WO2013053872A1 - Synthesis method and intermediates useful in the preparation of pyrrolobenzodiazepines - Google Patents

Synthesis method and intermediates useful in the preparation of pyrrolobenzodiazepines Download PDF

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Publication number
WO2013053872A1
WO2013053872A1 PCT/EP2012/070232 EP2012070232W WO2013053872A1 WO 2013053872 A1 WO2013053872 A1 WO 2013053872A1 EP 2012070232 W EP2012070232 W EP 2012070232W WO 2013053872 A1 WO2013053872 A1 WO 2013053872A1
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compound
group
formula
mmol
compound according
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PCT/EP2012/070232
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French (fr)
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Philip Wilson Howard
Arnaud Tiberghien
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Spirogen Sàrl
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Priority to EA201490550A priority Critical patent/EA029046B1/en
Priority to CA2850096A priority patent/CA2850096C/en
Priority to EP12772781.6A priority patent/EP2751076B1/en
Priority to AU2012322933A priority patent/AU2012322933B2/en
Priority to CN201280061966.5A priority patent/CN104011018B/en
Priority to JP2014535096A priority patent/JP6166728B2/en
Priority to US14/351,166 priority patent/US9102704B2/en
Priority to BR112014009070-0A priority patent/BR112014009070B1/en
Publication of WO2013053872A1 publication Critical patent/WO2013053872A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06008—Dipeptides with the first amino acid being neutral
    • C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic
    • C07K5/06034—Dipeptides with the first amino acid being neutral and aliphatic the side chain containing 2 to 4 carbon atoms
    • C07K5/06052—Val-amino acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/18—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member
    • C07D207/20—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/18—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member
    • C07D207/22—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D207/24—Oxygen or sulfur atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/18—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member
    • C07D207/22—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having one double bond between ring members or between a ring member and a non-ring member with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D207/24—Oxygen or sulfur atoms
    • C07D207/26—2-Pyrrolidones
    • C07D207/263—2-Pyrrolidones with only hydrogen atoms or radicals containing only hydrogen and carbon atoms directly attached to other ring carbon atoms
    • C07D207/27—2-Pyrrolidones with only hydrogen atoms or radicals containing only hydrogen and carbon atoms directly attached to other ring carbon atoms with substituted hydrocarbon radicals directly attached to the ring nitrogen atom
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D409/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms
    • C07D409/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings
    • C07D409/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D413/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D413/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings
    • C07D413/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D519/00—Heterocyclic compounds containing more than one system of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring system not provided for in groups C07D453/00 or C07D455/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F7/00—Compounds containing elements of Groups 4 or 14 of the Periodic Table
    • C07F7/02—Silicon compounds
    • C07F7/08—Compounds having one or more C—Si linkages
    • C07F7/18—Compounds having one or more C—Si linkages as well as one or more C—O—Si linkages
    • C07F7/1804—Compounds having Si-O-C linkages
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Definitions

  • the present invention relates to methods useful in the synthesis of pyrrolobenzodiazepin (PBDs), and in particular of pyrrolobenzodiazepines having C2 substitution and N10 protection.
  • PBDs pyrrolobenzodiazepin
  • PBDs are of the general structure
  • a diverse range cytotoxic compounds having an aryl group at the C2 position for example:
  • WO 2005/085251 discloses the synthesis of a number cytoxic compounds having an aromatic substituent or unsaturated substituent at C2 from the intermediate:
  • the C2-aryl or unsaturated group is installed by forming a C2-enol triflate followed by a palladium catalyzed Suzuki reaction. It is advantageous to add the C2 aryl or unsaturated substituents as late as possible in the synthesis by the methods described above, as this means a diverse range of substituents can be added to a single 'core'.
  • the protecting groups or linkers desired at N10 are labile under either the triflation conditions (e.g. substituted p-aminobenzyl or p- hydroxybenzyl carbamates) or palladium catalysis (e.g. Alloc), see below.
  • Such groups are preferably introduced at a point in the synthesis of the PBD before the B- ring is formed by cyclisation. Otherwise, these groups would need to be added via a chloroformate to the PBD's imine bond.
  • the cyclisation of PBD is best carried out where the pro-N 10 nitrogen has only a single hydrogen attached, i.e., where the protecting group is already in place.
  • WO 2005/085259 discloses PBD compounds useful as intermediates in the synthesis of PBD dimers. These intermediates have a hydroxy group at the C7 or C8 position.
  • the present invention comprises a compound of formula I:
  • R 8 is OProt°, where Prot° is a silicon-based oxygen protecting group orthogonal to R c ;
  • R 9 is selected from H, methyl and methoxy;
  • R s is selected from CF 3 , (CF 2 ) 3 CF 3 , CH 3 and ;
  • R c is selected from:
  • the compound of formula I is preferably substantially free of any of the corresponding compound of formula IB:
  • the present invention provides a method of synthesising a compound of formula I as defined in the first aspect of the invention from a compound of formula II:
  • R 7 , R 8 , R 9 and R c are as defined in the first aspect
  • R 2 is selected from:
  • Ci -5 alkyl group including a C2-5 alkenyl group and a C2-5 alkynyl group
  • R 2 is an optionally substituted C5-20 aryl group (a Suzuki coupling) carried out by palladium catalysed cross coupling of I with the appropriate aryl boron derivative.
  • the palladium catalyst may be any suitable catalyst, for example Pd(PPh 3 ) 4 , Pd(OCOCH 3 ) 2 , PdCI 2 , or Pd(dba) 3 .
  • Suzuki couplings may also be used to cross couple alkyl boron species.
  • the Gibbs group has reported (Mu, Y.Q and Gibbs, R.A, Tetrahedron Letters, 36, 5669-5672, 1995) the successful conversion of an enol triflate to a methyl substituted alkene (using a
  • Cyclopropyl Ml DA boronates have been successfully coupled to aryl chlorides under slow release conditions; intact Ml DA boronates are resistant to standards Suzuki conditions due to their pyramidal hybridization (Knapp et al. J. Am. Chem. Soc, 2009, 131, 6961-6963). These conditions are also be applicable to enoltriflates.
  • Molander Molander and Ito Org. Lett., 2001 , 3, 393-3966 has developed potassiu alkyltrifluoroborates as Suzuki coupling partners for aryl and vinyl triflates.
  • Suzuki couplings may also be used to cross couple alkenyl boron species. Suzuki and co workers have demonstrated (Suzuki et al., Synlett 1990, 221 ; J Org Chem 1993, 58, 2201 ) the coupling of cyclic triflates and benzodioxoleborole compounds
  • pinacol esters of acrylylboronic acids are known:
  • Alkynyl tin reagents have been coupled under Stille conditions to PBD triflates (Tiberghien et al., Bioorg. Med. Chem. Lett. 14, (2004), 5041 -504
  • Alkynyl substituents can also be installed via the Sonogishira reaction (Nishihara et al J. Org. Chem., 2000, 65, 1780-1787) :
  • palladium catalysed reactions include the use triakylindium reagents in cross- coupling reactions (Perez et al. Org. Lett, 1999, 1, 1267-1269) :
  • R D is selected from:
  • R D is orthogonal to Prot°.
  • Reprotection as an aceate can be achieved, for example, by reaction with acetyl chloride; reprotection as a benzoate can be achieved, for example, by reaction with benzoyl chloride; reprotection as a silyl ether can be achieved, for example, by reaction with the appropriate silyl chloride.
  • the reduction of the nitro group is then carried out as described above.
  • R 10 is a carbamate-based nitrogen protecting group, which is palladium-labile and/or the conditions used to add the group: , or contains a moiety which is palladium-labile and/or is labile under the conditions used to add the group:
  • simple nitrogen protecting groups can also be introduced as a chloroformate, fluoroformate or azidoformate.
  • the more complex nitrogen protecting groups, as well as the simple nitrogen protecting groups, can be introduced as O-succinamide carbonates, O- pentaflurophenyl carbonates and O-nitrophenyl carbonates.
  • TPAP/NMO TFAA/DMSO
  • S0 3 .Pyridine complex/DMSO PDC, PCC, BAIB/TEMPO or under Swern conditions
  • the conversion of V to VI may be achieved by initial removal of the silyl ether protecting group, for example using TBAF in THF, acetic acid in aqueous THF, CsF in DMF or HF in pyridine, followed by oxidation as described above.
  • the removal conditions must be selected such that they do not remove Prot°.
  • R E is selected from: 0-d -2 alkyl; 0-Si-(R Si1 )(R Si2 )(R Si3 ), where R Si1 , R Si2 , R Si3 are as defined above; 0-CH 2 OCH 3 (OMOM), 0-CH 2 OC 2 H 4 OCH 3 (OMEM), OC 5 H 10 (THP).
  • OMOM 0-CH 2 OCH 3
  • OMEM 0-CH 2 OC 2 H 4 OCH 3
  • THP OC 5 H 10
  • Prot° needs to be orthogonal to R E , and therefore R E needs to be selected with this in mind.
  • R E is converted to R E by conventional methods.
  • R E is 0-Ci -2 alkyl
  • the conversion may be carried out by rection with Ci -2 alkyl-l and silver oxide or (Ci -2
  • Deprotection of the phenolic hydroxy group is carried out by, for example, lithium acetate.
  • the removal conditions must be selected such that they do not remove R E .
  • Hal is selected from I, CI and Br, and n is from 1 to 12.
  • the conversion of VIII to IX is achieved by reaction with a compound of formula X: where Q is selected from I, CI and Br.
  • the reaction is carried out, for example, in refluxing acetone with a base, such as K 2 C0 3 .
  • An excess of the compound of formula X is required to achieve the desired product.
  • H The present invention also provides a method of linking compounds of formulae VIII and IX, to yield a compound of formula XI:
  • R 2 , R 7 , R 9 , R 10 and R E are independently selected from the same groups as R 2 , R 7 , R 9 , R 10 and R E respectively.
  • the reaction is carried out, for example, in refluxing acetone with a base, such as K 2 C0 3 .
  • the present invention also provides:
  • substantially free of means that there is less than 5% by weight of the compound of the undesired compound (i.e., B version) compared to the amount of the desired compound. More preferably, there is less than 1 % or 0.1 % by weight.
  • the present invention also provides
  • the defined substituent groups in the intermediate compounds are the same as those in the final products where designated in the same way (e.g. R 9 remains the same throughout the method of synthesis).
  • the methods may also comprise the intermediate steps as set out above.
  • Silican-based oxygen protecting groups are well known in the art. A large number of suitable groups are described on pages 1 13 to 148 of Greene, T.W. and Wuts, G.M., Protective Groups in Organic Synthesis, 3 Edition, John Wiley & Sons, Inc., 1999, which is incorporated herein by reference.
  • the groups form silyl ethers with the oxygen to which they are attached, and are generally of the formula:
  • 3 are independently selected from a Ci -6 saturated alkyl group and a phenyl group. Examples are:
  • a first protecting group which is orthogonal to a second protecting group means that the first protecting group can be removed from a compound bearing both protecting groups without the second protecting group being removed.
  • R D is orthogonal to Prot°
  • Prot° is orthogonal to R E .
  • Carbamate-based nitrogen protecting groups are well known in the art. A large number of suitable groups are described on pages 503 to 549 of Greene, T.W. and Wuts, G.M., Protective Groups in Organic Synthesis, 3 rd Edition, John Wiley & Sons, Inc., 1999, which is incorporated herein by reference.
  • the carbamate-based nitrogen protecting groups used may be those which are sensitive to palladium or sensitive to the conditions needed to add the group: 8 Table 1 provides examples of some such groups ('simple nitrogen protecting groups'):
  • the carbamate nitrogen protecting group is not Cbz (benzyl carbamate), PNZ (p-nitrobenzylcarbamate), or 3,4-dimethoxy-6-nitrobenzyl carbamate.
  • carbamate-based nitrogen protecting groups having a complex architecture are also suitable for use in the present invention. These groups may be or may comprise elements of the groups described, for example, in WO 00/12507. These carbamate-based nitrogen protecting groups may include elements that to palladium or the conditions used to add the group:
  • the carbamate-based nitrogen protecting group is a linker for connection to an antibody
  • R L where the asterisk indicates the point of attachment to the N10 position
  • G 1 is a functional group to form a connection to a cell binding agent
  • L 1 is a cleavable linker
  • the protecting group R L is removable from the N10 position thereby to provide the imine or carbinoalmine form of the PBD.
  • L 1 is preferably a cleavable linker, and may be referred to as a trigger for activation of the linker for cleavage.
  • L 1 and L 2 can vary widely. These groups are chosen on the basis of their cleavage characteristics, which may be dictated by the conditions at the site to which the PBD molecule is delivered, for example as an antibody drug conjugate. Those linkers that are cleaved by the action of enzymes are preferred, although linkers that are cleavable by changes in pH (e.g. acid or base labile), temperature or upon irradiation (e.g. photolabile) may also be used. Linkers that are cleavable under reducing or oxidising conditions may also find use in the present invention.
  • pH e.g. acid or base labile
  • temperature or upon irradiation e.g. photolabile
  • L 1 may comprise a contiguous sequence of amino acids.
  • the amino acid sequence may be the target substrate for enzymatic cleavage, thereby allowing release of R 10 from the N 10 position.
  • the L 1 is cleavable by the action of an enzyme.
  • the enzyme is an esterase or a peptidase.
  • the enzyme cleaves the bond between L 1 and L 2 .
  • L 1 and L 2 may be connected by a bond selected from:
  • An amino group of L 1 that connects to L 2 may be the N-terminus of an amino acid or may be derived from an amino group of an amino acid side chain, for example a lysine amino acid side chain.
  • An carboxyl group of L 1 that connects to L 2 may be the C-terminus of an amino acid or may be derived from a carboxyl group of an amino acid side chain, for example a glutamic acid amino acid side chain.
  • a hydroxy group of L 1 that connects to L 2 may be derived from a hydroxy group of an amino acid side chain, for example a serine amino acid side chain.
  • n is 0 to 3.
  • the phenylene ring is optionally substituted with one, two or three substituents as described herein. In one embodiment, the phenylene group is optionally substituted with halo, N0 2 , R or OR.
  • Y is NH
  • n is 0 or 1 .
  • n is 0.
  • the self-immolative linker may be referred to as a
  • PABC p-aminobenzylcarbonyl linker
  • the group L * is a linker L 1 as described herein, which may include a dipeptide group.
  • Each phenylene ring is optionally substituted with one, two or three substituents as described herein. In one embodiment, the phenylene ring having the Y substituent is optionally substituted and the phenylene ring not having the Y substituent is unsubstituted. In one embodiment, the phenylene ring having the Y substituent is unsubstituted and the phenylene ring not having the Y substituent is optionally substituted.
  • E is O, S or NR
  • D is N, CH, or CR
  • F is N, CH, or CR.
  • D is N.
  • D is CH.
  • E is O or S.
  • F is CH.
  • the linker is a cathepsin labile linker.
  • L 1 comprises a dipeptide
  • the dipeptide may be represented as -NH-X X 2 -CO-, where -NH- and -CO- represent the N- and C-terminals of the amino acid groups Xi and X 2 respectively.
  • the amino acids in the dipeptide may be any combination of natural amino acids.
  • the linker is a cathepsin labile linker
  • the dipeptide may be the site of action for cathepsin-mediated cleavage.
  • CO and NH may represent that side chain functionality.
  • the group -X ! -X 2 - in dipeptide, -NH-X X2-CO- is selected from:
  • the group -X X 2 - in dipeptide is selected from:
  • the group -X X 2 - in dipeptide is -Phe-Lys- or -Val-Ala-.
  • the group -X ! -X 2 - in dipeptide is -Val-Cit-.
  • dipeptide combinations may be used, including those described by Dubowchik et al. ⁇ Bioconjugate Chem. 2002, 13, 855-869), which is incorporated herein by reference.
  • the amino acid side chain is chemically protected, where appropriate.
  • the side chain protecting group may be a group as discussed below in relation to the group R L .
  • the present inventors have established that protected amino acid sequences are cleavable by enzymes.
  • the present inventors have established that a dipeptide sequence comprising a Boc side chain-protected Lys residue is cleavable by cathepsin.
  • the amino acids selected are those having no reactive side chain functionality.
  • the amino acids may be selected from: Ala, Gly, lie, Leu, Met, Phe, Pro, and Val.
  • the dipeptide is used in combination with a self-immolative linker.
  • the self-immolative linker may be connected to -X 2 -.
  • the dipeptide is used in combination with a self-immolative linker.
  • the self-immolative linker may be connected to -X 2 -.
  • -X 2 - is connected directly to the self-immolative linker.
  • the group -X 2 -CO- is connected to Y, where Y is NH, thereby forming the group -X 2 -CO-NH-.
  • -NH-Xr is connected directly to A.
  • A may comprise the functionality -CO- thereby to form an amide link with -X .
  • the PABC group is connected directly to the N10 position.
  • the self-immolative linker and the dipeptide together form the group -NH-Phe- Lys-CO-NH-PABC-, which is illustrated below:
  • the asterisk indicates the point of attachment to the N10 position
  • the wavy line indicates the point of attachment to the remaining portion of the linker L 1 or the point of attachment to A.
  • the wavy line indicates the point of attachment to A.
  • the side chain of the Lys amino acid may be protected, for example, with Boc, Fmoc, or Alloc, as described above.
  • the self-immolative linker and the dipeptide together form the group
  • n 0 to 3
  • Y is a covalent bond or a functional group
  • E is an activatable group, for example by enzymatic action or light, thereby to generate a self-immolative unit.
  • the phenylene ring is optionally further substituted with one, two or three substituents as described herein.
  • the phenylene group is optionally further substituted with halo, N0 2 , R or OR.
  • n is 0 or 1 , most preferably 0.
  • E is selected such that the group is susceptible to activation, e.g. by light or by the action of an enzyme.
  • E may be -N0 2 or glucoronic acid.
  • the former may be susceptible to the action of a nitroreductase, the latter to the action of a ⁇ -glucoronidase.
  • E * is the activated form of E
  • Y is as described above.
  • the group Y may be a covalent bond to L 1 .
  • the group Y may be a functional group selected from:
  • L 1 is a dipeptide
  • the dipeptide sequence need not be a substrate for an enzymatic activity.
  • G is intended to form a spacer group that is suitable for indirectly connecting L 1 to a cell binding agent.
  • L 1 and G 1 may be connected by a bond selected from:
  • the functional group G 1 is or comprises an amino, carboxylic acid, hydroxy, thiol, or maleimide group for reaction with an appropriate group on the cell binding agent.
  • G 1 comprises a maleimide group.
  • the group G 1 is an alkyl maleimide group. This group is suitable for reaction with thiol groups, particularly cysteine thiol groups, present in the cell binding agent, for example present in an antibody.
  • the group G 1 is:
  • n is 0 to 6. In one embodiment, n is 5.
  • the group G 1 is:
  • n is 0 to 6. In one embodiment, n is 5.
  • the group G 1 is:
  • n is 0 or 1
  • m is 0 to 30.
  • n is 1 and m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
  • the group G 1 is:
  • n is 0 or 1
  • m is 0 to 30.
  • n is 1 and m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
  • the maleimide-derived group is replaced with the group:
  • the maleimide group is replaced with a group selected from:
  • the group G 1 is:
  • n 0 to 6. In one embodiment, n is 0.
  • the group G 1 is:
  • m is 0 to 30. In a preferred embodiment, m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
  • the roup G 1 is:
  • m is 0 to 30. In a preferred embodiment, m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
  • G 1 is -NH 2 , -NHMe, -COOH, -OH or -SH.
  • G 1 is -NH 2 or -NHMe. Either group may be the N-terminal of an L 1 amino acid sequence. In one embodiment, where L 1 is present, G 1 is -NH 2 , and L 1 is an amino acid sequence -X X 2 -, as defined above in relation to R 10 . In one embodiment, where L 1 is present, G 1 is COOH. This group may be the C-terminal of an L 1 amino acid sequence.
  • G 1 is OH.
  • G 1 is SH.
  • the group G 1 may be convertable from one functional group to another. In one
  • G 1 is -NH 2 .
  • This group is convertable to another group G 1 comprising a maleimide group.
  • the group -NH 2 may be reacted with an acid or an activated acid (e.g. N-succinimide forms) of those G 1 groups comprising maleimide shown above.
  • R L is a group that is a precursor to the linker that is provided with a functional group.
  • G 1 is -NH 2 , -NHMe, -COOH, -OH or -SH.
  • these groups are provided in a chemically protected form.
  • the chemically protected form is therefore a precursor to the linker that is provided with a functional group.
  • G 1 is -NH 2 in a chemically protected form.
  • the group may be protected with a carbamate protecting group.
  • the carbamate protecting group may be selected from the group consisting of:
  • G 1 is -NH 2
  • it is protected with an Alloc or Fmoc group.
  • the nitrogen protecting group for one of the PBD units is a group R L and the nitrogen protecting group for the other of the PBD units is a capping group, Ca :
  • G 2 is a terminating group
  • L 3 is a covalent bond or a cleavable linker L 1
  • the protecting group R c is removable from the N10 position thereby to provide the imine or carbinolamine form of the PBD.
  • the term "capping group” is used to indicate that R c is prevented from reacting with a cell binding agent, such as an antibody. Instead, the group R L is for connection to a cell binding agent.
  • L 1 is as defined above in relation to R L .
  • L 2 is as defined above in relation to R L .
  • G 2 is Ac (acetyl) or Moc, or a carbamate protecting group selected from:
  • G 2 is different to G 1 . Furthermore, it is preferred that G 1 is removable under conditions that do not remove G 1 .
  • substituted refers to a parent group which may be unsubstituted or which may be substituted. Unless otherwise specified, the term “substituted” as used herein, pertains to a parent group which bears one or more substitutents.
  • substitutents refers to a chemical moiety which is covalently attached to, or if appropriate, fused to, a parent group. A wide variety of substituents are well known, and methods for their formation and introduction into a variety of parent groups are also well known.
  • C-1-12 alkyl refers to a monovalent moiety obtained by removing a hydrogen atom from a carbon atom of a hydrocarbon compound having from 1 to 12 carbon atoms, which may be aliphatic or alicyclic, and which may be saturated or unsaturated (e.g. partially unsaturated, fully unsaturated).
  • alkyl includes the sub-classes alkenyl, alkynyl, cycloalkyl, etc., discussed below.
  • saturated alkyl groups include, but are not limited to, methyl (Ci ), ethyl (C 2 ), propyl (C 3 ), butyl (C 4 ), pentyl (C 5 ), hexyl (C 6 ) and heptyl (C 7 ).
  • saturated linear alkyl groups include, but are not limited to, methyl (Ci), ethyl (C 2 ), n-propyl (C 3 ), n-butyl (C 4 ), n-pentyl (amyl) (C 5 ), n-hexyl (C 6 ) and n-heptyl (C 7 ).
  • saturated branched alkyl groups include iso-propyl (C 3 ), iso-butyl (C 4 ), sec-butyl (C 4 ), tert-butyl (C 4 ), iso-pentyl (C 5 ), and neo-pentyl (C 5 ).
  • C2-12 Alkenyl The term "C2-12 alkenyl" as used herein, pertains to an alkyl group having one or more carbon-carbon double bonds.
  • C2-12 alkynyl refers to an alkyl group having one or more carbon-carbon triple bonds.
  • unsaturated alkynyl groups include, but are not limited to, ethynyl (ethinyl, -C ⁇ CH) and 2-propynyl (propargyl, -CH 2 -C ⁇ CH).
  • C3-12 cycloalkyl refers to an alkyl group which is also a cyclyl group; that is, a monovalent moiety obtained by removing a hydrogen atom from an alicyclic ring atom of a cyclic hydrocarbon (carbocyclic) compound, which moiety has from 3 to 7 carbon atoms, including from 3 to 7 ring atoms.
  • cycloalkyl groups include, but are not limited to, those derived from:
  • C3-20 heterocyclyl refers to a monovalent moiety obtained by removing a hydrogen atom from a ring atom of a heterocyclic compound, which moiety has from 3 to 20 ring atoms, of which from 1 to 10 are ring heteroatoms.
  • each ring has from 3 to 7 ring atoms, of which from 1 to 4 are ring heteroatoms.
  • the prefixes e.g. C 3-2 o, C 3-7 , C 5-6 , etc.
  • the prefixes denote the number of ring atoms, or range of number of ring atoms, whether carbon atoms or heteroatoms.
  • C 5-6 heterocyclyl as used herein, pertains to a heterocyclyl group having 5 or 6 ring atoms.
  • Examples of monocyclic heterocyclyl groups include, but are not limited to, those derived from: aziridine (C 3 ), azetidine (C 4 ), pyrrolidine (tetrahydropyrrole) (C 5 ), pyrroline (e.g., 3-pyrroline, 2,5-dihydropyrrole) (C 5 ), 2H-pyrrole or 3H-pyrrole (isopyrrole, isoazole) (C 5 ), piperidine (C 6 ), dihydropyridine (C 6 ), tetrahydropyridine (C 6 ), azepine (C 7 );
  • Oi oxirane (C 3 ), oxetane (C 4 ), oxolane (tetrahydrofuran) (C 5 ), oxole (dihydrofuran) (C 5 ), oxane (tetrahyd ropy ran) (C 6 ), dihydropyran (C 6 ), pyran (C 6 ), oxepin (C 7 );
  • N 2 imidazolidine (C 5 ), pyrazolidine (diazolidine) (C 5 ), imidazoline (C 5 ), pyrazoline
  • NiOi tetrahydrooxazole (C 5 ), dihydrooxazole (C 5 ), tetrahydroisoxazole (C 5 ),
  • dihydroisoxazole C 5
  • morpholine C 6
  • tetrahydrooxazine C 6
  • dihydrooxazine C 6
  • oxazine C 6
  • OiSi oxathiole (C 5 ) and oxathiane (thioxane) (C 6 ); and,
  • NiOiSi: oxathiazine C 6
  • substituted monocyclic heterocyclyl groups include those derived from saccharides, in cyclic form, for example, furanoses (C 5 ), such as arabinofuranose, lyxofuranose, ribofuranose, and xylofuranse, and pyranoses (C 6 ), such as allopyranose, altropyranose, glucopyranose, mannopyranose, gulopyranose, idopyranose,
  • C5-20 aryl refers to a monovalent moiety obtained by removing a hydrogen atom from an aromatic ring atom of an aromatic compound, which moiety has from 3 to 20 ring atoms. Preferably, each ring has from 5 to 7 ring atoms.
  • the prefixes e.g. C 3-2 o, C 5-7 , C5-6, etc.
  • C 5-6 ary as used herein, pertains to an aryl group having 5 or 6 ring atoms.
  • the ring atoms may be all carbon atoms, as in "carboaryl groups".
  • carboaryl groups include, but are not limited to, those derived from benzene (i.e. phenyl) (C 6 ), naphthalene (Ci 0 ), azulene (Ci 0 ), anthracene (Ci 4 ), phenanthrene (Ci 4 ), naphthacene (Ci 8 ), and pyrene (Ci 6 ).
  • aryl groups which comprise fused rings, at least one of which is an aromatic ring include, but are not limited to, groups derived from indane (e.g.
  • the ring atoms may include one or more heteroatoms, as in "heteroaryl groups".
  • heteroaryl groups include, but are not limited to, those derived from:
  • Ni pyrrole (azole) (C 5 ), pyridine (azine) (C 6 );
  • NiOi oxazole (C 5 ), isoxazole (C 5 ), isoxazine (C 6 );
  • N1S1 thiazole (C 5 ), isothiazole (C 5 );
  • N 2 imidazole (1 ,3-diazole) (C 5 ), pyrazole (1 ,2-diazole) (C 5 ), pyridazine (1 ,2-diazine) (C 6 ), pyrimidine (1 ,3-diazine) (C 6 ) (e.g., cytosine, thymine, uracil), pyrazine (1 ,4-diazine) (C 6 );
  • heteroaryl which comprise fused rings, include, but are not limited to:
  • C 9 (with 2 fused rings) derived from benzofuran (O 1 ), isobenzofuran (O 1 ), indole (Ni), isoindole (N ⁇ , indolizine (N ⁇ , indoline (N ⁇ , isoindoline (N ⁇ , purine (N 4 ) (e.g., adenine, guanine), benzimidazole (N 2 ), indazole (N 2 ), benzoxazole ( ⁇ ), benzisoxazole ( ⁇ ), benzodioxole (0 2 ), benzofurazan (N 2 Oi), benzotriazole (N 3 ), benzothiofuran (Si), benzothiazole benzothiadiazole (N 2 S);
  • Cio (with 2 fused rings) derived from chromene (O 1 ), isochromene (O 1 ), chroman (O 1 ), isochroman (O 1 ), benzodioxan (0 2 ), quinoline (Ni), isoquinoline (Ni), quinolizine (Ni), benzoxazine (NiOi), benzodiazine (N 2 ), pyridopyridine (N 2 ), quinoxaline (N 2 ), quinazoline (N 2 ), cinnoline (N 2 ), phthalazine (N 2 ), naphthyridine (N 2 ), pteridine (N 4 ); On (with 2 fused rings) derived from benzodiazepine (N 2 );
  • Ci 3 (with 3 fused rings) derived from carbazole (Ni), dibenzofuran (Oi),
  • Ci 4 (with 3 fused rings) derived from acridine (N-i), xanthene (Oi), thioxanthene (Si), oxanthrene (0 2 ), phenoxathiin (O 1 S 1 ), phenazine (N 2 ), phenoxazine ( ⁇ ), phenothiazine thianthrene (S 2 ), phenanthridine (N-i), phenanthroline (N 2 ), phenazine (N 2 ).
  • N atoms in heterocyclyl and heteroaryl groups may be substituted by with CM 2 alkyl, where appropriate.
  • Ether -OR, wherein R is an ether substituent, for example, a Ci -7 alkyl group (also referred to as a Ci -7 alkoxy group, discussed below), a C 3-2 o heterocyclyl group (also referred to as a C 3-2 o heterocyclyloxy group), or a C 5-2 o aryl group (also referred to as a C 5 - 2 o aryloxy group), preferably a Ci -7 alkyl group.
  • R is an ether substituent, for example, a Ci -7 alkyl group (also referred to as a Ci -7 alkoxy group, discussed below), a C 3-2 o heterocyclyl group (also referred to as a C 3-2 o heterocyclyloxy group), or a C 5-2 o aryl group (also referred to as a C 5 - 2 o aryloxy group), preferably a Ci -7 alkyl group.
  • a Ci -7 alkyl group also
  • Alkoxy -OR, wherein R is an alkyl group, for example, a Ci -7 alkyl group.
  • C 1-7 alkoxy groups include, but are not limited to, -OMe (methoxy), -OEt (ethoxy), -O(nPr) (n-propoxy), -O(iPr) (isopropoxy), -O(nBu) (n-butoxy), -O(sBu) (sec-butoxy), -O(iBu) (isobutoxy), and -O(tBu) (tert-butoxy).
