.27-8.31 (m, 2H), 8.11 (s, 2H). MS: 230.4 (M+H) ⁺ .

Stage D

To a suspension of the title compound obtained in Step C above (0.3 g, 1.3 mmol) in acetonitrile (5 ml) at 0° C., tert-butyl nitrite (0.2 g, 1.95 mmol) was added via syringe. within 10 min. Copper(II) chloride (0.2 g, 1.48 mmol) was then added in portions at 0°C and the reaction mixture was stirred for 30 min. The reaction mixture was allowed to warm to 25°C for 1 hour and heated to 65°C for 4 hours. After completion of the reaction (under TLC monitoring), the solvent was evaporated under reduced pressure and the resulting residue was diluted with water (20 ml) and extracted with dichloromethane/ methanol (95:5) (3x20 ml). The combined organic layers were washed with brine (10 ml), dried over Na ₂ SO ₄ , filtered and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography (60-120) using dichloromethane/methanol (99:1) to obtain the title compound as an off-white solid (0.15 g, 46%).

1H-NMR (400 MHz, DMSO-de) δ=8.91 (d, J=2.40 Hz, 1H), 8.82 (d, J=1.60 Hz, 1H). MS: 250.9 (M+H) ⁺ .

Stage E.

To a solution of the title compound obtained in Step D above (0.18 g, 0.72 mmol) in dry dichloromethane (5 ml) at 0° C. was added triethylamine (0.3 ml, 2.16 mmol) and morpholine ( 74 mg, 0.85 mmol) and the reaction mixture was stirred at 25°C for 6 hours. After completion of the reaction (under TLC monitoring), the reaction mixture was concentrated under reduced pressure to obtain the crude product, which was purified by silica gel column chromatography (60- 120 mesh) using hexane/ethyl acetate (70:30) to give the title compound as an almost yellow solid (0.18 g, 83%).

- 39 044393

1H-NMR (400 MHz, DMSO-ph) δ=8.49 (d, J=2.00 Hz, 1H), 8.38 (d, J=1.60 Hz, 1H), 3.72-3 .74 (m, 4H),

3.61-3.62 (m, 4H).

MS: 302.0 (M+H)+.

Preparative example 27.

Br^N Cl KSCN HCI S N Cl tert-butyl nitrite Cl

IJ ------► h ₂ n4 Ύ Y ---------* ^and “ ^z

H ₂ N '= EtOH, 100°C Ν CuBr ₂ ,MeCN,

40-454 from 0°Сdoc.t., 2h

Stage A Stage B morpholine, Et ₃ N

CH ₂ C1 ₂

Stage C

C.I.

Stage A.

To a solution of 2-bromo-6-chloropyridin-3-amine (5 g, 24.1 mmol) and potassium thiocyanate (7 g, 72.3 mmol) in ethanol (50 ml) was added hydrochloric acid (37%, 100 ml) and the reaction mixture was stirred at 100°C for 40-45 hours. Completion of the reaction was confirmed by TLC. The reaction mixture was cooled to room temperature and concentrated to give a brown solid, which was partitioned in dichloromethane (150 ml) and aqueous 1 N. NaOH (50 ml). The solid was filtered and dried to obtain the crude title compound as a light yellow solid (3.5 g, 79%). The product was taken without further purification for the next step. MS: 186.1 (M+H) ⁺ .

Stage B.

To a suspension of the title compound obtained in Step A above (1.5 g, 8.08 mmol) in acetonitrile (25 ml) at 0° C., tert-butyl nitrite (1.4 ml, 12.12 mmol) was added via syringe. within 10 min. Copper(II) bromide (2.16 g, 9.69 mmol) was then added in portions. After 30 min at 0°C, the reaction mixture was allowed to warm to room temperature over 2 hours. The progress of the reaction was monitored by TLC. After completion of the reaction, the solvent was evaporated and diluted with water (20 ml) and dichloromethane/methanol (95:5) (3x20 ml). The combined organic layers were washed with brine (10 ml), dried over Na ₂ SO4, filtered and concentrated under reduced pressure. The crude compound was purified by silica gel column chromatography (60-120), eluting with dichloromethane/methanol (99:1) to obtain the title compound as a pale yellow solid (0.65 g, 32%). The product was taken without further purification for the next step. MS: 248.5 (M+H) ⁺ .

Stage C.

To a solution of the crude title compound obtained in Step B above (0.65 g, 2.61 mmol) in dry dichloromethane (5 mL) was added triethylamine (1.1 mL, 7.83 mmol) and morpholine (0.34 g, 3.91 mmol). The reaction mixture was then stirred at room temperature for 6 hours. The reaction mixture was concentrated in vacuo.

The crude compound was purified by silica gel column chromatography (60-120), eluting with petroleum ether/ethyl acetate (50/50) to obtain the title compound as an almost yellow solid (0.6 g, 90%).

1H-NMR (400 MHz, DMSO-ph) δ=7.83 (d, J=8.40 Hz, 1H), 7.41 (d, J=8.44 Hz, 1H), 3.72-3 .74 (m, 2H), 3.59-3.60 (m, 2H).

MS: 256.0 (M+H) ⁺ .

Preparative example 28.

S^^Br Stage A _ morpholine

Stage A.

Commercially available 6-bromo-2-chlorothiazolo[4,5-c]pyridine (50 mg, 0.20 mmol) and morpholine (3.5 mL, 40.1 mmol) were added to the microwave reaction tube. The tube was capped and stirred at room temperature for 10 min and then at 150°C for 10 min in a microwave reactor (Biotage). The solvent was removed under reduced pressure to give the title compound (0.60 g, 78%).

1H-NMR (400 MHz, DMSO-66) δ=8.48 (s, 1H), 8.05 (s, 1H), 3.61 (dd, 4H), 3.17-3.03 (m, 4H).

- 40 044393

Preparative example 29.

Stage A.

To a stirred solution of commercially available (2-fluorophenyl)hydrazine hydrochloride (10.0 g, 0.0615 mol) and tert-butyl-3-oxopiperidine-1-carboxylate (12.3 g, 0.0615 mol) in 1. 4-dioxane (100.0 ml) was added with concentrated H ₂ SO ₄ (10.0 ml) (at 0°C), then heated to 100°C for 12 hours under nitrogen atmosphere.

The reaction mixture was concentrated, the crude product was made alkaline using 30% aqueous NaOH (pH 9-10) and then extracted with dichloromethane. The dichloromethane layer was concentrated under reduced pressure to give the title compound (10.0 g, crude) as a brown oil.

The crude compound was used in the next step without further purification.

MS: 191.1 (M+H)+.

Stage B.

To a solution of the crude title compound obtained in Step A above (10.0 g, crude) in tetrahydrofuran (100.0 ml) was added triethylamine (2.83 ml, 0.0205 mol) and di-tert-butyl dicarbonate ( 1.88 ml, 0.00820 mol) at 0°C, then stirred at room temperature for 12 hours under a nitrogen atmosphere.

The progress of the reaction was monitored by TLC and LCMS. The reaction mixture was diluted with ethyl acetate (50 ml) and water (100 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0^90/10) to give the title compound (1.5 g, 10%) as a pale yellow solid .

1H-NMR (400 MHz, DMSO-d6) δ 11.32 (b.s., 1H), 7.22 (d, J=7.64 Hz, 1H), 6.86-6.88 (m, 2H ), 4.58 (s, 2H), 3.68 (t, J=5.60 Hz, 2H), 2.69 (t, J=5.00 Hz, 2H), 1.44 (s, 9H ). MS: 235.1 (M+H)-tert-butyl.

Stage C.

To a suspension of sodium hydride (0.575 g, 15.0 mmol) in THF (10.0 mL) was added dropwise a solution in THF (10.0 mL) of the title compound obtained in Step B above (1.50 g, 5 .00 mmol) at 0°C and the mixture was stirred at room temperature for 60 min. Tosyl chloride (1.20 g, 6.00 mol) was added dropwise at 0°C and then stirred at room temperature for 3 hours under a nitrogen atmosphere.

After completion of the reaction by TLC, the reaction mixture was quenched with ice water and then extracted with ethyl acetate (50.0 ml). The organic layer was separated, dried over sodium sulfate, filtered and then concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0^80/20) to give the title compound (1.6 g, 71.8%) as an off-white solid .

1H-NMR (400 MHz, DMSO-d6) δ 7.76 (d, J=7.52 Hz, 2H), 7.42 (d, J=8.08 Hz, 2H), 7.31 (d, J=7.20 Hz, 1H), 7.22-7.24 (m, 1H), 7.07-7.09 (m, 1H), 4.94 (s, 2H), 3.69 (w .s, 2H), 2.69 (b.s, 2H), 2.35 (s, 3H), 1.46 (s, 9H).

MS: 345.1 (M+H)+-Voc.

Stage D

To a stirred solution of the title compound obtained in Step C above (1.6 g, 3.59 mmol) in dichloromethane (10.0 ml) was added 4 M HCl in 1,4-dioxane (10.00 ml) at 0° C, then stirred for 2 hours at 0°C and warmed to room temperature.

After completion of the reaction by TLC, the reaction mixture was concentrated to give the title compound (1.3 g, 94.6%) as a gray solid.

The crude compound was used in the next step without further purification.

MS: 345.1 (M+H) ⁺ .

- 41 044393

Preparative example 30.

Stage A.

To a stirred solution of commercially available (2-fluorophenyl)hydrazine hydrochloride (10.0 g, 0.0615 mol) and tert-butyl-3-oxopiperidine-1-carboxylate (12.3 g, 0.0615 mol) in 1. 4-dioxane (100.0 ml) was added with concentrated H ₂ SO ₄ (10.0 ml) at 0°C. The reaction mixture was then heated at 100°C for 12 hours under a nitrogen atmosphere.

The reaction mixture was concentrated and the crude product was made alkaline using 30% aqueous NaOH (pH=9-10) and then extracted with dichloromethane. The dichloromethane layer was concentrated under reduced pressure to give the title compound (10.0 g, crude) as a brown oil.

The crude compound was used in the next step without further purification.

MS: 191.1 (M+H)+.

Stage B.

To a stirred solution of the title compound obtained in Step A (10.0 g, crude) in tetrahydrofuran (100.0 ml) was added triethylamine (2.83 ml, 0.0205 mol) and di-tert-butyl dicarbonate (1.88 ml , 8.20 mmol) at 0°C. The mixture was stirred at room temperature for 12 hours under a nitrogen atmosphere. The progress of the reaction was monitored by TLC and LCMS. The reaction mixture was diluted with ethyl acetate (50 ml) and water (100 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0^90/10) to give the title compound (1.5 g, 10%) as a pale yellow solid.

1H-NMR (400 MHz, DMSO-d ₆ ) δ 11.32 (b.s., 1H), 7.22 (d, J=7.64 Hz, 1H), 6.86-6.88 (m, 2H), 4.58 (s, 2H), 3.68 (t, J=5.60 Hz, 2H), 2.69 (t, J=5.00 Hz, 2H), 1.44 (s, 9H). MS: 235.1 (M+H)-tert-butyl.

Stage C.

To a suspension of sodium hydride (0.421 g, 0.0110 mol) in THF (10.0 ml) was added dropwise a solution in THF (10 ml) of the title compound obtained in step B (1.1 g, 3.66 mol ) at 0°C. The mixture was then stirred at room temperature for 60 minutes. Iodomethane (0.624 g, 4.39 mmol) was added at 0° C. and then the mixture was stirred at room temperature for 3 hours under a nitrogen atmosphere.

After completion of the reaction by TLC, the reaction mixture was quenched with ice water and then extracted with ethyl acetate (50.0 ml). The organic layer was separated, dried over sodium sulfate, filtered and then concentrated to give the title compound (1.10 g, 96.2%) as a pale brown solid.

The crude compound was used in the next step without further purification.

MS: 305.3 (M+H) ⁺ .

Stage D

To a stirred solution of the title compound obtained in Step C (1.10 g, 3.52 mol) in dichloromethane (5.0 ml) was added a solution of 4 N. HCl in dioxane (10.00 ml) at 0°C. The mixture was stirred for 2 hours at 0° and warmed to room temperature.

The reaction mixture was concentrated and the crude product was washed with diethyl ether (10.00 mL) and dried in vacuo to give the title compound (750 mg, 78.8%) as a gray solid.

1H-NMR (400 MHz, DMSO-d ₆ ) δ 9.94 (b.s., 2H), 7.29 (d, J=7.60 Hz, 1H), 6.97-6.98 (m, 2H), 4.41 (s, 2H), 3.82 (s, 3H), 3.37-3.38 (m, 2H), 2.92-2.93 (m, 2H).

MS: 205.0 (M+H) ⁺ .

- 42 044393

Preparative example 31.

Stage A.

To a solution of commercially available 2,5-dichloro-1,3-benzoxazole (1.00 g, 5.32 mmol) in dichloromethane (50.0 ml), triethylamine (1.61 g, 1.60 mol) and 1-methylpiperazine (0.693 g, 6.38 mmol). The mixture was then stirred at 25°C for 12 hours. The reaction mixture was diluted with water (50.0 ml) and dichloromethane (50.0 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a DCM/MeOH gradient (100/0 >90/10) to give the title compound (1.0 g, 73.9%) as a white solid. 1H-NMR (400 MHz, DMSO-d6) δ 7.41 (d, J=8.44 Hz, 1H), 7.33 (s, 1H), 7.02-7.03 (m, 1H), 3.59-3.61 (m, 4H), 2.41-2.42 (m, 4H), 2.23 (s, 3H). MS: 252.1 (M+H)+.

Preparative example 32.

Pd ₂ (dba) ₃ , xanthos, NaOtBu, TOLUENE 100°С 5 4

Stage A

Stage A.

Commercially available 2,5-dibromopyridine (1.0 g, 4.22 mmol) and N-methylpiperazine (0.55 g, 5.49 mmol) were dissolved in degassed toluene and filled with a nitrogen atmosphere. Tris(dibenzylideneacetone)dipalladium(0) (0.077 g, 0.084 mmol), xantphos (0.147 g, 0.253 mmol) and sodium tert-butoxide (0.609 g, 6.33 mmol) were then added and the mixture was heated to 100°C for 5 h.

The reaction mixture was filtered through celite, washed with dichloromethane and methanol, and the solvents were evaporated under reduced pressure. The crude material was purified on an HP-Sil column (Biotage) using a DCM/MeOH gradient (100/0>96/04) to give the title compound (0.46 g, 37%) as a pale yellow solid. 1H-NMR (400 MHz, MeOD) δ 8.14-8.14 (m, 1H), 7.63-7.64 (m, 1H), 6.79 (d, J=9.08 Hz, 1H ), 3.54-3.56 (m, 4H), 2.55-2.56 (m, 4H), 2.36 (s, 3H). MS: 258.1 (M+2H) ⁺ .

Preparative example 33.

Pd ₂ (dba) ₃ , xanthos, NaOtBu, toluene 12 h

Stage A

Stage A.

Commercially available 2,5-dibromopyrazine (0.500 g, 2.10 mmol) and 1-methylpiperazine (0.105 g, 1.05 mmol), xanthos (0.073 g, 0.126 mmol), tris-(dibenzylideneacetone) dipalladium(0) (0 .0385 g, 0.126 mmol) and sodium tert-butoxide (0.404 g, 4.20 mmol) were added to degassed and dry toluene (15.0 ml). The vessel was filled with argon gas, sealed and heated at 100°C for 12 hours. The reaction mixture was filtered through celite and the filtrate was concentrated under reduced pressure to give the title compound (0.165 g, 28%) as a pale brown tarry solid . The crude compound was used in the next step without further purification. MS: 259.0 (M+2H) ⁺ .

- 43 044393

Preparative example 34.

_Λ ΟΗ _Λ

Ο>ι<Ο S

Br

K ₂ CO ₃ DMF90°C 12 h

Stage A

Stage A.

To a solution of commercially available 5-bromo-2-fluoropyridine (0.3 g, 1.70 mmol) and 6-oxa-3azabicyclo[3.1.1]heptane was added 4-methylbenzenesulfonate (0.463 g, 1.7 mmol) in potassium carbonate in DMF (0.707 g, 5.11 mmol). The reaction mixture was heated to 90°C for 12 hours. The reaction mixture was diluted with ethyl acetate (30 ml) and water (30 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na2SO4, filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a 30/70 ethyl acetate/petroleum ether gradient to give the title compound (0.14 g, 31%) as a white solid.

1H-NMR (400 MHz, DMSO-d ₆ ) δ 8.20 (d, J=3.20 Hz, 1H), 7.71-7.73 (m, 1H), 6.63 (d, J= 11.60 Hz, 1H), 4.69-4.71 (m, 2H), 3.64-3.68 (m, 2H), 3.54-3.55 (m, 2H), 1.89 -1.90 (m, 2H).

MS: 255.1 (M+H) ⁺ .

Preparative example 35.

Vg

Vg

C.I.

K _g CO ₃ DMF90°C 12 h

Stage A

Stage A.

The carbonate potassium (0.677 g, 4.902 mmol) and the mixture was heated at 90°C for 12 hours. The reaction mixture was diluted with ethyl acetate (30 ml) and water (30 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a 30/70 ethyl acetate/petroleum ether gradient to give the title compound (0.25 g, 60%).

MS: 271.1 (M+2H)+.

Preparative example 36.

HN

Vg

Vg

Vg

P d ₂ (d bah, cantphos, NaOtBu, toluene 100'C 5 h

Stage A

Stage A.

Commercially available 2,5-dibromopyridine (0.5 g, 2.2 mmol) and (R)-2-methylmorpholine (0.234 g, 2.9 mmol) were added to the reaction tube and degassed toluene (10.0 mL) was added. Xanthos (0.073 g, 2.1 mol), tris-(dibenzylideneacetone)dipalladium(0) (0.024 g, 0.4 mmol) and sodium tert-butoxide (0.608 g, 6.3 mmol) were then added and the solution was heated at 100°C for 5 hours in a sealed tube filled with argon gas.