  • Acetal -CH(OR 1 )(OR 2 ), wherein R 1 and R 2 are independently acetal substituents, for example, a Ci -7 alkyl group, a C 3-2 o heterocyclyl group, or a C 5-2 o aryl group, preferably a
  • Ci -7 alkyl group or, in the case of a "cyclic" acetal group, R 1 and R 2 , taken together with the two oxygen atoms to which they are attached, and the carbon atoms to which they are attached, form a heterocyclic ring having from 4 to 8 ring atoms.
  • acetal groups include, but are not limited to, -CH(OMe) 2 , -CH(OEt) 2 , and -CH(OMe)(OEt).
  • Hemiacetal -CH(OH)(OR 1 ), wherein R 1 is a hemiacetal substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-20 aryl group, preferably a Ci -7 alkyl group.
  • R 1 is a hemiacetal substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-20 aryl group, preferably a Ci -7 alkyl group.
  • hemiacetal groups include, but are not limited to, -CH(OH)(OMe) and -CH(OH)(OEt).
  • Ketal -CR(OR 1 )(OR 2 ), where R 1 and R 2 are as defined for acetals, and R is a ketal substituent other than hydrogen, for example, a C 1-7 alkyl group, a C 3-2 o heterocyclyl group, or a C 5- 2o aryl group, preferably a C 1-7 alkyl group.
  • Examples ketal groups include, but are not limited to, -C(Me)(OMe) 2 , -C(Me)(OEt) 2 , -C(Me)(OMe)(OEt), -C(Et)(OMe) 2 ,
  • R 1 is as defined for hemiacetals, and R is a hemiketal substituent other than hydrogen, for example, a Ci -7 alkyl group, a C 3-2 o heterocyclyl group, or a C 5-2 o aryl group, preferably a Ci -7 alkyl group.
  • hemiacetal groups include, but are not limited to, -C(Me)(OH)(OMe), -C(Et)(OH)(OMe), -C(Me)(OH)(OEt), and -C(Et)(OH)(OEt).
  • Imino (imine): NR, wherein R is an imino substituent, for example, hydrogen, Ci -7 alkyl group, a C 3-2 o heterocyclyl group, or a C 5-2 o aryl group, preferably hydrogen or a Ci -7 alkyl group.
  • R is an acyl substituent, for example, a C 1-7 alkyl group (also referred to as Ci -7 alkylacyl or Ci -7 alkanoyl), a C 3-2 o heterocyclyl group (also referred to as C 3-20 heterocyclylacyl), or a C 5-2 o aryl group (also referred to as C 5 - 2 o arylacyl), preferably a Ci -7 alkyl group.
  • a C 1-7 alkyl group also referred to as Ci -7 alkylacyl or Ci -7 alkanoyl
  • C 3-2 o heterocyclyl group also referred to as C 3-20 heterocyclylacyl
  • C 5-2 o aryl group also referred to as C 5 - 2 o arylacyl
  • Carboxy (carboxylic acid): -C( 0)OH.
  • Thiocarboxy (thiocarboxylic acid): -C( S)SH.
  • Thiolocarboxy (thiolocarboxylic acid): -C( 0)SH.
  • Thionocarboxy (thionocarboxylic acid): -C( S)OH.
  • Acyloxy (reverse ester): -OC( 0)R, wherein R is an acyloxy substituent, for example, a Ci-7 alkyl group, a C 3-2 o heterocyclyl group, or a C5-20 aryl group, preferably a Ci -7 alkyl group.
  • R is an acyloxy substituent, for example, a Ci-7 alkyl group, a C 3-2 o heterocyclyl group, or a C5-20 aryl group, preferably a Ci -7 alkyl group.
  • Oxycarboyloxy: -OC( 0)OR, wherein R is an ester substituent, for example, a Ci -7 alkyl group, a C 3-2 o heterocyclyl group, or a C 5-2 o aryl group, preferably a Ci -7 alkyl group.
  • R 1 and R 2 are independently amino substituents, for example, hydrogen, a C 1 -7 alkyl group (also referred to as C 1-7 alkylamino or di-C 1 -7 alkylamino), a C 3-20 heterocyclyl group, or a C 5-2 o aryl group, preferably H or a Ci -7 alkyl group, or, in the case of a "cyclic" amino group, R 1 and R 2 , taken together with the nitrogen atom to which they are attached, form a heterocyclic ring having from 4 to 8 ring atoms.
  • R 1 and R 2 are independently amino substituents, for example, hydrogen, a C 1 -7 alkyl group (also referred to as C 1-7 alkylamino or di-C 1 -7 alkylamino), a C 3-20 heterocyclyl group, or a C 5-2 o aryl group, preferably H or a Ci -7 alkyl group, or, in the case of a "cyclic" amino group
  • Amino groups may be primary (-NH 2 ), secondary (-NHR 1 ), or tertiary (-NHR 1 R 2 ), and in cationic form, may be quaternary (- + NR 1 R 2 R 3 ).
  • Examples of amino groups include, but are not limited to, -NH 2 , -NHCH 3 , -NHC(CH 3 ) 2 , -N(CH 3 ) 2 , -N(CH 2 CH 3 ) 2 , and -NHPh.
  • Examples of cyclic amino groups include, but are not limited to, aziridino, azetidino, pyrrolidino, piperidino, piperazino, morpholino, and thiomorpholino.
  • R 1 and R 2 together with the nitrogen atom to which they are attached, form a heterocyclic structure as in, for example, piperidinocarbonyl, morpholinocarbonyl, thiomorpholinocarbonyl, and piperazinocarbonyl.
  • Thioamido (thiocarbamyl): -C( S)NR 1 R 2 , wherein R 1 and R 2 are independently amino substituents, as defined for amino groups.
  • Acylamido (acylamino): -NR 1 C( 0)R 2 , wherein R 1 is an amide substituent, for example, hydrogen, a Ci_ 7 alkyl group, a C 3-20 heterocyclyl group, or a C 5 - 2 o aryl group, preferably hydrogen or a Ci -7 alkyl group, and R 2 is an acyl substituent, for example, a Ci -7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-2 oaryl group, preferably hydrogen or a Ci -7 alkyl group.
  • R 1 and R 2 may together form a cyclic structure, as in for example, succinimidyl, maleimidyl, and phthalimidyl:
  • Ureido -N(R 1 )CONR 2 R 3 wherein R 2 and R 3 are independently amino substituents, as defined for amino groups, and R 1 is a ureido substituent, for example, hydrogen, a C 1-7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-20 aryl group, preferably hydrogen or a C 1-7 alkyl group.
  • R 1 is a ureido substituent, for example, hydrogen, a C 1-7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-20 aryl group, preferably hydrogen or a C 1-7 alkyl group.
  • ureido groups include, but are not limited to, -NHCONH 2 ,
  • Tetrazolyl a five membered aromatic ring having four nitrogen atoms and one carbon atom
  • Imino: NR, wherein R is an imino substituent, for example, for example, hydrogen, a C 1-7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-2 o aryl group, preferably H or a C 1-7 alkyl group.
  • amidine groups include, but are not limited to,
  • Isocyanato -NCO.
  • C 1-7 alkylthio groups include, but are not limited to, -SCH 3 and -SCH 2 CH 3 .
  • Disulfide -SS-R, wherein R is a disulfide substituent, for example, a Ci -7 alkyl group, a C3-20 heterocyclyl group, or a C 5- 2o aryl group, preferably a Ci -7 alkyl group (also referred to herein as Ci -7 alkyl disulfide).
  • R is a disulfide substituent, for example, a Ci -7 alkyl group, a C3-20 heterocyclyl group, or a C 5- 2o aryl group, preferably a Ci -7 alkyl group (also referred to herein as Ci -7 alkyl disulfide).
  • Ci -7 alkyl disulfide groups include, but are not limited to, -SSCH 3 and -SSCH 2 CH 3 .
  • Sulfine (sulfinyl, sulfoxide): -S( 0)R, wherein R is a sulfine substituent, for example, a Ci -7 alkyl group, a C 3-2 o heterocyclyl group, or a C5-20 aryl group, preferably a Ci -7 alkyl group.
  • R is a sulfine substituent, for example, a Ci -7 alkyl group, a C 3-2 o heterocyclyl group, or a C5-20 aryl group, preferably a Ci -7 alkyl group.
  • Sulfinic acid (sulfino): -S( 0)OH, -S0 2 H.
  • Sulfonic acid (sulfo): -S( 0) 2 OH, -S0 3 H.
  • Sulfinate (sulfinic acid ester): -S( 0)OR; wherein R is a sulfinate substituent, for example, a C 1 -7 alkyl group, a C3- 20 heterocyclyl group, or a C 5-2 o aryl group, preferably a Ci_ 7 alkyl group.
  • R is a sulfonate substituent, for example, a C 1-7 alkyl group, a C 3-2 o heterocyclyl group, or a C 5-2 o aryl group, preferably a C 1-7 alkyl group.
  • R is a sulfonyloxy substituent, for example, a Ci -7 alkyl group, a C3- 20 heterocyclyl group, or a C 5-2 o aryl group, preferably a Ci -7 alkyl group.
  • Sulfamyl (sulfamoyl; sulfinic acid amide; sulfinamide): -S( 0)NR 1 R 2 , wherein R 1 and R 2 are independently amino substituents, as defined for amino groups.
  • Sulfonamido sulfinamoyl; sulfonic acid amide; sulfonamide
  • R 1 is an amino substituent, as defined for amino groups, and R is a sulfonamino substituent, for example, a Ci_ 7 alkyl group, a
  • C 3-20 heterocyclyl group or a C 5-20 aryl group, preferably a C 1-7 alkyl group.
  • Sulfinamino: -NR 1 S( 0)R, wherein R 1 is an amino substituent, as defined for amino groups, and R is a sulfinamino substituent, for example, a C 1-7 alkyl group, a C 3-20 heterocyclyl group, or a C 5 - 2 o aryl group, preferably a Ci -7 alkyl group.
  • phosphino groups include, but are not limited to, -PH 2 , -P(CH 3 ) 2 , -P(CH 2 CH 3 ) 2 , -P(t-Bu) 2 , and -P(Ph) 2 .
  • Phospho: -P( 0) 2 .
  • R is a phosphinyl substituent, for example, a Ci -7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-2 o aryl group, preferably a Ci -7 alkyl group or a C 5-2 o aryl group.
  • R is a phosphonate substituent, for example, -H, a Ci -7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-2 o aryl group, preferably -H, a Ci -7 alkyl group, or a C 5-2 o aryl group.
  • Phosphate (phosphonooxy ester): -OP( 0)(OR) 2 , where R is a phosphate substituent, for example, -H, a Ci_ 7 alkyl group, a C3-20 heterocyclyl group, or a C 5 -2o aryl group, preferably -H, a C1-7 alkyl group, or a C 5 -2o aryl group.
  • Phosphorous acid -OP(OH) 2 .
  • Phosphite -OP(OR) 2 , where R is a phosphite substituent, for example, -H, a C 1-7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-20 aryl group, preferably -H, a C 1-7 alkyl group, or a C 5-2 o aryl group.
  • R is a phosphite substituent, for example, -H, a C 1-7 alkyl group, a C 3-20 heterocyclyl group, or a C 5-20 aryl group, preferably -H, a C 1-7 alkyl group, or a C 5-2 o aryl group.
  • Examples of phosphite groups include, but are not limited to, -OP(OCH 3 ) 2 , -OP(OCH 2 CH 3 ) 2 , -OP(0-t-Bu) 2 , and -OP(OPh) 2 .
  • Phosphoramidite -OP(OR 1 )-NR 2 2 , where R 1 and R 2 are phosphoramidite substituents, for example, -H, a (optionally substituted) Ci -7 alkyl group, a C 3-20 heterocyclyl group, or a C 5 - 2 o aryl group, preferably -H, a Ci -7 alkyl group, or a C 5-2 o aryl group.
  • Examples of phosphoramidite groups include, but are not limited to, -OP(OCH 2 CH 3 )-N(CH 3 ) 2 ,
  • substituents for example, -H, a (optionally substituted) Ci -7 alkyl group, a C 3-20 heterocyclyl group, or a C 5 - 2 o aryl group, preferably -H, a Ci -7 alkyl group, or a C 5 - 2 o aryl group.
  • C 3-12 alkylene The term "C 3- i 2 alkylene", as used herein, pertains to a bidentate moiety obtained by removing two hydrogen atoms, either both from the same carbon atom, or one from each of two different carbon atoms, of a hydrocarbon compound having from 3 to 12 carbon atoms (unless otherwise specified), which may be aliphatic or alicyclic, and which may be saturated, partially unsaturated, or fully unsaturated.
  • alkylene includes the sub-classes alkenylene, alkynylene, cycloalkylene, etc., discussed below.
  • linear saturated C 3- i 2 alkylene groups include, but are not limited to, -(CH 2 ) n - where n is an integer from 3 to 12, for example, -CH 2 CH 2 CH 2 - (propylene), -CH2CH2CH2CH2- (butylene), -CH2CH2CH2CH2CH2- (pentylene) and -CH 2 CH 2 CH 2 CH- 2CH2CH2CH2- (heptylene).
  • Examples of branched saturated C 3 -i 2 alkylene groups include, but are not limited to, -CH(CH 3 )CH 2 -, -CH(CH 3 )CH 2 CH 2 -, -CH(CH3)CH 2 CH 2 CH2-, -CH 2 CH(CH 3 )CH 2 -,
  • C 3- i 2 cycloalkylenes examples include, but are not limited to, cyclopentylene (e.g. cyclopent-1 ,3-ylene), and cyclohexylene
  • C 3- i 2 cycloalkylenes examples include, but are not limited to, cyclopentenylene (e.g. 4-cyclopenten-1 ,3-ylene), and cyclohexenylene (e.g. 2-cyclohexen-1 ,4-ylene; 3-cyclohexen-1 ,2-ylene; 2,5-cyclohexadien- 1 ,4-ylene).
  • cyclopentenylene e.g. 4-cyclopenten-1 ,3-ylene
  • cyclohexenylene e.g. 2-cyclohexen-1 ,4-ylene; 3-cyclohexen-1 ,2-ylene; 2,5-cyclohexadien- 1 ,4-ylene.
  • a reference to carboxylic acid (-COOH) also includes the anionic (carboxylate) form (-COO " ), a salt or solvate thereof, as well as conventional protected forms.
  • a reference to an amino group includes the protonated form (-N + HR 1 R 2 ), a salt or solvate of the amino group, for example, a hydrochloride salt, as well as conventional protected forms of an amino group.
  • a reference to a hydroxyl group also includes the anionic form (-0 " ), a salt or solvate thereof, as well as conventional protected forms.
  • Certain compounds may exist in one or more particular geometric, optical, enantiomeric, diasteriomeric, epimeric, atropic, stereoisomeric, tautomeric, conformational, or anomeric forms, including but not limited to, cis- and trans-forms; E- and Z-forms; c-, t-, and r- forms; endo- and exo-forms; R-, S-, and meso-forms; D- and L-forms; d- and l-forms; (+) and (-) forms; keto-, enol-, and enolate-forms; syn- and anti-forms; synclinal- and anticlinal-forms; a- and ⁇ -forms; axial and equatorial forms; boat-, chair-, twist-, envelope-, and halfchair- forms; and combinations thereof, hereinafter collectively referred to as "isomers” (or "isomeric forms").
  • compounds of formula VI have the following stereochemistry at the C1 1 position:
  • isomers are structural (or constitutional) isomers (i.e. isomers which differ in the connections between atoms rather than merely by the position of atoms in space).
  • a reference to a methoxy group, -OCH 3 is not to be construed as a reference to its structural isomer, a hydroxymethyl group, -CH 2 OH.
  • a reference to ortho-chlorophenyl is not to be construed as a reference to its structural isomer, meta- chlorophenyl.
  • Ci -7 alkyl includes n-propyl and iso-propyl; butyl includes n-, iso-, sec-, and tert-butyl; methoxyphenyl includes ortho-, meta-, and para- methoxyphenyl).
  • keto/enol (illustrated below), imine/enamine, amide/imino alcohol, amidine/amidine, nitroso/oxime,
  • H may be in any isotopic form, including 1 H, 2 H (D), and 3 H (T); C may be in any isotopic form, including 12 C, 13 C, and 14 C; O may be in any isotopic form, including 16 0 and 18 0; and the like.
  • a reference to a particular compound includes all such isomeric forms, including (wholly or partially) racemic and other mixtures thereof.
  • Methods for the preparation (e.g. asymmetric synthesis) and separation (e.g. fractional crystallisation and chromatographic means) of such isomeric forms are either known in the art or are readily obtained by adapting the methods taught herein, or known methods, in a known manner.
  • a reference to a particular compound also includes ionic, salt, solvate, and protected forms of thereof, for example, as discussed below.
  • a corresponding salt of the active compound for example, a pharmaceutically-acceptable salt.
  • a pharmaceutically-acceptable salt examples are discussed in Berge, et al., J. Pharm. Sci., 66, 1 -19 (1977).
  • a salt may be formed with a suitable cation.
  • suitable inorganic cations include, but are not limited to, alkali metal ions such as Na + and K + , alkaline earth cations such as Ca 2+ and Mg 2+ , and other cations such as ⁇ 3 .
  • Suitable organic cations include, but are not limited to, ammonium ion (i.e. NH 4 + ) and substituted ammonium ions (e.g. NH 3 R + , NH 2 R2 + , NHR 3 + , NR 4 + ).
  • suitable substituted ammonium ions are those derived from: ethylamine,
  • ethanolamine diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids, such as lysine and arginine.
  • An example of a common quaternary ammonium ion is N(CH 3 ) 4 + . If the compound is cationic, or has a functional group which may be cationic (e.g. -NH 2 may be -NH 3 + ), then a salt may be formed with a suitable anion.
  • suitable inorganic anions include, but are not limited to, those derived from the following inorganic acids: hydrochloric, hydrobromic, hydroiodic, sulfuric, sulfurous, nitric, nitrous, phosphoric, and phosphorous.
  • Suitable organic anions include, but are not limited to, those derived from the following organic acids: 2-acetyoxybenzoic, acetic, ascorbic, aspartic, benzoic,
  • suitable polymeric organic anions include, but are not limited to, those derived from the following polymeric acids: tannic acid, carboxymethyl cellulose.
  • solvate is used herein in the conventional sense to refer to a complex of solute (e.g. active compound, salt of active compound) and solvent. If the solvent is water, the solvate may be conveniently referred to as a hydrate, for example, a mono-hydrate, a di-hydrate, a tri-hydrate, etc.
  • oxidation for example using: TCCA and TEMPO; BAIB and TEMPO; TPAP; Dess-Martin conditions; or Swern conditions.
  • R 7 may be OR A , where R A is selected from Ci -4 saturated alkyl, optionally substituted by phenyl, pyridyl and furanyl.
  • R A is unsubstituted C 1-4 saturated alkyl, i.e. methyl, ethyl, propyl and butyl. It may be preferred that R A is methyl.
  • R A is C 1-4 saturated alkyl substituted by a group selected from phenyl, pyridyl and furanyl. It may be preferred that the substituent is phenyl. It may be further preferred that R A is phenyl-methyl (i.e. benzyl).
  • R 8 is OProt°
  • Prot° is a silicon-based oxygen protecting group, such that R D is orthogonal to Prot°.
  • Preferred Prot° groups include TIPS and TBS.
  • R 9 is selected from H, methyl and methoxy. In some embodiments, R 9 is hydrogen. R s
  • R s is selected from CF 3 , CH 3 and
  • R s is CF 3
  • the C2 group is triflate.
  • R s is CH 3
  • the C2 group is mesylate.
  • R s is (CF 2 ) 3 CF 3
  • the C2 group is nonaflate.
  • R s is preferably CF 3 .
  • R c is selected from:
  • R C1 is a saturated alkyl group, i.e. methyl, ethyl, propyl and butyl.
  • R C1 is preferably methyl or ethyl, and more preferably methyl.
  • R C2 is methyl
  • R c comprises an acetate group.
  • R C2 is phenyl
  • R c comprises a benzoate group.
  • it is preferred R C2 is methyl.
  • preferred silyl protecting groups are TBDMS and TBDPS, or which TBDMS is more preferred.
  • R C3 may be a saturated C 1-4 alkyl group, i.e. methyl, ethyl, propyl and butyl. In some embodiments, R C3 is preferably methyl or ethyl, and more preferably methyl. R 04 may be a saturated Ci -4 alkyl group, i.e. methyl, ethyl, propyl and butyl. In
  • R C4 is preferably methyl or ethyl, and more preferably methyl.
  • R c is selected from:
  • Y is O. In other embodiments, Y is S.
  • R c is -Si-(R Si1 )(R Si2 )(R Si3 ).
  • the compound of formula II is treated with the appropriate anhydride and anhydrous 2,6-lutidine or anhydrous 2-6- f Bu-pyridine at a temperature of -35°C or lower in a dry organic solvent under a inert atmosphere. Therefore, if the compound of formula I has a triflate group at C2, then the compound of formula II is treated with triflic anhydride. If the compound of formula I has a mesylate group at C2, then the compound of formula II is treated with mesyl anhydride. If the compound of formula I has a tosylate group at C2, then the compound of formula II is treated with tosyl anhydride. If the compound of formula I has a nonaflate group at C2, then the compound of formula II is treated with nonaflurobutane sulphonic anhydride.
  • the base may be 2,6-lutidine or 2-6- f Bu-pyridine. It may be preferred that the base is 2-6- lutidine.
  • the reaction may be carried out at a temperature of -35°C or lower.
  • the reaction is carried out at a temperature of -40°C, -45°C, or -50°C. In further embodiments, the reaction is carried out at -78°C.
  • R 2 is selected from:
  • Ci -5 alkyl group including a C2-5 alkenyl group and a C2-5 alkynyl group
  • R 2 is an optionally substituted C 5-2 o aryl group. In some of these embodiments, R 2 is preferably an optionally substituted C 5-7 aryl group, and most preferably an optionally substituted phenyl group.
  • R 2 is a C9-12 aryl group, for example naphth-1 -yl or naphth- 2-yl, preferably naphty-2-yl
  • C9-12 aryl groups include quinolinyl, for example, quinolin-2-yl, quinolin-3-yl and quinolin-6-yl.
  • R 2 is a C 5-7 heteroaryl group, for example furanyl, thiophenyl and pyridyl.
  • thiophenyl is preferred, for example, thiophen-2-yl and thiophen-3-yl.
  • the C5-20 aryl group may bear any substituent group. It preferably bears from 1 to 3 substituent groups, with 1 and 2 being more preferred, and singly substituted groups being most preferred.
  • Preferred C 5-2 o aryl substituents, particularly for phenyl include: halo (e.g. F, CI, Br); C 1-7 alkoxy (e.g. methoxy, ethoxy); di-C 1-4 alkylamino-C 1-7 alkoxy; C 1-7 alkyl (e.g. methyl, trifluoromethyl, ethyl, propyl, t-butyl); bis-oxy-alkylene (e.g.
  • C5-20 aryl groups include, but are not limited to, 4-methyl- phenyl, 4-methoxy-phenyl, 3-methoxyphenyl, 4-fluoro-phenyl, 3,4-bisoxymethylene-phenyl, 4-triflouoromethylphenyl, 4-methylthiophenyl, 4-cyanophenyl, 4-phenoxyphenyl, 4-N-methyl piperazinyl phenyl, 4-N,N-dimethyl amino propxyphenyl, 4-N,N-dimethyl amino
  • C5-20 aryl groups include, but are not limited to thiophen-2-yl, napth-2-yl, quinolin-3-yl and quinolin-6-yl.
  • the aryl groups can be installed with the appropriate boronic acid (or pinacol ester) at 0°C using tetrakis palladium triphenylphosphine as catalyst and sodium carbonate as base. Hindered or electron deficient boronic acids may require heating to 60°C, if higher temperatures or prolonged heating is required triethylamine is preferred to sodium carbonate as base.
  • R 2 is an optionally substituted C 1-5 alkyl group (including a C 2-5 alkenyl group and a C 2-5 alkynyl group).
  • R 2 is an optionally substituted Ci -5 (saturated) alkyl group (e.g. methyl, ethyl, propyl, butyl, pentyl), which may be cyclic (e.g. cyclopropyl, cyclobutyl, cyclopentyl).
  • Preferred groups may include methyl, ethyl and propyl, as well as cyclopropyl.
  • R 2 is an optionally substituted C2-5 alkenyl group, which may be cyclic.
  • R 2 is an optionally substituted C 2-5 alkynyl group.
  • the group may be fully unsaturated or partially unsaturated, and therefore may include ethynyl (-C ⁇ CH), propynyl (e.g. -C ⁇ C-CH 3 , -CH 2 -C ⁇ CH), butynyl (e.g. -C ⁇ C-CH 2 -CH 3 , -CH 2 -C ⁇ C- CH 3 , -CH 2 -CH 2 -C ⁇ CH), pentynyl (e.g.
  • the optional substitutents for the optionally substituted Ci -5 alkyl groups may include, in particular:
  • the amido group is as defined above.
  • the ester group is as defined above.
  • the ether group is as defined above.
  • the ether substituent is a Ci -7 alkyl group or a alkyl group, such that the ether group is, for example, -OCH 3 , -OCH 2 CH 3 and -OC(CH 3 ) 3 .
  • R 2 is H.
  • R 2 groups of particular interest include:
  • a weak acid e.g. 5% formic acid in ethanol.
  • R c is -C(-YR C3 )(-YR C4 )
  • this reduction may be carried out using Cd/Pb couple, sodium dithionite or tin II chloride.
  • the conversion of III to IV is achieved by first reducing of the ester and reprotection as an acetate or silyl ether.
  • the reduction can be achieved by standard means, for example with LiBH 4 , or LiEt 3 BH 4 .
  • Reprotection as an aceate can be achieved, for example, by reaction with acetyl chloride; reprotection as a benzoate can be achieved, for example, by reaction with benzoyl chloride; reprotection as a silyl ether can be achieved, for example, by reaction with the appropriate silyl chloride in DMF in the presence of imidazole.
  • R 10 is a carbamate-based nitrogen protecting group, which is palladium-labile, or contains a moiety which is palladium-labile. If R 10 is a simple nitrogen protecting group, it may be preferably selected from a group containing the moiety:
  • R 10 is a complex nitrogen protecting group, it may be preferably a group which is cathepsin labile.
  • the nitrogen protecting group is preferably introduced by reaction of IV with triphosgene to obtain the isocyanate followed by reaction with R 10 -OH.
  • the reaction of IV with triphosgene is preferably introduced by reaction of IV with triphosgene to obtain the isocyanate followed by reaction with R 10 -OH.
  • triphosgene should be carried in an anhydrous and non-hydroxylic organic solvent, which is preferably non-polar.
  • Suitable solvents include anhydrous dichloromethane and anhydrous toluene. The reaction may be carried out at room temperature. The subsequent reaction with R 10 -OH should be carried in an anhydrous and non-hydroxylic organic solvent, which is preferably non-polar.
  • Suitable solvents include anhydrous dichloromethane and anhydrous toluene. The reaction should be carried out in the presence of a base is present, and suitable bases include pyridine or TEA. The reaction may be carried out at 0°C, or at a higher temperature to increase the rate of reaction.
  • the PBD B-ring is preferably closed using Dess-Martin periodinane.
  • R E is selected from: 0-d -2 alkyl; 0-Si-(R Si1 )(R Si2 )(R Si3 ), where R Si1 , R Si2 , R Si3 are as defined above; 0-CH 2 OCH 3 (OMOM), 0-CH 2 OC 2 H 4 OCH3 (OMEM), OC 5 H 10 (THP). Furthermore, Prot° needs to be orthogonal to R E .
  • R E is 0-Si-(R Si1 )(R Si2 )(R Si3 )
  • Si-(R Si1 )(R Si2 )(R Si3 ) is a silcon-based oxygen protecting group as described above. In these embodiments, R E may preferably be O-TBS.
  • Hal and Q are the same, such that the compound of formula X is symmetrical. It is preferred that Hal is preferably I. It is also preferred that Q is I. Thus, in some embodiments, it is preferred that both Hal and Q are I. In some embodiments about 3 equialents of formula X are used relative to the compound of formula VIII.
  • R D is TBS and Prot° is TIPS. In other embodiments, R D is OAc and Prot° is either TIPS or TBS.
  • Prot° is TIPS and R E is OTBS.
  • Prot° is TIPS and R E is OMe, OMEM, THP or OMOM.
  • Prot° is TBS
  • R E is OMe, OMEM, THP or OMOM.
  • R D is TBS
  • Prot° is TIPS
  • R E is TBS.
  • Kieselgel 60 F254 silica gel with fluorescent indicator on aluminium plates. Visualisation of TLC was achieved with UV light or iodine vapour unless otherwise stated. Flash chromatography was performed using Merck Kieselgel 60 F254 silica gel. Extraction and chromatography solvents were bought and used without further purification from Fisher Scientific, U.K. All chemicals were purchased from Aldrich, Lancaster or BDH.
  • the LC/MS conditions were as follows: Method 1 (default when not speficied)
  • the HPLC Waters Alliance 2695
  • A mobile phase of water
  • B acetonitrile
  • Gradient initial composition 5% B over 1 .0 min then 5% B to 95% B within 3 min.
  • the composition was held for 0.5 min at 95% B, and then returned to 5% B in 0.3 minutes.
  • Total gradient run time equals 5 min.
  • Flow rate 3.0 mL/min, 400 ⁇ _ was split via a zero dead volume tee piece which passes into the mass spectrometer.
  • Wavelength detection range 220 to 400 nm.
  • Function type diode array (535 scans).
  • Method 2 The HPLC (Waters Alliance 2695) was run using a mobile phase of water (A) (formic acid 0.1 %) and acetonitrile (B) (formic acid 0.1 %). Gradient: initial composition 5% B over 1.0 min then 5% B to 95% B over 2.5 minute period. The composition was held for 0.5 min at 95% B, and then returned to 5% B in 0.1 minutes and held there for 0.9 minutes. Total gradient run time equals 5 min. Flow rate 3.0 mL/min, 400 ⁇ _ was split via a zero dead volume tee piece which passes into the mass spectrometer. Wavelength detection range: 220 to 400 nm. Function type: diode array (535 scans). Column: Phenomenex ® Onyx Monolithic C18 50 x 4.60 mm.
  • Neat triisopropylsilylchloride (86 mL) was added to a mixture of imidazole (54.7 g) and phenol (4 g) (ground together). The mixture was heated until the phenol and imidazole melted and went into solution (1 10°C). The reaction mixture was allowed to stir for 5 minutes and was then allowed to cool, whereupon a solid was observed to form at the bottom of the flask. The reaction mixture was loaded directly on to silica gel and the silica pad was washed with hexane. The desired product was eluted with 5 % ethyl acetate in hexane. Excess eluent was removed by rotary evaporation under reduced pressure to afford a quantitative amount of the product 1 (1 12.454 g, >100 %). LCMS RT 4.25 mins, m/z ES + 353.85
  • Triflic anhydride (27.7 mL) was injected in one portion to a vigorously stirred suspension of compound 4 (31 .86 g) in dry dichloromethane (900 mL) in the presence of 2,6-lutidine (25.6 mL, dried over sieves) at -50°C (acetone/ dry ice bath).
  • the reaction mixture was allowed to stir for 1.5 hours when LC-MS following a mini work-up (water/dichloromethane) revealed the reaction to be complete.
  • Water was added to the still cold reaction mixture and the organic layer was separated and washed with saturated sodium bicarbonate, brine and magnesium sulphate.
  • the organic phase was filtered and excess solvent was removed by rotary evaporation under reduced pressure.
  • the residue was subjected to column flash chromatography (silica gel; 10 % ethyl acetate in hexane), removal of excess eluent afforded the product 5 (45.50 g, 96 % yield)
  • Tetrakis(triphenylphosphine)palladium(0) (1.14 g) was added to triflate 5 (23.40 g), E 2- propenylboronic acid (4.05 g) and potassium phosphate tribasic (13.93 g) in a mixture of dry dioxane (100 mL) under an argon atmosphere. The solution turned black and the reaction went rapidly to completion after 30 minutes as monitored by LC/MS. The reaction mixture was partitioned between ethyl acetate and water and the organic layer was washed with brine and dried over magnesium sulphate. The organic phase was filtered and excess solvent was removed by rotary evaporation under reduced pressure.
  • the crude 8 was dissolved in a 3/1/1 mixture of acetic acid/water/tetrahydrofuran (30/10/10 mL) and allowed to stir at room temperature. Deprotection was slower than expected so a mixture of methanol/acetic acid/water (30/30/30 mL) was added causing the reaction to proceed faster. After 6 hours the reaction was found to be 90 % complete, to avoid acidic degradation the reaction mixture was stored at -20°C in a spark free freezer overnight. The reaction was continued at room temperature the following day until all the starting material had been consumed. The reaction mixture was diluted with ethyl acetate and washed sequentially with water (2 x 500 mL), saturated aqueous sodium bicarbonate (200 mL) and brine.
  • Dimethyl sulphoxide (1.75 mL) was injected into an oven dried sealed microwave vial containing a solution of oxalyl chloride (1 mL) in dry dichloromethane (50 mL) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 15 minutes a solution of 9 (5.36 g) in dry dichloromethane (40 mL) was added slowly with the temperature still at -78°C. After 15 mins triethylamine (6.87 mL, dried over 4A molecular sieves) was injected and the dry ice/acetone bath was removed. The reaction mixture was allowed to reach room
  • Lithium acetate (0.47 g) was added to a solution of compound 11 (3.05 g) in wet dimethylformamide (30 ml_, 50; 1 DMF/water). After 1 and three-quarter hours the reaction was observed to be complete and the reaction mixture was diluted with ethyl acetate and washed with citric acid, water (0.6 ml.) and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash
  • Triethylamine (0.745 mL) was added to a stirred solution of the amine 7 (1.4 g) and trisphosgene 260 mg) in dry tetrahydrofuran (30 mL) at 5°C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquot from the reaction mixture quenching with methanol and performing LC/MS analysis. Once the isocyanate formation was complete a solution of the alloc-Val-Ala-PABOH (1 .38 g) and triethylamine (0.5 mL) in dry tetrahydrofuran (50 mL) was rapidly added by injection to the freshly prepare isocyanate. The reaction mixture was allowed to stir at 30°C overnight.
  • the TBS ether 14 was allowed to stir in a 3/1/1 mixture of acetic acid, tetrahydrofuran and water for 3 hours at room temperature at which time the reaction was complete.
  • the reaction mixture was diluted with ethyl acetate and sequentially washed with water, saturated aqueous sodium bicarbonate and brine.
  • the organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure.
  • the resulting residue was subjected to column flash chromatography (silica gel; 80 %ethyl acetate in hexane).
  • Dimethyl sulphoxide (288 ⁇ _) was added dropwise to an oven dried sealed microwave vial containing a solution of oxalyl chloride (1.65 ⁇ _) in dry dichloromethane (20 ml.) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 20 minutes a solution of 15 (1.4 g) in dry dichloromethane (15 ml.) was added slowly with the temperature still at - 78°C. After 15 mins triethylamine (1 .13 ml_), dried over 4A molecular sieves) was added dropwise and the dry ice/acetone bath was removed.
  • the reaction mixture was diluted with dichloromethane (50 ml.) washed with saturated ammonium chloride (2 x 50 ml_), and extracted with water (50 ml_), aqueous sodium bicarbonate (50 ml.) and brine (30 ml.) .
  • the organic phase was dried over magnesium sulphate, filtered and excess was removed by rotary evaporation under reduced pressure.
  • the resulting residue was subjected to column flash chromatography (silica gel; gradient, 40 % ethyl acetate in hexane to 60 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 17 (580mg, 64 % yield).
  • Triphenylarsine (1.71 g, 5.60 mmol, 0.4 eq) was added to a mixture of triflate 5 (10.00 g, 14 mmol, 1 eq), methylboronic acid (2.94 g, 49.1 mmol, 3.5 eq), silver oxide (13 g, 56 mmol, 4 eq) and potassium phosphate tribasic (17.8 g, 84 mmol, 6 eq) in dry dioxane (80 mL) under an argon atmosphere. The reaction was flushed with argon 3 times and
  • Zinc powder (28 g, 430 mmol, 37 eq) was added to a solution of compound 20 (6.7 g, 1 1 .58 mmol) in 5% formic acid in ethanol v/v (70 ml.) at around 15°C.
  • the resulting exotherm was controlled using an ice bath to maintain the temperature of the reaction mixture below 30°C.
  • the reaction mixture was filtered through a pad of celite.
  • the filtrate was diluted with ethyl acetate and the organic phase was washed with water, saturated aqueous sodium bicarbonate and brine.
  • the organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure.
  • the resulting residue was subjected to flash column
  • Lithium acetate (87 mg, 0.85 mmol) was added to a solution of compound 25 (540 mg, 0.85 mmol) in wet dimethylformamide (6 mL, 50:1 DMF/water). After 4 hours, the reaction was complete and the reaction mixture was diluted with ethyl acetate (25 mL) and washed with aqueous citric acid solution (pH ⁇ 3), water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 25% to 75% ethyl acetate in hexane).
  • Triethylamine (2.23 mL, 18.04 mmol, 2.2 eq) was added to a stirred solution of the amine 21 (4 g, 8.20 mmol) and triphosgene (778 mg, 2.95 mmol, 0.36 eq) in dry tetrahydrofuran (40 mL) at 5 °C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquots from the reaction mixture and quenching with methanol and performing LC/MS analysis.
  • the TBS ether 28 (1 .32 g, 1 .38 mmol) was dissolved in a 7:1 :1 :2 mixture of acetic acid/methanol/tetrahydrofuran/water (14:2:2:4 ml.) and allowed to stir at room temperature. After 3 hours no more starting material was observed by LC/MS. The reaction mixture was diluted with ethyl acetate (25 ml.) and washed sequentially with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate filtered and excess ethyl acetate removed by rotary evaporation under reduced pressure.
  • Dimethyl sulphoxide (0.2 ml_, 2.75 mmol, 2.5 eq) was added dropwise to a solution of oxalyl chloride (0.1 1 ml_, 1.32 mmol, 1.2 eq) in dry dichloromethane (7 ml.) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 10 minutes a solution of 29 (920 mg, 1.10 mmol) in dry dichloromethane (5 ml.) was added slowly with the temperature still at -78°C.
  • the reaction was flushed with argon 3 times and stirred at ambient temperature for 12 hours.
  • the reaction was diluted with ethyl acetate and the organic phase was washed with water and brine and dried over magnesium sulphate.
  • the solvent was removed by rotary evaporation under reduced pressure.
  • the resulting residue was subjected to column flash chromatography (silica gel; 10 % ethyl acetate / hexane). Pure fractions were collected and combined, and excess eluent was removed by rotary evaporation under reduced pressure afforded the product 34 (19 g, 61 %).
  • Lithium acetate (31 1 mg, 3.04 mmol) was added to a solution of compound 39 (2.16 g, 3.04 mmol) in wet dimethylformamide (20 mL, 50:1 DMF/water). After 2 hours, the reaction was complete and the reaction mixture was diluted with ethyl acetate (25 mL) and washed with aqueous citric acid solution (pH ⁇ 3), water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 25% to 75% ethyl acetate in hexane).
  • Triethylamine (4.34 mL, 31 .15 mmol, 2.2 eq) was added to a stirred solution of the amine 35 (8.91 g, 14.16 mmol) and triphosgene (1 .51 g, 5.10 mmol, 0.36 eq) in dry tetrahydrofuran (80 mL) at 5 °C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquots from the reaction mixture and quenching with methanol and performing LC/MS analysis.
  • the TBS ether 43 (8.78 g, 8.51 mmol) was dissolved in a 7:1 :2 mixture of acetic acid/tetrahydrofuran/water (56:8:16 mL) and allowed to stir at room temperature. After 16 hours no more starting material was observed by LC/MS.
  • the reaction mixture was diluted with ethyl acetate (250 mL) and washed sequentially with water, saturated aqueous sodium bicarbonate and brine.
  • the organic phase was dried over magnesium sulphate filtered and excess ethyl acetate removed by rotary evaporation under reduced pressure.
  • the resulting residue was subjected to flash column chromatography (silica gel, 2% methanol to 5% in chloroform).
  • Lithium acetate (1 10 mg, 1.08 mmol) was added to a solution of compound 46 (1.1 1 g, 1.08 mmol) in wet dimethylformamide (10 mL, 50:1 DMF/water). After 4 hours, the reaction was complete and the reaction mixture was diluted with ethyl acetate and washed with citric acid (pH ⁇ 3), water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient, 50/50 to 0/100 v/v hexane/ethyl acetate).
  • LCMS data were obtained using an Agilent 1200 series LC/MS with an Agilent 61 10 quadrupole MS, with Electrospray ionisation.
  • Mobile phase A - 0.1 % Acetic acid in water.
  • Mobile Phase B - 0.1 % in acetonitrile.
  • Flow rate 1 .00ml/min.
  • the total run time is 5 minutes.
  • Optical rotations were measured on an ADP 220 polarimeter (Bellingham Stanley Ltd.) and concentrations (c) are given in g/100mL. Melting points were measured using a digital melting point apparatus (Electrothermal). IR spectra were recorded on a Perkin-Elmer Spectrum 1000 FT IR Spectrometer. 1 H and 13 C NMR spectra were acquired at 300 K using a Bruker Avance NMR spectrometer at 400 and 100 MHz, respectively.
  • EEDQ (5.51 g, 22.29 mmol, 1.05 eq) was added to a solution of p-aminobenzyl alcohol (2.74 g, 22.29 mmol, 1.05 eq) and acid I4 (5.78 g, 21 .23 mmol, 1 eq) in dry THF (100 ml_). and stirred at room temperature for 72 hours. The reaction mixture was then concentrated under reduced pressure and the resulting brown solid was triturated with diethyl ether and filtered with subsequent washing with an excess of diethyl ether to afford the product as an off-white solid (7.1 g, 88 %).
  • Tetrakis(triphenylphosphine)palladium(0) (0.41 g, 0.35 mmol, 0.03 eq) was added to a mixture of the triflate 5 (8.4 g, 1 1.8 mmol, 1 .0 eq), E-1-propene-1 -ylboronic acid (1 .42 g, 16.5 mmol, 1 .4 eq) and potassium phosphate (5.0 g, 23.6 mmol, 2.0 eq) in dry dioxane (60 ml.) under a nitrogen atmosphere. The mixture was stirred at 25°C for 120 mins when LC/MS indicated complete reaction.
  • the reaction mixture was filtered through a short bed of celite®.
  • the celite® was washed with ethyl acetate (65 ml.) and the combined organics were washed successively with water (35 ml_), saturated sodium bicarbonate (35 ml.) and brine (10 ml_).
  • the organic phase was dried over MgS0 4 , filtered and concentrated under reduced pressure.
  • the residue was purified by flash chromatography (silica gel, 90/10 v/v n-hexane/EtOAc) to afford the product as a pale yellow oil (3.6 g, 69.0 %).
  • the crude intermediate 8 (assumed 100% yield, 4.12 g, 6.25 mmol, 1.0 eq) was dissolved in a mixture of acetic acid (70 mL), methanol (10 mL), THF (10 mL) and water (20 mL) and allowed to stir at room temperature. After 6 hours the reaction mixture was diluted with ethyl acetate (500 mL) and washed successively with water (2 x 500 mL), saturated sodium bicarbonate (300 mL) and brine (50 mL). The organic phase was dried over MgS0 4 , filtered and concentrated under reduced pressure.
  • the reaction mixture was diluted with ethyl acetate (200 mL) and washed successively with 0.1 M citric acid (50 mL, pH 3), water (50 mL) and brine (10 mL), dried over MgS0 4 , filtered and concentrated under reduced pressure.
  • the residue was purified by flash chromatography (silica gel, 50/50 v/v n-hexane/EtOAc to 25/75 v/v n- hexane/EtOAc) to afford the product as a pale yellow solid (0.68g, 45 %).
  • LC/MS (3.352 min (ES + )), m/z: 501.1 [M+H] + .
  • Triethylamine (0.256 mL, 1 .84 mmol, 2.2 eq) was added to a stirred solution of the amine 7 (0.480 g, 0.835 mmol, 1 .0 eq) and triphosgene (0.089 g, 0.301 mmol, 0.36 eq) in dry THF (15 mL) at 5°C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquots from the reaction mixture and quenching with methanol and performing LCMS analysis.
  • silyl ether 14 (1.49 g, 1 .52 mmol, 1 .0 eq) was dissolved in a 7:1 :1 :2 mixture of acetic acid/ methanol/ tetrahydrofuran/ water (14:2:2:4 mL) and allowed to stir at room
  • Dry dimethyl sulfoxide (0.180 g, 2.3 mmol, 3.0 eq) was added dropwise to a solution of oxalyl chloride (0.147 g, 1.1 mmol, 1 .5 eq) in DCM (10 mL) at -78°C under an atmosphere of nitrogen. Maintaining the temperature at -78°C, after 20 minutes, a solution of the primary alcohol 15 (0.666 g, 0.77 mmol, 1.0 eq) in DCM (10 ml.) was added dropwise. After a further 15 minutes, dry triethylamine (0.390 g, 3.85 mmol, 5.0 eq) was added, and the reaction mixture allowed to warm to room temperature.
  • Lithium acetate dihydrate (0.010 g, 0.10 mmol, 1.0 eq) was added to a solution of the silyl ether 37 (0.100 g, 0.10 mmol, 1 .0 eq) in wet DMF (2 mL) at 25°C for 3 hours.
  • the reaction mixture was then diluted with ethyl acetate (20 mL) and washed successively with 0.1 M citric acid (20 mL, pH 3), water (20 mL) and brine (5 mL), dried over MgS0 4 , filtered and concentrated under reduced pressure.

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  • Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)

Abstract

A compound of formula I wherein: R7 is selected from: ORA, where RA is selected from C1-4 saturated alkyl, optionally substituted by phenyl, which may bear a chloro substituent, pyridyl and furanyl; chloro; NH2; -CH2-O-C(=O)Me; R8 is OProt°, where Prot° is a silicon-based oxygen protecting group orthogonal to Rc; R9 is selected from H, methyl and methoxy; Rs is selected from CF3, (CF2)3CF3, CH3 and (formula 2) and Rc is selected from: (i) -C(=O)-ORC1, where RC1 is a saturated C1-4 alkyl group; (ii) -CH2-O-C(=0)RC2, where RC2 is methyl or phenyl; (iii) -CH2-O-Si-(RSi1)(RSi2)(RSi3), where RSi1, RSi2, RSi3 are independently selected from C1-6 a saturated alkyl group and a phenyl group; and (iv) -C(-YRC3)(-YRC4) where each Y is independently O or S, and where RC3 and RC4 are independently a saturated C1-4 alkyl group, or together form a C2-3 alkylene.

Description

SYNTHESIS METHOD AND INTERMEDIATES USEFUL IN THE PREPARATION OF PYRROLOBENZODIAZEPINES
The present invention relates to methods useful in the synthesis of pyrrolobenzodiazepin (PBDs), and in particular of pyrrolobenzodiazepines having C2 substitution and N10 protection.
Background to the invention
PBDs are of the general structure
Figure imgf000002_0001
C2-substituted compounds
In WO 2004/043963, a diverse range cytotoxic compounds having an aryl group at the C2 position, for example:
Figure imgf000002_0002
were disclosed. The synthesis of these compounds was achieved via the following intermediate:
Figure imgf000002_0003
whose synthesis was described in detail in WO 00/12508. An analagous dimeric intermediate is disclosed in WO 2010/010347.
WO 2005/085251 discloses the synthesis of a number cytoxic compounds having an aromatic substituent or unsaturated substituent at C2 from the intermediate:
Figure imgf000003_0001
These intermediates are very useful for synthesis C2 substituted PBD compounds having a N10-C1 1 imine bond, or compounds readily derivable from these, e.g. bisulphite versions, as in WO 2010/010347. The N10 protecting groups used are not labile under the conditions used to add the C2 aryl substituents, for example, in a Suzuki coupling using a palladium catalyst.
The C2-aryl or unsaturated group is installed by forming a C2-enol triflate followed by a palladium catalyzed Suzuki reaction. It is advantageous to add the C2 aryl or unsaturated substituents as late as possible in the synthesis by the methods described above, as this means a diverse range of substituents can be added to a single 'core'.
However, there are compounds of interest where the protecting groups or linkers desired at N10 are labile under either the triflation conditions (e.g. substituted p-aminobenzyl or p- hydroxybenzyl carbamates) or palladium catalysis (e.g. Alloc), see below.
Such groups are preferably introduced at a point in the synthesis of the PBD before the B- ring is formed by cyclisation. Otherwise, these groups would need to be added via a chloroformate to the PBD's imine bond. In addition, the cyclisation of PBD is best carried out where the pro-N 10 nitrogen has only a single hydrogen attached, i.e., where the protecting group is already in place.
WO 2005/085259 discloses PBD compounds useful as intermediates in the synthesis of PBD dimers. These intermediates have a hydroxy group at the C7 or C8 position.
Disclosure of the invention
The present inventors have developed a key intermediate for the production of C2 substituted PBDs having a palladium sensitive N10 protecting group, and a method for its synthesis. In a first aspect, the present invention comprises a compound of formula I:
Figure imgf000004_0001
wherein:
R7 is selected from: ORA, where RA is selected from C1-4 saturated alkyl, optionally substituted by phenyl, which phenyl may bear a chloro substituent, pyridyl and furanyl; chloro; NH2; and -CH2-0-C(=0)Me;
R8 is OProt°, where Prot° is a silicon-based oxygen protecting group orthogonal to Rc; R9 is selected from H, methyl and methoxy;
Rs is selected from CF3, (CF2)3CF3, CH3 and
Figure imgf000004_0002
;
and
Rc is selected from:
(i) -C(=0)-ORc1, where RC1 is a saturated Ci-4 alkyl group;
(ii) -CH2-0-C(=0)RC2, where RC2 is methyl or phenyl;
(iii) -CH2-0-Si-(RSi1)(RSi2)(RSi3), where RSi1, RSi2, RSi3 are independently selected from a d-6 saturated alkyl group and a phenyl group; and
(iv) -C(-YRC3)(-YRC4) where each Y is independently O or S, and where RC3 and RC4 are independently a saturated Ci-4 alkyl group, or together form a C2-3 alkylene.
The compound of formula I is preferably substantially free of any of the corresponding compound of formula IB:
Figure imgf000004_0003
wherein R7, R8, R9 and Rc are as defined in the compound of formula I. "Substantially free" means that there is less than 5% by weight of the compound of formula IB compared to the amount of the compounds of formula I. More preferably, there is less than 1 % or 0.1 % by weight of the compound of formula IB. In a second aspect, the present invention provides a method of synthesising a compound of formula I as defined in the first aspect of the invention from a compound of formula II:
wherein R7, R8, R9 and Rc are as defined in the first aspect;
comprising treating II with the appropriate anhydride and anhydrous 2,6-lutidine or anhydrous 2,6-fBu-pyridine at a temperature of -35°C or lower in a dry organic solvent under an inert atmosphere. The following reaction has been previously disclosed (Kang, G-D., et ai, Chem.
Commun., 2003, -1689):
Figure imgf000005_0002
B
This reaction was carried out using (trifluoromethanesulfonyl)aminopyridine at -78°C, which followed treatment with either LDA (lithium diethylamide) or NaHMDS (sodium
hexamethyldisilazide). The method involving LDA gave a mixture of A to B in the ratio 1 :4, whilst the NHDMS method gave only B.
Compounds of formula I can be converted into N10 protected PBD compounds by the following steps. A.
Figure imgf000006_0001
in
where R2 is selected from:
(i) an optionally substituted C5-20 aryl group;
(ii) an optionally substituted Ci-5 alkyl group (including a C2-5 alkenyl group and a C2-5 alkynyl group); and
(iii) H.
These conversions are carried using palladium catalysis, and include: Suzuki couplings with an appropriate boron derivative; Heck coupling with alkenes, including acrylamides and acrylates; Stille couplings with organo tin reagents, such as alkyl tin reagents;
Sonagishira couplings with alkynes; and hydride transfer using triethyl silanes.
The conversion where R2 is an optionally substituted C5-20 aryl group (a Suzuki coupling) carried out by palladium catalysed cross coupling of I with the appropriate aryl boron derivative. The palladium catalyst may be any suitable catalyst, for example Pd(PPh3)4, Pd(OCOCH3)2, PdCI2, or Pd(dba)3.
Suzuki couplings may also be used to cross couple alkyl boron species. The Gibbs group has reported (Mu, Y.Q and Gibbs, R.A, Tetrahedron Letters, 36, 5669-5672, 1995) the successful conversion of an enol triflate to a methyl substituted alkene (using a
Pd(PHCN)2)CI2 catalyst):
MeB(OH)3
Figure imgf000006_0002
Dioxane
Deng's group successfully achieved the coupling of a cyclopropylboronic acid and a cyclic triflate using a Pd (MeCN)2CI2 catalyst (Yao and Deng, J. Org. Chem. 2000, 65, 1609).
Figure imgf000007_0001
Ag20/THF
Cyclopropyl Ml DA boronates have been successfully coupled to aryl chlorides under slow release conditions; intact Ml DA boronates are resistant to standards Suzuki conditions due to their pyramidal hybridization (Knapp et al. J. Am. Chem. Soc, 2009, 131, 6961-6963). These conditions are also be applicable to enoltriflates.
Figure imgf000007_0002
Knapp et al. J. Am. Chem. Soc, 2009, 131, 6961 -6963
Molander (Molander and Ito Org. Lett., 2001 , 3, 393-396) has developed potassiu alkyltrifluoroborates as Suzuki coupling partners for aryl and vinyl triflates.
9%
PdCl2(dppf)2.CH2Cl2
Figure imgf000007_0003
Suzuki couplings may also be used to cross couple alkenyl boron species. Suzuki and co workers have demonstrated (Suzuki et al., Synlett 1990, 221 ; J Org Chem 1993, 58, 2201 ) the coupling of cyclic triflates and benzodioxoleborole compounds
Figure imgf000008_0001
Alkenyl Boronic acids have also been coupled to PBD C-ring enoltriflates
(WO2005/085251 ):
Figure imgf000008_0002
The pinacol esters of acrylylboronic acids are known:
Figure imgf000008_0003
and are likely to ebe useful in Suzuki coupling reactions.
PBD enol triflates have been successfully coupled to acrylamides and acrylates under Heck conditions (Chen et a\.,Bioorg. Med. Chem. Lett. 14, (2004), 1547-1549):
Figure imgf000009_0001
Alkynyl tin reagents have been coupled under Stille conditions to PBD triflates (Tiberghien et al., Bioorg. Med. Chem. Lett. 14, (2004), 5041 -504
Figure imgf000009_0002
Alkynyl substituents can also be installed via the Sonogishira reaction (Nishihara et al J. Org. Chem., 2000, 65, 1780-1787) :
Figure imgf000009_0003
Cul, DMF
80°C
Other palladium catalysed reactions include the use triakylindium reagents in cross- coupling reactions (Perez et al. Org. Lett, 1999, 1, 1267-1269) :
Figure imgf000009_0004
THF, Reflux
The approach can also be used to introduce aryl, vinyl and alkynyl groups. Palladium catalysis can also be used in hydride transfer reactions to generate alkenes with triethylsilanes (Cramer et al. Angew. Chem. Int. Ed. 2005, 44, 820-822):
Figure imgf000010_0001
ΠΙ IV
where RD is selected from:
(ii) -CH2-0-C(=0)RC2, where RC2 is methyl or phenyl;
(iii) -CH2-0-Si-(RSi1)(RSi2)(RSi3), where RSi1, RSi2, RSi3 are independently selected from a d-6 saturated alkyl group and a phenyl group; and
(iv) -C(-YRC3)(-YRC4) where each Y is independently O or S, and where RC3 and RC4 are independently a saturated Ci-4 alkyl group, or together form a C2-3 alkylene,
and RD is orthogonal to Prot°.
If Rc is -CH2-0-C(=0)RC2 or -CH2-0-Si-(RSi1)(RSi2)(RSi3) then the conversion of III to IV is by reduction of the nitro group. If Rc is -C(=0)-ORc1, then the conversion of III to IV is achieved by first reducing of the ester and reprotection as an acetate or silyl ether. The reduction can be achieved by standard means, for example with LiAIH4, LiBH4 or NaBH4, but preferably LiBH4.
Reprotection as an aceate can be achieved, for example, by reaction with acetyl chloride; reprotection as a benzoate can be achieved, for example, by reaction with benzoyl chloride; reprotection as a silyl ether can be achieved, for example, by reaction with the appropriate silyl chloride. The reduction of the nitro group is then carried out as described above. C:
Figure imgf000011_0001
IV V
where R10 is a carbamate-based nitrogen protecting group, which is palladium-labile and/or the conditions used to add the group:
Figure imgf000011_0002
, or contains a moiety which is palladium-labile and/or is labile under the conditions used to add the group:
Figure imgf000011_0003
, i.e. liable to be removed in the presence of palladium or under the conditions the group:
Figure imgf000011_0004
. In this application these protecting groups are classified as 'simple' and 'complex'.
Conversion of IV to V is usually achieved by reaction of IV with triphosgene to obtain the isocyanate followed by reaction with R10-OH. This approach is described in WO
2005/023814.
Alternatively, simple nitrogen protecting groups can also be introduced as a chloroformate, fluoroformate or azidoformate. The more complex nitrogen protecting groups, as well as the simple nitrogen protecting groups, can be introduced as O-succinamide carbonates, O- pentaflurophenyl carbonates and O-nitrophenyl carbonates. D:
Figure imgf000012_0001
V VI
When RD is -CH2-0-C(=0)Me, the conversion of V to VI may be achieved by initial removal of the acetate protecting group, with potassium carbonate in aqueous methanol, or with lithium triethylborohydride. Oxidation with Dess-Martin periodinane (or alternatively
TPAP/NMO, TFAA/DMSO, S03.Pyridine complex/DMSO, PDC, PCC, BAIB/TEMPO or under Swern conditions) affords the ring closed product.
When RD is -CH2-0-C(=0)Ph, the conversion of V to VI may be achieved by initial removal of the benzoate protecting group, with sodium hydroxide in aqueous methanol, followed by oxidation as described above.
When RD is -CH2-0-Si-(RSi1)(RSi2)(RSi3), the conversion of V to VI may be achieved by initial removal of the silyl ether protecting group, for example using TBAF in THF, acetic acid in aqueous THF, CsF in DMF or HF in pyridine, followed by oxidation as described above. The removal conditions must be selected such that they do not remove Prot°.
When RD is -C(-YRC3)(-YRC4), the conversion of V to VI may be achieved by removal of the acetal or thioacetal protecting groups, respectively with acid or reaction with Hg (II) salts - the cyclisation occurs spontaneously
E:
Figure imgf000012_0002
VI VII
wherin RE is selected from: 0-d-2 alkyl; 0-Si-(RSi1)(RSi2)(RSi3), where RSi1, RSi2, RSi3 are as defined above; 0-CH2OCH3 (OMOM), 0-CH2OC2H4OCH3 (OMEM), OC5H10 (THP). Prot° needs to be orthogonal to RE, and therefore RE needs to be selected with this in mind. In some embodiments, RE may also be OC(=0)Me.
OH is converted to RE by conventional methods. For example, when RE is 0-Ci-2 alkyl, the conversion may be carried out by rection with Ci-2 alkyl-l and silver oxide or (Ci-2
alkyl)30+BF4 " in DCM in the presence of DIPEA. When RE is OMOM or OMEM, the conversion may be carried out by reaction with the appropriate chloride in DCM in the presence of DIPEA. When RE is THP, the conversion may be carried out by reaction with DHP in the presence of acid. When RE is 0-Si-(RSi1)(RSi2)(RSi3), the conversion may be carried out by reaction with the appropriate triflate in the presence of 2,6-lutidine.
F:
Figure imgf000013_0001
VII VIII
Deprotection of the phenolic hydroxy group is carried out by, for example, lithium acetate. The removal conditions must be selected such that they do not remove RE.
G:
Figure imgf000013_0002
VIII IX
where Hal is selected from I, CI and Br, and n is from 1 to 12.
The conversion of VIII to IX is achieved by reaction with a compound of formula X: where Q is selected from I, CI and Br. The reaction is carried out, for example, in refluxing acetone with a base, such as K2C03. An excess of the compound of formula X is required to achieve the desired product. H: The present invention also provides a method of linking compounds of formulae VIII and IX, to yield a compound of formula XI:
Figure imgf000014_0001
where R2 , R7 , R9 , R10 and RE are independently selected from the same groups as R2, R7, R9, R10 and RE respectively. The reaction is carried out, for example, in refluxing acetone with a base, such as K2C03.
The present invention also provides:
(a) a compound of formula III:
Figure imgf000014_0002
1V preferably substantially free of
(c) a compound of formula V:
Figure imgf000015_0001
Figure imgf000015_0002
VI preferably substantially free of
(e) a compound of formula VII:
Figure imgf000015_0003
vn preferably substantially free of
Figure imgf000015_0004
preferably substantially free of a compound of formula IX:
Figure imgf000016_0001
preferably substantially free of
Figure imgf000016_0002
IXB
The term 'substantially free of means that there is less than 5% by weight of the compound of the undesired compound (i.e., B version) compared to the amount of the desired compound. More preferably, there is less than 1 % or 0.1 % by weight.