- 44 044393

The reaction mixture was filtered through celite and the filtrate was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0>70/30) to give the title compound (0.3 g, 55%) as an off-white solid. 1H-NMR (400 MHz, DMSO-d ₆ ) δ 8.18 (d, J=3.20 Hz, 1H), 7.69-7.70 (m, 1H), 6.84 (d, J= 12.00 Hz, 1H), 3.87-3.88 (m, 3H), 3.48-3.49 (m, 2H), 2.74-2.76 (m, 1H), 1.14 (d, J=8.00 Hz, 3H). MS: 257.1 (M+H)+.

Preparative example 37.

Pd ₂ (dba) ₃ , KSZNTfoS, NaOtBu, toluene 100°C 5h

Stage A

Stage A.

Commercially available 2,5-dibromopyridine (0.5 g, 2.2 mmol) and (S)-2-methylmorpholine (0.234 g, 2.9 mmol) were added to the reaction tube and degassed toluene (10.0 mL) was added. Xanthos (0.073 g, 2.1 mol), tris-(dibenzylideneacetone)dipalladium(0) (0.024 g, 0.4 mmol) and sodium tert-butoxide (0.608 g, 6.3 mmol) were then added and the solution was heated at 100°C for 5 hours in a sealed tube filled with argon gas. The reaction mixture was filtered through celite and the filtrate was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0>70/30) to give the title compound (0.37 g, 68%) as an off-white solid. 1H-NMR (400 MHz, DMSOd ₆ ) δ 8.18 (d, J=2.32 Hz, 1H), 7.70-7.71 (m, 1H), 6.84 (d, J=9, 04 Hz, 1H), 3.90-3.91 (m, 3H), 3.52-3.53 (m, 2H), 2.52-2.53 (m, 1H), 1.15 (d , J=6.24 Hz, 3H). MS: 257.1 (M+H) ⁺ .

Preparative example 38.

Pd ₂ (dba) ₃ , xanthos, NaOtBu, toluene 100°С bh

Stage A

Stage A.

Commercially available 2,5-dibromopyridine (1 g, 4.22 mmol) and (R)-3-methylmorpholine (0.234 g, 2.9 mmol) were added to the reaction tube and degassed toluene (10.0 mL) was added. Xanthos (0.146 g, 0.253 mmol), tris-(dibenzylideneacetone)dipalladium(0) (0.048 g, 0.84 mmol) and sodium tert-butoxide (1.21 g, 12.66 mmol) were then added and the solution was heated at 100°C for 6 hours in a sealed tube filled with argon gas. The reaction mixture was filtered through celite and the filtrate was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether ethyl acetate gradient (100/0 >70/30) to give the title compound (0.17 g, 15%) as an off-white solid. The crude compound was used in the next step without further purification. MS: 257.1 (M+H) ⁺ .

Preparative example 39.

Stage A

Stage A.

To a solution of commercially available 5-bromo-2-fluoropyridine (0.5 g, 2.84 mmol) and 7-oxa-2azaspiro[3,5]nonane (0.36 g, 2.84 mol) in DMF (5 ml ) potassium carbonate (1.17 g, 8.52 mmol) was added and

- 45 044393 the mixture was heated to 90°C for 12 hours. The reaction mixture was diluted with ethyl acetate (30 ml) and water (30 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure to give the title compound (0.44 g, 55%).

¹ H-NMR (400 MHz, DMSO-d6) δ 8.12 (d, J=2.40 Hz, 1H), 7.64-7.65 (m, 1H), 6.36 (d, J= 11.60 Hz, 1H), 3.66-3.69 (m, 4H), 3.52-3.54 (m, 4H), 1.71-1.73 (m, 4H).

MS: 285.0 (M+H)+.

Preparative example 40.

^Br \. ^HN ^Br.

N CI ------------- N NX

K ₂ CO ₃ DMFAO'S 12 h ⁰

Stage A

Stage A.

Potassium carbonate (1.79 g, 13.02 mmol) and the mixture was heated to 90°C for 12 hours. The reaction mixture was diluted with ethyl acetate (50 ml) and water (30 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a 30/70 ethyl acetate/petroleum ether gradient to give the title compound (0.5 g, 42%).

¹ H-NMR (400 MHz, DMSO-d6) δ 8.14-8.15 (m, 1H), 7.63-7.64 (m, 1H), 6.85 (d, J=9.12 Hz, 1H), 3.86-3.87 (m, 2H), 3.39-3.40 (m, 1H), 3.27 (s, 3H), 3.14-3.15 (m, 2H), 1.84-1.85 (m, 2H), 1.35-1.36 (m, 2H).

MS: 273.1 (M+2H)+.

Preparative example 41.

Pd ₂ (dba) _3> xantphos, NaOtBu, toluene 100°C 6 h

Stage A

Stage A.

A solution of commercially available 2,5-dibromopyridine (0.5 g, 2.11 mmol) and ((1S,4S)-2-oxa-5azabicyclo[2.2.1]heptane (0.25 g, 2.53 mmol) was added into the reaction tube and add degassed toluene (10.0 ml).Then add xanthos (0.073 g, 0.127 mmol), tris-(dibenzylideneacetone)dipalladium(0) (0.024 g, 0.042 mmol) and sodium tert-butoxide (0.61 g, 6.3 mmol) and the solution was heated at 100°C for 6 hours in a sealed tube filled with argon gas. The reaction mixture was filtered through celite and the filtrate was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage), using a petroleum ether/ethyl acetate gradient (100/0^70/30) to give the title compound (0.3 g, 57%) as an off-white solid.

¹ H-NMR (400 MHz, DMSO-d6) δ 8.13 (s, 1H), 7.64-7.65 (m, 1H), 6.54 (d, J=8.80 Hz, 1H) , 4.80 (s, 1H), 4.65 (s, 1H), 3.76 (d, J=7.20 Hz, 1H), 3.61 (d, J=6.80 Hz, 1H) , 3.42 (d, J=10.00 Hz, 1H), 3.19 (d, J= 10.00 Hz, 1H), 1.86-1.89 (m, 2H). MS: 255.1 (M+H) ⁺ .

- 46 044393

Preparative example 42.

K ₂ CO ₃ DMF90°C 12 h

Stage A

Stage A.

To a solution of commercially available 5-bromo-2-fluoropyridine (0.23 g, 1.30 mmol) and ((1R,4R)-2-oxa5-azabicyclo[2.2.1]heptane (0.18 g, 1.30 mmol) in DMF (5 ml) was added potassium carbonate (0.54 g, 3.9 mmol) and the mixture was heated to 90°C for 12 hours. The reaction mixture was diluted with ethyl acetate (30 ml) and water (30 ml). the phase was separated and the aqueous phase was extracted with ethyl acetate two more times.The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure to give the title compound (0.32 g, 96%).1 HNMR (400 MHz, DMSO-d6) δ 8.13 (d, J=2.80 Hz, 1H), 7.64-7.64 (m, 1H), 6.54 (d, J=11.60 Hz, 1H), 4.81 (s, 1H), 4.65 (s, 1H), 3.76 (d, J=10.00 Hz, 1H), 3.61 (d, J=10.00 Hz, 1H), 3.43 (d, J=13.20 Hz, 1H), 3.20 (d, J=13.60 Hz, 1H), 1.83-1.86 (m, 2H).MS: 257.0 (M+2H)+.

Preparative example 43.

K ₂ CO ₃ DMF90°C 12 h

Stage A

Stage A.

Potassium carbonate ( 1.57 g, 11.43 mmol) and the mixture was heated to 90°C for 12 hours. The reaction mixture was diluted with ethyl acetate (50 ml) and water (30 ml).The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure.The crude product was purified on an HP-Sil column (Biotage) using a 30/70 ethyl acetate/petroleum ether gradient to give the title compound (0 .17 g, 12%).

1H-NMR (400 MHz, CDCl ₃ ) δ 8.20 (d, J=2.00 Hz, 1H), 7.54-7.54 (m, 1H), 6.54 (d, J=8, 80 Hz, 1H), 3.973.97 (m, 2H), 3.69-3.70 (m, 2H), 2.49-2.51 (m, 2H), 1.27 (d, J=6 ,40 Hz, 6H). MS: 271.1 (M+H) ⁺ .

Preparative example 44.

Pd ₂ (dba) ₃ , xanthos, NaOtBu, toluene 100'C 6 h

Stage A

Stage A.

A solution of commercially available 2,5-dibromopyridine (0.5 g, 2.11 mmol) and ((S)-3-methylmorpholine (0.213 g, 2.11 mmol) was added to the reaction tube and degassed toluene (10.0 mL) was added Xanthos (0.073 g, 0.12 mmol), tris-(dibenzylideneacetone)dipalladium(0) (0.038 g, 0.042 mmol) and sodium tert-butoxide (0.608 g, 6.33 mmol) were then added and the solution was heated at 100 °C for 6 hours in a sealed tube filled with argon gas. The reaction mixture was filtered through celite and the filtrate was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0^70 /30) to give the title compound (0.23 g, 42%) as a pale yellow solid.

- 47 044393

1H-NMR (400 MHz, CDCI3) δ 8.22-8.23 (m, 1H), 7.55-7.56 (m, 1H), 6.50 (d, J=9.04 Hz, 1H ), 4.23-4.24 (m,

1H), 4.01-4.02 (m, 1H), 3.76-3.77 (m, 3H), 3.59-3.60 (m, 1H), 3.16-3.17 ( m, 1H), 1.24 (d, J=6.72 Hz, 3H). MS:

259.1 (M+H)+.

Preparative example 45.

Pd ₂ (dba) ₃ , xanthos, NaOtBu, toluene YuO'S 6 h

Stage A

Stage A.

A solution of commercially available 2,5-dibromopyridine (0.5 g, 2.11 mmol) and 2-oxa-7azaspiro[3,5]nonane (0.268 g, 2.11 mmol) was added to the reaction tube and degassed toluene (10 ,0 ml). Xanthos (0.073 g, 0.12 mmol), tris-(dibenzylideneacetone)dipalladium(0) (0.038 g, 0.042 mmol) and sodium tert-butoxide (0.608 g, 6.33 mmol) were then added and the solution was heated at 100° C for 6 hours in a sealed tube filled with argon gas. The reaction mixture was filtered through celite and the filtrate was concentrated under reduced pressure to give the title compound (0.37 g, 62%) as an off-white solid. 1H-NMR (400 MHz, CDQ3) δ 8.19-8.19 (m, 1H), 7.52-7.53 (m, 1H), 6.59 (d, J=9.04 Hz, 1H ), 4.50 (s, 4H), 3.46-3.47 (m, 4H), 1.93-1.94 (m, 4H). MS: 285.0 (M+2H)+.

Preparative example 46.

K ₂ CO ₃ DMF90°C 12 h

Stage A

Stage A.

To a solution of commercially available 5-bromo-2-fluoropyridine (0.5 g, 3.34 mmol) and 3-oxa-8azabicyclo[3.2.1]octane (0.882 g, 5.0133 mmol) in DMF (5 ml) was added potassium carbonate (0.923 g, 6.6844 mmol) and the mixture was heated to 90°C for 12 hours. The reaction mixture was diluted with ethyl acetate (30 ml) and water (30 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure to give the title compound (0.5 g, 67%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ 8.17 (d, J=2.12 Hz, 1H), 7.67-7.68 (m, 1H), 6.80 (d, J= 8.92 Hz, 1H), 4.42 (s, 2H), 3.60-3.62 (m, 2H), 3.48-3.51 (m, 2H), 1.85-1.86 (m, 4H).

MS: 271.1 (M+2H)+.

Preparative example 47.

K ₂ CO ₃ DMFAYUO'S

Stage A

Stage A.

To a stirred suspension of the title compound obtained in Preparative Example 12 (0.370 g, 0.00199 mol) and commercially available 2,5-dichloro-1,3-benzoxazole (0.336 g, 0.00179 mol) in 15 ml DMF ( 15 ml) potassium carbonate (0.823 g, 0.00595 mol) was added and the mixture was heated at 100°C overnight. After completion of the reaction by TLC, water was added to obtain a solid. The solid was filtered and washed with hexane to give the title compound (0.4 g, crude).

MS: 338.1 (M+H) ⁺ .

- 48 044393

Preparative example 48.

Stage A

Stage A.

Potassium carbonate (0.445 g, 0.000756 mol) was added to a stirred suspension of the title compound prepared in Preparative Example 11 (0.350 g, 0.00107 mol) and commercially available (0.181 g, 0.000965 mol) in 15 ml DMF. and the reaction mixture was heated at 100°C overnight. After completion of the reaction by TLC, water was added to obtain a solid. The solid was filtered and washed with hexane to give the title compound (0.45 g, crude). MS: 478.1 (M+H)+

Preparative example 49.

Stage A

Stage A.

Triethylamine (0.516 g, 5 .0675 mmol) and the reaction mixture was heated at 80°C for 12 hours. The reaction mixture was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0 >70/30) to give the title compound (0.500 g, 64.93%).

1H-NMR (400 MHz, DMSO-d/ δ 7.56 (d, J=9.60 Hz, 1H), 7.40 (d, J=9.60 Hz, 1H), 4.18-4, 18 (m, 2H), 3.61-3.62 (m, 2H), 2.49-2.50 (m, 2H), 1.16 (d, J=6.40 Hz, 6H).

MS: 228.1 (M+H) ⁺ .

Preparative example 51.

Et ₃ N EtOH 80°С 12 h

Stage A

Stage A.

Triethylamine (0.509 g, 5.03 mmol) was added to a solution of commercially available 3,6-dichloropyridazine (0.500 g, 3.36 mmol) and (2S)-2methylmorpholine (0.339 g, 3.36 mmol) in ethanol (15 mL). and the reaction mixture was heated at 80°C for 12 hours. The reaction mixture was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0>70/30) to give the title compound (0.250 g, 34.5%).

1H-NMR (400 MHz, DMSO-d/ δ 7.57 (d, J=9.60 Hz, 1H), 7.40 (d, J=9.60 Hz, 1H), 4.08-4, 09 (m, 2H), 3.91-3.92 (m, 1H), 3.53-3.54 (m, 2H), 2.90-2.92 (m, 1H), 2.58- 2.61 (m, 1H), 1.16 (d, J=6.40 Hz, 3H) MS: 214.1 (M+n) ⁺ .

- 49 044393

Preparative example 52.

Et ₃ N EtOH ZO'S 12 H

Stage A

Stage A.

Triethylamine (0.509 g, 5.03 mmol) was added to a solution of commercially available 3,6-dichloropyridazine (0.500 g, 3.36 mmol) and (2R)-2methylmorpholine (0.339 g, 3.36 mmol) in ethanol (15 mL). and the reaction mixture was heated to 80°C for 12 hours. The reaction mixture was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0^70/30) to give the title compound (0.250 g, 34.5%).

1H-NMR (400 MHz, DMSO-d ₆ ) δ 7.57 (d, J=9.60 Hz, 1H), 7.40 (d, J=9.60 Hz, 1H), 3.91-3 .91 (m, 3H), 3.53-3.54 (m, 2H), 2.89-2.90 (m, 1H), 2.64-2.67 (m, 1H), 1.16 (d, J=6.40 Hz, 3H).

MS: 214.2 (M+H)+.

Preparative example 53.

Et ₃ N EtOH 9О°С 12 h

Stage A

Stage A.

To a solution of commercially available 3,6-dichloropyridazine (0.230 g, 1.54 mmol) and 4methoxypiperidine (0.265 g, 2.29 mmol) in ethanol (15 ml), triethylamine (0.301 g, 2.29 mmol) was added and the reaction mixture was heated to 90°C for 12 hours. The reaction mixture was concentrated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a petroleum ether/ethyl acetate gradient (100/0^70/30) to give the title compound (0.335 g, 95.44%).

1H-NMR (400 MHz, DMSOC) δ 7.50 (d, J=9.60 Hz, 1H), 7.41 (d, J=9.60 Hz, 1H), 3.92-3.93 ( m, 2H), 3.43-3.44 (m, 1H), 3.27-3.27 (m, 5H), 1.88-1.89 (m, 2H), 1.43-1, 44 (m, 2H).

MS: 228.1 (M+H) ⁺ .

Preparative example 54.

K ₂ CO ₃ DMF1OO°C

Stage A

Stage A.

To a stirred suspension of the title compound obtained in Preparative Example 12 (0.5 g, 2.24 mmol) and commercially available 2,6-dichloro-1,3-benzoxazole (0.43 g, 2.24 mmol) Potassium carbonate (0.93 g, 6.72 mmol) was added to DMF (15 ml) and the mixture was heated at 100°C overnight. After completion of the reaction by TLC, water was added to obtain a solid. The solid was filtered, washed with hexane to give the title compound (0.76 g, crude).

MS: 338.1 (M+H) ⁺ .

Examples 1-122.

Examples of the present invention were prepared in accordance with general procedures for Buchwald coupling, as shown in Schemes 5-7 above.

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Specific techniques used.

Method 1.

To a stirred solution of the tricyclic amine derivative (0.15 g, 1 eq.) in dry 1,4dioxane (5 ml) was added the appropriate bromo- or chlorine derivative (1 eq.), as indicated in the table. 1, and sodium tert-butoxide (3 eq.). The reaction mixture was degassed for 10 min under N2 atmosphere. Tris-(dibenzylideneacetone)dipalladium(0) (Pd ₂ (dba) ₃ ; 0.05 eq.) and 2-dicyclohexylphosphino2',6'-diisopropoxybiphenyl (Ru-Phos; 0.1 eq.) were then added and the reaction mixture was heated up to 100°C until the reaction is complete. After completion of the reaction (under LCMS monitoring), the reaction mixture was filtered through celite and washed with ethyl acetate. The filtrate was concentrated under reduced pressure to obtain the crude product. The crude material was purified by flash column chromatography or preparative HPLC to give the tosyl-protected compound. To a solution of the tosyl compound (1.0 eq.) in a mixture of dioxane:MeOH (1:1, 10 vol.) was added NaOtBu (3 eq.) and heated to 70°C for 6 hours. The reaction mixture was concentrated in vacuum and the crude the product was purified on a column to obtain the desired product. The crude material was purified by flash column chromatography or preparative HPLC to obtain the final compounds shown in table. 1.