The present invention also provides
(a) a method of making a compound of formula VIII, comprising the step of making a compound of formula I from a compound of formula II;
(b) a method of making a compound of formula IX, comprising the step of making a compound of formula I from a compound of formula II;
(c) a method of making a compound of formula X, comprising the step of making a compound of formula I from a compound of formula II.
In the above further aspects of the present invention, the defined substituent groups in the intermediate compounds are the same as those in the final products where designated in the same way (e.g. R9 remains the same throughout the method of synthesis).
In the above further aspects of the invention, the methods may also comprise the intermediate steps as set out above.
Definitions
Silicon-based oxygen protecting groups
Silican-based oxygen protecting groups are well known in the art. A large number of suitable groups are described on pages 1 13 to 148 of Greene, T.W. and Wuts, G.M., Protective Groups in Organic Synthesis, 3 Edition, John Wiley & Sons, Inc., 1999, which is incorporated herein by reference.
The groups form silyl ethers with the oxygen to which they are attached, and are generally of the formula:
pS
RSi2
RSi3
where RS|1, RS|2 and RS|3 are independently selected from a Ci-6 saturated alkyl group and a phenyl group. Examples are:
Figure imgf000017_0001
Orthogonal
A first protecting group which is orthogonal to a second protecting group means that the first protecting group can be removed from a compound bearing both protecting groups without the second protecting group being removed. As discussed above, RD is orthogonal to Prot°, and Prot° is orthogonal to RE.
Carbamate-based nitrogen protecting groups
Carbamate-based nitrogen protecting groups are well known in the art. A large number of suitable groups are described on pages 503 to 549 of Greene, T.W. and Wuts, G.M., Protective Groups in Organic Synthesis, 3rd Edition, John Wiley & Sons, Inc., 1999, which is incorporated herein by reference.
The carbamate-based nitrogen protecting groups used may be those which are sensitive to palladium or sensitive to the conditions needed to add the group: 8 Table 1 provides examples of some such groups ('simple nitrogen protecting groups'):
Figure imgf000018_0001
Figure imgf000019_0001
Table 1
In some emodiments of the present invention, the carbamate nitrogen protecting group is not Cbz (benzyl carbamate), PNZ (p-nitrobenzylcarbamate), or 3,4-dimethoxy-6-nitrobenzyl carbamate.
Also suitable for use in the present invention are carbamate-based nitrogen protecting groups having a complex architecture ('complex nitrogen protecting groups'). These groups may be or may comprise elements of the groups described, for example, in WO 00/12507. These carbamate-based nitrogen protecting groups may include elements that to palladium or the conditions used to add the group:
Figure imgf000019_0002
These elements may be any one of the structures listed in Table 1 above.
In one embodiment, the carbamate-based nitrogen protecting group is a linker for connection to an antibody, RL:
Figure imgf000019_0003
where the asterisk indicates the point of attachment to the N10 position, G1 is a functional group to form a connection to a cell binding agent, L1 is a cleavable linker, L2 is a covalent bond or together with -OC(=0)- forms a self-immolative linker.
In one embodiment, the protecting group RL is removable from the N10 position thereby to provide the imine or carbinoalmine form of the PBD.
L1 is preferably a cleavable linker, and may be referred to as a trigger for activation of the linker for cleavage.
The nature of L1 and L2, where present, can vary widely. These groups are chosen on the basis of their cleavage characteristics, which may be dictated by the conditions at the site to which the PBD molecule is delivered, for example as an antibody drug conjugate. Those linkers that are cleaved by the action of enzymes are preferred, although linkers that are cleavable by changes in pH (e.g. acid or base labile), temperature or upon irradiation (e.g. photolabile) may also be used. Linkers that are cleavable under reducing or oxidising conditions may also find use in the present invention.
L1 may comprise a contiguous sequence of amino acids. The amino acid sequence may be the target substrate for enzymatic cleavage, thereby allowing release of R10 from the N 10 position.
In one embodiment, the L1 is cleavable by the action of an enzyme. In one embodiment, the enzyme is an esterase or a peptidase.
In one embodiment, L2 is present and together with C(=0)0 forms a self-immolative linker.
In one embodiment, where L1 is cleavable by the action of an enzyme and L2 is present, the enzyme cleaves the bond between L1 and L2.
L1 and L2 may be connected by a bond selected from:
-C(=0)NH-,
-C(=0)0-
-NHC(=0)-,
-OC(=0)-,
-OC(=0)0- -NHC(=0)0- -OC(=0)NH-, and
-NHC(=0)NH-.
An amino group of L1 that connects to L2 may be the N-terminus of an amino acid or may be derived from an amino group of an amino acid side chain, for example a lysine amino acid side chain.
An carboxyl group of L1 that connects to L2 may be the C-terminus of an amino acid or may be derived from a carboxyl group of an amino acid side chain, for example a glutamic acid amino acid side chain.
A hydroxy group of L1 that connects to L2 may be derived from a hydroxy group of an amino acid side chain, for example a serine amino acid side chain.
In one embodiment, -C(=0 - and L2 together form the group:
Figure imgf000021_0001
where the asterisk indicates the point of attachment to the N10 position, the wavy line indicates the point of attachment to the linker L1, Y is -N(H)-, -0-, -C(=0)N(H)- or -C(=0)0-, and n is 0 to 3. The phenylene ring is optionally substituted with one, two or three substituents as described herein. In one embodiment, the phenylene group is optionally substituted with halo, N02, R or OR.
In one embodiment, Y is NH.
In one embodiment, n is 0 or 1 . Preferably, n is 0.
Where Y is NH and n is 0, the self-immolative linker may be referred to as a
p-aminobenzylcarbonyl linker (PABC).
The self-immolative linker will allow for release of the protected compound when a remote site is activated, proceeding along the lines shown below (for n=0):
Figure imgf000022_0001
where L is the activated form of the remaining portion of the linker. These groups have the advantage of separating the site of activation from the compound being protected. As described above, the phenylene group may be optionally substituted.
In one embodiment described herein, the group L* is a linker L1 as described herein, which may include a dipeptide group.
In another embodiment, -C(=0)0- and L2 together form a group selected from:
Figure imgf000022_0002
where the asterisk, the wavy line, Y, and n are as defined above. Each phenylene ring is optionally substituted with one, two or three substituents as described herein. In one embodiment, the phenylene ring having the Y substituent is optionally substituted and the phenylene ring not having the Y substituent is unsubstituted. In one embodiment, the phenylene ring having the Y substituent is unsubstituted and the phenylene ring not having the Y substituent is optionally substituted.
In another embodiment, -C(=0)0- and L2 together form a group selected from:
Figure imgf000023_0001
where the asterisk, the wavy line, Y, and n are as defined above, E is O, S or NR, D is N, CH, or CR, and F is N, CH, or CR. In one embodiment, D is N.
In one embodiment, D is CH.
In one embodiment, E is O or S.
In one embodiment, F is CH. In a preferred embodiment, the linker is a cathepsin labile linker.
In one embodiment, L1 comprises a dipeptide The dipeptide may be represented as -NH-X X2-CO-, where -NH- and -CO- represent the N- and C-terminals of the amino acid groups Xi and X2 respectively. The amino acids in the dipeptide may be any combination of natural amino acids. Where the linker is a cathepsin labile linker, the dipeptide may be the site of action for cathepsin-mediated cleavage.
Additionally, for those amino acids groups having carboxyl or amino side chain functionality, for example Glu and Lys respectively, CO and NH may represent that side chain functionality.
In one embodiment, the group -X!-X2- in dipeptide, -NH-X X2-CO-, is selected from:
-Phe-Lys-,
-Val-Ala-,
-Val-Lys-,
-Ala-Lys-,
-Val-Cit-,
-Phe-Cit-,
-Leu-Cit-,
-lle-Cit-,
-Phe-Arg-,
-Trp-Cit- where Cit is citrulline.
Preferably, the group -X X2- in dipeptide,
Figure imgf000024_0001
is selected from:
-Phe-Lys-,
-Val-Ala-,
-Val-Lys-,
-Ala-Lys-,
-Val-Cit-. Most preferably, the group -X X2- in dipeptide,
Figure imgf000024_0002
is -Phe-Lys- or -Val-Ala-. In some embodiments, the group -X!-X2- in dipeptide,
Figure imgf000024_0003
is -Val-Cit-.
Other dipeptide combinations may be used, including those described by Dubowchik et al. {Bioconjugate Chem. 2002, 13, 855-869), which is incorporated herein by reference.
In one embodiment, the amino acid side chain is chemically protected, where appropriate. The side chain protecting group may be a group as discussed below in relation to the group RL. The present inventors have established that protected amino acid sequences are cleavable by enzymes. For example, the present inventors have established that a dipeptide sequence comprising a Boc side chain-protected Lys residue is cleavable by cathepsin.
Protecting groups for the side chains of amino acids are well known in the art and are described in the Novabiochem Catalog. Additional protecting group strategies are set out in Protective Groups in Organic Synthesis, Greene and Wuts.
Possible side chain protecting groups are shown below for those amino acids having reactive side chain functionality:
Arg: Z, Mtr, Tos;
Asn: Trt, Xan;
Asp: Bzl, t-Bu;
Cys: Acm, Bzl, Bzl-OMe, Bzl-Me, Trt;
Glu: Bzl, t-Bu;
Gin: Trt, Xan;
His: Boc, Dnp, Tos, Trt; Lys: Boc, Z-CI, Fmoc, Z, Alloc;
Sen Bzl, TBDMS, TBDPS;
Thr: Bz;
Trp: Boc;
Tyr: Bzl, Z, Z-Br.
In other embodiments of the invention, the amino acids selected are those having no reactive side chain functionality. For example, the amino acids may be selected from: Ala, Gly, lie, Leu, Met, Phe, Pro, and Val.
In one embodiment, the dipeptide is used in combination with a self-immolative linker. The self-immolative linker may be connected to -X2-.
In one embodiment, the dipeptide is used in combination with a self-immolative linker. The self-immolative linker may be connected to -X2-.
Where a self-immolative linker is present, -X2- is connected directly to the self-immolative linker. Preferably the group -X2-CO- is connected to Y, where Y is NH, thereby forming the group -X2-CO-NH-. -NH-Xr is connected directly to A. A may comprise the functionality -CO- thereby to form an amide link with -X .
In one embodiment, L1 and L2 together with -OC(=0)- comprise the group
NH-X X2-CO-PABC-. The PABC group is connected directly to the N10 position.
Preferably, the self-immolative linker and the dipeptide together form the group -NH-Phe- Lys-CO-NH-PABC-, which is illustrated below:
Figure imgf000025_0001
where the asterisk indicates the point of attachment to the N10 position, and the wavy line indicates the point of attachment to the remaining portion of the linker L1 or the point of attachment to A. Preferably, the wavy line indicates the point of attachment to A. The side chain of the Lys amino acid may be protected, for example, with Boc, Fmoc, or Alloc, as described above. Alternatively, the self-immolative linker and the dipeptide together form the group
-NH-Val-Ala-CO-NH-PABC-, which is illustrated below:
Figure imgf000026_0001
where the asterisk and the wavy line are as defined above. In one embodiment, L2 together with -OC(=0)- represents:
Figure imgf000026_0002
where the asterisk indicates the point of attachment to the N10 position, the wavy line indicates the point of attachment to L1, n is 0 to 3, Y is a covalent bond or a functional group, and E is an activatable group, for example by enzymatic action or light, thereby to generate a self-immolative unit. The phenylene ring is optionally further substituted with one, two or three substituents as described herein. In one embodiment, the phenylene group is optionally further substituted with halo, N02, R or OR. Preferably n is 0 or 1 , most preferably 0. E is selected such that the group is susceptible to activation, e.g. by light or by the action of an enzyme. E may be -N02 or glucoronic acid. The former may be susceptible to the action of a nitroreductase, the latter to the action of a β-glucoronidase.
In this embodiment, the self-immolative linker will allow for release of the protected compound when E is activated, proceeding along the lines shown below (for n=0):
Figure imgf000027_0001
where the asterisk indicates the point of attachment to the N10 position, E* is the activated form of E, and Y is as described above. These groups have the advantage of separating the site of activation from the compound being protected. As described above, the phenylene group may be optionally further substituted.
The group Y may be a covalent bond to L1.
The group Y may be a functional group selected from:
-C(=0)- -NH-
-O-
-C(=0)NH-,
-C(=0)0-,
-NHC(=0)-,
-OC(=0)-,
-OC(=0)0-,
-NHC(=0)0-,
-OC(=0)NH-,
-NHC(=0)NH-,
-NHC(=0)NH,
-C(=0)NHC(=0)-, and
-S-.
Where L1 is a dipeptide, it is preferred that Y is -NH- or -C(=0)-, thereby to form an amide bond between L1 and Y. In this embodiment, the dipeptide sequence need not be a substrate for an enzymatic activity.
In another embodiment, G is intended to form a spacer group that is suitable for indirectly connecting L1 to a cell binding agent. L1 and G1 may be connected by a bond selected from:
-C(=0)NH-,
-C(=0)0-,
-NHC(=0)-,
-OC(=0)-,
-OC(=0)0-,
-NHC(=0)0-,
-OC(=0)NH-, and
-NHC(=0)NH-.
In one embodiment, the functional group G1 is or comprises an amino, carboxylic acid, hydroxy, thiol, or maleimide group for reaction with an appropriate group on the cell binding agent. In a preferred embodiment, G1 comprises a maleimide group.
In one embodiment, the group G1 is an alkyl maleimide group. This group is suitable for reaction with thiol groups, particularly cysteine thiol groups, present in the cell binding agent, for example present in an antibody.
In one embodiment, the group G1 is:
Figure imgf000028_0001
where the asterisk indicates the point of attachment to L1 and n is 0 to 6. In one embodiment, n is 5.
In one embodiment, the group G1 is:
Figure imgf000028_0002
where the asterisk indicates the point of attachment to L1 and n is 0 to 6. In one embodiment, n is 5.
In one embodiment, the group G1 is:
Figure imgf000029_0001
where the asterisk indicates the point of attachment to L1 , n is 0 or 1 , and m is 0 to 30. In a preferred embodiment, n is 1 and m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
In one embodiment, the group G1 is:
Figure imgf000029_0002
where the asterisk indicates the point of attachment to L1 , n is 0 or 1 , and m is 0 to 30. In a preferred embodiment, n is 1 and m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
In each of the embodiments above, an alternative functionality may be used in place of the maleimide group shown below:
Figure imgf000029_0003
where asterisk indicates the bond to the remaining portion of the G group.
In one embodiment, the maleimide-derived group is replaced with the group:
Figure imgf000029_0004
where the asterisk indicates the bond to the remaining portion of the G group.
In one embodiment, the maleimide group is replaced with a group selected from:
-C(=0)OH,
-OH,
-NH2, -SH,
-C(=0)CH2X, where X is CI, Br or I,
-CHO,
-NHIMH2
-C≡CH, and
-IM3 (azide).
In particular embodiments, the maleimide group may be replaced by -C(=0)CH2l. Thus, in particular embodiments, the group G1 is:
Figure imgf000030_0001
where the asterisk indicates the point of attachment to L1 and n is 0 to 6. In one embodiment, n is 0.
In particular embodiments, the group G1 is:
Figure imgf000030_0002
where the asterisk indicates the point of attachment to L1 , m is 0 to 30. In a preferred embodiment, m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
In particular embodiments, the roup G1 is:
Figure imgf000030_0003
where the asterisk indicates the point of attachment to L1 , m is 0 to 30. In a preferred embodiment, m is 0 to 10, 1 to 7, preferably 3 to 7, and most preferably 3 or 7.
In one embodiment, where L1 is present, G1 is -NH2, -NHMe, -COOH, -OH or -SH.
In one embodiment, where L1 is present, G1 is -NH2 or -NHMe. Either group may be the N-terminal of an L1 amino acid sequence. In one embodiment, where L1 is present, G1 is -NH2, and L1 is an amino acid sequence -X X2-, as defined above in relation to R10. In one embodiment, where L1 is present, G1 is COOH. This group may be the C-terminal of an L1 amino acid sequence.
In one embodiment, where L1 is present, G1 is OH.
In one embodiment, where L1 is present, G1 is SH.
The group G1 may be convertable from one functional group to another. In one
embodiment, where L1 is present, G1 is -NH2. This group is convertable to another group G1 comprising a maleimide group. For example, the group -NH2 may be reacted with an acid or an activated acid (e.g. N-succinimide forms) of those G1 groups comprising maleimide shown above.
The group G1 may therefore be converted to a functional group that is more appropriate for reaction with a cell binding agent. In other embodiments, RL is a group that is a precursor to the linker that is provided with a functional group.
As noted above, in one embodiment, where L1 is present, G1 is -NH2, -NHMe, -COOH, -OH or -SH. In a further embodiment, these groups are provided in a chemically protected form. The chemically protected form is therefore a precursor to the linker that is provided with a functional group.
In one embodiment, G1 is -NH2 in a chemically protected form. The group may be protected with a carbamate protecting group. The carbamate protecting group may be selected from the group consisting of:
Alloc, Fmoc, Boc, Troc, Teoc, Cbz and PNZ.
Preferably, where G1 is -NH2, it is protected with an Alloc or Fmoc group. In one embodiment, particularly where the compound is a dimer, the nitrogen protecting group for one of the PBD units is a group RL and the nitrogen protecting group for the other of the PBD units is a capping group, Ca :
Figure imgf000032_0001
where the asterisk indicates the point of attachment to the N10 position, G2 is a terminating group, L3 is a covalent bond or a cleavable linker L1 , L2 is a covalent bond or together with OC(=0) forms a self-immolative linker.
The protecting group Rc is removable from the N10 position thereby to provide the imine or carbinolamine form of the PBD. The term "capping group" is used to indicate that Rc is prevented from reacting with a cell binding agent, such as an antibody. Instead, the group RL is for connection to a cell binding agent.
Where L3 and L2 are both covalent bonds, G2 and -OC(=0)- together form a carbamate protecting group such as shown above in Table 1 .
L1 is as defined above in relation to RL.
L2 is as defined above in relation to RL. In one embodiment L3 is a cleavable linker L1 , and L2, together with OC(=0), forms a self- immolative linker. In this embodiment, G2 is Ac (acetyl) or Moc, or a carbamate protecting group selected from:
Alloc, Fmoc, Boc, Troc, Teoc, Psec, Cbz and PNZ. In one embodiment L3 is a cleavable linker L1 , and L2, together with OC(=0), forms a self-immolative linker. In this embodiment, G2 together with OC(=0), is a carbamate protecting group selected from Table 1 above.
G2 is different to G1. Furthermore, it is preferred that G1 is removable under conditions that do not remove G1.
Substituents
The phrase "optionally substituted" as used herein, pertains to a parent group which may be unsubstituted or which may be substituted. Unless otherwise specified, the term "substituted" as used herein, pertains to a parent group which bears one or more substitutents. The term "substituent" is used herein in the conventional sense and refers to a chemical moiety which is covalently attached to, or if appropriate, fused to, a parent group. A wide variety of substituents are well known, and methods for their formation and introduction into a variety of parent groups are also well known.
Examples of substituents are described in more detail below.
C-1-12 alkyl: The term "C1-12 alkyl" as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a carbon atom of a hydrocarbon compound having from 1 to 12 carbon atoms, which may be aliphatic or alicyclic, and which may be saturated or unsaturated (e.g. partially unsaturated, fully unsaturated). Thus, the term "alkyl" includes the sub-classes alkenyl, alkynyl, cycloalkyl, etc., discussed below.
Examples of saturated alkyl groups include, but are not limited to, methyl (Ci ), ethyl (C2), propyl (C3), butyl (C4), pentyl (C5), hexyl (C6) and heptyl (C7). Examples of saturated linear alkyl groups include, but are not limited to, methyl (Ci), ethyl (C2), n-propyl (C3), n-butyl (C4), n-pentyl (amyl) (C5), n-hexyl (C6) and n-heptyl (C7).
Examples of saturated branched alkyl groups include iso-propyl (C3), iso-butyl (C4), sec-butyl (C4), tert-butyl (C4), iso-pentyl (C5), and neo-pentyl (C5).
C2-12 Alkenyl: The term "C2-12 alkenyl" as used herein, pertains to an alkyl group having one or more carbon-carbon double bonds.
Examples of unsaturated alkenyl groups include, but are not limited to, ethenyl (vinyl, -CH=CH2), 1-propenyl (-CH=CH-CH3), 2-propenyl (allyl, -CH-CH=CH2), isopropenyl (1-methylvinyl, -C(CH3)=CH2), butenyl (C4), pentenyl (C5), and hexenyl (C6).
C2-12 alkynyl: The term "C2-12 alkynyl" as used herein, pertains to an alkyl group having one or more carbon-carbon triple bonds. Examples of unsaturated alkynyl groups include, but are not limited to, ethynyl (ethinyl, -C≡CH) and 2-propynyl (propargyl, -CH2-C≡CH).
C3-12 cycloalkyl: The term "C3-12 cycloalkyl" as used herein, pertains to an alkyl group which is also a cyclyl group; that is, a monovalent moiety obtained by removing a hydrogen atom from an alicyclic ring atom of a cyclic hydrocarbon (carbocyclic) compound, which moiety has from 3 to 7 carbon atoms, including from 3 to 7 ring atoms.
Examples of cycloalkyl groups include, but are not limited to, those derived from:
saturated monocyclic hydrocarbon compounds:
cyclopropane (C3), cyclobutane (C4), cyclopentane (C5), cyclohexane (C6), cycloheptane (C7), methylcyclopropane (C4), dimethylcyclopropane (C5), methylcyclobutane (C5), dimethylcyclobutane (C6), methylcyclopentane (C6), dimethylcyclopentane (C7) and methylcyclohexane (C7);
unsaturated monocyclic hydrocarbon compounds:
cyclopropene (C3), cyclobutene (C4), cyclopentene (C5), cyclohexene (C6),
methylcyclopropene (C4), dimethylcyclopropene (C5), methylcyclobutene (C5),
dimethylcyclobutene (C6), methylcyclopentene (C6), dimethylcyclopentene (C7) and methylcyclohexene (C7); and
saturated polycyclic hydrocarbon compounds:
norcarane (C7), norpinane (C7), norbornane (C7).
C3-20 heterocyclyl: The term "C3-20 heterocyclyl" as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a ring atom of a heterocyclic compound, which moiety has from 3 to 20 ring atoms, of which from 1 to 10 are ring heteroatoms. Preferably, each ring has from 3 to 7 ring atoms, of which from 1 to 4 are ring heteroatoms.
In this context, the prefixes (e.g. C3-2o, C3-7, C5-6, etc.) denote the number of ring atoms, or range of number of ring atoms, whether carbon atoms or heteroatoms. For example, the term "C5-6heterocyclyl", as used herein, pertains to a heterocyclyl group having 5 or 6 ring atoms.
Examples of monocyclic heterocyclyl groups include, but are not limited to, those derived from: aziridine (C3), azetidine (C4), pyrrolidine (tetrahydropyrrole) (C5), pyrroline (e.g., 3-pyrroline, 2,5-dihydropyrrole) (C5), 2H-pyrrole or 3H-pyrrole (isopyrrole, isoazole) (C5), piperidine (C6), dihydropyridine (C6), tetrahydropyridine (C6), azepine (C7);
Oi: oxirane (C3), oxetane (C4), oxolane (tetrahydrofuran) (C5), oxole (dihydrofuran) (C5), oxane (tetrahyd ropy ran) (C6), dihydropyran (C6), pyran (C6), oxepin (C7);
Si: thiirane (C3), thietane (C4), thiolane (tetrahydrothiophene) (C5), thiane
(tetrahydrothiopyran) (C6), thiepane (C7);
02: dioxolane (C5), dioxane (C6), and dioxepane (C7);
03: trioxane (C6);
N2: imidazolidine (C5), pyrazolidine (diazolidine) (C5), imidazoline (C5), pyrazoline
(dihydropyrazole) (C5), piperazine (C6);
NiOi: tetrahydrooxazole (C5), dihydrooxazole (C5), tetrahydroisoxazole (C5),
dihydroisoxazole (C5), morpholine (C6), tetrahydrooxazine (C6), dihydrooxazine (C6), oxazine (C6);
Ν^ι: thiazoline (C5), thiazolidine (C5), thiomorpholine (C6);
N2Oi: oxadiazine (C6);
OiSi: oxathiole (C5) and oxathiane (thioxane) (C6); and,
NiOiSi: oxathiazine (C6). Examples of substituted monocyclic heterocyclyl groups include those derived from saccharides, in cyclic form, for example, furanoses (C5), such as arabinofuranose, lyxofuranose, ribofuranose, and xylofuranse, and pyranoses (C6), such as allopyranose, altropyranose, glucopyranose, mannopyranose, gulopyranose, idopyranose,
galactopyranose, and talopyranose.
C5-20 aryl: The term "C5-20 aryl", as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from an aromatic ring atom of an aromatic compound, which moiety has from 3 to 20 ring atoms. Preferably, each ring has from 5 to 7 ring atoms. In this context, the prefixes (e.g. C3-2o, C5-7, C5-6, etc.) denote the number of ring atoms, or range of number of ring atoms, whether carbon atoms or heteroatoms. For example, the term "C5-6 ary as used herein, pertains to an aryl group having 5 or 6 ring atoms.
The ring atoms may be all carbon atoms, as in "carboaryl groups". Examples of carboaryl groups include, but are not limited to, those derived from benzene (i.e. phenyl) (C6), naphthalene (Ci0), azulene (Ci0), anthracene (Ci4), phenanthrene (Ci4), naphthacene (Ci8), and pyrene (Ci6). Examples of aryl groups which comprise fused rings, at least one of which is an aromatic ring, include, but are not limited to, groups derived from indane (e.g. 2,3-dihydro-1 H- indene) (C9), indene (C9), isoindene (C9), tetraline (1 ,2,3,4-tetrahydronaphthalene (C10), acenaphthene (C12), fluorene (C13), phenalene (C13), acephenanthrene (C15), and aceanthrene (C16).
Alternatively, the ring atoms may include one or more heteroatoms, as in "heteroaryl groups". Examples of monocyclic heteroaryl groups include, but are not limited to, those derived from:
Ni: pyrrole (azole) (C5), pyridine (azine) (C6);
Oi: furan (oxole) (C5);
Si: thiophene (thiole) (C5);
NiOi: oxazole (C5), isoxazole (C5), isoxazine (C6);
N2Oi: oxadiazole (furazan) (C5);
N3Oi: oxatriazole (C5);
N1S1: thiazole (C5), isothiazole (C5);
N2: imidazole (1 ,3-diazole) (C5), pyrazole (1 ,2-diazole) (C5), pyridazine (1 ,2-diazine) (C6), pyrimidine (1 ,3-diazine) (C6) (e.g., cytosine, thymine, uracil), pyrazine (1 ,4-diazine) (C6);
N3: triazole (C5), triazine (C6); and,
N4: tetrazole (C5).
Examples of heteroaryl which comprise fused rings, include, but are not limited to:
C9 (with 2 fused rings) derived from benzofuran (O1), isobenzofuran (O1), indole (Ni), isoindole (N^, indolizine (N^, indoline (N^, isoindoline (N^, purine (N4) (e.g., adenine, guanine), benzimidazole (N2), indazole (N2), benzoxazole (Ν^Οι), benzisoxazole (Ν^Οι), benzodioxole (02), benzofurazan (N2Oi), benzotriazole (N3), benzothiofuran (Si), benzothiazole
Figure imgf000036_0001
benzothiadiazole (N2S);
Cio (with 2 fused rings) derived from chromene (O1), isochromene (O1), chroman (O1), isochroman (O1), benzodioxan (02), quinoline (Ni), isoquinoline (Ni), quinolizine (Ni), benzoxazine (NiOi), benzodiazine (N2), pyridopyridine (N2), quinoxaline (N2), quinazoline (N2), cinnoline (N2), phthalazine (N2), naphthyridine (N2), pteridine (N4); On (with 2 fused rings) derived from benzodiazepine (N2);
Ci3 (with 3 fused rings) derived from carbazole (Ni), dibenzofuran (Oi),
dibenzothiophene (Si), carboline (N2), perimidine (N2), pyridoindole (N2); and,
Ci4 (with 3 fused rings) derived from acridine (N-i), xanthene (Oi), thioxanthene (Si), oxanthrene (02), phenoxathiin (O1S1), phenazine (N2), phenoxazine (Ν^Οι), phenothiazine thianthrene (S2), phenanthridine (N-i), phenanthroline (N2), phenazine (N2).
The above groups, whether alone or part of another substituent, may themselves optionally be substituted with one or more groups selected from themselves and the additional substituents listed below.
N atoms in heterocyclyl and heteroaryl groups may be substituted by with CM2 alkyl, where appropriate. Halo: -F, -CI, -Br, and -I.
Hydroxy: -OH.
Ether: -OR, wherein R is an ether substituent, for example, a Ci-7 alkyl group (also referred to as a Ci-7 alkoxy group, discussed below), a C3-2o heterocyclyl group (also referred to as a C3-2o heterocyclyloxy group), or a C5-2o aryl group (also referred to as a C5-2o aryloxy group), preferably a Ci-7alkyl group.
Alkoxy: -OR, wherein R is an alkyl group, for example, a Ci-7 alkyl group. Examples of C1-7 alkoxy groups include, but are not limited to, -OMe (methoxy), -OEt (ethoxy), -O(nPr) (n-propoxy), -O(iPr) (isopropoxy), -O(nBu) (n-butoxy), -O(sBu) (sec-butoxy), -O(iBu) (isobutoxy), and -O(tBu) (tert-butoxy).
Acetal: -CH(OR1)(OR2), wherein R1 and R2 are independently acetal substituents, for example, a Ci-7 alkyl group, a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably a
Ci-7 alkyl group, or, in the case of a "cyclic" acetal group, R1 and R2, taken together with the two oxygen atoms to which they are attached, and the carbon atoms to which they are attached, form a heterocyclic ring having from 4 to 8 ring atoms. Examples of acetal groups include, but are not limited to, -CH(OMe)2, -CH(OEt)2, and -CH(OMe)(OEt). Hemiacetal: -CH(OH)(OR1), wherein R1 is a hemiacetal substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-20 aryl group, preferably a Ci-7 alkyl group. Examples of hemiacetal groups include, but are not limited to, -CH(OH)(OMe) and -CH(OH)(OEt).
Ketal: -CR(OR1)(OR2), where R1 and R2 are as defined for acetals, and R is a ketal substituent other than hydrogen, for example, a C1-7 alkyl group, a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably a C1-7 alkyl group. Examples ketal groups include, but are not limited to, -C(Me)(OMe)2, -C(Me)(OEt)2, -C(Me)(OMe)(OEt), -C(Et)(OMe)2,
-C(Et)(OEt)2, and -C(Et)(OMe)(OEt).
Hemiketal: -CR(OH)(OR1), where R1 is as defined for hemiacetals, and R is a hemiketal substituent other than hydrogen, for example, a Ci-7 alkyl group, a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group. Examples of hemiacetal groups include, but are not limited to, -C(Me)(OH)(OMe), -C(Et)(OH)(OMe), -C(Me)(OH)(OEt), and -C(Et)(OH)(OEt).
Oxo (keto, -one): =0. Thione (thioketone): =S.
Imino (imine): =NR, wherein R is an imino substituent, for example, hydrogen, Ci-7 alkyl group, a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably hydrogen or a Ci-7 alkyl group. Examples of ester groups include, but are not limited to, =N H, =NMe, =NEt, and =NPh.
Formyl (carbaldehyde, carboxaldehyde): -C(=0)H.
Acyl (keto): -C(=0)R, wherein R is an acyl substituent, for example, a C1-7 alkyl group (also referred to as Ci-7 alkylacyl or Ci-7 alkanoyl), a C3-2o heterocyclyl group (also referred to as C3-20 heterocyclylacyl), or a C5-2o aryl group (also referred to as C5-2o arylacyl), preferably a Ci-7 alkyl group. Examples of acyl groups include, but are not limited to, -C(=0)CH3 (acetyl), -C(=0)CH2CH3 (propionyl), -C(=0)C(CH3)3 (t-butyryl), and -C(=0)Ph (benzoyl, phenone). Carboxy (carboxylic acid): -C(=0)OH. Thiocarboxy (thiocarboxylic acid): -C(=S)SH. Thiolocarboxy (thiolocarboxylic acid): -C(=0)SH. Thionocarboxy (thionocarboxylic acid): -C(=S)OH. Imidic acid: -C(=NH)OH.
Hydroxamic acid: -C(=NOH)OH.
Ester (carboxylate, carboxylic acid ester, oxycarbonyl): -C(=0)OR, wherein R is an ester substituent, for example, a Ci_7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group. Examples of ester groups include, but are not limited to, -C(=0)OCH3, -C(=0)OCH2CH3, -C(=0)OC(CH3)3, and -C(=0)OPh.
Acyloxy (reverse ester): -OC(=0)R, wherein R is an acyloxy substituent, for example, a Ci-7 alkyl group, a C3-2o heterocyclyl group, or a C5-20 aryl group, preferably a Ci-7 alkyl group. Examples of acyloxy groups include, but are not limited to, -OC(=0)CH3 (acetoxy), -OC(=0)CH2CH3, -OC(=0)C(CH3)3, -OC(=0)Ph, and -OC(=0)CH2Ph.
Oxycarboyloxy: -OC(=0)OR, wherein R is an ester substituent, for example, a Ci-7 alkyl group, a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group.
Examples of ester groups include, but are not limited to, -OC(=0)OCH3,
-OC(=0)OCH2CH3, -OC(=0)OC(CH3)3, and -OC(=0)OPh.
Amino: -NR1 R2, wherein R1 and R2 are independently amino substituents, for example, hydrogen, a C1 -7 alkyl group (also referred to as C1-7 alkylamino or di-C1 -7 alkylamino), a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably H or a Ci-7 alkyl group, or, in the case of a "cyclic" amino group, R1 and R2, taken together with the nitrogen atom to which they are attached, form a heterocyclic ring having from 4 to 8 ring atoms. Amino groups may be primary (-NH2), secondary (-NHR1), or tertiary (-NHR1 R2), and in cationic form, may be quaternary (-+NR1 R2R3). Examples of amino groups include, but are not limited to, -NH2, -NHCH3, -NHC(CH3)2, -N(CH3)2, -N(CH2CH3)2, and -NHPh. Examples of cyclic amino groups include, but are not limited to, aziridino, azetidino, pyrrolidino, piperidino, piperazino, morpholino, and thiomorpholino.
Amido (carbamoyl, carbamyl, aminocarbonyl, carboxamide): -C(=0)NR1R2, wherein R1 and R2 are independently amino substituents, as defined for amino groups. Examples of amido groups include, but are not limited to, -C(=0)NH2, -C(=0)NHCH3, -C(=0)N(CH3)2,
-C(=0)NHCH2CH3, and -C(=0)N(CH2CH3)2, as well as amido groups in which R1 and R2, together with the nitrogen atom to which they are attached, form a heterocyclic structure as in, for example, piperidinocarbonyl, morpholinocarbonyl, thiomorpholinocarbonyl, and piperazinocarbonyl.
Thioamido (thiocarbamyl): -C(=S)NR1R2, wherein R1 and R2 are independently amino substituents, as defined for amino groups. Examples of amido groups include, but are not limited to, -C(=S)NH2, -C(=S)NHCH3, -C(=S)N(CH3)2, and -C(=S)NHCH2CH3.
Acylamido (acylamino): -NR1C(=0)R2, wherein R1 is an amide substituent, for example, hydrogen, a Ci_7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably hydrogen or a Ci-7 alkyl group, and R2 is an acyl substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2oaryl group, preferably hydrogen or a Ci-7 alkyl group. Examples of acylamide groups include, but are not limited to, -NHC(=0)CH3 ,
-NHC(=0)CH2CH3, and -NHC(=0)Ph. R1 and R2 may together form a cyclic structure, as in for example, succinimidyl, maleimidyl, and phthalimidyl:
Figure imgf000040_0001
succinimidyl maleimidyl phthalimidyl
Aminocarbonyloxy: -OC(=0)NR1 R2, wherein R1 and R2 are independently amino
substituents, as defined for amino groups. Examples of aminocarbonyloxy groups include, but are not limited to, -OC(=0)NH2, -OC(=0)NHMe, -OC(=0)NMe2, and -OC(=0)NEt2.
Ureido: -N(R1)CONR2R3 wherein R2 and R3 are independently amino substituents, as defined for amino groups, and R1 is a ureido substituent, for example, hydrogen, a C1-7 alkyl group, a C3-20 heterocyclyl group, or a C5-20 aryl group, preferably hydrogen or a C1-7 alkyl group. Examples of ureido groups include, but are not limited to, -NHCONH2,
-NHCONHMe, -NHCONHEt, -NHCONMe2, -NHCONEt2, -NMeCONH2, -NMeCONHMe, -NMeCONHEt, -NMeCONMe2, and -NMeCONEt2. Guanidino: -NH-C(=NH)NH2.
Tetrazolyl: a five membered aromatic ring having four nitrogen atoms and one carbon atom,
Figure imgf000041_0001
Imino: =NR, wherein R is an imino substituent, for example, for example, hydrogen, a C1-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably H or a C1-7alkyl group. Examples of imino groups include, but are not limited to, =NH, =NMe, and =NEt. Amidine (amidino): -C(=NR)NR2, wherein each R is an amidine substituent, for example, hydrogen, a Ci-7 alkyl group, a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably H or a Ci_7 alkyl group. Examples of amidine groups include, but are not limited to,
-C(=NH)NH2, -C(=NH)NMe2, and -C(=NMe)NMe2. Nitro: -N02.
Nitroso: -NO.
Azido: -N3.
Cyano (nitrile, carbonitrile): -CN. Isocyano: -NC. Cyanato: -OCN.
Isocyanato: -NCO. Thiocyano (thiocyanato): -SCN.
Isothiocyano (isothiocyanato): -NCS. Sulfhydryl (thiol, mercapto): -SH.
Thioether (sulfide): -SR, wherein R is a thioether substituent, for example, a C -7 alkyl group (also referred to as a C1-7alkylthio group), a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably a C1 -7 alkyl group. Examples of C1-7 alkylthio groups include, but are not limited to, -SCH3 and -SCH2CH3.
Disulfide: -SS-R, wherein R is a disulfide substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group (also referred to herein as Ci-7 alkyl disulfide). Examples of Ci-7 alkyl disulfide groups include, but are not limited to, -SSCH3 and -SSCH2CH3.
Sulfine (sulfinyl, sulfoxide): -S(=0)R, wherein R is a sulfine substituent, for example, a Ci-7 alkyl group, a C3-2o heterocyclyl group, or a C5-20 aryl group, preferably a Ci-7 alkyl group. Examples of sulfine groups include, but are not limited to, -S(=0)CH3 and
-S(=0)CH2CH3.
Sulfone (sulfonyl): -S(=0)2R, wherein R is a sulfone substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group, including, for example, a fluorinated or perfluorinated Ci-7 alkyl group. Examples of sulfone groups include, but are not limited to, -S(=0)2CH3 (methanesulfonyl, mesyl), -S(=0)2CF3 (triflyl), -S(=0)2CH2CH3 (esyl), -S(=0)2C4F9 (nonaflyl), -S(=0)2CH2CF3 (tresyl),
-S(=0)2CH2CH2NH2 (tauryl), -S(=0)2Ph (phenylsulfonyl, besyl), 4-methylphenylsulfonyl (tosyl), 4-chlorophenylsulfonyl (closyl), 4-bromophenylsulfonyl (brosyl), 4-nitrophenyl (nosyl), 2-naphthalenesulfonate (napsyl), and 5-dimethylamino-naphthalen-1 -ylsulfonate (dansyl).
Sulfinic acid (sulfino): -S(=0)OH, -S02H. Sulfonic acid (sulfo): -S(=0)2OH, -S03H. Sulfinate (sulfinic acid ester): -S(=0)OR; wherein R is a sulfinate substituent, for example, a C1-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci_7 alkyl group. Examples of sulfinate groups include, but are not limited to, -S(=0)OCH3
(methoxysulfinyl; methyl sulfinate) and -S(=0)OCH2CH3 (ethoxysulfinyl; ethyl sulfinate).
Sulfonate (sulfonic acid ester): -S(=0)2OR, wherein R is a sulfonate substituent, for example, a C1-7 alkyl group, a C3-2o heterocyclyl group, or a C5-2o aryl group, preferably a C1-7 alkyl group. Examples of sulfonate groups include, but are not limited to, -S(=0)2OCH3 (methoxysulfonyl; methyl sulfonate) and -S(=0)20CH2CH3 (ethoxysulfonyl; ethyl sulfonate).
Sulfinyloxy: -OS(=0)R, wherein R is a sulfinyloxy substituent, for example, a C1-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group.
Examples of sulfinyloxy groups include, but are not limited to, -OS(=0)CH3 and
-OS(=0)CH2CH3.
Sulfonyloxy: -OS(=0)2R, wherein R is a sulfonyloxy substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group.
Examples of sulfonyloxy groups include, but are not limited to, -OS(=0)2CH3 (mesylate) and -OS(=0)2CH2CH3 (esylate).
Sulfate: -OS(=0)2OR; wherein R is a sulfate substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group. Examples of sulfate groups include, but are not limited to, -OS(=0)2OCH3 and -SO(=0)2OCH2CH3. Sulfamyl (sulfamoyl; sulfinic acid amide; sulfinamide): -S(=0)NR1R2, wherein R1 and R2 are independently amino substituents, as defined for amino groups. Examples of sulfamyl groups include, but are not limited to, -S(=0)NH2, -S(=0)NH(CH3), -S(=0)N(CH3)2, -S(=0)NH(CH2CH3), -S(=0)N(CH2CH3)2, and -S(=0)NHPh. Sulfonamido (sulfinamoyl; sulfonic acid amide; sulfonamide): -S(=0)2NR1R2, wherein R1 and R2 are independently amino substituents, as defined for amino groups. Examples of sulfonamido groups include, but are not limited to, -S(=0)2NH2, -S(=0)2NH(CH3),
-S(=0)2N(CH3)2, -S(=0)2NH(CH2CH3), -S(=0)2N(CH2CH3)2, and -S(=0)2NHPh. Sulfamino: -NR1S(=0)2OH, wherein R1 is an amino substituent, as defined for amino groups. Examples of sulfamino groups include, but are not limited to, -NHS(=0)2OH and -N(CH3)S(=0)2OH. Sulfonamino: -NR1S(=0)2R, wherein R1 is an amino substituent, as defined for amino groups, and R is a sulfonamino substituent, for example, a Ci_7 alkyl group, a
C3-20 heterocyclyl group, or a C5-20 aryl group, preferably a C1-7 alkyl group. Examples of sulfonamino groups include, but are not limited to, -NHS(=0)2CH3 and -N(CH3)S(=0)2C6H5. Sulfinamino: -NR1S(=0)R, wherein R1 is an amino substituent, as defined for amino groups, and R is a sulfinamino substituent, for example, a C1-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group. Examples of sulfinamino groups include, but are not limited to, -NHS(=0)CH3 and -N(CH3)S(=0)C6H5. Phosphino (phosphine): -PR2, wherein R is a phosphino substituent, for example, -H, a Ci_7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably -H, a Ci-7 alkyl group, or a C5-2o aryl group. Examples of phosphino groups include, but are not limited to, -PH2, -P(CH3)2, -P(CH2CH3)2, -P(t-Bu)2, and -P(Ph)2. Phospho: -P(=0)2.
Phosphinyl (phosphine oxide): -P(=0)R2, wherein R is a phosphinyl substituent, for example, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably a Ci-7 alkyl group or a C5-2o aryl group. Examples of phosphinyl groups include, but are not limited to, -P(=0)(CH3)2, -P(=0)(CH2CH3)2, -P(=0)(t-Bu)2, and -P(=0)(Ph)2.
Phosphonic acid (phosphono): -P(=0)(OH)2.
Phosphonate (phosphono ester): -P(=0)(OR)2, where R is a phosphonate substituent, for example, -H, a Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably -H, a Ci-7 alkyl group, or a C5-2o aryl group. Examples of phosphonate groups include, but are not limited to, -P(=0)(OCH3)2, -P(=0)(OCH2CH3)2, -P(=0)(0-t-Bu)2, and -P(=0)(OPh)2.
Phosphoric acid (phosphonooxy): -OP(=0)(OH)2. Phosphate (phosphonooxy ester): -OP(=0)(OR)2, where R is a phosphate substituent, for example, -H, a Ci_7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably -H, a C1-7 alkyl group, or a C5-2o aryl group. Examples of phosphate groups include, but are not limited to, -OP(=0)(OCH3)2, -OP(=0)(OCH2CH3)2, -OP(=0)(0-t-Bu)2, and
-OP(=0)(OPh)2.
Phosphorous acid: -OP(OH)2.
Phosphite: -OP(OR)2, where R is a phosphite substituent, for example, -H, a C1-7 alkyl group, a C3-20 heterocyclyl group, or a C5-20 aryl group, preferably -H, a C1-7 alkyl group, or a C5-2o aryl group. Examples of phosphite groups include, but are not limited to, -OP(OCH3)2, -OP(OCH2CH3)2, -OP(0-t-Bu)2, and -OP(OPh)2.
Phosphoramidite: -OP(OR1)-NR2 2, where R1 and R2 are phosphoramidite substituents, for example, -H, a (optionally substituted) Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably -H, a Ci-7 alkyl group, or a C5-2o aryl group. Examples of phosphoramidite groups include, but are not limited to, -OP(OCH2CH3)-N(CH3)2,
-OP(OCH2CH3)-N(i-Pr)2, and -OP(OCH2CH2CN)-N(i-Pr)2. Phosphoramidate: -OP(=0)(OR1)-NR2 2, where R1 and R2 are phosphoramidate
substituents, for example, -H, a (optionally substituted) Ci-7 alkyl group, a C3-20 heterocyclyl group, or a C5-2o aryl group, preferably -H, a Ci-7 alkyl group, or a C5-2o aryl group.
Examples of phosphoramidate groups include, but are not limited to, -OP(=0)(OCH2CH3)- N(CH3)2, -OP(=0)(OCH2CH3)-N(i-Pr)2, and -OP(=0)(OCH2CH2CN)-N(i-Pr)2.
Alkylene
C3-12 alkylene: The term "C3-i2 alkylene", as used herein, pertains to a bidentate moiety obtained by removing two hydrogen atoms, either both from the same carbon atom, or one from each of two different carbon atoms, of a hydrocarbon compound having from 3 to 12 carbon atoms (unless otherwise specified), which may be aliphatic or alicyclic, and which may be saturated, partially unsaturated, or fully unsaturated. Thus, the term "alkylene" includes the sub-classes alkenylene, alkynylene, cycloalkylene, etc., discussed below.
Examples of linear saturated C3-i2 alkylene groups include, but are not limited to, -(CH2)n- where n is an integer from 3 to 12, for example, -CH2CH2CH2- (propylene), -CH2CH2CH2CH2- (butylene), -CH2CH2CH2CH2CH2- (pentylene) and -CH2CH2CH2CH- 2CH2CH2CH2- (heptylene).
Examples of branched saturated C3-i2 alkylene groups include, but are not limited to, -CH(CH3)CH2-, -CH(CH3)CH2CH2-, -CH(CH3)CH2CH2CH2-, -CH2CH(CH3)CH2-,
-CH2CH(CH3)CH2CH2-, -CH(CH2CH3)-, -CH(CH2CH3)CH2-, and -CH2CH(CH2CH3)CH2-.
Examples of linear partially unsaturated C3-12 alkylene groups (C3-12 alkenylene, and alkynylene groups) include, but are not limited to, -CH=CH-CH2-, -CH2-CH=CH2-,
-CH=CH-CH2-CH2-, -CH=CH-CH2-CH2-CH2-, -CH=CH-CH=CH-, -CH=CH-CH=CH-CH2-, -CH=CH-CH=CH-CH2-CH2-, -CH=CH-CH2-CH=CH-, -CH=CH-CH2-CH2-CH=CH-, and
Figure imgf000046_0001
Examples of branched partially unsaturated C3-i2 alkylene groups (C3-i2 alkenylene and alkynylene groups) include, but are not limited to, -C(CH3)=CH-, -C(CH3)=CH-CH2-, -CH=CH-CH(CH3)- and -C≡C-CH(CH3)-.
Examples of alicyclic saturated C3-i2 alkylene groups (C3-i2 cycloalkylenes) include, but are not limited to, cyclopentylene (e.g. cyclopent-1 ,3-ylene), and cyclohexylene
(e.g. cyclohex-1 ,4-ylene).
Examples of alicyclic partially unsaturated C3-i2 alkylene groups (C3-i2 cycloalkylenes) include, but are not limited to, cyclopentenylene (e.g. 4-cyclopenten-1 ,3-ylene), and cyclohexenylene (e.g. 2-cyclohexen-1 ,4-ylene; 3-cyclohexen-1 ,2-ylene; 2,5-cyclohexadien- 1 ,4-ylene).
Includes Other Forms
Unless otherwise specified, included in the above are the well known ionic, salt, solvate, and protected forms of these substituents. For example, a reference to carboxylic acid (-COOH) also includes the anionic (carboxylate) form (-COO"), a salt or solvate thereof, as well as conventional protected forms. Similarly, a reference to an amino group includes the protonated form (-N+HR1 R2), a salt or solvate of the amino group, for example, a hydrochloride salt, as well as conventional protected forms of an amino group. Similarly, a reference to a hydroxyl group also includes the anionic form (-0"), a salt or solvate thereof, as well as conventional protected forms. Isomers, Salts and Solvates
Certain compounds may exist in one or more particular geometric, optical, enantiomeric, diasteriomeric, epimeric, atropic, stereoisomeric, tautomeric, conformational, or anomeric forms, including but not limited to, cis- and trans-forms; E- and Z-forms; c-, t-, and r- forms; endo- and exo-forms; R-, S-, and meso-forms; D- and L-forms; d- and l-forms; (+) and (-) forms; keto-, enol-, and enolate-forms; syn- and anti-forms; synclinal- and anticlinal-forms; a- and β-forms; axial and equatorial forms; boat-, chair-, twist-, envelope-, and halfchair- forms; and combinations thereof, hereinafter collectively referred to as "isomers" (or "isomeric forms").
Preferably compounds of formula VI have the following stereochemistry at the C1 1 position:
Figure imgf000047_0001
Note that, except as discussed below for tautomeric forms, specifically excluded from the term "isomers", as used herein, are structural (or constitutional) isomers (i.e. isomers which differ in the connections between atoms rather than merely by the position of atoms in space). For example, a reference to a methoxy group, -OCH3, is not to be construed as a reference to its structural isomer, a hydroxymethyl group, -CH2OH. Similarly, a reference to ortho-chlorophenyl is not to be construed as a reference to its structural isomer, meta- chlorophenyl. However, a reference to a class of structures may well include structurally isomeric forms falling within that class (e.g. Ci-7 alkyl includes n-propyl and iso-propyl; butyl includes n-, iso-, sec-, and tert-butyl; methoxyphenyl includes ortho-, meta-, and para- methoxyphenyl).
The above exclusion does not pertain to tautomeric forms, for example, keto-, enol-, and enolate-forms, as in, for example, the following tautomeric pairs: keto/enol (illustrated below), imine/enamine, amide/imino alcohol, amidine/amidine, nitroso/oxime,
thioketone/enethiol, N-nitroso/hyroxyazo, and nitro/aci-nitro.
Figure imgf000048_0001
keto enol enolate
Note that specifically included in the term "isomer" are compounds with one or more isotopic substitutions. For example, H may be in any isotopic form, including 1H, 2H (D), and 3H (T); C may be in any isotopic form, including 12C, 13C, and 14C; O may be in any isotopic form, including 160 and 180; and the like.
Unless otherwise specified, a reference to a particular compound includes all such isomeric forms, including (wholly or partially) racemic and other mixtures thereof. Methods for the preparation (e.g. asymmetric synthesis) and separation (e.g. fractional crystallisation and chromatographic means) of such isomeric forms are either known in the art or are readily obtained by adapting the methods taught herein, or known methods, in a known manner.
Unless otherwise specified, a reference to a particular compound also includes ionic, salt, solvate, and protected forms of thereof, for example, as discussed below.
It may be convenient or desirable to prepare, purify, and/or handle a corresponding salt of the active compound, for example, a pharmaceutically-acceptable salt. Examples of pharmaceutically acceptable salts are discussed in Berge, et al., J. Pharm. Sci., 66, 1 -19 (1977).
For example, if the compound is anionic, or has a functional group which may be anionic (e.g. -COOH may be -COO"), then a salt may be formed with a suitable cation. Examples of suitable inorganic cations include, but are not limited to, alkali metal ions such as Na+ and K+, alkaline earth cations such as Ca2+ and Mg2+, and other cations such as Α 3.
Examples of suitable organic cations include, but are not limited to, ammonium ion (i.e. NH4 +) and substituted ammonium ions (e.g. NH3R+, NH2R2+, NHR3 +, NR4 +). Examples of some suitable substituted ammonium ions are those derived from: ethylamine,
diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine,
ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids, such as lysine and arginine. An example of a common quaternary ammonium ion is N(CH3)4 +. If the compound is cationic, or has a functional group which may be cationic (e.g. -NH2 may be -NH3 +), then a salt may be formed with a suitable anion. Examples of suitable inorganic anions include, but are not limited to, those derived from the following inorganic acids: hydrochloric, hydrobromic, hydroiodic, sulfuric, sulfurous, nitric, nitrous, phosphoric, and phosphorous.
Examples of suitable organic anions include, but are not limited to, those derived from the following organic acids: 2-acetyoxybenzoic, acetic, ascorbic, aspartic, benzoic,
camphorsulfonic, cinnamic, citric, edetic, ethanedisulfonic, ethanesulfonic, fumaric, glucheptonic, gluconic, glutamic, glycolic, hydroxymaleic, hydroxynaphthalene carboxylic, isethionic, lactic, lactobionic, lauric, maleic, malic, methanesulfonic, mucic, oleic, oxalic, palmitic, pamoic, pantothenic, phenylacetic, phenylsulfonic, propionic, pyruvic, salicylic, stearic, succinic, sulfanilic, tartaric, toluenesulfonic, and valeric. Examples of suitable polymeric organic anions include, but are not limited to, those derived from the following polymeric acids: tannic acid, carboxymethyl cellulose.
It may be convenient or desirable to prepare, purify, and/or handle a corresponding solvate of the active compound. The term "solvate" is used herein in the conventional sense to refer to a complex of solute (e.g. active compound, salt of active compound) and solvent. If the solvent is water, the solvate may be conveniently referred to as a hydrate, for example, a mono-hydrate, a di-hydrate, a tri-hydrate, etc.
General synthetic routes
Figure imgf000050_0001
by oxidation for example using: TCCA and TEMPO; BAIB and TEMPO; TPAP; Dess-Martin conditions; or Swern conditions.
Compounds of formula A may be synthesised by coupling appropriate compounds of formulae B and C, or activated derivatives thereof:
Figure imgf000050_0002
Compound of formulae B and C are generally commercially available or readily
synthesisable.
If compound B is a dimer, then this may be synthesised as described in WO 00/12508. Further Embodiments
The following further embodiments may be combined with each other as appropriate.
R7 is selected from: ORA, where RA is selected from Ci-4 saturated alkyl, optionally substituted by phenyl, which phenyl may bear a chloro substituent, pyridyl and furanyl; chloro; NH2; and -CH2-0-C(=0)Me; or R7 and R8 together form a group -0-(CH2)m-0-, where m is 1 or 2.
In some embodiments, R7 may be ORA, where RA is selected from Ci-4 saturated alkyl, optionally substituted by phenyl, pyridyl and furanyl.
In some of these embodiments, RA is unsubstituted C1-4 saturated alkyl, i.e. methyl, ethyl, propyl and butyl. It may be preferred that RA is methyl.
In other of these embodiments, RA is C1-4 saturated alkyl substituted by a group selected from phenyl, pyridyl and furanyl. It may be preferred that the substituent is phenyl. It may be further preferred that RA is phenyl-methyl (i.e. benzyl).
R8
R8 is OProt°, and Prot° is a silicon-based oxygen protecting group, such that RD is orthogonal to Prot°. Preferred Prot° groups include TIPS and TBS.
R!
R9 is selected from H, methyl and methoxy. In some embodiments, R9 is hydrogen. Rs
Rs is selected from CF3, CH3 and
Figure imgf000051_0001
When Rs is CF3, the C2 group is triflate. When Rs is CH3, the C2 group is mesylate.
When Rs
Figure imgf000051_0002
the C2 group is tosylate.
When Rs is (CF2)3CF3, the C2 group is nonaflate. In some embodiments, Rs is preferably CF3.
Rc is selected from:
(i) -C(=0)-ORc1 , where RC1 is a saturated Ci-4 alkyl group;
(ii) -CH2-0-C(=0) RC2, where RC2 is methyl or phenyl;
(iii) -CH2-0-Si-(RSi1)(RSi2)(RSi3), where RSi1 , RSi2, RSi3 are independently selected from C1-6 a saturated alkyl group and a phenyl group; and
(iv) -C(-YRC3) (-YRC4) where each Y is independently O or S, and where RC3 and R04 are independently a saturated C1-4 alkyl group, or together form a C2-3 alkylene.
RC1 is a saturated
Figure imgf000052_0001
alkyl group, i.e. methyl, ethyl, propyl and butyl. In some
embodiments, RC1 is preferably methyl or ethyl, and more preferably methyl. When RC2 is methyl, Rc comprises an acetate group. When RC2 is phenyl, Rc comprises a benzoate group. In some embodiments, it is preferred RC2 is methyl.
-Si-(RSl1)(RSl2)(RSl3) is a silyl protecting group, which are well known in the art. Groups of particular interest in the present invention are:
Figure imgf000052_0002
In some embodiments, preferred silyl protecting groups are TBDMS and TBDPS, or which TBDMS is more preferred.
RC3 may be a saturated C1-4 alkyl group, i.e. methyl, ethyl, propyl and butyl. In some embodiments, RC3 is preferably methyl or ethyl, and more preferably methyl. R04 may be a saturated Ci-4 alkyl group, i.e. methyl, ethyl, propyl and butyl. In
embodiments, RC4 is preferably methyl or ethyl, and more preferably methyl.
When RC3 and RC4 together form a C2- alkylene group, Rc is selected from:
Figure imgf000053_0001
*
In some embodiments, Y is O. In other embodiments, Y is S.
In some embodiments, it is preferred that Rc is -Si-(RSi1)(RSi2)(RSi3). Second aspect
In the method of the second aspect, the compound of formula II is treated with the appropriate anhydride and anhydrous 2,6-lutidine or anhydrous 2-6-fBu-pyridine at a temperature of -35°C or lower in a dry organic solvent under a inert atmosphere. Therefore, if the compound of formula I has a triflate group at C2, then the compound of formula II is treated with triflic anhydride. If the compound of formula I has a mesylate group at C2, then the compound of formula II is treated with mesyl anhydride. If the compound of formula I has a tosylate group at C2, then the compound of formula II is treated with tosyl anhydride. If the compound of formula I has a nonaflate group at C2, then the compound of formula II is treated with nonaflurobutane sulphonic anhydride.
The base may be 2,6-lutidine or 2-6-fBu-pyridine. It may be preferred that the base is 2-6- lutidine.
The reaction may be carried out at a temperature of -35°C or lower. In some
embodiments, the reaction is carried out at a temperature of -40°C, -45°C, or -50°C. In further embodiments, the reaction is carried out at -78°C. A
R2 is selected from:
(i) an optionally substituted C5-20 aryl group;
(ii) an optionally substituted Ci-5 alkyl group (including a C2-5 alkenyl group and a C2-5 alkynyl group); and
(iii) H.
In some embodiments, R2 is an optionally substituted C5-2o aryl group. In some of these embodiments, R2 is preferably an optionally substituted C5-7 aryl group, and most preferably an optionally substituted phenyl group.
In other of these embodiments, R2 is a C9-12 aryl group, for example naphth-1 -yl or naphth- 2-yl, preferably naphty-2-yl Further examples of C9-12 aryl groups include quinolinyl, for example, quinolin-2-yl, quinolin-3-yl and quinolin-6-yl.
In other of these embodiments, R2 is a C5-7 heteroaryl group, for example furanyl, thiophenyl and pyridyl. Of these thiophenyl is preferred, for example, thiophen-2-yl and thiophen-3-yl.