Method 2.

To a stirred solution of the tricyclic amine derivative (0.15 g, 1 eq.) in dry 1,4dioxane (5 ml) was added the appropriate bromo- or chlorine derivative (1 eq.), as indicated in Table 1, and sodium tert-butoxide (3 eq.). The reaction mixture was degassed for 10 min under N2 atmosphere. Then tris-(dibenzylideneacetone)dipalladium(0) (Pd ₂ (dba) ₃ ; 0.05 eq.) and 2-dicyclohexylphosphino-2',6'-diisopropoxybiphenyl (Ru-Phos; 0.1 eq.) were added and the reaction the mixture was heated to 100°C until the reaction was complete. After completion of the reaction (under LCMS monitoring), the reaction mixture was filtered through celite and washed with ethyl acetate. The filtrate was concentrated under reduced pressure to obtain the crude product. The crude material was purified by flash column chromatography or preparative HPLC to obtain the final compounds shown in table. 1.

Method 3.

Pd(OAc) ₂ (0.1 eq) and xantphos (0.3 eq) were added to the reaction vessel and degassed dioxane (4 ml) was added. The vessel was filled with argon gas and hermetically sealed. The suspension was heated at 110°C for 1 min, then the tricyclic amine derivative (70 mg, 1 eq.), bromo- or chlorine derivative (1.1 eq.) and Cs ₂ CO ₃ (3.5 eq.) and solution heated at 100°C for 18 hours. The reaction mixture was diluted with ethyl acetate (30 ml) and water (30 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil column (Biotage) using a DCM/MeOH gradient (100/0^95/05) to give the tosyl-protected compound. Tosyl derivatives (50 mg, 1 eq.), Cs ₂ CO ₃ (3 eq.), followed by MeOH (ratio: 1.000, volume: 4 ml) and degassed THF (ratio: 2,000, volume: 8 ml). The reaction mixture was heated at 110°C for 30 min in a microwave reactor and cooled at room temperature. Solvents were removed under reduced pressure and the residue was purified on HP-Sil SNAP cartridges using the Biotage Isolera One purification system using a DCM/MeOH gradient (100/0^90/10) to obtain the final compounds shown in Table. 1.

Method 4.

To a stirred solution of the tricyclic amine derivative (0.15 g, 1 eq.) in dry 1,4dioxane (5 ml) was added the appropriate bromo- or chlorine derivative (1 eq.), as indicated in Table 1, and sodium Cs ₂ CO ₃ ( 3 eq.). The reaction mixture was degassed for 10 min under N2 atmosphere. Then Pd(OAc) ₂ was added; 0.1 eq.) and 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (XPhos; 0.3 eq.) and the reaction mixture was heated to 100°C until the reaction was complete. After completion of the reaction (under LCMS monitoring), the reaction mixture was filtered through celite and washed with ethyl acetate. The filtrate was concentrated under reduced pressure to obtain the crude product. The crude material was purified by flash column chromatography or preparative HPLC to give the tosyl-protected compound. To a solution of the tosyl compound (1.0 eq.) in a mixture of dioxane:MeOH (1:1, 10 vol.) was added NaOtBu (3 eq.) and heated to 70°C for 6 hours. The reaction mixture was concentrated in vacuum and the crude the product was purified on a column to obtain the desired product. The crude material was purified by flash column chromatography or preparative HPLC to obtain the final compounds as indicated in the table. 1.

Method 5.

To a stirred solution of the tricyclic amine derivative (150 mg, 1 eq.) in dry dioxane (5 ml) was added the appropriate bromo- or chlorine derivative (1 eq.), as indicated in the table. 1. Add sodium tert-butoxide (3 eq.) and degas for 10 min under nitrogen. Ruphos G4 Pd (0.3 eq) was added to this reaction mixture and heated to 100°C until the reaction was complete. The reaction mixture was filtered through a pad of celite and washed with EtOAc. The filtrate was concentrated and the crude product was purified by column chromatography or preparative HPLC to give the compound examples shown in Table. 1.

Table 1

Example Tricyclic amine derivative Bromine or chlorine derivative, aqueous Product Example 1. Exit; % 2. ¹ H-NMR 3. MH ⁺ (ESI) 4. Synthesis procedure 1 OnAn Ν I ^CI ^ ^N ^ N <4 ^N I 1. 11% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ) b = 7.57 (s, 1H), 7.39-7.40 (m, ZN), 7.09-7, 11 (m, 1H), 6.98-7.00 (m, 1H), 4.44 (s, 2H), 3.72-3.73 (m, 7H), 3.58-3.60 ( m, 2H), 3.48 (b.s., 4H), 2.82 (b.s., 2H). 3. 405.0 4. Method 2 2 SASPn Ν I AIN » _Ν / 1. 11% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.53 (s, 1H), 7.35-7.37 (m, ZN), 7.09-7, 11 (m, 1H), 6.98-7.00 (m, 1H), 4.42 (s, 2H), 3.71 (s, ZN), 3.56-3.57 (m, 2H) , 3.11 (s, 6H), 2.82 (b.s, 2H). 3. 363.2 4. Method 2

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3 E. €^Vz ^NH N 1 _O W> _ Qm-O-n ' 'Μ; ϊ 1 1. 19% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ) δ = 7.397.43 (m, 1 Η), 7.27-7.31 (μ, 2Η), 7.20 (d , J = 8.80 Hz, 1Η), 6.99 (dd, J = 2.40, 8.40 Hz, 1 Η), 6.82-6.87 (m, 1 Η), 4.40 ( sh.s, 2Η), 3.69-3.73 (μ, 7Η), 3.52-3.58 (μ, 6Η), 2.51-2.81 (μ, 2Η). 3. 407.0 4. Method 2 4 E. N 1 F θΆ~ ^Ν 0° 1. 19% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 7.397.42 (m, 1 Η), 7.28-7.31 (μ, 1Η), 6.93-6 .98 (μ, 2Η), 6.82-6.87 (μ, 2Η), 4.39 (sh.s., 2Η), 3.683.73 (μ, 7Η), 3.54-3.57 (μ , 2Η), 2.89-2.91 (μ, 4Η), 2.78-2.79 (m, 2Η). 3. 384.2 4. Method 2 5 R__ Cav3 ^nh N Tos F ^ci ~Oo F hl H 1. 19% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.92 (b.s., 1H), 7.28-7.32 (m, 1H), 7.13- 7.16 (m, 1H), 6.83-6.97 (m, ZN), 6.76-6.78 (m, 1H), 4.36 (sh.s, 2H), 3.71 -3.73 (m, 4H), 3.57-3.60 (m, 2H), 2.88-2.90 (m, 4H), 2.73-2.75 (m, 2H). 3. 370.2 4. Method 1 6 fX2wZ? ^nh N 1 οι^ΟζΙ-ιθ, Vr~N ¹ SA Q 1. 20% 2. ¹ H-NMR (400 MHz, DMSO-cfe) δ = 7.57 (d, J = 2.00 Hz, 1H), 7.447.39 (m,2H), 7.22-7 .15 (m, 2H), 6.97-6.91 (m, 1H), 4.43 (s, 2H), 3.743.72 (m, 7H), 3.59 (t, J = 5.20 Hz, 2H), 3.49 (t, J = 4.80 Hz, 4H), 2.79 (t, J = 4.80 Hz, 2H). 3. 423.2 4. Method 2

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7 Ν' 1 β ι ⁴ o \ 2-N Cl'' f-^WD ⁿ --ci NV ^s S 1 ^N 1. 24% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ) δ = 7.61 (d, J = 8.40 Hz, 1 Η), 7.397.43 (m, 1 Η), 7 .28-7.31 (μ, 1Η), 7.24 (d, J = 2.40 Hz, 1Η), 6.97 (dd, J = 2.40, 8.80 Hz, 1 Η), 6 ,83-6.87 (m, 1 Η), 4.46 (sh.s., 2Η), 3.70-3.75 (m, 7Η), 3.62-3.64 (μ, 2Η), 3.50-3.53 (μ.4Η), 2.79-2.81 (μ.2Η). 3. 423.0 4. Method 2 8 \Uk? ^nh N 1 ^ci <yo ΟαΓ/ν-ζλ N ^V N Ύθ 1. 21% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 8.05 (d, J = 2.80 Hz, 1 Η), 7.497.52 (μ, 1 Η), 7 .40-7.43 (μ, 1Η), 7.18-7.21 (μ, 1Η), 6.91-6.96 (μ, 1 Η), 6.81-6.83 (μ, 1 Η), 4.36 (s, 2Η), 3.70-3.72 (μ, 7Η), 3.48 (τ, J = 5.20 Hz, 2Η), 3.29-3.32 (m , 4H), 2.75-2.76 (m,2H). 3. 367.1 4. Method 2 9 CYYYnh 1 FQoCl' F\_ QYq-f 1. 29% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.407.44 (m, 1H), 7.30 (dd, J = 2.00, 10.40 Hz, 1H) , 7.04-7.09 (m, 1H), 6.83-6.90 (m, 2H), 6.59-6.62 (m, 1H), 4.41 (sh.s, 2H) , 3.87 (s, ZN), 3.69 (s, ZN), 3.56-3.59 (m, 2H), 2.79-2.82 (m, 2H). 3. 329.0 4. Method 2

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eleven I jZ AND / AN %\ ^F \_θ ^- Ong\ n VAs ¹ “V 1 1. 13% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 7.57 (d, J = 8.40 Hz, 1 Η), 7.407.44 (m, 1 Η), 7 ,18-7.22 (μ, 2Η), 6.91-6.96 (μ, 2Η), 4.47 (s, 2Η), 3.12 (s, 3Η), 3.63 (τ, J = 5.60 Hz, 2Η), 3.13 (s, 6Η), 2.79 (d, J = 4.80 Hz, 2Η). 3. 381.2 4. Method 2 12 E^cYY?nh I Cl^ G AN ⁴ F ₄ ___ Tsrchh w/ 1. 18% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.397.42 (m, 1H), 7.26 (d, J = 2.40 Hz, 1H), 7.177.20 (m, 1H), 3.52 (d, J = 8.40 Hz, 1H), 6.906.96 (m, 2H), 3.12 (s, 2H), 3.70 (s, ZN), 3 .54 (t, J = 5.60 Hz, 2H), 3.08 (s, 6H), 2.76 (t, J = 4.80 Hz, 2H). 3. 365.1 4. Method 2 13 F. QO N I szh r-Q ' L An F _x ___ ΟΖΖ'-ΓΆ N Jo ¹ ^N- 1 1. 12% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.407.44 (m, 1H), 7.25 (d, J = 8.64 Hz, 1H), 7.19 (dd, J = 2.48, 9.80 Hz, 1H), 7.06 (d, J = 2.40 Hz, 1H), 6.91-6.96 (m, 1H), 6.77 ( dd, J ⁼ 2.44, 8.72 Hz, 1H), 4.41 (b.s., 2H), 3.72 (s, ZN), 3.55-3.57 (m, 2H),3 .11 (s, 6H), 2.76-2.78 (m, 2H). 3. 365.2 4. Method 2

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14 ABOUT Ν' I _C i^4>~n I o _h Tsr-Qs I nA Ό 1. 25% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 7.60 (d, J = 8.80 Hz, 1 Η), 7.30 (d, J = 8.80 Hz, 1 Η), 7.23 (s, 1H), 6.93-6.98 (m, 2H), 6.74 (dd, J = 2.00, 8.60 Hz, 1H), 4, 45 (sh.s, 2N), 3.76 (s, ZN), 3.72-3.76 (m, 4N), 3.69 (s, ZN), 3.62-3.65 (m, 2H), 3.50-3.52 (m, 4H), 2.78-2.80 (m, 2H). 3. 435.2 4. Method 2 15 / Ο ΓΗν ^η I _C| <xDo I o ODD-'/CH N M% ¹ ЪA 1. 35% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.56 (d, J = 2.80 Hz, 1H), 7.40 (d, J = 8.80 Hz , 1H), 7.30 (d, J = 8.80 Hz, 1H), 7.16 (dd, J = 2.40, 8.80 Hz, 1H), 6.94 (d, J = 2, 40 Hz, 1H), 6.74 (dd, J = 2.40, 8.80 Hz, 1H), 4.41 (sh.s, 2H), 3.76 (s, ZN), 3.72- 3.74 (m, 4H), 3.68 (s, ZN), 3.57-3.60 (m, 2H), 3.47-3.50 (m, 4H), 2.78-2, 80 (m, 2H). 3. 435.2 4. Method 2 16 /o I ciAACvp °P>Oo ι nA Q 1. 12% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.277.31 (m, 2H), 7.09 (d, J = 2.40 Hz, 1H), 6.93 (d, J = 2.40 Hz, 1H), 6.82 (dd, J = 2.80, 8.60 Hz, 1H), 6.73 (dd, J = 2.40, 8.80 Hz, 1Н), 4.39 (sh.s, 2Н), 3.76 (s, ZN), 3.71-3.73 (m, 4Н), 3.67 (s, ZN), 3.55-3 .57 (m, 6H), 2.76-2.79 (m, 2H). 3. 419.2 4. Method 2

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17 / O I A / L HF <N \ MeO _s ___ ΟτΟ-ΓΎ N' W^S ^{1 n4} n / 1. 11% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 7.56 (d, J = 8.40 Hz, 1H), 7.30 (d, J = 9.20 Hz , 1H), 7.21 (s, 1H), 6.90-6.93 (m, 2H), 6.73 (dd, J = 2.00, 8.80 Hz, 1H), 4.44 ( sh.s, 2N), 3.76 (s, ZN), 3.69 (s, ZN), 3.613.64 (m, 2N), 3.12 (s, 6N), 2.78-2.79 (m,2H). 3. 393.2 4. Method 2 18 / O ΟίΝΗ I A 01-^^ F: MeO (Shl V N/ 1. 19% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 7.52 (d, J = 2.00 Hz, 1H), 7.297.37 (m, 2H), 7.12 (dd, J = 2.40, 8.80 Hz, 1H), 6.93 (d, J = 2.40 Hz, 1H), 6.73 (dd, J ⁼ 2.40, 9.00 Hz, 1Н), 4.39 (sh.s, 2Н), 3.76 (s, ZN), 3.67 (s, ZN), 3.55-3.58 (m, 2Н), 3.10 (s , 6H), 2.79-2.82 (m, 2H). 3. 393.2 4. Method 2 19 ό ΟγΌ ^η Ν I A ci-^^ Γ ΎΝ MeO / ^U N^ / 1. 13% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ) δ = 7.267.31 (m, 2H), 7.14-7.16 (m, 1H), 6.93-6, 97 (m, 2H), 6.74 (d, J = 8.80 Hz, 1H), 4.38 (sh.s, 2H), 3.76 (s, ZN), 3.67 (s, ZN ), 3.55-3.55 (m, 2H), 3.10 (s, 6H), 2.782.80 (m, 2H). 3. 377.2 4. Method 2

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20 o ΟγΌ ^η Ν I _Clxryo -o OrOo ν Oo 1. 15% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ) b = 7.277.31 (m, 2H), 7.20 (d, J = 8.80 Hz, 1H), 6,977.00 (m, 1H), 3.52 (d, J = 2.00 Hz, 1H), 6.726.75 (m, 1H), 3.12 (s, 2H), 3.76 (s, ZH), 3 .72 (t, J = 4.80 Hz, 4H), 3.67 (s, ZN), 3.52-3.58 (m, 6H), 2.78 (s, 2H). 3. 419.2 4. Method 2 21 / O oyNH I UYVn _c| A^nx MeO Q-Ohty 'N Vi^O ¹ 1 1. 22% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 7.247.31 (m, 2H), 7.05 (d, J = 2.12 Hz, 1H), 6.93 (d, J = 2.20 Hz, 1H), 6.726.77 (m, 2H), 4.38 (sh.s, 2H), 3.76 (s, ZN), 3.67 (s, ZN) , 3.54-3.57 (m, 2H), 3.11 (s, 6H), 2.77-2.79 (m, 2H). 3. 377.1 4. Method 2 22 XiXiang N ² Tos Cl^^ 4 k 0 1. 13% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.94 (b.s., 1H), 7.60 (d, J = 8.80 Hz, 1H), 7.297 .32 (m, 1H), 7.11-7.16 (m, 2H), 6.83-6.92 (m, 2H), 3.12 (s, 2H), 3.72 (t, J = 5.20 Hz, 4H), 3.66 (t, J = 5.60 Hz, 2H), 3.51 (t, J = 4.80 Hz, 4H), 2.75 (s, 2H). 3. 409.2 4. Method 1

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23 Tos οι-Α/ζΧίθ, 'γγΟΌΐ 1. 11% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.94 (br.s, 1H), 7.49 (s, 1H), 7.39 (d, J = 8.80 Hz, 1H), 7.29-7.32 (m, 1H), 7.16 (d, J = 10.00 Hz, 1H), 7.07-7.09 (m, 1H), 6.87 (t, J = 8.40 Hz, 1H), 4.38 (s, 2H), 3.73 (t, J = 4.80 Hz, 4H), 3.60 (d, J = 4 .80 Hz, 2H), 3.48 (t, J = 4.40 Hz, 4H), 2.78 (s, 2H). 3. 409.2 4. Method 1 24 F^ sUumn Ν Tos _ο <ϊΗ> ^F vvC ^N ^hl ⁰ o H <A> 1. 18% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.94 (b.s., 1H), 7.29-7.32 (m, 1H), 7.14- 7.21 (m, ZN), 6.84-6.93 (m, 2H), 4.36 (s, 2H), 3.12 (t, J = 4.80 Hz, 4H), 3.59 (t, J = 5.60 Hz, 2H), 3.53 (t, J = 4.40 Hz, 4H), 2.76 (s, 2H). 3. 393.1 4. Method 1 25 F_ CtC-nh Tos _/ ______. CI-Aj'lMj' ITr M~N 1^0 H 1. 23% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.93 (b.s., 1H), 7.26-7.31 (m, 2H), 7.15 ( d, J = 2.00 Hz, 1H), 3.52 (d, J = 2.40 Hz, 1H), 6.836.88 (m, 1H), 6.74-6.76 (m, 1H), 4.35 (s, 2H), 3.71 (t, J = 4.80 Hz, 4H), 3.54-3.60 (m, 6H), 2.75 (s, 2H). 3. 393.2 4. Method 1