The C5-20 aryl group may bear any substituent group. It preferably bears from 1 to 3 substituent groups, with 1 and 2 being more preferred, and singly substituted groups being most preferred. Preferred C5-2o aryl substituents, particularly for phenyl, include: halo (e.g. F, CI, Br); C1-7 alkoxy (e.g. methoxy, ethoxy); di-C1-4 alkylamino-C1-7 alkoxy; C1-7 alkyl (e.g. methyl, trifluoromethyl, ethyl, propyl, t-butyl); bis-oxy-alkylene (e.g. bis-oxy-methylene, -0-CH2-0-); and N-C1-4 alkyl piperazinyl. Particularly preferred substituted C5-20 aryl groups include, but are not limited to, 4-methyl- phenyl, 4-methoxy-phenyl, 3-methoxyphenyl, 4-fluoro-phenyl, 3,4-bisoxymethylene-phenyl, 4-triflouoromethylphenyl, 4-methylthiophenyl, 4-cyanophenyl, 4-phenoxyphenyl, 4-N-methyl piperazinyl phenyl, 4-N,N-dimethyl amino propxyphenyl, 4-N,N-dimethyl amino
ethoxyphenyl, 6-methoxynaphth-2-yl, and 6-ethoxynaphth-2-yl. Particularly preferred unsubstituted C5-20 aryl groups include, but are not limited to thiophen-2-yl, napth-2-yl, quinolin-3-yl and quinolin-6-yl.
The aryl groups can be installed with the appropriate boronic acid (or pinacol ester) at 0°C using tetrakis palladium triphenylphosphine as catalyst and sodium carbonate as base. Hindered or electron deficient boronic acids may require heating to 60°C, if higher temperatures or prolonged heating is required triethylamine is preferred to sodium carbonate as base. In some embodiments, R2 is an optionally substituted C1-5 alkyl group (including a C2-5 alkenyl group and a C2-5 alkynyl group).
In some of these embodiments, R2 is an optionally substituted Ci-5 (saturated) alkyl group (e.g. methyl, ethyl, propyl, butyl, pentyl), which may be cyclic (e.g. cyclopropyl, cyclobutyl, cyclopentyl). Preferred groups may include methyl, ethyl and propyl, as well as cyclopropyl.
In others of these embodiments, R2 is an optionally substituted C2-5 alkenyl group, which may be cyclic. The group may be fully unsaturated or partially unsaturated, and therefore may include ethenyl (-CH=CH2), propenyl (e.g. -CH=CH-CH3, -CH2-CH=CH2, - C(=CH2)-CH3), butenyl (e.g. -CH=CH-CH2-CH3, -CH2-CH=CH-CH3, -CH2-CH2-CH=CH2, - C(=CH2)-CH2-CH3), butadienyl (e.g. -CH=CH-CH=CH2, -C(=CH2)-CH=CH2), pentenyl (e.g. - CH=CH-CH2-CH2-CH3, -CH2-CH=CH-CH2-CH3, -CH2-CH2-CH=CH-CH3, -CH2-CH2-CH2- CH=CH2, -C(=CH2)-CH2-CH2-CH3), pentadienyl (e.g. -CH=CH-CH=CH-CH3, -CH2-CH=CH- CH=CH2, -C(=CH2)-CH=CH-CH3) and cyclopentenyl.
In others of these embodiments, R2 is an optionally substituted C2-5 alkynyl group. The group may be fully unsaturated or partially unsaturated, and therefore may include ethynyl (-C≡CH), propynyl (e.g. -C≡C-CH3, -CH2-C≡CH), butynyl (e.g. -C≡C-CH2-CH3, -CH2-C≡C- CH3, -CH2-CH2-C≡CH), pentynyl (e.g. -C≡C-CH2-CH2-CH3, -CH2-C≡C-CH2-CH3, -CH2-CH2- C≡C-CH3, -CH2-CH2-CH2-C≡CH), and cyclopentynyl.
For groups above that contain unsaturation, at least one of the unsaturated bonds may be conjugated to the double bond present between C2 and C3 in the PBD's C-ring. It may be the case that all the unsaturated bonds are conjugated. The optional substitutents for the optionally substituted Ci-5 alkyl groups (including C2-5 alkenyl group and C2-5 alkynyl groups), may include, in particular:
(i) C5-20 aryl groups, which themselves may be substituted as discussed above for R2 groups in themselves;
(ii) cyano (-CN);
(iiii) amido (-C(=0)-NR1 R2);
(iv) ester (-C(=0)-OR).
(v) -OR;
(vi) boronic groups suitable for coupling a further group, such as:
Figure imgf000056_0001
(vii) tin groups, suitable for couplinga further group, such as -Sn(R)3 where R is a C1-7 alkyl group.
The amido group is as defined above. In some embodiments, the amino substituents may preferably be selected from Ci-7 alkyl or more preferably Ci-4 alkyl, such that the amido group is, for example, -C(=0)NH2, -C(=0)NHCH3, -C(=0)N(CH3)2, -C(=0)NHCH2CH3, and -C(=0)N(CH2CH3)2
The ester group is as defined above. In some embodiments, the ester substituent is a Ci-7 alkyl group or a
Figure imgf000056_0002
alkyl group, such that the ester group is, for example, -C(=0)OCH3, -C(=0)OCH2CH3 and -C(=0)OC(CH3)3.
The ether group is as defined above. In some embodiments, the ether substituent is a Ci-7 alkyl group or a
Figure imgf000056_0003
alkyl group, such that the ether group is, for example, -OCH3, -OCH2CH3 and -OC(CH3)3.
In some embodiments, R2 is H.
R2 groups of particular interest include:
Figure imgf000057_0001
B
If Rc is -CH2-0-C(=0)RC2 or -CH2-0-Si-(RSi1)(RSi2)(RSi3) then the conversion of III to IV is by reduction of the nitro group. If Rc is -CH2-0-C(=0)RC2, this reduction is carried out by using zinc in acetic acid. Alternatives include Cd/Pb couple, sodium dithionite or tin II chloride. If Rc is -CH2-0-Si-(RSi1)(RSi2)(RSi3), then this reduction is carried out using zinc in a weak acid, e.g. 5% formic acid in ethanol. If Rc is -C(-YRC3)(-YRC4), this reduction may be carried out using Cd/Pb couple, sodium dithionite or tin II chloride. If Rc is -C(=0)-ORc1 , then the conversion of III to IV is achieved by first reducing of the ester and reprotection as an acetate or silyl ether. The reduction can be achieved by standard means, for example with LiBH4, or LiEt3BH4. Reprotection as an aceate can be achieved, for example, by reaction with acetyl chloride; reprotection as a benzoate can be achieved, for example, by reaction with benzoyl chloride; reprotection as a silyl ether can be achieved, for example, by reaction with the appropriate silyl chloride in DMF in the presence of imidazole.
C
R10 is a carbamate-based nitrogen protecting group, which is palladium-labile, or contains a moiety which is palladium-labile. If R10 is a simple nitrogen protecting group, it may be preferably selected from a group containing the moiety:
.A, , such as Alloc, Ipaoc, Coc, Noc, Paloc, and may more preferably be Alloc. If R10 is a complex nitrogen protecting group, it may be preferably a group which is cathepsin labile.
The nitrogen protecting group is preferably introduced by reaction of IV with triphosgene to obtain the isocyanate followed by reaction with R10-OH. The reaction of IV with
triphosgene should be carried in an anhydrous and non-hydroxylic organic solvent, which is preferably non-polar. Suitable solvents include anhydrous dichloromethane and anhydrous toluene. The reaction may be carried out at room temperature. The subsequent reaction with R10-OH should be carried in an anhydrous and non-hydroxylic organic solvent, which is preferably non-polar. Suitable solvents include anhydrous dichloromethane and anhydrous toluene. The reaction should be carried out in the presence of a base is present, and suitable bases include pyridine or TEA. The reaction may be carried out at 0°C, or at a higher temperature to increase the rate of reaction.
D
The PBD B-ring is preferably closed using Dess-Martin periodinane. E
RE is selected from: 0-d-2 alkyl; 0-Si-(RSi1)(RSi2)(RSi3), where RSi1 , RSi2, RSi3 are as defined above; 0-CH2OCH3 (OMOM), 0-CH2OC2H4OCH3 (OMEM), OC5H10 (THP). Furthermore, Prot° needs to be orthogonal to RE. When RE is 0-Si-(RSi1)(RSi2)(RSi3), Si-(RSi1)(RSi2)(RSi3) is a silcon-based oxygen protecting group as described above. In these embodiments, RE may preferably be O-TBS.
G
In some embodiments. Hal and Q are the same, such that the compound of formula X is symmetrical. It is preferred that Hal is preferably I. It is also preferred that Q is I. Thus, in some embodiments, it is preferred that both Hal and Q are I. In some embodiments about 3 equialents of formula X are used relative to the compound of formula VIII.
H
The preferences expressed are applicable to this aspect of the invention. Othogonal protecting groups
In some embodiments of the present invention, RD is TBS and Prot° is TIPS. In other embodiments, RD is OAc and Prot° is either TIPS or TBS.
In some embodiments of the present invention, Prot° is TIPS and RE is OTBS.
In other embodiments of the present invention, Prot° is TIPS and RE is OMe, OMEM, THP or OMOM.
In further embodiments of the present invention, Prot° is TBS, and RE is OMe, OMEM, THP or OMOM. In particularly preferred embodiments of the present invention, RD is TBS, Prot° is TIPS and RE is TBS.
Examples
General Information
Reaction progress was monitored by thin-layer chromatography (TLC) using Merck
Kieselgel 60 F254 silica gel, with fluorescent indicator on aluminium plates. Visualisation of TLC was achieved with UV light or iodine vapour unless otherwise stated. Flash chromatography was performed using Merck Kieselgel 60 F254 silica gel. Extraction and chromatography solvents were bought and used without further purification from Fisher Scientific, U.K. All chemicals were purchased from Aldrich, Lancaster or BDH.
The LC/MS conditions were as follows: Method 1 (default when not speficied) The HPLC (Waters Alliance 2695) was run using a mobile phase of water (A) (formic acid 0.1 %) and acetonitrile (B) (formic acid 0.1 %). Gradient: initial composition 5% B over 1 .0 min then 5% B to 95% B within 3 min. The composition was held for 0.5 min at 95% B, and then returned to 5% B in 0.3 minutes. Total gradient run time equals 5 min. Flow rate 3.0 mL/min, 400μΙ_ was split via a zero dead volume tee piece which passes into the mass spectrometer. Wavelength detection range: 220 to 400 nm. Function type: diode array (535 scans). Column: Phenomenex® Onyx Monolithic C18 50 x 4.60 mm.
Method 2: The HPLC (Waters Alliance 2695) was run using a mobile phase of water (A) (formic acid 0.1 %) and acetonitrile (B) (formic acid 0.1 %). Gradient: initial composition 5% B over 1.0 min then 5% B to 95% B over 2.5 minute period. The composition was held for 0.5 min at 95% B, and then returned to 5% B in 0.1 minutes and held there for 0.9 minutes. Total gradient run time equals 5 min. Flow rate 3.0 mL/min, 400μΙ_ was split via a zero dead volume tee piece which passes into the mass spectrometer. Wavelength detection range: 220 to 400 nm. Function type: diode array (535 scans). Column: Phenomenex® Onyx Monolithic C18 50 x 4.60 mm.
Example 1
Figure imgf000061_0001
The synthesis of the starting material is described in JP 2008050321 A.
(a) Compound 1
Neat triisopropylsilylchloride (86 mL) was added to a mixture of imidazole (54.7 g) and phenol (4 g) (ground together). The mixture was heated until the phenol and imidazole melted and went into solution (1 10°C). The reaction mixture was allowed to stir for 5 minutes and was then allowed to cool, whereupon a solid was observed to form at the bottom of the flask. The reaction mixture was loaded directly on to silica gel and the silica pad was washed with hexane. The desired product was eluted with 5 % ethyl acetate in hexane. Excess eluent was removed by rotary evaporation under reduced pressure to afford a quantitative amount of the product 1 (1 12.454 g, >100 %). LCMS RT 4.25 mins, m/z ES+ 353.85
(b) Compound 2
A solution of sodium chlorite (60.6 g) and sodium dihydrogenphosphate (53.2 g)
(NaH2P04) in water (800 mL) was added to a solution of compound 1 (1 12.4 g) in tetrahydrofuran (500 mL) at room temperature. Hydrogen peroxide (60 % w/w, 179.02 mL) was immediately added to the vigorously stirred biphasic mixture. The reaction was initiated by addition of 1 M hydrochloric acid to lower the pH of the reaction mixture to between 5 and 6. The reaction mixture evolved gas (oxygen), the starting material dissolved and the temperature of the reaction mixture rose to 40°C. As the reaction proceeded the pH continued to drop and was kept between 5 and 6 by addition of solid sodium dihydrogenphosphate (1 - 2 g). After 1 hour LC/MS revealed that the reaction was complete. The reaction mixture was cooled in an ice bath and hydrochloric acid (1 M) was added to lower the pH to between 1 and 2. The reaction mixture was then extracted with ethyl acetate (1 L) and the organic phases washed with brine (2 x 100 mL) and dried over magnesium sulphate. The organic phase was filtered and excess solvent removed by rotary evaporation under reduced pressure to afford the product 2 in quantitative yield. LC/MS 3.93 mins, m/z ES"367.74
(c) Compound 3
A solution of acid 2 (58.60 g), hydroxybenzotriazole (26.59 g) and DCC (39.12 g) in dichloromethane (200 mL) was allowed to stir for 30 mins at which time a solution of C-ring (36.70 g) and triethylamine (33 mL) in dichloromethane (100 mL) was added at 0°C under argon. The coupling reaction proceeded rapidly and went to completion inside 1 hour, as monitored by LC/MS. The reaction mixture was filtered and diluted with dichloromethane (150 mL). The organic phase was washed sequentially with cold dilute hydrochloric acid (0.1 M), saturated aqueous sodium bicarbonate and brine. The organic layer was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure. The residue was subjected to column flash chromatography (silica gel; gradient 40 % ethyl acetate in hexane to 80 % ethyl acetate in hexane). Excess solvent was removed by rotary evaporation under reduced pressure. A white solid was observed in the resulting residue which was removed by dissolving the residue up in diethyl ether. Removal of diethyl ether by rotary evaporation under reduced pressure afforded the pure product 3, (38.80 g, 42 % yield). LC/MS 4.43 mins, m/z ES+582.92
(d) Compound 4
TCCA (8.82 g) was added to a stirred solution of 3 (31 .67 g) and TEMPO (0.85 g) in dry dichloromethane (250 mL) at 0°C. The reaction mixture was vigorously stirred for 20 minutes at room temperature, at which point TLC (50/50 ethyl acetate/hexane) revealed complete consumption of the starting material. The reaction mixture was filtered through celite and the filtrate washed with aqueous saturated sodium bicarbonate (100 mL), sodium thiosulphate (9 g in 300 mL), brine (100 mL) and dried over magnesium sulphate. Rotary evaporation under reduced pressure afforded product 4 in quantitative yield. LC/MS 4.52 mins, m/z ES+ 581.08
(e) Compound 5
Triflic anhydride (27.7 mL) was injected in one portion to a vigorously stirred suspension of compound 4 (31 .86 g) in dry dichloromethane (900 mL) in the presence of 2,6-lutidine (25.6 mL, dried over sieves) at -50°C (acetone/ dry ice bath). The reaction mixture was allowed to stir for 1.5 hours when LC-MS following a mini work-up (water/dichloromethane) revealed the reaction to be complete. Water was added to the still cold reaction mixture and the organic layer was separated and washed with saturated sodium bicarbonate, brine and magnesium sulphate. The organic phase was filtered and excess solvent was removed by rotary evaporation under reduced pressure. The residue was subjected to column flash chromatography (silica gel; 10 % ethyl acetate in hexane), removal of excess eluent afforded the product 5 (45.50 g, 96 % yield)
LC/MS RT 4.32 mins, m/z ES 712.89 (f) Compound 6
Tetrakis(triphenylphosphine)palladium(0) (1.14 g) was added to triflate 5 (23.40 g), E 2- propenylboronic acid (4.05 g) and potassium phosphate tribasic (13.93 g) in a mixture of dry dioxane (100 mL) under an argon atmosphere. The solution turned black and the reaction went rapidly to completion after 30 minutes as monitored by LC/MS. The reaction mixture was partitioned between ethyl acetate and water and the organic layer was washed with brine and dried over magnesium sulphate. The organic phase was filtered and excess solvent was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; 5 % ethyl acetate/10 % hexane). Pure fractions were collected and combined, and excess eluent was removed by rotary evaporation under reduced pressure afforded the product 6 (30.5 g, 82 % yield).
LC/MS 4.43 mins, m/z ES+ 605.1 1.
(f) Compound 7
Zinc powder (106 g) was added to a solution of compound 6 (27.06 g) in 5 % formic acid in ethanol (270 ml_). The resulting exotherm was controlled using an ice bath to maintain the temperature of the reaction mixture at 20°C. After 20 minutes the reaction mixture was filtered through celite. The filtrate was diluted with ethyl acetate and the organic phase was washed with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; 10 % ethyl acetate in hexane). Pure fractions were collected and combined and excess was removed by rotary evaporation under reduced pressure to afford the product 7 (14.63 g, 48 % yield). LC/MS RT 4.42 mins (g) Compound 8
Allyl chloroformate (1 .75 ml.) was added to a solution of amine 7 (8.602 g) in the presence of dry pyridine(2.66 g) in dry dichloromethane (200 ml.) at -78°C (acetone/ dry ice bath). After addition was complete the bath was removed and the reaction mixture was allowed to warm to room temperature. The reaction mixture was washed sequentially with dilute hydrochloric acid (0.01 N), saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure to afford the product 8 which was used directly in the next reaction.
LC/MS RT 4.08 mins (h) Compound 9
The crude 8 was dissolved in a 3/1/1 mixture of acetic acid/water/tetrahydrofuran (30/10/10 mL) and allowed to stir at room temperature. Deprotection was slower than expected so a mixture of methanol/acetic acid/water (30/30/30 mL) was added causing the reaction to proceed faster. After 6 hours the reaction was found to be 90 % complete, to avoid acidic degradation the reaction mixture was stored at -20°C in a spark free freezer overnight. The reaction was continued at room temperature the following day until all the starting material had been consumed. The reaction mixture was diluted with ethyl acetate and washed sequentially with water (2 x 500 mL), saturated aqueous sodium bicarbonate (200 mL) and brine. The organic phase was dried over magnesium sulphate filtered and excess ethyl acetate removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel, gradient; 40/ 60 to 50/50 ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to afford the desired product 9 (1 g, 70 %).
(i) Compound 10
Dimethyl sulphoxide (1.75 mL) was injected into an oven dried sealed microwave vial containing a solution of oxalyl chloride (1 mL) in dry dichloromethane (50 mL) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 15 minutes a solution of 9 (5.36 g) in dry dichloromethane (40 mL) was added slowly with the temperature still at -78°C. After 15 mins triethylamine (6.87 mL, dried over 4A molecular sieves) was injected and the dry ice/acetone bath was removed. The reaction mixture was allowed to reach room
temperature and was extracted with cold hydrochloric acid (0.1 M), saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess dichloromethane was removed by rotary evaporation under reduced pressure to afford product 10 in a quantitative yield. LC/MS RT 3.68 mins, m/z ES+543.03.
(j) Compound 11
7e/t-butyldimethylsilyltriflate (4.713 mL) was added to a solution of compound 10 (3.713 g) and 2,6-lutidine (3.187 mL) in dry dichloromethane (40 mL) and cooled to 0°C under argon. After 1 hour, the reaction mixture was extracted with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient, 5 % ethyl acetate in hexane to 20 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 11 (3.05 g, 68 % yield). LC/MS RT 4.05 mins, ES+ 657.16. (k) Compound 12
Lithium acetate (0.47 g) was added to a solution of compound 11 (3.05 g) in wet dimethylformamide (30 ml_, 50; 1 DMF/water). After 1 and three-quarter hours the reaction was observed to be complete and the reaction mixture was diluted with ethyl acetate and washed with citric acid, water (0.6 ml.) and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash
chromatography (silica gel; gradient, 25 % ethyl acetate to 75 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 12 (2.12 g, 91 % yield). LC/MS RT 3.55 mins, ES+ 501.12.
(I) Compound 13
Diiodopentane (377 μί) was added to a solution of compound 12 (254 mg) in acetone (2 mL) over potassium carbonate (70 mg) under an argon atmosphere. The reaction mixture was heated at 60°C for 2 hours at which time TLC indicated that the reaction was complete. Excess acetone was removed by rotary evaporation under reduced pressure. The residue (including inorganics) was subjected to column flash chromatography (silica gel; gradient 20 % ethyl acetate in hexane to 33 % ethyl acetate in hexane). Pure fractions were collected and combined and removal of excess eluent by rotary evaporation under reduced pressure afforded the product 13 (282 mg, 80 % yield). LC/MS RT 4.03 mins, m/z ES+ 696.73
Figure imgf000067_0001
(a) Compound 14
Triethylamine (0.745 mL) was added to a stirred solution of the amine 7 (1.4 g) and trisphosgene 260 mg) in dry tetrahydrofuran (30 mL) at 5°C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquot from the reaction mixture quenching with methanol and performing LC/MS analysis. Once the isocyanate formation was complete a solution of the alloc-Val-Ala-PABOH (1 .38 g) and triethylamine (0.5 mL) in dry tetrahydrofuran (50 mL) was rapidly added by injection to the freshly prepare isocyanate. The reaction mixture was allowed to stir at 30°C overnight. Excess solvent was removed by rotary evaporation under reduced pressure and the residue absorbed onto silica gel and dry loaded onto a chromatographic column. Gradient elution (20 % ethyl acetate in hexane to 50 % ethyl acetate in hexane) afforded the product 14 (1.72 g, 72 % yield). LC/MS RT 4.37 mins, ES+958.30
(b) Compound 15
The TBS ether 14 was allowed to stir in a 3/1/1 mixture of acetic acid, tetrahydrofuran and water for 3 hours at room temperature at which time the reaction was complete. The reaction mixture was diluted with ethyl acetate and sequentially washed with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; 80 %ethyl acetate in hexane). Pure fractions were collected and combined and removal of excess eluent by rotary evaporation under reduced pressure afforded the product 15 (1 .4 g, 93 %). LC/MS RT 3.75 mins, ES+ 864.26 (c) Compound 16
Dimethyl sulphoxide (288 μΙ_) was added dropwise to an oven dried sealed microwave vial containing a solution of oxalyl chloride (1.65 μΙ_) in dry dichloromethane (20 ml.) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 20 minutes a solution of 15 (1.4 g) in dry dichloromethane (15 ml.) was added slowly with the temperature still at - 78°C. After 15 mins triethylamine (1 .13 ml_), dried over 4A molecular sieves) was added dropwise and the dry ice/acetone bath was removed. The reaction mixture was allowed to reach 20°C and was extracted with cold hydrochloric acid (0.1 M), saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess dichloromethane was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient 50 % ethyl acetate in hexane to 74 % ethyl acetate in hexane). Pure fractions were collected and combined and removal of excess eluent by rotary evaporation under reduced pressure afforded the product 16 (930 mg, 70 % yield). LC/MS RT 3.60 mins, m/z ES+ 862.08.
(d) Compound 17
7~e/t-butyldimethylsilyltriflate (744 μΙ_) was added to a solution of compound 16 (930 mg) and 2,6-lutidine (502 μΙ_) in dry dichloromethane (10 ml.) and cooled to 0°C under argon. TLC (ethyl acetate revealed c. 25 % product formation after 45 minutes. Additional 2,6 lutidine (502 μΙ_ )and TBS triflate (744 μΙ_) were injected and the reaction mixture allowed to warm to room temperature. The reaction mixture was allowed to stir for 30 mins at room temperature at which time TLC indicated that the reaction was complete. The reaction mixture was diluted with dichloromethane (50 ml.) washed with saturated ammonium chloride (2 x 50 ml_), and extracted with water (50 ml_), aqueous sodium bicarbonate (50 ml.) and brine (30 ml.) . The organic phase was dried over magnesium sulphate, filtered and excess was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient, 40 % ethyl acetate in hexane to 60 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 17 (580mg, 64 % yield). LC/MS RT 5.37 mins, ES+ 976.42.
(e) Compound 18
Lithium acetate (59.5 mg) was added to a solution of compound 17 (570 mg) in wet dimethylformamide (10 ml_, 50; 1 DMF/water). After 3 hours the reaction was observed to be complete and the reaction mixture was diluted with ethyl acetate and washed with citric acid, water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient, 50 % ethyl acetate in hexane to 80 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 18 (476 mg, quantitative yield). LC/MS RT 3.52 mins, ES+ 820.23.
Figure imgf000070_0001
Figure imgf000070_0002
Compound 19
A solution of 13 (70 mL) and the phenol 18 (82.4 mL) over potassium carbonate (13.88 mg) in acetone (2 mL) was heated at 65°C for 10 hours under argon. The reaction mixture was evaporated to dryness by rotary evaporation under reduced pressure. The residue
(including inorganics) was subjected to column flash chromatography (silica gel; gradient 60 % ethyl acetate in hexane to 75 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent removed by rotary evaporation under reduced pressure to afford the product 19 (86 mg, 62 % yield). LC/MS RT 4.33 mins, m/z ES+ 1389.63 Exam le 4
Figure imgf000071_0001
(a) Compound 20 - (S)-(2-(((tert-butyldimethylsilyl)oxy)methy^
pyrrol-1-yl)(5-methoxy-2-nitro-4-((triisopropylsilyl)oxy)phenyl)methanone
Triphenylarsine (1.71 g, 5.60 mmol, 0.4 eq) was added to a mixture of triflate 5 (10.00 g, 14 mmol, 1 eq), methylboronic acid (2.94 g, 49.1 mmol, 3.5 eq), silver oxide (13 g, 56 mmol, 4 eq) and potassium phosphate tribasic (17.8 g, 84 mmol, 6 eq) in dry dioxane (80 mL) under an argon atmosphere. The reaction was flushed with argon 3 times and
bis(benzonitrile)palladium(ll) chloride (540 mg, 1 .40 mmol, 0.1 eq) was added. The reaction was flushed with argon 3 more times before being warmed instantaneously to 1 10°C (the drysyn heating block was previously warmed to 1 10°C prior addition of the flask). After 10 mins the reaction was cooled to room temperature and filtered through a pad celite. The solvent was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; 10 % ethyl acetate / hexane). Pure fractions were collected and combined, and excess eluent was removed by rotary evaporation under reduced pressure afforded the product 20 (4.5 g, 55 %). LC/MS, method 2, 4.27 min (ES+) m/z (relative intensity) 579.18 {[M + Hf, 100); 1H NMR (400 MHz, CDCI3) δ 7.70 (s, 1 H), 6.77 (s, 1 H), 5.51 (d, J = 1.7 Hz, 1 H), 4.77 - 4.59 (m, 1 H), 3.89 (s, 3H), 2.92 - 2.65 (m, 1 H), 2.55 (d, J = 14.8 Hz, 1 H), 1 .62 (d, J = 1.1 Hz, 3H), 1.40 - 1 .18 (m, 3H), 1.1 1 (s, 9H), 1.10 (s, 9H), 0.90 (s, 9H), 0.1 1 (d, J = 2.3 Hz, 6H). (b) Compound 21 - (S)-(2-amino-5-methoxy-4-((triisopropylsilyl)oxy)phenyl)(2-(((tert- butyldimethylsilyl)oxy)methyl)-4-methyl-2,3-dihydro-1H-pyrrol- 1-yl)methanone
Zinc powder (28 g, 430 mmol, 37 eq) was added to a solution of compound 20 (6.7 g, 1 1 .58 mmol) in 5% formic acid in ethanol v/v (70 ml.) at around 15°C. The resulting exotherm was controlled using an ice bath to maintain the temperature of the reaction mixture below 30°C. After 30 minutes the reaction mixture was filtered through a pad of celite. The filtrate was diluted with ethyl acetate and the organic phase was washed with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column
chromatography (silica gel; 10 % ethyl acetate in hexane). The pure fractions were collected and combined and excess solvent was removed by rotary evaporation under reduced pressure to afford the product 21 (5.1 g, 80 %). LC/MS, method 2, 4.23 min (ES+) m/z (relative intensity) 550.21 {[M + Hf, 100); 1H NMR (400 MHz, CDCI3) δ 7.28 (s, 1 H), 6.67 (s, 1 H), 6.19 (s, 1 H), 4.64 - 4.53 (m, J = 4.1 Hz, 1 H), 4.17 (s, 1 H), 3.87 (s, 1 H), 3.77 - 3.69 (m, 1 H), 3.66 (s, 3H), 2.71 - 2.60 (m, 1 H), 2.53 - 2.43 (m, 1 H), 2.04 - 1 .97 (m, J = 1 1 .9 Hz, 1 H), 1 .62 (s, 3H), 1.26 - 1 .13 (m, 3H), 1.08 - 0.99 (m, 18H), 0.82 (s, 9H), 0.03 - - 0.03 (m, J = 6.2 Hz, 6H). (c) Compound 22 - (S)-allyl (2-(2-(((tert-butyldimethylsilyl)oxy)methyl)-4-methy^^
1H-pyrrole- 1-carbonyl)-4-methoxy-5-((triisopropylsilyl)oxy)phenyl)carbamate
Allyl chloroformate (0.30 ml_, 3.00 mmol, 1 .1 eq) was added to a solution of amine 21 (1 .5 g, 2.73 mmol) in the presence of dry pyridine (0.48 ml_, 6.00 mmol, 2.2 eq) in dry dichloromethane (20 ml.) at -78°C (acetone/dry ice bath). After 30 minutes, the bath was removed and the reaction mixture was allowed to warm to room temperature. The reaction mixture was diluted with dichloromethane and saturated aqueous copper sulphate was added. The organic layer was then washed sequentially with saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure to afford the product 22 which was used directly in the next reaction. LC/MS, method 2, 4.45 min (ES+) m/z (relative intensity) 632.91 {[M + Hf, 100) (d) Compound 23 - (S)-allyl (2-(2-(hydroxymethyl)-4-methyl-2,3-dihydro-1 H-pyrrole-1 - carbonyl)-4-methoxy-5-((triisopropylsilyl)oxy)phenyl)carbamate
The crude 22 was dissolved in a 7:1 :1 :2 mixture of acetic
acid/methanol/tetrahydrofuran/water (28:4:4:8 mL) and allowed to stir at room temperature. After 3h, complete disappearance of starting material was observed by LC/MS. The reaction mixture was diluted with ethyl acetate and washed sequentially with water (2 x 500 mL), saturated aqueous sodium bicarbonate (200 mL) and brine. The organic phase was dried over magnesium sulphate filtered and excess ethyl acetate removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel, 25% ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to afford the desired product 23 (1 g, 71 %). LC/MS, method 2, 3.70 min (ES+) m/z (relative intensity) 519.13 {[M + Hf, 95); 1 H NMR (400 MHz, CDCI3) δ 8.34 (s, 1 H), 7.69 (s, 1 H), 6.78 (s, 1 H), 6.15 (s, 1 H), 5.95 (ddt, J = 17.2, 10.5, 5.7 Hz, 1 H), 5.33 (dq, J = 17.2, 1 .5 Hz, 1 H), 5.23 (ddd, J = 10.4, 2.6, 1.3 Hz, 1 H), 4.73 (tt, J = 7.8, 4.8 Hz, 1 H), 4.63 (dt, J = 5.7, 1.4 Hz, 2H), 4.54 (s, 1 H), 3.89 - 3.70 (m, 5H), 2.87 (dd, J = 16.5, 10.5 Hz, 1 H), 2.19 (dd, J = 16.8, 4.6 Hz, 1 H), 1 .70 (d, J = 1.3 Hz, 3H), 1.38 - 1.23 (m, 3H), 1 .12 (s, 10H), 1.10 (s, 8H). (e) Compound 24 - (1 1S, 11aS)-allyl 11 -hydroxy-7-methoxy-2-methyl-5-oxo-8-
((triisopropylsilyl)oxy)- 1 1, 11a-dihydro- 1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)- carboxylate
Dimethyl sulphoxide (0.35 mL, 4.83 mmol, 2.5 eq) was added dropwise to a solution of oxalyl chloride (0.2 mL, 2.32 mmol, 1 .2 eq) in dry dichloromethane (10 mL) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 10 minutes a solution of 23 (1 g, 1.93 mmol) in dry dichloromethane (8 mL) was added slowly with the temperature still at - 78°C. After 15 minutes triethylamine (1.35 mL, dried over 4A molecular sieves, 9.65 mmol, 5 eq) was added dropwise and the dry ice/acetone bath was removed. The reaction mixture was allowed to reach room temperature and was extracted with cold hydrochloric acid (0.1 M), saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess dichloromethane was removed by rotary evaporation under reduced pressure to afford product 24 (658 mg, 66%). LC/MS, method 2, 3.52 min (ES+) m/z (relative intensity) 517.14 {[M + Hf, 100); 1H NMR (400 MHz, CDCIs) δ 7.20 (s, 1 H), 6.75 - 6.63 (m, J = 8.8, 4.0 Hz, 2H), 5.89 - 5.64 (m, J = 9.6, 4.1 Hz, 2H), 5.23 - 5.03 (m, 2H), 4.68 - 4.38 (m, 2H), 3.84 (s, 3H), 3.83 - 3.77 (m, 1 H), 3.40 (s, 1 H), 3.05 - 2.83 (m, 1 H), 2.59 (d, J = 17.1 Hz, 1 H), 1 .78 (d, J = 1 .3 Hz, 3H), 1 .33 - 1.16 (m, 3H), 1 .09 (d, J = 2.2 Hz, 9H), 1 .07 (d, J = 2.1 Hz, 9H).
(f) Compound 25 - (1 1S, 11aS)-allyl 11-((tert-butyldimethylsilyl)oxy)-7-methoxy-2-methyl-5- oxo-8-((triisopropylsilyl)oxy)-11, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)-carboxylate
7~e/t-butyldimethylsilyltriflate (0.70 mL, 3.00 mmol, 3 eq) was added to a solution of compound 24 (520 mg, 1.00 mmol) and 2,6-lutidine (0.46 mL, 4.00 mmol, 4 eq) in dry dichloromethane (40 mL) at 0°C under argon. After 10 minutes, the cold bath was removed and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was extracted with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 10 % ethyl acetate in hexane to 20 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 25 (540 mg, 85 %). LC/MS, method 2, 4.42 min (ES+) m/z (relative intensity) 653.14 {[M + Naf, 100); 1H NMR (400 MHz, CDCIs) δ 7.20 (s, 1 H), 6.71 - 6.64 (m, J = 5.5 Hz, 2H), 5.83 (d, J = 9.0 Hz, 1 H), 5.80 - 5.68 (m, J = 5.9 Hz, 1 H), 5.14 - 5.06 (m, 2H), 4.58 (dd, J = 13.2, 5.2 Hz, 1 H), 4.36 (dd, J = 13.3, 5.5 Hz, 1 H), 3.84 (s, 3H), 3.71 (td, J = 10.1 , 3.8 Hz, 1 H), 2.91 (dd, J = 16.9, 10.3 Hz, 1 H), 2.36 (d, J = 16.8 Hz, 1 H), 1.75 (s, 3H), 1.31 - 1.16 (m, 3H), 1.12 - 1.01 (m, J = 7.4, 2.1 Hz, 18H), 0.89 - 0.81 (m, 9H), 0.25 (s, 3H), 0.19 (s, 3H).
(g) Compound 26 - (1 1S, 11aS)-allyl 11 -((tert-butyldimethylsilyl)oxy)-8-hydroxy-7-methoxy- 2-methyl-5-oxo- 1 1, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][ 1,4]diazepine-10(5H)- carboxylate
Lithium acetate (87 mg, 0.85 mmol) was added to a solution of compound 25 (540 mg, 0.85 mmol) in wet dimethylformamide (6 mL, 50:1 DMF/water). After 4 hours, the reaction was complete and the reaction mixture was diluted with ethyl acetate (25 mL) and washed with aqueous citric acid solution (pH ~ 3), water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 25% to 75% ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 26 (400 mg, quantitative). LC/MS, method 2, (3.33 min (ES+) m/z (relative intensity) 475.26 ([M+H]+, 100).
(h) Compound 27 - (1 1S, 11aS)-allyl 11 -((tert-butyldimethylsilyl)oxy)-8-((5-iodopentyl)oxy)- 7-methoxy-2-methyl-5-oxo- 11, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)- carboxylate
Diiodopentane (0.63 ml_, 4.21 mmol, 5 eq) and potassium carbonate (1 16 mg, 0.84 mmol, 1 eq) were added to a solution of phenol 26 (400 mg, 0.84 mmol) in acetone (4 ml_, dried over molecular sieves). The reaction mixture was then warmed to 60°C and stirred for 6 hours. Acetone was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; 50/50, v/v, hexane/ethyl acetate,). Pure fractions were collected and combined and excess eluent was removed to provide 27 in 90% yield. LC/MS, method 2, 3.90 min (ES+) m/z (relative intensity) 670.91 ([M]+, 100). 1H NMR (400 MHz, CDCI3) δ 7.23 (s, 1 H), 6.69 (s, 1 H), 6.60 (s, 1 H), 5.87 (d, J = 8.8 Hz, 1 H), 5.83 - 5.68 (m, J = 5.6 Hz, 1 H), 5.15 - 5.01 (m, 2H), 4.67 - 4.58 (m, 1 H),
4.45 - 4.35 (m, 1 H), 4.04 - 3.93 (m, 2H), 3.91 (s, 3H), 3.73 (td, J = 10.0, 3.8 Hz, 1 H), 3.25 - 3.14 (m, J = 8.5, 7.0 Hz, 2H), 2.92 (dd, J = 16.8, 10.3 Hz, 1 H), 2.38 (d, J = 16.8 Hz, 1 H), 1.95 - 1.81 (m, 4H), 1.77 (s, 3H), 1 .64 - 1 .49 (m, 2H), 0.88 (s, 9H), 0.25 (s, 3H), 0.23 (s, 3H).
Exam le 5
Figure imgf000076_0001
(a) Compound 28 - allyl 3-(2-(2-(4-((((2-((S)-2-(((tert-butyldimethylsilyl)oxy)m
methyl-2,3-dihydro- 1 H-pyrrole- 1 -carbonyl) -4-methoxy-5- ((triisopropylsilyl)oxy)phenyl)carbamoyl)oxy)methyl)phenyl)hydrazinyl)propanam
methyl-2-oxopentanoate
Triethylamine (2.23 mL, 18.04 mmol, 2.2 eq) was added to a stirred solution of the amine 21 (4 g, 8.20 mmol) and triphosgene (778 mg, 2.95 mmol, 0.36 eq) in dry tetrahydrofuran (40 mL) at 5 °C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquots from the reaction mixture and quenching with methanol and performing LC/MS analysis. Once the isocyanate formation was complete a solution of the alloc-Val-Ala-PABOH (4.12 g, 12.30 mmol, 1 .5 eq) and triethylamine (1.52 mL, 12.30 mmol, 1 .5 eq) in dry tetrahydrofuran (40 mL) was rapidly added by injection to the freshly prepared isocyanate. The reaction mixture was allowed to stir at 40 °C for 4 hours. Excess solvent was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 1 % methanol to 5% methanol in dichloromethane). (Alternative chromatography conditions using EtOAc and Hexane have also been successful). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 28 (3.9 g, 50%). LC/MS, method 2, 4.23 min (ES+) m/z (relative intensity) 952.36 {[M + H]+ , 100); 1H NMR (400 MHz, CDCI3) δ 8.62 (br s, 1 H), 8.46 (s, 1 H), 7.77 (br s, 1 H), 7.53 (d, J = 8.4 Hz, 2H), 7.32 (d, J = 8.5 Hz, 2H), 6.76 (s, 1 H), 6.57 (d, J = 7.6 Hz, 1 H), 6.17 (s, 1 H), 6.03 - 5.83 (m, 1 H), 5.26 (dd, J = 33.8, 13.5 Hz, 3H), 5.10 (s, 2H), 4.70 - 4.60 (m, 2H), 4.58 (dd, J = 5.7, 1.3 Hz, 2H), 4.06 - 3.99 (m, 1 H), 3.92 (s, 1 H), 3.82 - 3.71 (m, 1 H), 3.75 (s, 3H), 2.79 - 2.64 (m, 1 H), 2.54 (d, J = 12.9 Hz, 1 H), 2.16 (dq, J = 13.5, 6.7 Hz, 1 H), 1 .67 (s, 3H), 1.46 (d, J = 7.0 Hz, 3H), 1.35 - 1 .24 (m, 3H), 1.12 (s, 9H), 1 .10 (s, 9H), 0.97 (d, J = 6.8 Hz, 3H), 0.94 (d, J = 6.8 Hz, 3H), 0.87 (s, 9H), 0.07 - -0.02 (m, 6H).
(b) Compound 29 - allyl 3-(2-(2-(4-((((2-((S)-2-(hydroxymethyl)-4-methyl-^
pyrrole- 1 -carbonyl)-4-methoxy-5-
((triisopropylsilyl)oxy)phenyl)carbamoyl)oxy)methyl)phenyl)hydrazinyl)p^
methyl-2-oxopentanoate
The TBS ether 28 (1 .32 g, 1 .38 mmol) was dissolved in a 7:1 :1 :2 mixture of acetic acid/methanol/tetrahydrofuran/water (14:2:2:4 ml.) and allowed to stir at room temperature. After 3 hours no more starting material was observed by LC/MS. The reaction mixture was diluted with ethyl acetate (25 ml.) and washed sequentially with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate filtered and excess ethyl acetate removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel, 2% methanol in dichloromethane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to afford the desired product 29 (920 mg, 80%). LC/MS, method 2, 3.60 min (ES+) m/z (relative intensity) 838.18 ([M+H]+ , 100).1 H NMR (400 MHz, CDCI3) δ 8.55 (s, 1 H), 8.35 (s, 1 H), 7.68 (s, 1 H), 7.52 (d, J = 8.1 Hz, 2H), 7.31 (d, J = 8.4 Hz, 2H), 6.77 (s, 1 H), 6.71 (d, J = 7.5 Hz, 1 H), 6.13 (s, 1 H), 5.97 - 5.82 (m, J = 5.7 Hz, 1 H), 5.41 - 5.15 (m, 3H), 5.10 (d, J = 3.5 Hz, 2H), 4.76 - 4.42 (m, 5H), 4.03 (t, J = 6.6 Hz, 1 H), 3.77 (s, 5H), 2.84 (dd, J = 16.7, 10.4 Hz, 1 H), 2.26 - 2.08 (m, 2H), 1.68 (s, 3H), 1.44 (d, J = 7.0 Hz, 3H), 1.30 (dt, J = 14.7, 7.4 Hz, 3H), 1.12 (s, 9H), 1.10 (s, 9H), 0.96 (d, J = 6.8 Hz, 3H), 0.93 (d, J = 6.8 Hz, 3H). (c) Compound 30 - (11 S, 11 aS)-4-(2-(1 -((1 -(allyloxy)-4-methyl-1 ,2-dioxopentan-3-yl)amino)- 1 -oxopropan-2-yl)hydrazinyl)benzyl 11 -hydroxy-7-methoxy-2-methyl-5-oxo-8- ((triisopropylsilyl)oxy)- 1 1, 11a-dihydro- 1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)- carboxylate
Dimethyl sulphoxide (0.2 ml_, 2.75 mmol, 2.5 eq) was added dropwise to a solution of oxalyl chloride (0.1 1 ml_, 1.32 mmol, 1.2 eq) in dry dichloromethane (7 ml.) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 10 minutes a solution of 29 (920 mg, 1.10 mmol) in dry dichloromethane (5 ml.) was added slowly with the temperature still at -78°C. After 15 minutes triethylamine (0.77 ml_, dried over 4A molecular sieves, 5.50 mmol, 5 eq) was added dropwise and the dry ice/acetone bath was removed. The reaction mixture was allowed to reach room temperature and was extracted with cold hydrochloric acid (0.1 M), saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess dichloromethane was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient 2% methanol to 5 % methanol in
dichloromethane). Pure fractions were collected and combined and removal of excess eluent by rotary evaporation under reduced pressure afforded the product 30 (550 mg, 60%). LC/MS, method 2, 3.43 min (ES+) m/z (relative intensity) 836.01 ([M]+ , 100). 1H NMR (400 MHz, CDCI3) δ 8.39 (s, 1 H), 7.52 - 7.40 (m, 2H), 7.21 - 7.08 (m, J = 1 1 .5 Hz, 2H), 6.67 (s, 1 H), 6.60 - 6.47 (m, J = 7.4 Hz, 1 H), 5.97 - 5.83 (m, 1 H), 5.79 - 5.66 (m, 1 H), 5.38 - 4.90 (m, 6H), 4.68 - 4.52 (m, J = 18.4, 5.5 Hz, 4H), 4.04 - 3.94 (m, J = 6.5 Hz, 1 H), 3.87 - 3.76 (m, 5H), 3.00 - 2.88 (m, 1 H), 2.66 - 2.49 (m, 2H), 2.21 - 2.08 (m, 2H), 1 .76 (s, 3H), 1.45 (d, J = 7.0 Hz, 3H), 1 .09 - 0.98 (m, J = 8.9 Hz, 18H), 0.96 (d, J = 6.7 Hz, 3H), 0.93 (d, J = 6.9 Hz, 3H).
(d) Compound 31 - (1 1 S, 1 1 aS)-4-(2-(1 -((1 -(allyloxy)-4-methyl- 1 ,2-dioxopentan-3-yl)amino)- 1 -oxopropan-2-yl)hydrazinyl)benzyl 11-((tert-butyldimethylsilyl)oxy)-7-methoxy-2-methyl-5- oxo-8-((triisopropylsilyl)oxy)-11, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)-carboxylate
7~e/t-butyldimethylsilyltriflate (0.38 ml_, 1 .62 mmol, 3 eq) was added to a solution of compound 30 (450 mg, 0.54 mmol) and 2,6-lutidine (0.25 ml_, 2.16 mmol, 4 eq) in dry dichloromethane (5 ml.) at 0°C under argon. After 10 minutes, the cold bath was removed and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was extracted with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; 50/50 v/v hexane/ethyl acetate). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 31 (334 mg, 65%). LC/MS, method 2, 4.18 min (ES+) m/z (relative intensity) 950.50 ([Mf, 100). 1H NMR (400 MHz, CDCI3) δ 8.53 (s, 1 H), 8.02 (s, 1 H), 7.44 (d, J = 7.6 Hz, 2H), 7.21 (s, 1 H), 7.08 (d, J = 8.2 Hz, 2H), 6.72 - 6.61 (m, J = 8.9 Hz, 2H), 6.16 (s, 1 H), 5.97 - 5.79 (m, J = 24.4, 7.5 Hz, 2H), 5.41 - 5.08 (m, 5H), 4.86 (d, J = 12.5 Hz, 1 H), 4.69 - 4.60 (m, 1 H), 4.57 (s, 1 H), 4.03 (t, J = 6.7 Hz, 1 H), 3.87 (s, 3H), 3.74 (td, J = 9.6, 3.6 Hz, 1 H), 2.43 - 2.09 (m, J = 34.8, 19.4, 1 1 .7 Hz, 3H), 1 .76 (s, 3H), 1.43 (d, J = 6.9 Hz, 3H), 1 .30 - 1.21 (m, 3H), 0.97 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 8.4 Hz, 3H), 0.84 (s, 9H), 0.23 (s, 3H), 0.12 (s, 3H).
(e) Compound 32 - (1 1 S, 1 1 aS)-4-(2-(1 -((1 -(allyloxy)-4-methyl- 1 ,2-dioxopentan-3-yl)amino)- 1-oxopropan-2-yl)hydrazinyl)benzyl 11 -((tert-butyldimethylsilyl)oxy)-8-hydroxy-7-methoxy- 2-methyl-5-oxo- 1 1, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)- carboxylate
Lithium acetate (50 mg, 0.49 mmol) was added to a solution of compound 31 (470 mg, 0.49 mmol) in wet dimethylformamide (4 mL, 50:1 DMF/water). After 4 hours, the reaction was complete and the reaction mixture was diluted with ethyl acetate and washed with citric acid (pH ~ 3), water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient, 50/50 to 25/75 v/v hexane/ethyl acetate). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 32 (400 mg, quantitative). LC/MS, method 2, 3.32 min (ES+) m/z
(relative intensity) 794.18 ([M+H]+ , 100). 1 H NMR (400 MHz, CDCI3) δ 8.53 (s, 1 H), 8.02 (s, 1 H), 7.44 (d, J = 7.6 Hz, 2H), 7.21 (s, 1 H), 7.08 (d, J = 8.2 Hz, 2H), 6.72 - 6.61 (m, J = 8.9 Hz, 2H), 6.16 (s, 1 H), 5.97 - 5.79 (m, J = 24.4, 7.5 Hz, 2H), 5.41 - 5.08 (m, 5H), 4.86 (d, J = 12.5 Hz, 1 H), 4.69 - 4.60 (m, 1 H), 4.57 (s, 1 H), 4.03 (t, J = 6.7 Hz, 1 H), 3.87 (s, 3H), 3.74 (td, J = 9.6, 3.6 Hz, 1 H), 2.43 - 2.09 (m, J = 34.8, 19.4, 1 1 .7 Hz, 3H), 1 .76 (s, 3H), 1 .43 (d, J = 6.9 Hz, 3H), 1.30 - 1 .21 (m, 3H), 0.97 (d, J = 6.7 Hz, 3H), 0.92 (t, J = 8.4 Hz, 3H), 0.84 (s, 9H), 0.23 (s, 3H), 0.12 (s, 3H). Example 6
Ala-Val-Alloc
Figure imgf000080_0001
Compound 33 - (1 1S)-allyl 8-((5-(((11S)-10-(((4-(2-(1-((1-(allyloxy)-4-me^
dioxopentan-3-yl)amino)-1-oxopropan-2-yl)hydrazinyl)benzyl)oxy)carbonyl)- 1 1-((tert- butyldimethylsilyl)oxy)-7-methoxy-2-methyl-5-oxo-5, 10, 1 1, 11a-tetrahydro-1 H- benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepin-8-yl)oxy)pentyl)oxy)-1 1 -((tert-butyldimethylsilyl)oxy)-7- methoxy-2-methyl-5-oxo- 11, 1 1a-dihydro- 1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)- carboxylate
Potassium carbonate (70 mg, 0.504 mmol, 1 eq) was added to a solution of 27 (370 mg, 0.552 mmol, 1.2 eq) and phenol 32 (400 mg, 0.504 mmol) in dry acetone (25 ml_). The reaction was stirred 8 hours at 70°C. The LC/MS showed that all the starting material was not consumed, so the reaction was allowed to stir overnight at room temperature and stirred for an additional 2 hours the next day. Acetone was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column
chromatography (silica gel; 80% ethyl acetate in hexane to 100% ethyl acetate). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 33 (385 mg, 57%). LC/MS, method 2, 4.07 min (ES+) m/z (relative intensity) 1336.55 ([M+H]+ , 50). Exam le 7
Figure imgf000081_0001
(a) Compound 34 - (S)-(2-(((tert-butyldimethylsilyl)oxy)methyl)-4-(4-fluoroph
dihydro- 1 H-pyrrol- 1 -yl)(5-methoxy-2-nitro-4-((t isopropylsilyl)oxy)phenyl)methanone Tetrakis(triphenylphosphine) palladium(O) (1.65 g, 1.43 mmol, 0.03 eq) was added to a mixture of triflate 5 (34.00 g, 47.6 mmol, 1 eq), 4-fluorophenylboronic acid (8.00 g, 57.17 mmol, 1 .2 eq) and potassium phosphate tribasic (20.2 g, 95.16 mmol, 2 eq) in dry dioxane (170 ml.) under an argon atmosphere. The reaction was flushed with argon 3 times and stirred at ambient temperature for 12 hours. The reaction was diluted with ethyl acetate and the organic phase was washed with water and brine and dried over magnesium sulphate. The solvent was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; 10 % ethyl acetate / hexane). Pure fractions were collected and combined, and excess eluent was removed by rotary evaporation under reduced pressure afforded the product 34 (19 g, 61 %). LC/MS, method 2, 4.28 min (ES+) m/z (relative intensity) 659.17 {[M + H]+ , 60) (b) Compound 35 - (S)-(2-amino-5-methoxy-4-((triisopropylsilyl)oxy)phenyl)(2-(((tert- butyldimethylsilyl)oxy)methyl)-4-(4-fluorophenyl)-2,3-dihydro- 1 H-pyrrol- 1 -yl)methanone Zinc powder (44.2 g, 676 mmol, 37 eq) was added to a solution of compound 34 (12 g, 18.21 mmol) in 5% formic acid in ethanol v/v (120 ml.) at around 15°C. The resulting exotherm was controlled using an ice bath to maintain the temperature of the reaction mixture below 30°C. After 30 minutes the reaction mixture was filtered through a pad of celite. The filtrate was diluted with ethyl acetate and the organic phase was washed with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; 5 % ethyl acetate in hexane). The pure fractions were collected and combined and excess solvent was removed by rotary evaporation under reduced pressure to afford the product 35 (1 1 g, 96 %). LC/MS, method 2, 4.32 min (ES+) m/z (relative intensity) 630.19 {[M + Hf, 100)
(c) Compound 36 - (S)-allyl (2-(2-(((tert-butyldimethylsilyl)oxy)methyl)-4-(4-fluorophenyl)- 2,3-dihydro-1H-pyrrole-1-carbonyl)-4-methoxy-5-((triisopropylsilyl)oxy)phenyl)carbamate Allyl chloroformate (0.9 ml_, 8.75 mmol, 1 .1 eq) was added to a solution of amine 35 (5.0 g, 7.95 mmol) in the presence of dry pyridine (1.4 ml_, 17.5 mmol, 2.2 eq) in dry
dichloromethane (100 ml.) at -78°C (acetone/dry ice bath). After 30 minutes, the bath was removed and the reaction mixture was allowed to warm to room temperature. The reaction mixture was diluted with dichloromethane and saturated aqueous copper sulphate was added. The organic layer was then washed sequentially with saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent removed by rotary evaporation under reduced pressure to afford the product 36 which was used directly in the next reaction. LC/MS, method 2, 4.52 min (ES+) m/z (relative intensity) 712.86 {[M + Hf, 100).
(d) Compound 37 - (S)-allyl (2-(4-(4-fluorophenyl)-2-(hydroxymethyl)-2,3-dihydro- 1H- pyrrole- 1 -carbonyl)-4-methoxy-5-((triisopropylsilyl)oxy)phenyl)carbamate
The crude 36 was dissolved in a 7:1 :1 :2 mixture of acetic
acid/methanol/tetrahydrofuran/water (28:4:4:8 ml.) and allowed to stir at room temperature. After 3 hours, complete disappearance of starting material was observed by LC/MS. The reaction mixture was diluted with ethyl acetate and washed sequentially with water (2 x 500 mL), saturated aqueous sodium bicarbonate (200 mL) and brine. The organic phase was dried over magnesium sulphate filtered and excess ethyl acetate removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel, 25% ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to afford the desired product 37 (3.0 g, 64 % over 2 steps). LC/MS, method 2, 3.85 min (ES+) m/z (relative intensity) 598.69 {[M + H]+ , 95)
(e) Compound 38 - (1 1S, 11aS)-allyl 2-(4-fluorophenyl)- 11 -hydroxy-7-methoxy-5-oxo-8- ((triisopropylsilyl)oxy)-11, 1 1 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)- carboxylate
Dimethyl sulphoxide (0.89 mL, 12.54 mmol, 2.5 eq) was added dropwise to a solution of oxalyl chloride (0.51 mL, 6.00 mmol, 1.2 eq) in dry dichloromethane (30 mL) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 10 minutes a solution of 37 (3 g, 4.98 mmol) in dry dichloromethane (24 mL) was added slowly with the temperature still at - 78°C. After 15 minutes triethylamine (3.48 mL, dried over 4A molecular sieves, 25.02 mmol, 5 eq) was added dropwise and the dry ice/acetone bath was removed. The reaction mixture was allowed to reach room temperature and was extracted with cold hydrochloric acid (0.1 M), saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess dichloromethane was removed by rotary evaporation under reduced pressure to afford product 38 (2.4 g, 80%). LC/MS, method 2, 3.70 min (ES+) m/z (relative intensity) 596.67 {[M + H]+ , 100).
(f) Compound 39 - (1 1S, 11aS)-allyl 11 -((tert-butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-7- methoxy-5-oxo-8-((triisopropylsilyl)oxy)-1 1 , 1 1 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2- a][1 ,4]diazepine-10(5H)-carboxylate
7~ert-butyldimethylsilyltriflate (3.0 mL, 13.08 mmol, 3 eq) was added to a solution of compound 38 (2.6 g, 4.36 mmol) and 2,6-lutidine (2.0 mL, 17.44 mmol, 4 eq) in dry dichloromethane (40 mL) at 0°C under argon. After 10 minutes, the cold bath was removed and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was extracted with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 10 % ethyl acetate in hexane to 20 % ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 39 (2.23 g, 72 %). LC/MS, method 2, 4.52 min (ES+) m/z (relative intensity) 710.82 {[M + Hf, 100)
(g) Compound 40 - (1 1S, 11aS)-allyl 11 -((tert-butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-8- hydroxy-7-methoxy-5-oxo- 1 1, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)- carboxylate
Lithium acetate (31 1 mg, 3.04 mmol) was added to a solution of compound 39 (2.16 g, 3.04 mmol) in wet dimethylformamide (20 mL, 50:1 DMF/water). After 2 hours, the reaction was complete and the reaction mixture was diluted with ethyl acetate (25 mL) and washed with aqueous citric acid solution (pH ~ 3), water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 25% to 75% ethyl acetate in hexane). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 40 (1.7 g, quantitative). LC/MS, method 2, (3.57 min (ES+) m/z (relative intensity) 554.93 ([M+H]+, 100).
(h) Compound 41 - (1 1S, 11aS)-allyl 11 -((tert-butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-8- ((5-iodopentyl)oxy)-7-methoxy-5-oxo-1 1 , 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2- a][1 ,4]diazepine-10(5H)-carboxylate
Diiodopentane (1.07 mL, 7.2 mmol, 5 eq) and potassium carbonate (200 mg, 1.44 mmol, 1 eq) were added to a solution of phenol 40 (800 mg, 1 .44 mmol) in acetone (8 mL, dried over molecular sieves). The reaction mixture was then warmed to 60°C and stirred for 6 hours. Acetone was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; 50/50, v/v, hexane/ethyl acetate,). Pure fractions were collected and combined and excess eluent was removed to provide 41 (800 mg, 74%). LC/MS, method 2, 4.00 min (ES+) m/z (relative intensity) 750.66 ([M+H]+, 100). (i) Compound 42 - (11S, 1 1aS)-allyl 1 1-((tert-butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-8-(3- iodopropoxy)-7-methoxy-5-oxo-11 , 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)-carboxylate
Diiodopropane (0.52 mL, 4.50 mmol, 5 eq) and potassium carbonate (124 mg, 0.90 mmol, 1 eq) were added to a solution of phenol 40 (500 mg, 0.90 mmol) in acetone (5 mL, dried over molecular sieves). The reaction mixture was then warmed to 60°C and stirred for 3 hours. Acetone was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; 50/50, v/v, hexane/ethyl acetate,). Pure fractions were collected and combined and excess eluent was removed to provide 42 (450 mg, 70% with some impurities due to elimination reaction). LC/MS, method 2, 3.90 min (ES+) m/z (relative intensity) 722.33 ([M+H]+, 100).
Example 8
Figure imgf000085_0001
(a) Compound 43 - allyl ((S)-1-(((S)- 1-((4-((((2-((S)-2-(((tert-butyW^
4-(4-fluorophenyl)-2,3-dihydro-1 H-pyrrole-1-carbonyl)-4-methoxy-5- ((triisopropylsilyl)oxy)phenyl)carbamoyl)oxy)methyl)phenyl)amino)-1-oxopropan-2 yl)amino)-3-methyl- 1 -oxobutan-2-yl)carbamate
Triethylamine (4.34 mL, 31 .15 mmol, 2.2 eq) was added to a stirred solution of the amine 35 (8.91 g, 14.16 mmol) and triphosgene (1 .51 g, 5.10 mmol, 0.36 eq) in dry tetrahydrofuran (80 mL) at 5 °C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquots from the reaction mixture and quenching with methanol and performing LC/MS analysis. Once the isocyanate formation was complete a solution of the alloc-Val-Ala-PABOH (8.02 g, 21.24 mmol, 1 .5 eq) and triethylamine (2.96 mL, 21 .24 mmol, 1.5 eq) in dry tetrahydrofuran (80 mL) was rapidly added by injection to the freshly prepared isocyanate. The reaction mixture was allowed to stir at 40 °C for 4 hours. Excess solvent was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; gradient, 1 % methanol to 5% methanol in dichloromethane). (Alternative chromatography conditions using EtOAc and Hexane have also been successful). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 43 (8.78 g, 60%). : LC/MS, method 2, 4.28 min (ES+) m/z (relative intensity) 1032.56 {[M + H]+, 100). (b) Compound 44 - allyl ((S)- 1-(((S)-1-((4-((((2-((S)-4-(4-fluorophenyl)-2^
2,3-dihydro-1 H-pyrrole- 1 -carbonyl)-4-methoxy-5-
((triisopropylsilyl)oxy)phenyl)carbamoyl)oxy)methyl)phenyl)amino)- 1-oxopropan-2- yl)amino)-3-methyl- 1 -oxobutan-2-yl)carbamate
The TBS ether 43 (8.78 g, 8.51 mmol) was dissolved in a 7:1 :2 mixture of acetic acid/tetrahydrofuran/water (56:8:16 mL) and allowed to stir at room temperature. After 16 hours no more starting material was observed by LC/MS. The reaction mixture was diluted with ethyl acetate (250 mL) and washed sequentially with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate filtered and excess ethyl acetate removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel, 2% methanol to 5% in chloroform). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to afford the desired product 44 (3.43 mg, 44%). LC/MS, method 2, 3.75 min (ES+) m/z (relative intensity) 918.01 ([M+H]+ , 100). (c) Compound 45 - (11S, 11aS)-4-((S)-2-((S)-2-(((allyloxy)carbonyl)amino)-3- methylbutanamido)propanamido)benzyl 2-(4-fluorophenyl)- 1 1 -hydroxy-7-methoxy-5-oxo-8- ((triisopropylsilyl)oxy)- 1 1, 11a-dihydro- 1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)- carboxylate
Dimethyl sulphoxide (0.60 mL, 8.50 mmol, 2.5 eq) was added dropwise to a solution of oxalyl chloride (0.35 mL, 4.08 mmol, 1.2 eq) in dry dichloromethane (30 mL) at -78°C (dry ice /acetone bath) under an atmosphere of argon. After 10 minutes a solution of 44 (3.12 g, 3.40 mmol) in dry dichloromethane (40 mL) was added slowly with the temperature still at - 78°C. After 15 minutes triethylamine (2.37 mL, dried over 4A molecular sieves, 17.01 mmol, 5 eq) was added dropwise and the dry ice/acetone bath was removed. The reaction mixture was allowed to reach room temperature and was extracted with cold hydrochloric acid (0.1 M), saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess dichloromethane was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient 0% to 2% methanol in chloroform). Pure fractions were collected and combined and removal of excess eluent by rotary evaporation under reduced pressure afforded the product 45 (2.27 g, 66%). LC/MS, method 2, 3.58 min (ES+) m/z (relative intensity) 916.17 ([Mf, 100).