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26 CjvJnh Tos £LH ci^^ ^{s F} \-X Q-O'^cy Ν 'ΓΑν ^H / 1. 17% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.93 (b.s., 1 Η), 7.45 (d, J = 2.40 Hz, 1H), 7.337.36 (m, 1H), 7.29-7.31 (m, 1H), 7.14-7.17 (m, 1H), 7.03-7.06 (m, 1H), 6. 83-6.89 (m, 1H), 4.36 (s, 2H), 3.58 (t, J = 5.60 Hz, 2H), 3.10 (s, 6H), 2.77 (s , 2H). 3. 367.1 4. Method 1 27 StS-MI γ Tos GT JN _C| >^ox E^O-Oita. N VZ^N ^H °% / 1. 13% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.95 (b.s., 1H), 7.29-7.30 (m, 1H), 7.13- 7.18 (m, ZN), 6.84-6.86 (m, 2H), 4.34 (s, 2H), 3.55-3.57 (m, 2H), 3.09 (s, 6H), 2.76-2.77 (m, 2H). 3. 351.2 4. Method 1 28 A-x CjvJnh Tos Г T />-N α>^Ν x F. Ο-θ-ττ n vzo ^H A / 1. 23% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.95 (b.s., 1H), 7.23-7.32 (m, 2H), 7.14- 7.16 (m, 1H), 6.96 (s, 1H), 6.846.88 (m, 1H), 6.68-6.71 (m, 1H), 4.35 (sh.s, 2H) , 3.58-3.59 (m, 2H), 3.10 (s, 6H), 2.68-2.75 (m, 2H). 3. 351.2 4. Method 1

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29 F _x ___ hT I N / G X ach OIDA'S ^x F ₄ ___ Qb ^N A, rr VO ¹ sA ^b N 1 1. 21% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 7.53 (d, J = 2.40 Hz, 1 Η), 7.407.41 (m, 1 Η), 7 .36 (d, J = 8.80 Hz, 1H), 7.20 (dd, J = 2.40, 9.60 Hz, 1H), 7.13 (dd, J = 2.80, 8.80 Hz, 1H), 6.916.91 (m, 1H), 4.41 (s, 2H), 3.71 (s, ZN), 3.56 (t, J = 5.60 Hz, 2H), 3, 11 (s, 6H), 2.77-2.79 (m, 2H). 3. 381.2 4. Method 2 thirty _ £/Nh Tos c—CAr ^ ^N ^O° __ Οχ ^ν π NW? ^HnA Ao 1. 19% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.57 (b.s., 1H), 7.51 (d, J = 11.60 Hz, 1H), 7 .13 (d, J= 10.80 Hz, 1H), 7.03 (s, 1H), 6.81-6.83 (m, 2H), 6.59 (d, J = 10.40 Hz, 1Н), 4.32 (s, 2Н), 3.66 (sh.s, 6Н), 3.58 (sh.s, ZN), 3.42 (sh.s, ZN), 2.67 (sh .s, ZN). 3. 421.1 4. Method 1 31 V^NH (A\^N Tos £ 1L An P 0|Ά>~3 -0. A>q ^H Sa Ao 1. 27% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.66 (b.s., 1H), 7.49 (d, J = 2.40 Hz, 1H), 7 .39 (d, J = 8.80 Hz, 1H), 7.21 (d, J = 8.80 Hz, 1H), 7.067.07 (m, 1H), 6.90 (d, J = 2, 40 Hz, 1H), 6.666.66 (m, 1H), 4.36 (s, 2H), 3.72-3.73 (m, 8H), 3.59-3.60 (m, 2H), 3.47-3.48 (m, 4H), 2.78-2.79 (m, 2H). 3. 421.1 4. Method 1

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32 N.H. TXX tos “-Olr <-0 -o Co 1. 17% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.66 (b.s., 1H), 7.17-7.18 (m, ZN), 6.89- 6.89 (m, 2H), 6.66-6.66 (m, 1H), 4.34 (s, 2H), 3.75 (s, ZH), 3.71-3.72 (m, 4H), 3.57-3.59 (m, 2H), 3.52-3.53 (m, 4H), 2.76-2.77 (m, 2H). 3. 405.2 4. Method 1 33 J^NH TjV tos ci—£%? -0. Tsr-Qo £-0 1. 29% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.65 (b.s., 1H), 7.27 (d, J = 8.80 Hz, 1H), 7 .20 (d, J = 8.80 Hz, 1H), 6.99 (d, J = 2.40 Hz, 1H), 6.89 (d, J = 2.40 Hz, 1H), 6.73 -6.74 (m, 1H), 6.65-6.66 (m, 1H), 4.34 (s, 2H), 3.70-3.71 (m, 4H), 3.55-3 .56 (m, 7H), 2.71 (sh.s, 2H), 2.51 (sh.s, 2H). 3. 405.2 4. Method 1 34 y~NH IXV tos Г £ H CI^^N ⁴ -4 Tsr-Qs " X / 1. 13% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.66 (b.s., 1H), 7.55 (d, J = 8.80 Hz, 1H), 7 .21 (d, J = 8.40 Hz, 1H), 7.08 (d, J = 2.00 Hz, 1H), 6.836.84 (m, 2H), 6.65-6.66 (m, 1H), 4.39 (s, 2H), 3.74 (s, ZN), 3.64-3.65 (m, 2H), 3.11 (s, 6H), 2.74-2.75 (m, 2H). 3. 379.2 4. Method 1

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35 .0^7^NH tos ^-CH, H 1. 11% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.66 (b.s., 1 Η), 7.45 (d, J = 2.40 Hz, 1H), 7.35 (d, J = 8.80 Hz, 1H), 7.20 (d, J = 8.40 Hz, 1H), 7.037.04 (m, 1H), 6.90 (d, J = 2 .00 Hz, 1H), 6.666.67 (m, 1H), 4.34 (s, 2H), 3.75 (s, ZN), 3.573.58 (m, 2H), 3.10 (s, 6H ), 2.77-2.78 (m,2H). 3. 379.2 4. Method 1 36 JNH TXV Tos ^ci Or, O^N rOr N Rn ^H ° ^Λ Ν- 1 1. 29% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.64 (b.s., 1H), 7.11-7.14 (m, ZN), 6.85- 6.86 (m, 2H), 6.64-6.65 (m, 1H), 4.31 (s, 2H), 3.74 (s, ZN), 3.54-3.56 (m, 2H), 3.08 (s, 6H), 2.74 (b.s, 2H). 3. 363.3 4. Method 1 37 /R ⁴ NH ( Jr- ⁷ \Rn Tos ^C| -O~l ^N 'O Op^n H 1. 25% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.85 (b.s., 1H), 7.60 (d, J = 8.80 Hz, 1H), 7 .39 (d, J = 7.60 Hz, 1H), 7.33 (d, J = 7.60 Hz, 1H), 7.12 (s, 1H), 7.02-7.04 (m, 1H), 6.97 (d, J = 7.20 Hz, 1H), 6.90-6.92 (m, 1H), 4.43 (s, 2H), 3.673.68 (m, 6H), 3.51-3.52 (m, 4H), 2.79 (sh.s, 2H). 3. 391.2 4. Method 1

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38 /νΎ^ΝΗ ίχν Tos A AVCh LTV ΑΛνΑ aa-n\A H 1. 27% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.84 (b.s., 1 Η), 7.49 (d, J = 2.40 Hz, 1H), 7.317.41 (m, ZN), 7.02-7.10 (m, 2H), 6.94-6.98 (m, 1H), 4.38 (s, 2H), 3.723.74 (m, 4H), 3.60-3.62 (m, 2H), 3.47-3.49 (m, 4H), 2.80-2.82 (m, 2H). 3. 391.2 4. Method 1 39 Χλ /2. X ⁴ ΝΗ τχν χ^ν Tos ^Вг Хд°1 H 1. 18% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.187.24 (m, 2H), 6.87-6.94 (m, 2H), 6.64-6, 69 (m, 2H), 3.52 (s, 2H), 3.74 (s, ZH), 3.12 (t, J = 5.20 Hz, 2H), 3.09 (s, 6H), 2.74 (s, 2H). 3. 363.2 4. Method 1 40 ΌΧ Tos „ λύ-νΏρ An ^- ⁷ H 1. 17% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.94 (b.s., 1H), 7.95 (d, J = 2.80 Hz, 1H), 7.417 .44 (m, 1H), 7.27-7.30 (m, 1H), 7.12-7.15 (m, 1H), 6.79-6.87 (m, 2H), 4.29 (s, 2H), 3.69 (t, J = 5.20 Hz, 4H), 3.49 (t, J = 5.60 Hz, 2H), 3.28 (t, J = 4.80 Hz , 4H), 2.73 (t, J = 5.20 Hz, 2H). 3. 353.3 4. Method 1

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41 NH w tos HClP) ^B nA ^Ρ νΥ^ ^Ν Λ^ ^Νν - ⁷ ° ]| γ -/ N-^ H 1. Yield: 16% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ 10.95 (b.s., 1 Η), 7.86 (d, J = 2.80 Hz, 1H) , 7.337.36 (m, 1Η), 7.27-7.29 (m, 1Η), 7.12-7.15 (μ, 1Η), 3.12 (d, J = 9.20 Hz, 1Η), 6.83-6.88 (μ, 1Η), 4.66 (s, 2Η), 3.83 (τ, J = 5.60 Hz, 2Η), 3.72 (τ, J = 4 .80 Hz, 4Η), 2.97 (τ, J = 4.80 Hz, 4Η), 2.72 (τ, J = 5.20 Hz, 2Η). 3. 353.2 4. Method 1 42 Υ-ΤΛ,ΝΗ N ^? 'Tos bY\. Y^-s ^N= V I F _x ___ O'-Αγ N 'YS ^H 1 1. 13% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.94 (b.s., 1 Η), 7.55 (d, J = 8.40 Hz, 1H), 7.297.32 (m, 1H), 7.09-7.16 (m, 2H), 6.83-6.88 (m, 2H), 3.12 (s, 2H), 3.65 (t, J = 5.60 Hz, 2H), 3.11 (s, 6H), 2.76 (d, J = 5.20 Hz, 2H). 3. 367.2 4. Method 1 43 yyy at Tos bYY n-AnJo OtO'-ay ka O H'—/ 1. 21% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.85 (b.s., 1H), 7.86 (d, J = 2.80 Hz, 1H), 7.297 .31 (m, ZN), 7.00-7.01 (m, 1H), 6.93-6.95 (m, 2H), 4.67 (s, 2H), 3.833.84 (m, 2H ), 3.71-3.73 (m, 4H), 2.96-2.97 (m, 4H), 2.74-2.76 (m, 2H). 3. 335.2 4. Method 1

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44 QHnh Tos input w^V and Χ ^Ν Ο> 1. 23% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ) δ = 10.82 (b.s., 1H), 7.96 (d, J = 2.84 Hz, 1H), 7.307 .44 (m, ZN), 7.01-7.03 (m, 1H), 6.95-6.97 (m, 1H), 6.80 (d, J = 9.12 Hz, 1H), 4.30 (s, 2H), 3.68-3.70 (m, 4H), 3.48-3.51 (m, 2H), 3.27-3.30 (m, 4H), 2. 76-2.77 (m, 2H). 3. 335.2 4. Method 1 45 1 βΧλ '—χ n-AnJo ΟτΟ'ΎΧ Ν Ν-ΧΝ^ρ 1. 24% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 7.89 (d, J = 2.80 Hz, 1H), 7.357.43 (m, ZN), 7.08 -7.10 (m, 1H), 6.99-7.03 (m, 2H), 4.69 (s, 2H), 3.833.86 (m, 2H), 3.70-3.75 (m , 7H), 3.70-3.72 (m, 7H), 2.97-3.00 (m, 4H), 2.77-2.79 (m, 2H). 3. 349.2 4. Method 2 46 θ ^- Osh 1 ^Βγ Ά_ υΛΟ COO'-hu Ν W^^J ⁰ 1. 28% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 8.05 (d, J = 2.80 Hz, 1H), 7.497.52 (m, 1H), 7.40 -7.43 (m, 2H), 7.09-7.13 (m, 1H), 6.98-7.02 (m, 1H), 6.82 (d, J = 9.20 Hz, 1H ), 4.37 (s, 2H), 3.703.72 (m, 7H), 3.48-3.51 (m, 2H), 3.29-3.34 (m, 4H), 2.79- 2.80 (m, 2H). 3. 349.2 4. Method 2

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47 ΎγΑ ^ΝΗ Tos Br—(~\-П Ъ N—' Υγ^ΑΑΟ Η 1. 14% 2. ¹ H-NMR (400 MHz, CD ₃ OD) δ = 7.76 (s, 1H), 7.20-7.23 (m,2H), 7.14-7.17 ( m, 1H), 6.74-6.79 (m, 2H), 5.22 (d, J = 5.20 Hz, 1H), 4.84-4.87 (m, 1H), 3.78 (t, J = 9.20 Hz, 4H), 3.34-3.38 (m, 1H), 2.95 (t, J = 6.00 Hz, 4H), 2.32-2.49 ( m, ZN), 2.02-2.07 (m, 1H), 1.79-1.86 (m, 1H). 3. 379.2 4. Method 1 48 voL ^nh Tos F ^ΒΓ Ύν WL ^N Tt ή n % 1. 13% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.96 (b.s., 1H), 7.84 (d, J = 1.44 Hz, 1H), 7.457 .45 (m, 1H), 7.28-7.30 (m, 1H), 7.14-7.15 (m, 1H), 6.84-6.84 (m, 1H), 4.40 (s, 2H), 3.71-3.72 (m, 4H), 3.58-3.60 (m, 2H), 3.15-3.16 (m, 4H), 2.75-2 .76 (m, 2H). 3. 371.2 4. Method 1 49 F _x ___ С^-С^и Tos ^0 m Αργν n sA ^b Ν'/ %Ο 1. 28% 2. ^1H NMR (400 MHz, DMSO-s) δ 10.97 (s, 1H), 8.34 (d, J = 0.7 Hz, 1H), 7.49 (d, J = 0.9 Hz, 1H), 7.29(dd, J = 8.8, 4.5 Hz, 1H), 7.14 (dd, J = 9.9, 2.6 Hz, 1H), 6 .95 - 6.71 (m, 1H), 4.74 (s, 2H), 3.90 (t, J = 5.7 Hz, 2H), 3.73 (t, J = 4.9 Hz, 4H), 3.51 (t, J = 4.9 Hz, 4H), 2.76 (d, J = 5.9 Hz, 2H). 3. 410.15 4. Method 3 and 5

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50 Tos C~ ^Q nY ^Br 4Y '-N /-0 Uy/x \UG\)*-nU\ NW 1. 25% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.93 (bs, 1H), 7.88 (s, 1H), 7.76 (s, 1H) , 7.29-7.30 (m, 1H), 7.17 (d, J = 9.60 Hz, 1H), 6.846.87 (m, 2H), 4.47 (s, 2H), 3, 74-3.75 (m,4H), 3.67-3.68 (m, 2H), 3.17-3.18 (m,4H), 2.77 (sh.s, 2H). 3. 353.2 4. Method 1 51 fO~L ^nh Tos FW^O f-CHUY Ν 1. 17% 2. ¹ H-NMR (400 MHz, DMSO-cfe) δ = 10.96 (b.s., 1H), 7.84 (d, J = 1.52 Hz, 1H), 7.46 (dd, J = 2.44, 15.42 Hz, 1H), 7.36-7.38 (m, 1H), 7.11 (dd, J = 2.28, 10.20 Hz, 1H), 6.796.80 (m, 1H), 4.38 (s, 2H), 3.71-3.72 (m, 4H), 3.58-3.59 (m, 2H), 3.15-3, 16 (m, 4H), 2.75-2.77 (m, 2H). 3. 371.2 4. Method 1 52 Tos F Br-Z=< vy-NQo ρΌπΟ'ΎΥ. VZ-N Η УУ 1. 29% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.94 (b.s., 1H), 7.84 (d, J = 1.52 Hz, 1H), 7.447 .45 (m, 1H), 7.35-7.37 (m, 1H), 7.08-7.10 (m, 1H), 6.78-6.80 (m, 2H), 4.39 (s, 2H), 3.72-3.73 (m, 4H), 3.56-3.57 (m, 2H), 3.16-3.17 (m, 4H), 2.75-2 .78 (m, 2H). 3. 370.2 4. Method 1

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53 F _x ___ Tos Br^ ^{S b} N\ Uo 1. 2% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ 10.96 (s, 1H), 8.19 (d, J = 2.8 Hz, 1H), 8.00 ( d, J = 2.8 Hz, 1H), 7.31 (dd, J = 8.7, 4.6 Hz, 1H), 7.15 (td, J = 9.0, 8.2, 2, 6 Hz, 1H), 6.87 (td, J = 9.2, 2.6 Hz, 1H), 4.42 (s, 2H), 3.74 (kv, J = 5.6, 5.1 Hz, 4H), 3.63 (kv, J = 5.6, 4.9 Hz, 2H), 3.59 - 3.50 (m, 4H), 2.79 (t, J = 5.6 Hz , 2H). 3. 410.15 4. Method 3 54 Tos ^Вг -/Л №=/ N %A ^S N^ %0 OtOal n^ nAn ^H 1. 22% 2. ¹ H NMR (400 MHz, DMSO-s/b) δ 10.97 (s, 1H), 7.69 (d, J = 8.9 Hz, 1H), 7.30 (dd , J = 8.7, 4.5 Hz, 1H), 7.15 (dd, J = 9.9, 2.6 Hz, 1H), 6.99 (d, J = 8.9 Hz, 1H) , 6.86 (td, J = 9.2, 2.6 Hz, 1H), 4.73 (s, 2H), 3.91 (t, J = 5.6 Hz, 2H), 3.72 ( dd, J = 5.8, 3.9 Hz, 4H), 3.48 (t, J = 4.9 Hz, 4H), 2.83-2.67 (m, 2H). 3. 410.14 4. Method 3 55 R_ Tos UAnQo __ Ql?nY\ /A H 4 1. 27% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.92 (b.s., 1H), 7.28-7.29 (m, 1H), 7.14 ( dd, J = 2.48, 9.86 Hz, 1H), 6.96-6.98 (m, 2H), 6.83-6.83 (m, ZN), 4.29 (s, 2H) , 3.71-3.72 (m, 4H), 3.51 (t, J = 5.52 Hz, 2H), 2.94-2.97 (m, 4H), 2.73 (t, J = 5.28 Hz, 2H). 3. 352.2 4. Method 1

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56 N.H. Tos ^Вг ~Г\ /-Х K/ ^N O F R Ον/ΥΎ ^-x ⁴ A-^“ ^N /o HF P 1. 20% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.92 (b.s., 1H), 7.30 (dd, J = 4.40, 8.80 Hz, 1H), 7.15 (dd, J = 2.40, 9.60 Hz, 1H), 6.976.99 (m, 1H), 6.81-6.82 (m, 2H), 6.68 (dd , J = 2.40, 8.80 Hz, 1H), 4.19 (s, 2H), 3.71-3.72 (m, 4H), 3.34-3.35 (m, 2H), 3.03-3.04 (m, 4H), 2.71-2.72 (m, 2H). 3. 370.2 4. Method 1 57 F-Vxk ^NH N' Tos ^Br Y~V --ι f-Yo4 ⁿ YV ^~l V b H <У 1. 33% 2. ¹ H-NMR (400 MHz, DMSO-s/b) δ = 10.92 (b.s., 1H), 7.35-7.36 (m, 1H), 7.10 ( dd, J = 2.16, 10.22 Hz, 1H), 6.92-6.96 (m, 2H), 6.87-6.89 (m, 2H), 6.79-6.79 ( m, 1H), 4.27 (s, 2H), 3.71-3.73 (m, 4H), 3.49-3.50 (m, 2H), 3.08-3.09 (m, 4H), 2.74-2.75 (m, 2H). 3. 352.3 4. Method 4 58 V/N N Tos __F ^V P yi.N'Zt kAN b H'—' 1. 16% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.82 (b.s., 1H), 7.39 (d, J = 7.64 Hz, 1H), 7 .32 (d, J = 7.96 Hz, 1H), 7.04 (t, J = 7.64 Hz, 1H), 6.896.93 (m, ZN), 6.78 (d, J = 8, 72 Hz, 1H), 4.36 (s, 2H), 3.72-3.73 (m, 4H), 3.58-3.59 (m, 2H), 2.89-2.90 (m , 4H), 2.77 (b.s., 2H). 3. 352.3 4. Method 1

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59 QP ^NH Tos Vg-UCH UUnQo Ova-'/'v Vn b H'—' 1. 15% 2. Ή-NMR (400 MHz, DMSO-c/s) δ = 10.81 (b.s., 1H), 7.39 (d, J = 7.68 Hz, 1H), 7.31 (d, J = 7.92 Hz, 1H), 7.017.03 (m, 1H), 6.96-6.96 (m, ZN), 6.87-6.89 (m, 2H), 4.29 (s , 2H), 3.723.73 (m, 4H), 3.51-3.52 (m, 2H), 2.97-2.98 (m, 4H), 2.77 (b.s., 2H). 3. 334.2 4. Method 1 62 pP ^nh H vg-oo ypOO ^- Η 18%; 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ = 10.92 (b.s, 1H), 7.34-7.36 (m, 1H), 7.10 (dd, J = 2.12, 10.16 Hz, 1H), 6.94-6.96 (m, 2H), 6.78-6.79 (m, ZN), 4.26 (s, 2H), 3.48 -3.49 (m, 2Н), 3.01 (sh.s, 4Н), 2.75 (sh.s, 2Н), 2.67 (sh.s, 2Н), 2.33 (sh.s , 2H), 2.23 (s, ZN). 3. 365.2 4. Method 4 67 hn3° OPAsR° 1. 43.55% 2. 1. 43.55% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 7.45 (t, J = 7.28 Hz, 2H), 7. 20 (d, J = 8.56 Hz, 1H), 7.127.13 (m, 2H), 7.02 (t, J = 7.20 Hz, 1H), 6.816.82 (m, 1H), 4, 88 (s, 2H), 3.95 (t, J = 5.56 Hz, 2H), 3.74-3.75 (m, 4H), 3.71 (s, ZN), 3.053.06 (m , 4H), 2.87 (t, J = 5.12 Hz, 2H). 3. 389.2 4. Method 3

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68 Ts N-Λν^\ ^N o> %<XN-^^==1 U. X ^{n N} o> 1. 12.9% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.85 (b.s., 1 Η), δ 7.38 (τ, J = 6.00 Hz, 2Н),7.30(d, J = 8.00 Hz, 1 Η), 7.23 (d, J = 10.00 Hz, 1Н), 7.01-7.02 (m, 1Н), 6 .96 (t, J = 6.80 Hz, 1H), 4.71 (s, 2H), 3.80 (t, J = 5.60 Hz, 2H), 3.673.69 (m, 4H), 3 .28-3.29 (m, 4H), 2.74-2.75 (m, 2H). 3. 336.1 4. Method 2 69 SI> Ts Br-V> ^~ ^N CCo . N-V% N^' ft D η ν-Όγ 1. 61% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.89 (s, 1H), 7.92 (d, J = 2.80 Hz, 1H), 7.38 -7.39 (m, 2H), 7.31 (d, J = 8.00 Hz, 1H), 7.01-7.01 (m, 1H), 6.93-6.94 (m, 1H ), 6.51 (d, J = 8.80 Hz, 1H), 4.26 (s, 2H), 3.833.85 (m, 2H), 3.50-3.51 (m, 2H), 3 .41-3.43 (m, 4H), 3.25-3.26 (m, 2H), 2.96-2.97 (m, 2H), 2.75-2.76 (m, 2H) . 3. 361.3 4. Method 3

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70 C? ^nh Ν Ts g/m 'MUH \\ A /t^ xf 1. 44% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 10.83 (s, 1H), 7.95 (d, J = 2.80 Hz, 1H), 7.39 -7.41 (m, 2H), 7.31 (d, J = 8.00 Hz, 1H), 7.03 (t, J = 7.60 Hz, 1H), 6.96 (t, J = 7.20 Hz, 1H), 6.53 (d, J = 8.80 Hz, 1H), 4.29 (s, 2H), 4.23 (d, J = 6.00 Hz, 2H),4 .18(d, J = 10.40 Hz, 2H), 3.62 (d, J = 10.40 Hz, 2H), 3.49-3.50 (m, 2H), 2.77-2, 78 (m, 2H), 2.61-2.63 (m, 1H), 1.78 (d, J = 8.00 Hz, 1H). 3. 347.1 4. Method 3 71 MUMUN Ν Ts Ml th'^ to ΜνΌα ^Ν \ X ' 1. 87% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 10.83 (s, 1H), 7.97 (d, J = 2.44 Hz, 1H), 7.45 -7.46 (m, 1H), 7.39 (d, J = 7.68 Hz, 1H), 7.31 (d, J = 7.68 Hz, 1H), 7.03 (t, J = 7.64 Hz, 1H), 6.95 (t, J = 7.00 Hz, 1H), 6.61 (d, J = 9.12 Hz, 1H), 4.69 (d, J = 6, 28 Hz, 2H), 4.27 (s, 2H), 3.643.67 (m, 2H), 3.46-3.49 (m, 4H), 3.08-3.09 (m, 1H), 2.77 (b.s., 2H), 1.89 (d, J = 8.40 Hz, 1H). 3. 347.3 4. Method 3

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74 C3 ^nh N Ts ^Вг -/^ VnA' ^N u> Υ=ς~ί n^/=^ 1. 50.3% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.83 (s, 1H), 7.95 (d, J = 2.64 Hz, 1H), 7 .37-7.39 (m, 2H), 7.31 (d, J = 7.96 Hz, 1H), 7.03 (t, J = 7.32 Hz, 1H), 6.95 (t, J = 7.56 Hz, 1H), 6.81 (d, J = 9.16 Hz, 1H), 4.29 (s, 2H), 3.83-3.87 (m, ZN), 3, 48-3.49 (m, 4H), 2.76 (sh.s, 2H), 2.302.33 (m, 2H), 1.14 (d, J = 6.16 Hz, ZN). 3. 349.3 4. Method 3 75 ^Q~x? ^nh N Ts ^N u> rCY x=\X \\ A ^HN V 1. 52% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.83 (s, 1H), 7.96 (s, 1H), 7.307.32 (m, ZH), 6 ,94-6.95 (m, 2H), 6.81 (d, J = 8.00 Hz, 1H), 4.30 (s, 2H), 3.83-3.86 (m, ZN), 3.50-3.55 (m, 4H), 2.66-2.76 (m, ZN), 2.34 (sh.s, 1H), 1.14 (d, J = 4.00 Hz, ZN). 3. 349.3 4. Method 3

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76 Ν Ts ъ0 .—z /ZI And 1. 38% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.83 (s, 1H), 7.95 (d, J = 2.80 Hz, 1H), 7.38 -7.40 (m, 2H), 7.31 (d, J = 8.00 Hz, 1H), 7.03 (t, J = 7.60 Hz, 1H), 6.95 (t, J = 7.20 Hz, 1H), 6.81 (d, J = 9.20 Hz, 1H), 4.29 (s, 2H), 3.96 (d, J = 11.60 Hz, 2H), 3.603 .61 (m, 2H), 3.49 (t, J = 5.60 Hz, 2H), 2.77 (sh.s, 2H), 2.27 (t, J = 10.80 Hz, 2H) , 1.14 (d, J = 6.00 Hz, 6H). 3. 363.4 4. Method 3 77 N Ts Br-^/=^ <-—. ^'Cf 1. 55% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.83 (s, 1H), 7.96 (d, J = 2.80 Hz, 1H), 7.38 -7.40 (m, 2H), 7.31 (d, J = 8.00 Hz, 1H), 6.98-7.01 (m, 2H), 6.97 (d, J = 6.80 Hz, 1H), 4.30 (s, 2H), 3.60-3.61 (m, 2H), 3.50 (t, J = 5.60 Hz, 2H), 3.31 (s, 2H ), 3.22-3.23 (m, 2H), 2.77 (t, J = 5.60 Hz, 2H), 1.931.93 (m, 4H), 1.68-1.69 (m, 2H). 3. 375.2 4. Method 3

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78 Cxx? ^nh Ν Ts Br^/^ = n^ ^/ η XA 1. 54% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.83 (s, 1H), 7.96 (d, J = 2.44 Hz, 1H), 7.30 -7.32 (m, ZN), 6.93-6.95 (m, 2H), 6.73 (d, J = 9.12 Hz, 1H), 4.13 (s, 2H), 4.094, 10 (m, 1H), 3.88-3.89 (m, 1H), 3.62-3.64 (m, ZN), 3.45-3.47 (m, ZN), 2.98- 3.00 (m, 1H), 2.95-2.97 (m, 2H), 1.04 (d, J = 6.52 Hz, ZN). 3. 349.3 4. Method 3 79 QA'n ^h N Ts Br^/^ 1 \\ \ A. ^{n n} 0° 1. 65% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.81 (s, 1H), 7.96 (d, J = 2.80 Hz, 1H), 7.37 -7.39 (m, 2H), 7.30 (d, J = 8.00 Hz, 1H), 7.02 (t, J = 7.20 Hz, 1H), 6.95 (t, J = 7.20 Hz, 1H), 6.72 (d, J = 8.80 Hz, 1H), 4.28 (s, 2H), 4.16-4.17 (m, 1H), 3.88- 3.89 (m, 1H), 3.61-3.63 (m, ZN), 3.48-3.49 (m, ZN), 2.94-2.96 (m, 1H), 2, 76 (b.s., 2H), 1.03 (d, J = 6.40 Hz, ZN). 3. 349.3 4. Method 3

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80 N Ts AAJ FVzz ⁿ ZZ 1. 57% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 10.81 (s, 1H), 7.89 (d, J = 2.80 Hz, 1H), 7.38 -7.39 (m, 2H), 7.31 (d, J = 8.00 Hz, 1H), 7.01-7.01 (m, 1H), 6.96 (t, J = 7.20 Hz, 1H), 6.37 (d, J = 8.80 Hz, 1H), 4.26 (s, 2H), 3.63 (sh.s, 4H), 3.53-3.54 (m , 4H), 3.45 (t, J = 5.60 Hz, 2H), 2.76 (t, J = 5.20 Hz, 2H), 1.71 (t, J = 5.20 Hz, 4H ). 3. 375.2 4. Method 3 82 Cx~C ^nh N Ts Vg-xAD N L to Z zzzz \=<Т ν~ζζ ^Η % ^А О ΖΖό 1. 71% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 10.82 (s, 1H), 7.94 (d, J = 2.96 Hz, 1H), 7.38 -7.38 (m, 2H), 7.31 (d, J = 7.96 Hz, 1H), 7.01-7.02 (m, 1H), 6.93-6.94 (m, 1H ), 6.81 (d, J = 9.12 Hz, 1H), 4.28 (d, J = Hz, 2H), 3.79-3.80 (m, 2H), 3.48 (t, J = 5.60 Hz, 2H), 3.32-3.33 (m, 1H), 3.26 (s, ZN), 2.97-2.99 (m, 2H), 2.75-2 .76 (m, 2H), 1.86-1.87 (m, 2H), 1.39-1.40 (m, 2H). 3. 363.3 4. Method 3

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83 N Ts /=<~ί N ⁿ y ^χο 1. 65% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.82 (s, 1H), 7.93 (d, J = 2.76 Hz, 1H), 7.37 -7.38 (m, 2H), 7.31 (d, J = 8.04 Hz, 1H), 7.03 (t, J = 7.84 Hz, 1H), 6.95 (t, J = 7.16 Hz, 1H), 6.81 (d, J = 9.12 Hz, 1H), 4.33 (s, 4H), 4.28 (s, 2H), 3.47 (t, J = 5.40 Hz, 2H), 3.30-3.32 (m, 4H), 2.76 (t, J = 5.04 Hz, 2H), 1.80 (t, J = 5.32 Hz, 4H). 3. 375.2 4. Method 3 85 Ts GUi \=ζ~ί ⁿ 1. 69% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.82 (s, 1H), 7.90 (d, J = 2.80 Hz, 1H), 7.38 -7.39 (m, 2H), 7.31 (d, J = 8.00 Hz, 1H), 7.03 (t, J = 8.00 Hz, 1H), 6.95 (t, J = 7.20 Hz, 1H), 6.52 (d, J = 9.20 Hz, 1H), 4.71 (s, 1H), 4.60 (s, 1H), 4.26 (s, 2H) , 3.74 (d, J = 7.20 Hz, 1H), 3.62 (d, J = 7.20 Hz, 1H), 3.433.45 (m, ZN), 3.16 (d, J = 10.00 Hz, 1H), 2.76 (b.s., 2H), 1.88 (d, J = 9.60 Hz, 1H), 1.81 (d, J = 9.60 Hz, 1H) . 3. 347.3 4. Method 3

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86 CXx? ^nh Ν Ts ν ^λ φ= OXA _B ά 1. 38% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): 400 MHz, DMSO-s16: δ 10.83 (s, 1H), 7.91 (d, J = 2.84 Hz , 1H), 7.40-7.41 (m, 2H), 7.31 (d, J = 7.96 Hz, 1H), 7.03 (t, J = 7.92 Hz, 1H), 6 .95 (t, J = 7.44 Hz, 1H), 6.52 (d, J = 8.96 Hz, 1H), 4.71 (s, 1H), 4.60 (s, 1H), 4 .26 (s, 2H), 3.74 (d, J = 7.08 Hz, 1H), 3.62 (d, J = 7.20 Hz, 1H), 3.413.43 (m, ZN), 3 .15-3.17 (m, 1H), 2.77 (t, J = 5.24 Hz, 2H), 1.88 (d, J = 9.44 Hz, 1H), 1.81 (d, J = 9.60 Hz, 1H). 3. 347.3 4. Method 3 87 F Y' NH ΎΥ-NTs ryO Br-^^N XY-nh 1. 71% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 11.18 (s, 1H), 7.99 (d, J = 2.92 Hz, 1H), 7.43 -7.43 (m, 1H), 7.13 (d, J = 8.08 Hz, 1H), 6.96-6.97 (m, 1H), 6.81 (d, J = 9.08 Hz, 1H), 6.70-6.72 (m, 1H), 4.38 (s, 2H), 3.693.70 (m, 4H), 3.53 (t, J = 5.60 Hz, 2H ), 3.283.30 (m, 4H), 2.87 (t, J = 5.12 Hz, 2H). 3. 353.2 4. Method 3

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88 F NH VJ-NTs rR ⁰ NH 1. 19% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): 11.15 (s, 1H), 7.88 (d, J = 2.92 Hz, 1H), 7.33 ( dd, J = 3.04, 9.12 Hz, 1H), 7.12 (d, J = 8.08 Hz, 1H), 6.91-6.93 (m,2H), 6.69-6 .71 (m, 1H), 4.71 (s, 2H), 3.88 (t, J = 5.64 Hz, 2H), 3.713.73 (m, 4H), 2.96-2.97 ( m, 4H), 2.85 (t, J = 5.44 Hz, 2H). 3. 353.3 4. Method 5 89 ^nh VJ—NTs o j 0 0 °A --O 1. 45% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 11.17 (s, 1H), 7.27 (d, J = 8.68 Hz, 1H), 7.13 (d, J = 8.04 Hz, 1H), 6.976.98 (m, 2H), 6.70-6.72 (m, 2H), 4.44 (s, 2H), 3.70-3, 71 (m, 4H), 3.55-3.56 (m, 6H), 2.87-2.89 (m, 2H). 3. 393.2 4. Method 2 90 I CH h r 0 1. 49% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 7.247.26 (m, 2H), 7.01-7.02 (m, 2H), 6.75-6, 76 (m, 2H), 4.45 (s, 2H), 3.653.70 (m, 9H), 3.54-3.55 (m, 4H), 2.90 (s, 2H). 3. 407.1 4. Method 2

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91 Γνη\ ^xF ci-4^4nQ _q JYY°v _(j^nY > Q4 F 1. 66% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 7.237.25 (m, 2H), 7.11-7.12 (m, 1H), 6.84-6, 85 (μ, 3Η), 4.41 (s, 2Η), 3.88 (s, 3Η), 3.71-3.72 (μ, 4Η), 3.56-3.57 (μ, 6Η) , 2.79 (sh.s., 2Η). 3. 407.3 4. Method 2 92 ^NH χΥ-NTs F ΓΥθ Вг-Ύν ΖΎθ Γ ν-^,ν N.H. F 1. 76.1% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 11.33 (s, 1H), 7.96 (d, J = 3.20 Hz, 1H), 7 ,42-7.43 (m, 1H), 7.22 (d, J = 7.60 Hz, 1H), 6.87-6.88 (m, ZN), 4.31 (s, 2H), 3.703.71 (m, 4H), 3.45-3.47 (m, 2H), 3.28-3.30 (m, 4H), 2.68-2.76 (m, 2H). 3. 353.1 4. Method 3 93 ^NH YJ~-NTs F θΥΥν.'\ -X3~°V '\ ^Γ)-ύαΛ ^ν Οο ^NH F 1. 50.0% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 11.32 (s, 1H), 7.28 (d, J = 8.72 Hz, 1H), 7 .22 (d, J = 7.60 Hz, 1H), 6.99 (d, J = 2.40 Hz, 1H), 6.876.87 (m, 2H), 6.75 (dd, J = 2, 44, 8.74 Hz, 1H), 4.36 (s, 2H), 3.70-3.71 (m, 4H), 3.55-3.56 (m, 6H), 2.78 (t , J = 5.44 Hz, 2H). 3. 393.1 4. Method 3

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94 GHn _Αν- Ν^Χ^-Ν J 1 // ΒΓΧ^Ν WITH -x Г ⁴ N - Пг^ / n-^Vn rV ⁷ Λ-Ν \ 1. 17.6% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ 8.13 (s, 1H), 7.93 (s, 1H), 7.41 (τ, J = 6 ,80 Hz, 2Η), 7.10 (τ, J = 7.20 Hz, 1Η), 7.00 (τ, J = 7.20 Hz, 1Η), 4.67 (s, 2Η), 3, 83 (sh.s, 2H), 3.70 (s, ZN), 3.33 (sh.s, 4N), 2.77 (sh.s, 2N), 2.41 (sh.s, 4N) , 2.19 (s, ZN). 4. Method 5 95 / ⁴ ΝΗ \ _ Αν Χγ ^ν α Br^VN WITH _ ΑνΧΑ Γ νΧ^ν λ-Ν\ 1. 13.4% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 8.03 (s, 1H), 7.44 (t, J = 7.60 Hz, ZH), 6 .98-7.00 (m, 2H), 6.81 (d, J = 8.80 Hz, 1H), 4.36 (s, 2H), 3.69 (s, ZN), 3.47 ( t, J = 5.60 Hz, 2H), 3.29 (b.s., 4H), 2.73 (b.s., 2H), 2.37-2.38 (m.4H), 2.16 (s, ZN). 3. 362.2 4. Method 5 96 Χνη \ Br-XjT WITH 0 1. 31.8% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 7.49 (d, J = 10.00 Hz, ZN), 7.407.41 (m, 1H), 7 .30 (d, J = 10.00 Hz, 1H), 7.11 (t, J = 7.60 Hz, 1H), 7.00 (t, J = 7.20 Hz, 1H), 4.78 (s, 2H), 3.85 (t, J = 5.60 Hz, 2H), 3.71-3.72 (m, 7H), 3.33-3.35 (m, ZN), 2, 29-2.33 (m, 2H). 3. 350.2 4. Method 5

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97 N.H. AX-ni\ O I? r° c __________________________1__________________________________________________________________________________________ A L st 0 / 1. 50.3% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 7.41 (τ, J = 7.60 Hz, 2Η), 7.26 (d, J = 8, 68 Hz, 1 Η), 7.087.10 (m, 2Η), 7.00 (τ, J = 7.12 Hz, 1 Η), 6.81 (d, J = 8.68 Hz, 1 Η), 4.42 (s, 2H), 3.71 (s, ZN), 3.57 (sh.s, 6H), 2.80 (s, 2H), 2.40-2.41 (m, 4H) , 2.22 (s, ZN). 3. 402.3 4. Method 3 98 Γ^ΝΗ VA~NTs / 0 A* k? O o i yy c / 1. 25.7% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.83 (s, 1H), 7.39 (d, J = 7.60 Hz, 1H), 7 .31 (d, J = 8.00 Hz, 1H), 7.25 (d, J = 8.80 Hz, 1H), 6.946.95 (m, ZN), 6.72-6.73 (m, 1H), 4.35 (s, 2H), 3.56-3.57 (m, 6H), 2.78 (s, 2H), 2.40-2.41 (m, 4H), 2.22 (s, ZN). 3. 388.2 4. Method 3 99 v=<K^ ^NH N Ts <=/Ar° OriAN/YrO H 1. 21% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.84 (b.s., 1H), 7.39 (d, J = 7.68 Hz, 1H), 7.277 .29 (m,2H), 6.94-6.96 (m, ZN), 6.77 (d, J = 7.96 Hz, 1H), 4.37 (s, 2H), 3.70- 3.71 (m, 4H), 3.55-3.56 (m, 6H), 2.79 (sh.s, 2H). 3. 375.2 4. Method 2

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100 Γ^ΝΗ ^0| ΆΎ>ν_ ₀ ΟμΓ 1. 30.5% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 7.42 (t, J = 7.32 Hz, 1H), 7.28 (d, J = 8, 72 Hz, 1H), 7.097.10 (m, 1H), 7.00 (t, J = 7.28 Hz, 1H), 6.826.82 (m, 1H), 4.43 (s, 2H), 3 .71-3.73 (m, 7H), 3.55-3.56 (m, 6H), 2.81 (s, 2H). 3. 389.2 4. Method 4 101 ηνΟ C.I. Gu^ΥλΥ ο Ζ-Ν \-ο 1. 25.3% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 7.447.45 (m, 2H), 7.29-7.29 (m, 1H), 7.14 ( t, J = 7.20 Hz, 1H), 7.017.03 (m, 1H), 6.94-6.94 (m, 1H), 6.65-6.65 (m, 1H), 4.91 (s, 2H), 3.97 (t, J = 5.40 Hz, 2H), 3.71-3.72 (m, 7H), 3.05-3.06 (m, 4H), 2, 88 (sh.s., 2N). 3. 389.3 4. Method 4 102 Α° ηνΆ , ^ν Όα οο ^ΝΗ ΝΗ 1. 46.7% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.96 (s, 1H), 7.42 (d, J = 7.76 Hz, 1H), 7 .34 (d, J = 8.04 Hz, 1H), 7.18 (d, J = 8.52 Hz, 1H), 7.127.13 (m, 1H), 7.06 (t, J = 7, 84 Hz, 1H), 6.98 (t, J = 7.16 Hz, 1H), 6.796.80 (m, 1H), 4.81 (s, 2H), 3.95 (t, J = 5, 60 Hz, 2H), 3.73-3.74 (m, 4H), 3.04-3.05 (m, 4H), 2.86-2.87 (m, 2H). 3. 375.2 4. Method 4

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103 Γ^ΝΗ 1>-ΝΗ F ^Z jArA = ¹ N ^N __,0 β ^Ν Χ ^λΝ Λο Α^ ^νη F ^Z 1. 33.4% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.94 (s, 1H), 7.35-7.36 (m, 1H), 7.27 (d, J = 8.68 Hz, 1H), 7.09-7.10 (m, 1 Η), 7.00 (d, J = 2.24 Hz, 1H), 6.79-6.79 (m, 1 Η), 6.73-6.74 (m, 1 Η), 4.34 (s, 2Η), 3.70-3.71 (μ, 4Η), 3.54-3.55 (μ, 6Η), 2.77 (τ, J = 5.08 Hz, 2Η). 3. 393.3 4. Method 4 104 F.___ _NH N I c^yO /-L Ν θ __/ \ ^N NJ 1.7%; 2. ¹ Η NMR (400 MHz, DMSO-c/s): δ 7.387.42 (m, 1 Η), 7.277.31 (m, 2H), 7.09 (d, J = 2.40 Hz, 1H ), 6.806.87 (m, 2H), 4.40 (sh.s., 2H), 3.70-3.73 (m, 4H), 3.68 (s, ZN), 3.55-3, 57 (m, 6H), 2.75-2.79 (m, 2H). 3. 407.2 4 Method 2 105 Λ ^{4 nh} PU ax ⁿ O° Br-AX or 1. 11%; 2. ^1H NMR (400 MHz, DMSO-s/b): δ 8.12 (d, J = 3.20 Hz, 1H), 7.85 (d, J = 2.80 Hz, 1H), 7.247 .24 (m, 1H), 7.09 (d, J = 6.80 Hz, 1H), 6.876.88 (m, 1H), 6.76 (d, J = 9.20 Hz, 1H), 3.673 .69 (m, 9H), 3.30-3.31 (m, 4H), 2.85-2.86 (m, 2H), 1.87 (sh.s, 2H). 3. 350.2 4. Method 2

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107 F ζ Ha l ⁿ I co 2-==4. 0 F. СНО-сл ^N y 1. 23%; 2. Ή-NMR (400 MHz, DMSO-c/s): δ 8.16 (d, J = 1.12 Hz, 1H), 7.94 (d, J = 2.12 Hz, 1 Η), 7.287.28 (m, 1Η), 7.16-7.16 (μ, 1Η), 6.66 (d, J = 8.96 Hz, 1Η), 3.93 (sh.s, 2Η), 3.71-3.72 (m, 7Η), 3.01-3.02 (μ, 4Η), 2.89-2.90 (m, 2Η), 2.08 (sh.s, 2Η). 3. 368.2 4. Method 2 108 NH ^Br №X_ 4 O lU 4n \ lx _p AsA~NH 1. 28%; 2. Ή NMR (400 MHz, DMSO-c/s): δ 10.94 (b.s, 1 Η), 7.96 (d, J = 3.00 Hz, 1 Η), 7.427.43 (μ, 1 Η), 7.35-7.37 (μ, 1Η), 7.09-7.09 (μ, 1Η), 6.79-6.79 (μ, 1 Η), 4.28 (s, 2Η), 3.69-3.70 (μ, 4Η), 3.49 (sh.s, 2Η), 3.28-3.29 (μ,4Η), 2.75 (sh.s, 1Η) . 3. 353.2 4. Method 2 109 _p a)P ⁴ - Ts N-^V_7 F F 44 p u. 1.29%; 2. Ή NMR (400 MHz, DMSO-c/s): δ 10.99 (b.s., 1 Η), 7.86 (d, J = 2.80 Hz, 1 Η), 7.337.34 (μ, 2Η), 7.08-7.08 (μ, 1Η), 6.94 (d, J = 9.08 Hz, 1Η), 6.786.78 (μ, 1Η), 4.64 (s, 2Η), 3.81-3.82 (μ,2Η), 3.71-3.73 (μ, 4Η), 2.96-2.97 (μ, 4Η), 2.74 (sh.s, 2Η). 3. 353.2

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110 Ν I V ^Ν \ ^Ν \^y° рХ-ТТ/ТТ Z-i V 7N ' 7 ^/° 1.26%; 2. ¹ H NMR (400 MHz, DMSO-s/ ₆ ): δ 8.04 (b.s, 1H), 7.48-7.49 (m, 1H), 7.38-7.40 (m , 1H), 7.30 (d, J = 10.40 Hz, 1H), 6.81-6.83 (m, 2H), 4.35 (s, 2H), 3.673.70 (m, 7H) , 3.47-3.48 (m, 2H), 3.30-3.31 (m, 4H), 2.77 (sh.s., 2H). 3. 367.2 4. Method 2 111 Ν I 0 0 to her Ν Ν—Ζ~~ ^Ν ^ ^o 1. 25%; 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 7.88 (d, J = 2.88 Hz, 1H), 7.357.35 (m, 2H), 7.28-7.29 (m, 1H), 7.01 (d, J = 9.20 Hz, 1H), 6.816.82 (m, 1H), 4.66 (s, 2H), 3.81-3.83 (m, 2H), 3.72-3.73 (m, 4H), 3.68 (s, ZN), 2.97-2.98 (m, 4H), 2.76 (sh.s, 2H). 3. 367.2 4. Method 2 112 Q—C ^nh Ν I ^Br vd ν A-gn <N V/-CN 1. 16% 2. ¹ H-NMR (400 MHz, DMSO-s/ ₆ ): δ 8.63 (d, J = 2.64 Hz, 1H), 7.81 (d, J = 8.80 Hz , 1H), 7.577.58 (m, 1H), 7.41-7.43 (m, 2H), 7.13 (t, J = 7.56 Hz, 1H), 7.02 (t, J = 7.28 Hz, 1H), 4.73 (s, 2H), 3.87 (t, J = 5.48 Hz, 2H), 3.73 (s, ZH), 2.85 (sh.s, 2H). 3. 289.2 4. Method 2

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113 N Ts v a-cn A OlXa ln ^N H ^N 1. 12% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.91 (b.s., 1 Η), 8.54 (d, J = 2.56 Hz, 1H), 7.79 (d, J = 8.84 Hz, 1H), 7.337.35 (m, ZN), 6.96-6.97 (m, 2H), 4.66 (s, 2H), 3.89 (t, J = 5.40 Hz, 2H), 2.85 (sh.s, 2H). 3. 275.1 4. Method 2 117 Caa? ^nh N Ts θ'ΎΧ id\ ^N ~N\N ₀ q^Oo. N NAVX \ 1. 20% 2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.90 (s, 1H), 7.38-7.39 (m, 2H), 7.27-7, 30 (m, 2H), 7.01-7.01 (m, 1H), 6.93-6.93 (m, 1H), 4.72 (s, 2H), 3.78-3.80 ( m, 4H), 3.36-3.37 (m, 1H), 3.27 (s, ZN), 3.03-3.04 (m, 2H), 2.75-2.77 (m, 2H), 1.88-1.88 (m, 2H), 1.43-1.44 (m, 2H). 3. 364.3 4. Method 2 118 SN> N Ts ^c Ak ά n-LO OtOaTy ah ⁿ ~n Vyj 1. 47% 2. ¹ H-NMR (400 MHz, DMSO-s/b): 400 MHz, DMSO-06: δ 10.90 (s, 1H), 7.37-7.39 (m, 2H) , 7.27-7.29 (m, 2H), 7.03-7.03 (m, 2H), 4.72 (s, 2H), 3.83-3.84 (m, 5H), 3 .58-3.58 (m, 2N), 2.72-2.73 (m, ZN), 2.44-2.46 (m, 1H), 1.15-1.16 (m, ZN) . 3. 350.1 4. Method 2

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119 Sh)n Ts ABOUT υ -Z G ZI 0 1. 11% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.89 (s, 1H), 7.43 (d, J = 9.60 Hz, 1H), 7.38 (d, J = 7.60 Hz, 1H), 7.277.29 (m, 2H), 7.01-7.01 (m, 1H), 6.93-6.93 (m, 1H), 4, 64 (s, 2H), 3.883.89 (m, 5H), 3.58-3.58 (m, 2H), 2.75-2.76 (m, ZN), 2.43-2.44 ( m, 1H), 1.15 (d, J = 6.40 Hz, ZN). 3. 350.2 4. Method 2 120 Ts O Vr -Z I ^zx o 1.30% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.90 (s, 1H), 7.37-7.39 (m, 2H), 7.27-7, 30 (m, 2H), 6.95-6.95 (m, 2H), 4.73 (s, 2H), 3.95-3.97 (m, 2H), 3.81-3.83 ( m, 2H), 3.62-3.63 (m, 2H), 2.77-2.78 (m, 2H), 2.33-2.35 (m, 2H), 1.14 (d, J = 6.00 Hz, 6H). 3. 364.3 4. Method 2 121 F ) NH N Tos Q NY ^in, ~P F O F ₄ NY Hn hr H 1. 38% 2. ¹ H-NMR (400 MHz, DMSO-c/s): δ 10.95 (s, 1H), 7.88 (d, 1H), 7.57 (d, 1H), 7 .33 (dd, 1H), 7.18 (m, 2H), 6.98 6.77 (m, ZN), 4.54 (s, 2H), 3.78 (t, 2H), 3.72 (t, 4H), 3.60 (t, 4H), 2.79 (t, 2H). 3. 403.21 4. Method 3

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1. 25%

2. ¹ H-NMR (400 MHz, DMSO-s/b): δ 10.96 (s, 1H), 7.96 (d, 1H), 7.53 (s, 2H), 7.31 (dd , 1H), 7.25 - 7.03 (m, ZN), 6.87 (t, 1H), 4.45 (s, 2H), 3.70 (m, 6H), 3.56 (m, 4H), 2.89 -2.68 (m, 2H).

3.403.19

4. Method 3

Example 123.

Stage A.

Sodium hydride (60%) (0.0412 g, 1.79 mmol) was slowly added to a solution of Example 44 in THF (15.0 mL) at 0°C, then stirred for 1 hour at 25°C. Ethyl iodide (0.305 g, 0.00179 mol) in 15.0 mL THF was added slowly at 0°C, then stirred for 2 hours at 25°C. The reaction mixture was monitored by LCMS, and the reaction mixture was diluted with water (50.0 ml). The organic phase was separated and the aqueous phase was extracted with ethyl acetate two more times. The combined organic phase was dried over Na ₂ SO ₄ , filtered and the solvents were evaporated under reduced pressure. The crude product was purified on an HP-Sil (biotage) column using a 50/50 ethyl acetate/petroleum ether gradient to give the title compound as an off-white solid.

1H-NMR (400 MHz, DMSO-d ₆ ): δ 8.05 (d, J=2.88 Hz, 1H), 7.49-7.50 (m, 1H), 7.42 (d, J =8.20 Hz, 2H), 7.08-7.08 (m, 1H), 6.97-6.97 (m, 1H), 6.81 (d, J=9.08 Hz, 1H) , 4.38 (s, 2H), 4.18 (kv, J=7.08 Hz, 2H), 3.69-3.70 (m, 4H), 3.50 (t, J=5.60 Hz, 2H), 3.29-3.30 (m, 4H), 2.78 (t, J=5.52 Hz, 2H), 1.26 (T, J=7.12 Hz, 3H). MS: 363.2 (M+H)+.

HCl salt of the compounds of the invention.

General methodology.

To a solution of the compound of example (0.1 g) in dry DCM (10 ml), cooled to 0°C, was added 1.M HCl in ether (5 eq.) or 4M HCl in dioxane (5 eq.) and stirred for 15 minutes. The reaction mixture was concentrated in vacuo and triturated with diethyl ether to obtain the desired product shown in table. 2.

Examples.

Following the procedure for preparing the hydrochloride salt described in the general procedure above, the following compounds were prepared.

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table 2

Example Original free base Product 1. Output 2. ¹ H-NMR 3. MH ⁺ (ESI) 123 HCl o o 0 y* z^ VAz^ O 1. 83% 2. Ή-NMR (400 MHz, DMSOd ₆ ): δ 8.21 (d, J = 8.36 Hz, 1H), 7.63 (b.s., 1H), 7.37-7 .40 (m, ZN), 6.99-7.01 (m, 2H), 4.53 (s, 2H), 4.20-4.22 (m, 2H), 3.743.75 (m, 4H ), 3.57-3.58 (m, 6H), 2.83 (s,2H), 1.28 (t, J = 7.16 Hz, ZN). 3. 363.2 108 HCl g- guO _p XX~NH α ауС ⁰ Г II V·^ HCI _f A~nh 1.65% 2. Ή-NMR (400 MHz, DMSOd ₆ ) δ 10.83 (b.s., 1H), 7.92-7.93 (m, 1H), 7.41-7.43 (m, 1H ), 7.357.37 (m, 1H), 7.05-7.07 (m, 1H), 6.77-6.79 (m, 1H), 4.25 (s, 2H), 3.68- 3.69 (m, 4H), 3.52 (b.s., 2H), 3.25-3.27 (m, 4H), 2.83 (b.s., 1H). 3. 353.1

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109 HCl jYCV N- ^{7 F} ^H U\ /^\— Λ N-Λ Г X_/ g ιΓ?- ^{7 F} Η HCl 1. 85% 2. ¹ H-NMR (400 MHz, DMSO-c/s) δ 10.85 (b.s., 1 Η), 7.91-7.93 (m, 1H), 7.40- 7.42 (m, 1H), 7.367.37 (m, 1H), 7.03-7.05 (m, 1H), 6.77-6.79 (m, 1H), 4.27 (s, 2H), 3.65-3.67 (m, 4H), 3.57 (b.s., 2H), 3.25-3.27 (m, 4H), 2.85 (b.s., 1H) . 3. 353.1 110 HCl % LL jTV ⁿ O° , ^ν Ά ^λ ι H ^HCI _f A>-n 1. 82% 2. Ή-NMR (400 MHz, DMSOd ₆ ): 400 MHz, DMSO-s/ ₆ : δ 8.20 (d, J = 7.88 Hz, 1H), 7.62 (s, 1H ), 7.38-7.41 (m, ZN), 6.836.84 (m, 1H), 4.48 (s, 2H), 3.693.73 (m, 7H), 3.56-3.59 ( m, 6H), 2.82 (s, 2H). 3. 367.2 111 HCl UX'O' / nA / χΆ J A G L r^xvO / nA J nv ⁿ _F JU% HCl 1. 79% 2. Ή-NMR (400 MHz, DMSOc/s): δ 8.08 (d, J = 8.72 Hz, 1H), 7.60 (d, J = 9.64 Hz, 1H), 7.447.46 (m, 2H), 7.31-7.32 (m, 1H), 6.85-6.86 (m, 1H), 4.93 (s, 2H), 4.02 (t, J = 5 ,16 Hz, 2H), 3.70 (sh.s, 7H), 3.11 (sh.s, 4H), 2.87 (s, 2H). 3. 367.2 80 HCl \=(~ί II 1 ^H A-O/d Ύ> αα,,ν^/h \\ A ^H HCl 1.0 1. 74% 2. Ή-NMR (400 MHz, DMSOd ₆ ): δ 10.91 (s, 1H), 8.14-8.15 (m, 1H), 7.40 (d, J = 7, 72 Hz, 1H), 7.33 (d, J = 7.96 Hz, 1H), 7.05 (t, J = 7.60 Hz, 2H), 6.906.92 (m, 2H), 4.37 (sh.s, 2H), 3.97 (sh.s, ZN), 3.53-3.54 (m, 7N), 2.79 (s, 2H), 1.77-1.78 (m , 4H). 3. 375.2 69 HCl th ιΛ<ο h Am HCl \/ ^U 1. 87% 2. Ή-NMR (400 MHz, DMSOd ₆ ): δ 10.93 (s, 1H), 8.17-8.18 (m, 1H), 7.33-7.35 (m, ZN), 7.117.13 (m, 1H), 6.99-7.03 (m, 2H), 4.38 (s, 2H), 3.73-3.75 (m, 4H), 3.59 -3.61 (m, 4H), 3.48-3.49 (m, 2H), 3.12-3.13 (m, 2H), 2.80 (s, 2H). 3. 361.3

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86 HCl \=ς~ί Αν VI H _z Nx/4 Parts l Ν HCl φ 1.89% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.93 (s, 1 Η), 8.16-8.17 (μ, 1 Η), 7.41 (d, J = 7.68 Hz, 1 Η), 7.34 (d, J = 7.72 Hz, 2Η), 7.21 (d, J = 9.84 Hz, 1 Η), 7.05 (t, J = 7.96 Hz, 1 Η), 6.97 (t, J = 7.44 Hz, 1 Η), 5.17 (s, 1H), 4.78 (s, 1H), 4.38 (s, 2H), 3.74 -3.76 (m, 2H), 3.59-3.60 (m, ZN), 3.393.41 (m, 1H), 2.81 (s, 2H), 1.98 (s, 2H). ). 3. 347.1 85 HCl ΛΥ-Ζ7 N^'''\\A “ A/ V<1 H ΙΑ Ν HCl 1.78% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.92 (s, 1H), 8.16-8.17 (m, 1H), 7.41 (d, J = 8.00 Hz, 1H) , 7.34 (d, J = 8.00 Hz, 2H), 7.21 (d, J = 9.60 Hz, 1H), 7.037.04 (m, 1H), 6.95-6.95 ( m, 1H), 5.17 (s, 1H), 4.79 (s, 1H), 4.39 (s, 2H), 3.75-3.77 (m, 2H), 3.593.60 (m , ZN), 3.39-3.41 (m, 1H), 2.81-2.82 (m, 2H), 1.99 (s, 2H). 3. 347.1 82 HCl ^H O. / <^/0 A/ ⁴ Wx H _χ Ν 4Ζ HCI Αο ⁷ 1.78% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.92 (s, 1H), 8.16-8.17 (m, 1H), 7.41 (d, J = 8.00 Hz, 1H) , 7.34 (d, J = 8.00 Hz, 2H), 7.21 (d, J = 9.60 Hz, 1H), 7.037.04 (m, 1H), 6.95-6.95 ( m, 1H), 5.17 (s, 1H), 4.79 (s, 1H), 4.39 (s, 2H), 3.75-3.77 (m, 2H), 3.593.60 (m , ZN), 3.39-3.41 (m, 1H), 2.81-2.82 (m, 2H), 1.99 (s, 2H). 3. 347.1 44 HCl CZ/-l N w ^A O QZ NH ^ ^Ν ΖΧ HCI Ό 1. 88% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.98 (s, 1H), 8.18-8.19 (m, 1H), 7.34-7.35 (m, 4H), 6.966, 98 (m, 2H), 4.46 (s, 2H), 3.613.62 (m, JH), 2.84 (s, 2H). 3. 335.3

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46 HCI From O'CHA. C ng Ή 1 ΑχΓΥ z -z Y X-N HCI JO 1.85% 2. Ή-NMR (400 MHz, DMSOd6): δ 8.24-8.24 (m, 1 Η), 7.67 (s, 1 Η), 7.39-7.41 (m, ZN), 7.12 (τ, J = 7.20 Hz, 1H), 7.01 (t, J = 7.20 Hz, 1H), 4.52 (s, 2H), 3.623.64 (m, 13H), 2.85 (sh.s., 2H). 3. 349.3 45 HCI ζ-λ N M-HD_o (O NG 1 ?ίΓι z- - ⁷ V HCI 0 1. 78% 2. Ή-NMR (400 MHz, DMSOd6): δ 8.07 (b.s., 1H), 7.61-7.63 (m, 1H), 7.43-7.45 (m, ZN), 7.15 (t, J = 7.76 Hz, 1H), 7.04 (t, J = 7.00 Hz, 1H), 4.95 (s, 2H), 4.024.03 (m, 2H), 3.71-3.72 (m, 7H), 3.11-3.12 (m, 4H), 2.89-2.90 (m, 2H). 3. 349.2 83 HCI « XZ^ ^H A ', ¹ —Ύο ci H hci '^A. ^0 1.88% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.91 (s, 1H), 8.16 (d, J = 8.28 Hz, 1H), 7.33-7.35 (m, 4H) , 7.05 (t, J = 7.04 Hz, 1H), 6.97 (t, J = 7.08 Hz, 1H), 4.41 (s, 2H), 3.72 (sh.s, 2H), 3.63 (s, 6H), 3.40 (br.s, 2H), 2.81 (b.s, 2H), 1.56-1.58 (m, 4H). 3. 375.2 79 HCI A-/>/^Χί.Ν'ν' ⁵⁵ '! 1 ft XX ^HN (V H HCI 1. 81% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.94 (s, 1H), 8.16 (d, J = 9.04 Hz, 1H), 7.60 (s, 1H), 7.42 (d, J = 7.72 Hz, 1H), 7.34 (d, J = 8.00 Hz, 2H), 7.06 (t, J = 7.24 Hz, 1H), 6.98 (t , J = 7.56 Hz, 1H), 4.45 (s, 2H), 3.97-4.00 (m, 1H), 3.66-3.67 (m, 4H), 3.323.38 ( m, 4H), 2.84 (s, 2H), 1.191.21 (m, ZN). 3. 349.3

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76 HCl --Z /zz And t=4~X z ^N MX ft У /-м ^H ιΜ HCl X' 1.69% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.92 (s, 1H), 8.15 (d, J = 7.68 Hz, 1H), 7.52 (brs, 1H), 7 .41 (d, J = 7.56 Hz, 2H), 7.34 (d, J = 7.96 Hz, 1H), 7.06 (t, J = 7.68 Hz, 1H), 6.97 (t, J = 7.16 Hz, 1H), 4.44 (s, 2H), 4.08-4.11 (m, 2H), 3.66 (sh.s, 4H), 2.83 ( sh.s, 2H), 2.65-2.67 (m, 2H), 1.16 (d, J = 6.16 Hz, 6H). 3. 363.2 75 HCl ft X z—y ^HN V \=4~£ nmm ^H О hci X-° 1. 85% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.90 (s, 1H), 8.09 (br.s, 1H), 7.59 (b.s, 1H), 7.41 (d, J = 7.60 Hz, 1H), 7.34 (d, J = 8.00 Hz, 2H), 7.06 (t, J = 7.20 Hz, 1H), 6.98 (t, J = 7.60 Hz, 1H), 4.44 (s, 2H), 4.04-4.07 (m, 1H), 3.94-3.96 (m, 2H), 3.88-3.91 (m, 4H), 3.03-3.05 (m, 1H), 2.682.73 (m, ZN), 1.16 (d, J = 6.00 Hz, ZN). 3. 349.2 74 HCl n^ ^/ ft X -—/ rVx/ \=(~ί ^N MM ft X zM' n ^ir ^N I n° HCl 1.79% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.90 (s, 1H), 8.10 (br.s, 1H), 7.53 (b.s, 1H), 7.41 (d, J = 7.60 Hz, 1H), 7.34 (d, J = 8.00 Hz, 2H), 7.06 (t, J = 7.20 Hz, 1H), 6.98 (t, J = 8.00 Hz, 1H), 4.44 (s, 2H), 3.94-3.96 (m, ZN), 3.38-3.40 (m, 4H) 2.68-2.68 ( m, 4H), 1.16 (d, J = 6.40 Hz, ZN). 3. 349.3 71 HCl ft X ^H ft 1 η ν ^Λ 0 HCl 1. 82% 2. Ή-NMR (400 MHz, DMSOd6): δ 10.93 (s, 1H), 8.09 (s, 1H), 7.41-7.43 (m, ZH), 7.34 (d, J = 8.00 Hz, 1H), 7.06 (t, J = 6.80 Hz, 1H), 6.98 (t, J = 7.20 Hz, 1H), 4.44-4.47 ( m, 2H), 4.004.10 (m, 4H), 3.65-3.66 (m, 4H), 2.83 (sh.s., 2H), 2.03-2.04 (m, 2H) . 3. 347.1 105 HCl r/>O Α,ί f Ir ^N \ g l-O nA-// r Tr /—N N2XHCl 1.71% 2. Ή-NMR (400 MHz, DMSOd6): 400 MHz, flMCO-d6: δ 8.23 (s, 1H), 7.96 (s, 1H), 7.35 (d, J = 10.80 Hz , ZN), 7.01-7.02 (m, 1N), 3.59-3.68 (m, 13N), 2.91 (sh.s., 2N), 1.91 (sh.s., 2N ). 3. 350.1 107 HCl /-\ rvO _c J N-Λ J '—' YTV ^N /—\rntO ⁰ _l J νΆ J YtV' ^N M' ^N _x 2 X HCl 1.82% 2. Ή-NMR (400 MHz, DMSOd6): δ 8.24 (s, 1H), 8.00 (d, J = 9.20 Hz, 1H), 7.60 (s, 1H), 7.39 (t, J = 7.60 Hz, 1H), 7.20 (d, J = 9.60 Hz, 1H), 3.91-3.98 (m, 2H), 3.74-3.76 ( m, 7H), 3.16 (s, 4H), 2.96-2.97 (m, 2H), 2.06 (b.s, 2H). 3. 368.3

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Description of biological analysis

Full-length tau (flTau) disaggregation assay using thioflavin T (ThT).

The longest isoform of human tau protein (2N4R; 441 amino acids) was expressed in bacteria and purified. To analyze the disaggregation of tau protein by ThT, 35 μM recombinant full-length (fl)Tau in PBS was aggregated for 24 h at 37°C in the presence of 50 μM heparin (SigmaAldrich) and 10 mM DTT (Sigma-Aldrich) under shaking conditions at 750 rpm Compounds were dissolved in anhydrous dimethyl sulfoxide (DMSO, Sigma-Aldrich) to achieve a concentration of 10 mM. flTau aggregates and serial dilutions of compounds were mixed together in PBS (50 μL volume) to a final concentration of 2 μM flTau aggregates and 160 to 0.04 μM compounds. The mixture was incubated for 30 min at room temperature (RT), then 40 μl of this mixture was transferred to a black 384-well assay plate (Perkin-Elmer) and mixed with 10 μl of 100 μM ThT in 250 mM glycine (both from Sigma-Aldrich) in PBS. Fluorescence (relative fluorescence units; RFU) was measured in one or two replicates on a Tesap reader (excitation: 440 nm; emission: 485 nm). The percentage of flTau disaggregation was then calculated and the half maximum effective concentration (EC ₅₀ ) was determined using GraphPad Prism version 5 (GraphPad Software), assuming a single-binding site selection model.

Analysis of Tau K18 disaggregation by ThT.

The Tau K18 fragment, comprising amino acids 244 to 372 of the longest isoform (2N4R) of human Tau441, was expressed in bacteria and purified or purchased from SignalChem. To analyze the disaggregation of K18 by ThT, 35 μM recombinant K18 in PBS was aggregated for 24 h at 37°C in the presence of 50 μM heparin (Sigma-Aldrich) and 10 mM DTT (Sigma-Aldrich) under shaking conditions at 750 rpm. Compounds were dissolved in anhydrous dimethyl sulfoxide (DMSO, SigmaAldrich) to achieve a concentration of 10 mM. K18 aggregates and serial dilutions of compounds were mixed together in PBS (50 μL volume) to a final concentration of 2 μM K18 aggregates and 160 to 0.04 μM compounds. The mixture was incubated for 30 min at room temperature (RT), then 40 μl of this mixture was transferred to a black 384-well assay plate (Perkin-Elmer) and mixed with 10 μl of 100 μM ThT in 250 mM glycine (both from Sigma-Aldrich) in PBS. Fluorescence (relative fluorescence units; RFU) was measured in one or two replicates on a Tesap reader (excitation: 440 nm; emission: 485 nm). The percentage of K18 disaggregation was then calculated and the half maximum effective concentration (EC ₅₀ ) was determined using GraphPad Prism version 5 (GraphPad Software), assuming a single-binding site selection model.

The following examples of compounds were measured.

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- 97 044393

- 98 044393

97 ++ 98 + 99 +++ 100 +++ 101 +++ 102 +++ 103 +++ 104 +++ 112 +++ 113 +++ 117 +++ 118 +++ 119 +++ 120 +++ 121 ++ 122 +++ 80 HCl ++ 69 HCl +++ 86 HCl +++ 85 HCl ++ 82 HCl +++ 107 HCl + 123 HCl +++ 108 HCl +++ 109 HCl +++ 110 HCl +++ 111 HCl +++ 44 HCl +++ 46 HCl ++ 45 HCl +++ 83 HCl + 79 HCl ++ 76 HCl +++ 75 HCl +++ 74 HCl +++ 71 HCl ++ 105 HCl +

Legend: +++ EC5 ₀ < 10 µM;

++ EC5 ₀ 10<x<25 µM;

+ EC ₅₀ 25<x<50 µM.

Reduced intracellular aggregation of tau protein.

A human neuroblastoma cell line overexpressing full-length human tau carrying the P301L mutation was cultured in complete medium [DMEM-F12 4.5 g/L Glu

- 99 044393 tamax (Invitrogen), 15% FBS (Biochrom), 1% penicillin/streptomycin (Invitrogen) with the addition of 2.5 μg/ml selective antibiotic G418 (Sigma-Aldrich)]. The day before the experiment, ^5x105 cells/well were seeded into a 6-well plate in 3 ml of complete medium. The next day, cells were incubated with DMSO or a compound of this invention at 5 μM for an additional 24 hours at 37°C. After incubation, cells were trypsinized, resuspended in 100 μl homogenization buffer [25 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1 mM EDTA, 1 mM EGTA-containing phosphatase inhibitors (30 mM NaF, 0.2 mM Na ₃ VO ₄ , 1 nM okadaic acid, 1 mM PMSF, 5 mM Na ₄ P ₂ O ₇ ) and protease inhibitor cocktail (Complete™, Roche)] and then physically lysed using three rapid freeze-thaw cycles. The samples were then directly tested in the AlphaLISA assay.

The AlphaLisa method quantified phosphorylated, aggregated and total tau protein using the following antibody pairs:

HT7-acceptor granules + biotin (BT)-tau13-donor granules: total tau protein,

HT7-acceptor granules + biotin(BT)-HT7-donor granules: aggregated human tau protein,

Tau13 (Abcam) was biotinylated using the EZ-Link® NHS-PEO Solid Phase Biotinylation Kit (Thermo Scientific), with HT7-biotin purchased from a commercial supplier (Thermo Scientific).

For each pair of antibodies, the concentration of acceptor beads and biotinylated antibodies was optimized. All samples were first tested in a series of dilutions in PBS to determine the linear range and optimal dilution for each sample and assay. For the final protocol, the following reagents were added to a 384-well white OptiPlate (PerkinElmer):

µl of test diluted sample.

µl of a mixture of biotin-mAb acceptor beads at the following final concentrations:

HT7-BT at a concentration of 1.25 nM in combination with HT7-Ac granules at a concentration of 10 μg/ml, Tau13-BT at a concentration of 5 nM in combination with HT7-Ac granules at a concentration of 2.5 μg/ml. After incubating this mixture at room temperature for 1 h, 25 μl of streptavidin donor beads (Perkin Elmer) at a concentration of 25 μg/ml was added in the dark. Plates were analyzed after 30 min of incubation using the EnSpire Alpha instrument and EnSpire workstation version 3.00. Data for aggregated tau were normalized to total tau and then expressed as a percentage of DMSO-treated cells.

The following examples of compounds were measured.

Legend: +++%>50;

+% 50<x<25;

+% 25<x<10.

Reduction of intracellular tau protein misfolding by immunocytochemistry.

A human neuroblastoma cell line overexpressing full-length human tau carrying the P301L mutation was cultured in complete medium [DMEM-F12 4.5 g/L Glutamax (Invitrogen), 15% FBS (Biochrom), 1% penicillin/streptomycin (Invitrogen ) with the addition of 2.5 μg/ml selective antibiotic G418 (Sigma-Aldrich)].

To induce the accumulation of intracellular misfolded tau protein, cells were differentiated in vitro from neuroblastoma cells to neuron-like cells. To do this, cells were seeded in a 96-well plate at a density of 2500 cells/well in 100 μl of complete medium supplemented with 10 μM retinoic acid (RA; Sigma, R2625) for 1 week. Every 2-3 days the medium was changed and fresh retinoic acid was added. To evaluate the ability of the compounds of this invention to reduce levels of misfolded intracellular tau protein, the compounds were applied to cells at

- 100 044393 concentrations in the range between 0.1 and 10 nM for 24 hours. After incubation with the compounds, cells were fixed in 4% PFA for 15 minutes and washed 3 times with PBS. Cells were then blocked in 10% pure goat serum (NGS), 0.25% Triton X-100 in PBS for 2 h at room temperature. Fixed permeabilized cells were then incubated overnight in 5% NGS/0.25% Triton X100 in PBS with anti-mouse MC1 monoclonal antibody (provided by Professor Peter Davis, Albert Einstein College of Medicine, New York, New York, USA) at a dilution of 1:4000 for detection of misfolded tau and polyclonal total tau to rabbit (Abcam; ab64193) and diluted 1:400 to detect total tau. After incubation with primary antibodies, cells were washed 3 times in PBS and then incubated for 30 min with FITC-labeled goat-anti-mouse (Abcam ab6785) and Alexa Fluor 594-labeled goat-anti-rabbit secondary antibodies (Abcam 150080). Cells were then washed 3 times in PBS and imaged using Incucyte. The misfolded tau signal was normalized to the total tau signal, and the reduction in tau misfolding was expressed as a percentage compared to vehicle-treated cells. Data represent the average of at least 3 images per well. When assessed for the ability to reduce intracellular tau misfolding, Example 44 was effective at low nM concentrations as illustrated in FIG. 1.

Analysis of inhibition of full-length tau (flT) aggregation by ThaT.

The human tau protein isoform (2N4R; 441 amino acids) was purchased from Biotechne (USA). The protein was expressed in E coli bacteria, purified and concentrated in PBS to a final concentration of 50 μM.

To induce tau aggregation, 4 μM monomeric flTau was incubated for 72 h at 37°C with agitation cycles including both orbital shaking and the stir function using a Hula Mixer (Life Technologies) with paired-helical tau filaments (PHF) enriched from the autopsied brain of one Alzheimer's disease (AD) patient obtained from an external source (Tissue Solutions) were diluted 1:200. The enrichment methodology was prepared based on Jicha et al., 1997 (Journal of Neuroscience Research 48:128-132 (1997)) and Rostagno and Ghiso, 2009 (Current protocols in cell biology (2009), Chapter 3, Unit 3.33 3.33.1- 33). Briefly, approximately 9 g of human AD brain sample was thawed on ice and homogenized with 50 ml homogenization buffer [0.75 M NaCl in RAB buffer (100 mM 2-(N-morpholino)ethanesulfonic acid (MES), 1 mM EGTA , 0.5 mM MgSO ₄ , 2 mM DTT, pH 6.8) with added protease inhibitors (Complete; Roche 11697498001)] in a glass Dounce homogenizer. The homogenate was then incubated at 4°C for 20 min to allow any residual microtubules to depolymerize before being transferred to polycarbonate centrifuge vials (16 x 76 mm; Beckman 355603) and centrifuged at 11,000 g (12,700 rpm) in an ultracentrifuge (Beckman, XL100K ) for 20 min at 4°C using a pre-cooled 70.1 rotor (Beckman, 342184). The pellets were stored on ice. The supernatants were collected in polycarbonate vials and centrifuged again at 100,000 g (38,000 rpm) for 1 h at 4°C in a 70.1 Ti rotor to isolate PHF-rich pellets, leaving soluble tau protein in the supernatants. Pellets from the first and second centrifugation were resuspended in 120 ml of extraction buffer [10 mM Tris-HCl pH 7.4, 10% sucrose, 0.85 M NaCl, 1% protease inhibitor (Calbiochem 539131), 1 mM EGTA, 1% phosphatase inhibitor (Sigma P5726 and P0044)]. The solution was then transferred to polycarbonate centrifuge vials (16 x 76 mm; Beckman 355603) and centrifuged at 15,000 g (14,800 rpm) in an ultracentrifuge (Beckman, XL100K) for 20 min at 4°C using a 70.1 Ti rotor. In the presence of 10% sucrose and low-speed centrifugation, most of the PHF remained in the supernatant, while intact or fragmented NFTs and larger PHF aggregates precipitated. The pellets were discarded. 20% Sarkosyl (Sigma L7414-10ML) was added to the supernatants to a final concentration of 1% and stirred at room temperature for 1 hour. This solution was then centrifuged in polycarbonate vials at 100,000 g (38,000 rpm) for 1 hour at 4°C With 70.1 Ti in the rotor, the pellets containing the PHF-rich material were resuspended in a total final volume of 1.5 ml PBS, aliquoted and stored at 80°C.

To test the ability of the compounds of this invention to inhibit tau protein aggregation, serial dilutions of the compounds in DMSO were added to the flTau/PHF monomer mixture prior to incubation. After incubation, 40 μl of the mixture was transferred to a black 384-well assay plate (Perkin-Elmer) and mixed with 10 μl of 100 μM ThT in 250 mM glycine (both from Sigma-Aldrich, Buchs, Switzerland) in PBS. Fluorescence was measured in duplicate on a Tecan Spark reader using a filter (excitation at 448 nm/BW 7 nm, emission at 485 nm/BW 20 nm). The type of dose-response curve was obtained from two independent experiments, each with technical duplicates, and IC ₅₀ was calculated using GraphPad Prism 7.03.

The flTau aggregation inhibition IC ₅₀ values of Example 44 and Example 46 are shown in the table below.

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Claims (22)

CLAIM 1. Compound of formula (Ia) or its stereoisomers, racemic mixtures, tautomers, pharmaceutically acceptable salts; B represents NR ^a ; E and V are independently selected from the group consisting of N, NR ⁵ , O and S; G is selected from the group consisting of a benzene ring and a pyridine ring; J is selected from the group consisting of O, NR ¹ and CH2 or J is selected from the group consisting of CH or C if J is attached to R ² ; Y, Y ¹ , Y2 and Y ³ represent CZ; Z is independently selected from the group consisting of H, halogen, O-(C1- _C6 alkyl), C1- _C6 alkyl and CN; R is independently selected from the group consisting of and -NR ³ R ⁴ ; R ^a is selected from the group consisting of H and C ₁ -C ₆ alkyl; R ^d , R ^e , R ^f and R ^g are H or one of R ^d and R ^e , and one of R ^f , R ^g can be joined to form a 5-7 membered ring; where when A represents or . R ^j is independently selected from the group, co- consisting of -halogen, -O-(C1-C ₆ alkyl), -NR ³ R ⁴ , -CN, where a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom and carbon atom c or d or a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom b and carbon atom c or d; And - 102 044393 when A represents Rj is independently selected from the group consisting of -F, -O-(C1-C ₆ alkyl), -NR ³ R ⁴ , -CN, wherein a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom and carbon atom c or d, or where a bridge or bond containing a C _1-2 carbon atom may be present between carbon atom b and carbon atom c or d; R1 is selected from the group consisting of H and C1-C6 alkyl; R2 is independently selected from the group consisting of C1-C6 alkyl or -O-( _C1 - _C6 alkyl), and if two R2 are geminal, they can be joined to form a 3-6 membered ring; R ³ and R ⁴ are independently selected from the group consisting of H and C1-C6 alkyl; R ⁵ is selected from the group consisting of H and C1-C6 alkyl; n is 0, 1, 2, 3 or 4; r and s are independently equal to 0, 1, 2 or 3; and t and u are independently equal to 1, 2 or 3. 2. The compound according to claim 1, characterized in that A is attached to the N atom in any accessible position, and the substituents Rj and Rj are as defined in claim 1. and may be replaced by one or more 3. A compound according to any one of claims 1 and 2, which is a compound of formula (Ib) where R ^a , R j and Z are as defined in paragraph 1, and p is equal to 1 or 2. 4. A connection selected from the group consisting of - 103 044393 - 104 044393 - 105 044393 5. A pharmaceutical composition containing a therapeutically effective amount of a compound according to any one of claims. 1-4 and a pharmaceutically acceptable carrier or excipient. 6. Use of a compound according to any one of claims 1 to 4 for the treatment, amelioration or prevention of a disorder or disorder associated with tau protein aggregates. 7. A method of treating, preventing, or alleviating a disorder associated with tau protein aggregates, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof. 8. Use of a compound according to any one of claims 1 to 4 in the manufacture of a medicament for the treatment of a disorder or disorder associated with tau protein aggregates. 9. Use of a compound according to any one of claims 1 to 4 in the manufacture of a medicinal product for the treatment of Alzheimer's disease. 10. Use of a compound according to any one of claims 1 to 4 for the manufacture of a medicament for the treatment of progressive supranuclear palsy (PSP). 11. A method of treating, preventing or alleviating Alzheimer's disease, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof. 12. A method of treating, preventing or alleviating PSP, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof. - 106 044393 13. A method of reducing tau protein aggregation, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof. 14. A method for preventing the formation of tau protein aggregates and/or inhibiting tau protein aggregation, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof. 15. A method of intracellular interaction with tau protein aggregates, comprising administering an effective amount of a compound according to any one of claims 1 to 4 to a subject in need thereof. 16. Method according to any one of claims 11 to 15, characterized in that the subject is an animal or a human. 17. A combination pharmaceutical composition containing a therapeutically effective amount of a compound according to any one of claims 1 to 4 and a therapeutically effective amount of at least one additional biologically active compound selected from a therapeutic agent other than a compound according to any one of claims 1 to 4, pharmaceutically acceptable carrier, diluent and excipient. 18. The composition according to claim 17, characterized in that the additional biologically active compound is a compound used in the treatment of amyloidosis. 19. The composition according to claim 17 or 18, characterized in that the additional biologically active compound is selected from the group consisting of compounds against oxidative stress, anti-apoptotic compounds, metal chelators, DNA repair inhibitors, 3-amino-1-propanesulfonic acid (3APS) , 1,3-propane disulfonate (1,3PDS), α-secretase activators, tau β- and γ-secretase inhibitors, neurotransmitters, β-sheet disruptors, cellular component attractants, beta-amyloid scavenging/depleting agents, N-truncated inhibitors end of amyloid beta, including amyloid beta pyroglutamate 3-42, anti-inflammatory compounds or cholinesterase (ChEl) inhibitors, M1 agonists, amyloid or tau-modifying drugs and dietary supplements, antibodies, including any functionally equivalent antibody or functional parts thereof or vaccine . 20. Composition according to any one of claims 17 to 19, characterized in that the compound and/or additional biologically active compound is/are present in a therapeutically effective amount. 21. Use according to claim 6 or 8, characterized in that the disorder is selected from Alzheimer's disease (AD), familial AD, primary age-related tauopathy (PART), Creutzfeldt-Jakob disease, Boxer dementia, Down syndrome, Gerstmann-Straussler-Scheinker disease (GSS), inclusion body myositis, prion cerebral amyloid angiopathy, traumatic brain injury (TBI), amyotrophic lateral sclerosis (ALS), parkinsonian dementia (Guam syndrome), non-Guam motor neuron disease with neurofibrillary tangles, a disease characterized by the appearance of argyrophilic grains , corticobasal degeneration (CBD), diffuse neurofibrillary tangles with calcification, frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), Hallerwarden-Spatz disease, multiple system atrophy (MSA), Niemann-Pick disease type C, pallido -ponto-nigral degeneration, Pick's disease (PiD), progressive subcortical gliosis, progressive supranuclear palsy (PSP), subacute sclerosing panencephalitis, tangle-predominant dementia, postencephalitic parkinsonism, myotonic dystrophy, subacute sclerosing panencephalitis, mutations in LRRK2, chronic traumatic encephalopathy ( CTE), familial British dementia, familial Danish dementia, other frontotemporal lobar degenerations, Guadalupean parkinsonism, neurodegeneration with brain iron accumulation associated with SLC9A6 mental retardation, white matter tauopathies with globular glial inclusions, epilepsy, dementia with Lewy bodies (LBD), mild cognitive impairment (MCI), multiple sclerosis, Parkinson's disease, HIV-associated dementia, midlife diabetes, senile cardiac amyloidosis, glaucoma, ischemic stroke, AD psychosis and Huntington's disease. 22. The method according to claim 7, characterized in that the disorder is selected from Alzheimer's disease (AD), familial AD, primary age-related tauopathy (PART), Creutzfeldt-Jakob disease, Boxer dementia, Down syndrome, Gerstmann-Straussler-Scheinker disease (GSS ), inclusion body myositis, prion cerebral amyloid angiopathy, traumatic brain injury (TBI), amyotrophic lateral sclerosis (ALS), parkinsonian dementia (Guam syndrome), non-Guam motor neuron disease with neurofibrillary tangles, a disease characterized by the appearance of argyrophilic grains, corticobasal degeneration (CBD), diffuse neurofibrillary tangles with calcification, frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), Hallerwarden-Spatz disease, multiple system atrophy (MSA), Niemann-Pick disease type C, pallidoponto -nigral degeneration, Pick's disease (PiD), progressive subcortical gliosis, progressive supranuclear palsy (PSP), subacute sclerosing panencephalitis, tangle-predominant dementia, postencephalitic parkinsonism, myotonic dystrophy, - 107 -