(d) Compound 46 - (1 1S, 1 1aS)-4-((S)-2-((S)-2-(((allyloxy)carbonyl)amino)-3- methylbutanamido)propanamido)benzyl 1 1-((tert-butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-
7- methoxy-5-oxo-8-((triisopropylsilyl)oxy)-11 , 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2- a][1 ,4]diazepine-10(5H)-carboxylate
7~e/t-butyldimethylsilyltriflate (1.61 mL, 6.99 mmol, 3 eq) was added to a solution of compound 45 (2.13 g, 2.33 mmol) and 2,6-lutidine (1 .09 mL, 9.32 mmol, 4 eq) in dry dichloromethane (35 mL) at 0°C under argon. After 10 minutes, the cold bath was removed and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was extracted with water, saturated aqueous sodium bicarbonate and brine. The organic phase was dried over magnesium sulphate, filtered and excess solvent was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; 50/50 v/v hexane/ethyl acetate). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 46 (1.12 g, 47%). : LC/MS, method 2, 4.30 min (ES+) m/z (relative intensity) 1030.27 ([M]+ , 100). (e) Compound 47 - (1 1S, 1 1aS)-4-((S)-2-((S)-2-(((allyloxy)carbonyl)amino)-3- methylbutanamido)propanamido)benzyl 1 1 -((tert-butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-
8- hydroxy-7-methoxy-5-oxo-11 , 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)-carboxylate
Lithium acetate (1 10 mg, 1.08 mmol) was added to a solution of compound 46 (1.1 1 g, 1.08 mmol) in wet dimethylformamide (10 mL, 50:1 DMF/water). After 4 hours, the reaction was complete and the reaction mixture was diluted with ethyl acetate and washed with citric acid (pH ~ 3), water and brine. The organic layer was dried over magnesium sulphate filtered and excess ethyl acetate was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to column flash chromatography (silica gel; gradient, 50/50 to 0/100 v/v hexane/ethyl acetate). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 47 (940 mg, quantitative). LC/MS, method 2, 3.52 min (ES+) m/z (relative intensity) 874.1 1 ([M+Hf, 100). Example 9
(a) Compound 48 - (1 1S, 1 1aS)-allyl 8-((5-(((1 1S, 1 1aS)-10-(((4-(2-(2- (((allyloxy)carbonyl)amino)-3-methylbutanamido)propanamido)benzyl)oxy)carbonyl)- 1 1 - ((tert-butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-7-methoxy-5-oxo-5, 10, 1 1, 11 a-tetrahydro- 1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepin-8-yl)oxy)pentyl)oxy)-11 -((tert-butyldimethylsilyl)oxy)- 2-(4-fluorophenyl)-7-methoxy-5-oxo- 1 1, 1 1 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2- a][1 ,4]diazepine-10(5H)-carboxylate
Figure imgf000088_0001
Potassium carbonate (63 mg, 0.458 mmol, 1 eq) was added to a solution of 41 (429 mg, 0.572 mmol, 1 eq) and phenol 47 (500 mg, 0.572 mmol) in dry acetone (5 ml_). The reaction was stirred 48 hours at 60°C. Acetone was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column chromatography (silica gel; 66% ethyl acetate in hexane to 100% ethyl acetate). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 48 (323 mg, 38%). LC/MS, method 2, 4.22 min (ES+) m/z (relative intensity) 1497.16 ([M+Hf, 100). (b) Compound 49 - (1 1S, 1 1aS)-allyl 8-(3-(((1 1S, 1 1aS)-10-(((4-(2-(1-((1-(allyloxy)-4-methyl- 1 ,2-dioxopentan-3-yl)amino)-1 -oxopropan-2-yl)hydrazinyl)benzyl)oxy)carbonyl)-1 1 -((tert- butyldimethylsilyl)oxy)-2-(4-fluorophenyl)-7-methoxy-5-oxo-5, 10, 11, 1 1 a-tetrahydro- 1H- benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepin-8-yl)oxy)propoxy)-1 1 -((tert-butyldimethylsilyl)oxy)-2-(4- fluorophenyl)-7-methoxy-5-oxo-11 , 11a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine- 10(5H)-carboxylate
loc
Figure imgf000089_0001
49
Potassium carbonate (50.6 mg, 0.366 mmol, 0.8 eq) was added to a solution of 42 (497 mg, 0.686 mmol, 1.5 eq) and phenol 47 (400 mg, 0.457 mmol) in dry acetone (40 mL). The reaction was stirred 8 hours at 70°C. The LC/MS showed that all the starting material was not consumed, so the reaction was allowed to stir overnight at room temperature and stirred for an additional 2 hours the next day. Acetone was removed by rotary evaporation under reduced pressure. The resulting residue was subjected to flash column
chromatography (silica gel; 80% ethyl acetate in hexane to 100% ethyl acetate). Pure fractions were collected and combined and excess eluent was removed by rotary evaporation under reduced pressure to give the product 49 (440 mg, 66%). LC/MS, method 2, 4.17 min (ES+) m/z (relative intensity) 1467.69 ([M+H]+ , 50). General Experiment for following examples
LCMS data were obtained using an Agilent 1200 series LC/MS with an Agilent 61 10 quadrupole MS, with Electrospray ionisation. Mobile phase A - 0.1 % Acetic acid in water. Mobile Phase B - 0.1 % in acetonitrile. Flow rate of 1 .00ml/min. Gradient from 5% B rising up to 95% B over 3 minutes, remaining at 95% B for 1 minute and then back down to 5% B over 6 seconds. The total run time is 5 minutes. Column: Phenomenex Gemini-NX 3μηη C18, 30 x 2.00mm. Chromatograms based on UV detection at 254nm. Mass Spectra were achieved using the MS in positive mode. Proton NMR chemical shift values were measured on the delta scale at 400 MHz using a Bruker AV400. The following abbreviations have been used: s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; br, broad. Coupling constants are reported in Hz. Unless otherwise stated, column chromatography (by the flash procedure) were performed on Merck Kieselgel silica (Art. 9385). Mass spectroscopy (MS) data were collected using a Waters Micromass LCT instrument coupled to a Waters 2795 HPLC separations module. Thin Layer Chromatography (TLC) was performed on silica gel aluminium plates (Merck 60, F254). All other chemicals and solvents were purchased from Sigma-Aldrich or Fisher Scientific and were used as supplied without further purification.
Optical rotations were measured on an ADP 220 polarimeter (Bellingham Stanley Ltd.) and concentrations (c) are given in g/100mL. Melting points were measured using a digital melting point apparatus (Electrothermal). IR spectra were recorded on a Perkin-Elmer Spectrum 1000 FT IR Spectrometer. 1H and 13C NMR spectra were acquired at 300 K using a Bruker Avance NMR spectrometer at 400 and 100 MHz, respectively. Chemical shifts are reported relative to TMS (δ = 0.0 ppm), and signals are designated as s (singlet), d (doublet), t (triplet), dt (double triplet), dd (doublet of doublets), ddd (double doublet of doublets) or m (multiplet), with coupling constants given in Hertz (Hz). Mass spectroscopy (MS) data were collected using a Waters Micromass ZQ instrument coupled to a Waters 2695 HPLC with a Waters 2996 PDA. Waters Micromass ZQ parameters used were:
Capillary (kV), 3.38; Cone (V), 35; Extractor (V), 3.0; Source temperature (°C), 100;
Desolvation Temperature (°C), 200; Cone flow rate (L/h), 50; De-solvation flow rate (L/h), 250. High-resolution mass spectroscopy (HRMS) data were recorded on a Waters Micromass QTOF Global in positive W-mode using metal-coated borosilicate glass tips to introduce the samples into the instrument. Thin Layer Chromatography (TLC) was performed on silica gel aluminium plates (Merck 60, F254), and flash chromatography utilised silica gel (Merck 60, 230-400 mesh ASTM). Except for the HOBt (NovaBiochem) and solid-supported reagents (Argonaut), all other chemicals and solvents were purchased from Sigma-Aldrich and were used as supplied without further purification. Anhydrous solvents were prepared by distillation under a dry nitrogen atmosphere in the presence of an appropriate drying agent, and were stored over 4A molecular sieves or sodium wire. Petroleum ether refers to the fraction boiling at 40-60°C.
General LC/MS conditions: The HPLC (Waters Alliance 2695) was run using a mobile phase of water (A) (formic acid 0.1 %) and acetonitrile (B) (formic acid 0.1 %). Gradient: initial composition 5% B over 1.0 min then 5% B to 95% B within 3 min. The composition was held for 0.5 min at 95% B, and then returned to 5% B in 0.3 minutes. Total gradient run time equals 5 min. Flow rate 3.0 mL/min, 400μΙ_ was split via a zero dead volume tee piece which passes into the mass spectrometer. Wavelength detection range: 220 to 400 nm. Function type: diode array (535 scans). Column: Phenomenex® Onyx Monolithic C18 50 x 4.60 mm
Synthesis of Key Intermediates
i)
Figure imgf000091_0001
(a) (S)-2-(allyloxycarbonylamino)-3-methylbutanoic acid (12)
Allyl chloroformate (36.2 ml, 340.59 mmol, 1.2 eq) was added dropwise to a stirred solution of L-valine (I1 )(33.25 g, 283.82 mmol, 1 .0 eq) and potassium carbonate (59.27 g, 425.74 mmol, 1 .5 eq) in water (650 ml.) and THF (650 ml_). The reaction mixture was stirred at room temperature for 18 hours, then the solvent was concentrated under reduced pressure and the remaining solution extracted with diethyl ether (3 x 100 ml_). The aqueous portion was acidified to pH 2 with cone. HCI and extracted with DCM (3 x 100 ml_). The combined organics were washed with brine, dried over MgS04, filtered and concentrated under reduced pressure to afford the product as a colourless oil (57.1 g, assumed 100% yield). LC/MS (1 .966 min (ES+)), m/z: 202.1 [M+H]+. 1H NMR (400 MHz, DMSO-d6) δ 12.57 (br s, 1H), 7.43 (d, 1H, J=8.6 Hz), 5.96-5.86 (m, 1H), 5.30 (ddd, 1H, J= 17.2, 3.4, 1.7 Hz), 5.18 (ddd, 1H, J= 10.4, 2.9, 1.6 Hz), 4.48 (dt, 2H, J= 5.3, 1.5 Hz), 3.85 (dd, 1H, J= 8.6, 6.0 Hz), 2.03 (oct, 1H, J = 6.6 Hz), 0.89 (d, 3H, J= 6.4 Hz), 0.87 (d, 3H, J= 6.5 Hz), (b) (S)-2,5-dioxopyrrolidin-1 -yl 2-(allyloxycarbonylamino)-3-methylbutanoate (13)
To a stirred solution of the protected acid 12 (60.6 g, 301.16 mmol, 1.0 eq) and N- hydroxysuccinimide (34.66 g, 301.16 mmol, 1.0 eq) in dry THF (800 ml_)was added dicyclohexylcarbodiimide (62.14 g, 301.16 mmol, 1 eq). The reaction was stirred for 18 hours at room temperature. The reaction mixture was then filtered, the solid washed with THF and the combined filtrate was concentrated under reduced pressure. The residue was re-dissolved in DCM and left to stand at 0°C for 30 minutes. The suspension was filtered and washed with cold DCM. Concentration of the filtrate under reduced pressure afforded the product as a viscous colourless oil (84.7 g, assumed 100% yield) which was used in the next step without further purification. LC/MS (2.194 min (ES+)), m/z: 321.0 [M+Na]+. 1H NMR (400 MHz, DMSO-d6) δ 8.0 (d, 1 H, J = 8.3 Hz), 5.97 - 5.87 (m, 1 H), 5.30 (ddd, 1H, J= 17.2, 3.0, 1.7 Hz), 5.19 (ddd, 1H, J= 10.4, 2.7, 1.4 Hz), 4.52 (dt, 2H, J= 5.3, 1.4 Hz), 4.32 (dd, 1H, J= 8.3, 6.6 Hz), 2.81 (m, 4H), 2.18 (oct, 1H, J= 6.7 Hz), 1.00 (d, 6H, J= 6.8 Hz), (c) (S)-2-((S)-2-(allyloxycarbonylamino)-3-methylbutanamido)propanoic acid (14)
A solution of succinimide ester 13(12.99 g, 43.55 mmol, 1.0 eq) in THF (50 mL) was added to a solution of L-alanine (4.07 g, 45.73 mmol, 1.05 eq) and NaHC03 (4.02 g, 47.90 mmol, 1.1 eq) in THF (100 mL) and H20 (100 mL). The mixture was stirred at room temperature for 72 hours when the THF was removed under reduced pressure. The pH was adjusted to 3-4 with citric acid to precipitate a white gum. After extraction with ethyl acetate (6 x 150 mL), the combined organics were washed with H20 (200 mL), dried over MgS04, filtered and concentrated under reduced pressure. Trituration with diethyl ether afforded the product as a white powder which was collected by filtration and washed with diethyl ether (5.78 g, 49%). LC/MS (1.925 min (ES+)), m/z: 273.1 [M+H]+. 1H NMR (400 MHz, DMSO-d6) δ 12.47 (brs, 1H), 8.17 (d, 1H, J=6.8 Hz), 7.16 (d, 1H, J=9.0 Hz), 5.95- 5.85 (m, 1H), 5.29 (dd, 1H, J= 17.2, 1.7 Hz), 5.17 (dd, 1H, J= 10.4, 1.5 Hz), 4.46 (m, 2H), 4.18 (quin, 1 H, J = 7.2 Hz), 3.87 (dd, 1 H, J = 9.0, 7.1 Hz), 1.95 (oct, 1 H, J = 6.8 Hz), 1.26 (d, 3H, J = 7.3 Hz), 0.88 (d, 3H, J= 6.8 Hz), 0.83 (d, 3H, J= 6.8 Hz), (d) Allyl (S)-1-((S)- 1 -(4-(hydroxymethyl)phenylamino)- 1 -oxopropan-2-ylamino)-3-methyl- 1 - oxobutan-2-ylcarbamate (15)
EEDQ (5.51 g, 22.29 mmol, 1.05 eq) was added to a solution of p-aminobenzyl alcohol (2.74 g, 22.29 mmol, 1.05 eq) and acid I4 (5.78 g, 21 .23 mmol, 1 eq) in dry THF (100 ml_). and stirred at room temperature for 72 hours. The reaction mixture was then concentrated under reduced pressure and the resulting brown solid was triturated with diethyl ether and filtered with subsequent washing with an excess of diethyl ether to afford the product as an off-white solid (7.1 g, 88 %). LC/MS (1 .980 min (ES+)), m/z: 378.0 [M+H]+. 1H NMR (400 MHz, DMSO-d6) δ 9.89 (br s, 1 H), 8.13 (d, 1 H, J = 7.0 Hz), 7.52 (d, 2H, J = 8.5 Hz), 7.26 (m, 1 H), 7.23 (d, 2H, J = 8.5 Hz), 5.91 (m, 1 H), 5.30 (m, 1 H), 5.17 (m, 1 H), 4.46 (m, 2H), 5.09 (t, 1 H, J = 5.6 Hz), 4.48 (m, 2H), 4.42 (m, 3H), 3.89 (dd, 1 H, J = 8.6, 6.8 Hz), 1 .97 (m, 1 H), 1.30 (d, 3H, J = 7.1 Hz), 0.88 (d, 3H, J = 6.8 Hz), 0.83 (d, 3H, J = 6.7 Hz),
(ϋ)
Figure imgf000093_0001
1-iodo-2-oxo-6,9, 12, 15-tetraoxa-3-azaoctadecan-18-oic acid (17)
A solution of iodoacetic anhydride (0.250 g, 0.706 mmol, 1.1 eq) in dry DCM (1 ml.) was added to amino-PEG(4)-acid I6 (0.170 g, 0.642 mmol, 1 .0 eq) in DCM (1 ml_). The mixture was stirred in the dark at room temperature overnight. The reaction mixture was washed with 0.1 M HCI, water, dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 3% MeOH and 0.1 % formic acid in chloroform to 10% MeOH and 0.1 % formic acid in chloroform) to afford the product as an orange oil (0.1 18 g, 42%). LC/MS (1 .623 min (ES+)), m/z: 433..98
[M+H]+. 1 H NMR (400 MHz, CDCI3) δ 8.069 (s, 1 H), 7.22 (br s, 1 H), 3.79 (t, 2H, J = 5.8 Hz), 3.74 (s, 2H), 3.72 - 3.58 (m, 14H), 3.50 - 3.46 (m, 2H), 2.62 (t, 2H, J = 5.8 Hz). Exam le 10
Figure imgf000094_0001
Figure imgf000094_0002
(a) (S)-5-((tert-butyldimethylsilyloxy)methyl)- 1 -(5-methoxy-2-nitro-4- (triisopropylsilyloxy)benzoyl)-4,5-dihydro-1H-pyrrol-3-yl trifluoromethanesulfonate (5) Triflic anhydride (28.4 g, 100.0 mmol, 3.0 eq) was added dropwise, over 25 mins, to a vigorously stirred solution of the ketone 4 (19.5 g, 30.0 mmol, 1 .0 eq) in DCM (550 mL) containing 2,6-lutidine (14.4 g, 130.0 mmol, 4.0 eq) at -50°C. The reaction mixture was stirred for 1.5 hours when LC/MS indicated complete reaction. The organic phase was washed successively with water (100 mL), saturated sodium bicarbonate (150 mL), brine (50 mL), and the organic phase was dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 90/10 v/v n-hexane/EtOAc) to afford the product as a pale yellow oil (19.5 g, 82 %). LC/MS (4.391 min (ES+)), m/z: 713.25 [M+H]+. 1H NMR (400 MHz, CDCI3) δ 7.68 (s, 1 H), 6.72 (s, 1 H), 6.02 (t, 1 H, J = 1 .9 Hz), 4.75 (m, 1 H), 4.05 (m, 2H), 3.87 (s, 3H), 3.15 (ddd, 1 H, J = 16.2, 10.3, 2.3 Hz), 2.96 (ddd, 1 H, J = 16.2, 4.0, 1.6 Hz), 1 .28 - 1 .21 (m, 3H), 1.07 (d, 18H, J = 7.2 Hz), 0.88 (s, 9H), 0.09 (s, 3H), 0.08 (s, 3H). (b) (S,E)-(2-((tert-butyldimethylsilyloxy)methyl)-4-(prop- 1 -enyl)-2,3-dihydro- 1 H-pyrrol- 1 - yl)(5-methoxy-2-nitro-4-(triisopropylsilyloxy)phenyl)methanone (6)
Tetrakis(triphenylphosphine)palladium(0) (0.41 g, 0.35 mmol, 0.03 eq) was added to a mixture of the triflate 5 (8.4 g, 1 1.8 mmol, 1 .0 eq), E-1-propene-1 -ylboronic acid (1 .42 g, 16.5 mmol, 1 .4 eq) and potassium phosphate (5.0 g, 23.6 mmol, 2.0 eq) in dry dioxane (60 ml.) under a nitrogen atmosphere. The mixture was stirred at 25°C for 120 mins when LC/MS indicated complete reaction. Ethyl acetate (120 ml.) and water (120 ml.) were added, the organic phase was removed, washed with brine (20 ml_), dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 95/5 v/v n-hexane/EtOAc to 90/10 v/v n-hexane/EtOAc) to afford the product as a yellow foam (4.96 g, 70 %). LC/MS (4.477 min (ES+)), m/z: 605.0 [M+H]+. 1 H NMR (400 MHz, CDCI3) δ 7.67 (s, 1 H), 6.74 (s, 1 H), 5.93 (d, 1 H, J = 15.4 Hz), 5.67 (s, 1 H), 4.65 (m, 1 H), 4.04 (m, 2H), 3.86 (s, 3H), 2.85 (m, 1 H), 2.71 (m, 1 H), 1 .72 (dd, 3H, J = 6.8, 1 .0 Hz), 1 .30 - 1.22 (m, 3H), 1.07 (d, 18H, J = 7.2 Hz), 0.87 (s, 9H), 0.08 (s, 3H), 0.07 (s, 3H).
(c) (S,E)-(2-amino-5-methoxy-4-(triisopropylsilyloxy)phenyl)(2-((tert- butyldimethylsilyloxy)methyl)-4-(prop- 1 -enyl)-2,3-dihydro-1 H-pyrrol- 1 -yl)methanone (7) Zinc dust (22.0 g, 0.33 mol, 37 eq) was added, in portions over 20 mins, to a solution of the propenyl intermediate 6 (5.5 g, 9.1 mmol, 1.0 eq) in 5% v/v formic acid / ethanol (55 ml_), using an ice bath to maintain the temperature between 25-30°C. After 30 mins, the reaction mixture was filtered through a short bed of celite®. The celite® was washed with ethyl acetate (65 ml.) and the combined organics were washed successively with water (35 ml_), saturated sodium bicarbonate (35 ml.) and brine (10 ml_). The organic phase was dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 90/10 v/v n-hexane/EtOAc) to afford the product as a pale yellow oil (3.6 g, 69.0 %). LC/MS (4.439 min (ES+)), m/z: 575.2 [M+H]+. 1H NMR (400 MHz, CDCI3) δ 6.75 (m, 1 H), 6.40 (br s, 1 H), 6.28 (m, 1 H), 6.1 1 (d, 1 H, J = 15.4 Hz), 5.53 (m, 1 H), 4.67 (m, 1 H), 4.36 (m, 2H), 3.93 (br s, 1 H), 3.84 (br s, 1 H), 3.73 (s, 3H), 2.86 (dd, 1 H, J = 15.7, 10.4 Hz), 2.73 (dd, 1 H, J = 15.9, 4.5 Hz), 1 .80 (dd, 3H, J = 6.8, 1 .3 Hz), 1.35 - 1.23 (m, 3H), 1 .12 (d, 18H, J = 7.3 Hz), 0.89 (s, 9H), 0.08 (s, 3H), 0.07 (s, 3H). (d) (S,E)-allyl 2-(2-((tert-butyldimethylsilyloxy)methyl)-4-(prop-1-enyl)-2,3-d^ pyrrole- 1 -carbonyl) -4-methoxy-5-(triisopropylsilyloxy)phenylcarbamate (8)
Allyl chloroformate (0.83 g, 6.88 mmol, 1 .1 eq) was added to a solution of the amine 7 (3.6 g, 6.26 mmol, 1 .0 eq) in dry DCM (80 ml.) containing dry pyridine (1.09 g, 13.77 mmol, 2.2 eq) at -78°C. The dry ice was removed and the reaction mixture allowed to warm to room temperature. After stirring for a further 15 minutes, LC/MS indicated complete reaction. The organic phase was washed successively with 0.01 N HCI (50 ml_), saturated sodium bicarbonate (50 ml_), brine (10 ml_), dried over MgS04, filtered and concentrated under reduced pressure to leave a pale yellow oil which was used in the next step without further purification (4.12g, assumed 100% yield). LC/MS (4.862 min (ES+)), m/z: 659.2 [M+H]+.
(e) (S,E)-allyl 2-(2-(hydroxymethyl)-4-(prop-1 -enyl)-2,3-dihydro-1 H-pyrrole-1 -carbonyl)-4- methoxy-5-(triisopropylsilyloxy)phenylcarbamate (9)
The crude intermediate 8 (assumed 100% yield, 4.12 g, 6.25 mmol, 1.0 eq) was dissolved in a mixture of acetic acid (70 mL), methanol (10 mL), THF (10 mL) and water (20 mL) and allowed to stir at room temperature. After 6 hours the reaction mixture was diluted with ethyl acetate (500 mL) and washed successively with water (2 x 500 mL), saturated sodium bicarbonate (300 mL) and brine (50 mL). The organic phase was dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 1/99 v/v methanol/DCM to 5/95 v/v methanol/DCM) to afford the product as a yellow oil and a further 1 g of unreacted starting material was recovered. This material was subjected to the same reaction conditions as above, but was left stirring for 16 hours. After work up and purification, additional product was isolated (2.7 g, 79%, 2 steps) LC/MS (3.742 min (ES+)), m/z: 545.2 [M+H]+. 1H NMR (400 MHz, CDCI3) δ 8.38 (m, 1 H), 7.72 (m, 1 H), 6.81 (s, 1 H), 6.37 (m, 1 H), 6.10 (d, 1 H, J = 15.8 Hz), 5.97 (m, 1 H), 5.53 (m, 1 H), 5.36 (ddd, 1 H, J = 17.2, 3.1 , 1 .5 Hz), 5.25 (ddd, 1 H, J = 10.4, 2.5, 1.3 Hz), 4.78 (m, 1 H), 4.65 (dt, 2H, J = 5.7, 1.3 Hz), 3.84 (m, 3H), 3.79 (s, 3H), 3.04 (dd, 1 H, J = 16.7, 10.5 Hz), 2.40 (dd, 1 H, J = 16.0, 4.5 Hz), 1 .82 (dd, 3H, J = 6.8, 1 .0 Hz), 1.36 - 1.26 (m, 3H), 1.14 (d, 18H, J = 7.3 Hz).
(f) (11S, 1 1aS)-allyl 1 1-hydroxy-7-methoxy-5-oxo-2-((E)-prop-1-enyl)-8-(triisopropylsilyloxy)- 11, 1 1 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)-carboxylate (10)
Dry dimethyl sulfoxide (1 .16 g, 14.87 mmol, 3.0 eq) was added dropwise to a solution of oxalyl chloride (0.94 g, 7.43 mmol, 1 .5 eq) in DCM (25 mL) at -78°C under an atmosphere of nitrogen. Maintaining the temperature at -78°C, after 10 mins a solution of the primary alcohol 9 (2.7 g, 4.96 mmol, 1 .0 eq) in DCM (20 ml.) was added dropwise. After a further 15 mins, dry triethylamine (2.5g, 24.78 mmol, 5.0 eq) was added, and the reaction mixture allowed to warm to room temperature. The reaction mixture was washed successively with cold 0.1 N HCI (50 ml_), saturated sodium hydrogen carbonate (50 ml.) and brine (10 ml.) and the organic layer was dried over MgS04, filtered and concentrated under reduced pressure to afford the product as a yellow oil which was used in the next step without further purification (2.68g, assumed 100% yield). LC/MS (3.548 min (ES+)), m/z: 543.2 [M+H]+.
(g) (11S, 11aS)-allyl 1 1-(tert-butyldimethylsilyloxy)-7-methoxy-5-oxo-2-((E)-prop-1-enyl)-8- (triisopropylsilyloxy)- l 1 , 1 1 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)- carboxylate (11)
7ert-butyldimethylsilyltrifluoromethane sulfonate (3.93 g, 14.87 mmol, 3.0 eq) was added to a solution of the carbinolamine 10 (assumed 100% yield, 2.68 g, 4.96 mmol, 1 .0 eq) and 2,6-lutidine (2.12 g, 19.83 mmol, 4.0 eq) in dry DCM (40 ml.) at 0°C under an atmosphere of nitrogen. After 10 minutes, the reaction mixture was allowed to warm to room
temperature and stirred for a further 60 minutes. The organic phase was washed successively with water (10 ml_), saturated sodium bicarbonate (10 ml.) and brine (5 ml_), dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, chloroform to 2/98 v/v Methanol/chloroform) to afford the product as a yellow oil (2.0g, 63%, 2 steps). LC/MS (4.748 min (ES+)), m/z:
657.2 [M+H]+. 1H NMR (400 MHz, CDCI3) δ 7.19 (s, 1 H), 6.86 (m, 1 H), 6.66 (s, 1 H), 6.22 (d, 1 H, J = 15.4 Hz), 5.81 (d, 1 H, J = 8.8 Hz), 5.78 (m, 1 H), 5.48 (m, 1 H), 5.1 1 (d, 1 H, J = 5.0 Hz), 5.08 (m, 1 H), 4.58 (dd, 1 H, J = 13.4, 5.4 Hz), 4.35 (dd, 1 H, J = 13.2, 5.7 Hz), 3.83 (s, 3H), 3.76 (s, 1 H), 3.00 (dd, 1 H, J = 15.6, 1 1.0 Hz), 2.53 (m, 1 H), 1 .81 (dd, 3H, J = 6.8, 0.9 Hz), 1.30 - 1 .18 (m, 3H), 1 .08 (d, 9H, J = 2.3 Hz), 1.06 (d, 9H, J = 2.3 Hz), 0.86 (s, 9H), 0.25 (s, 3H), 0.18 (s, 3H). (h) (11S, 11aS)-allyl 1 1-(tert-butyldimethylsilyloxy)-8-hydroxy-7-methoxy-5-oxo
1-enyl)-11, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)-carboxylate (12) Lithium acetate dihydrate (0.31 g, 3.04 mmol, 1 .0 eq) was added to a solution of the diazepine 11 (2.0 g, 3.04 mmol, 1 .0 eq) in wet DMF (20 mL) at 25°C and stirred for 4 hours. The reaction mixture was diluted with ethyl acetate (200 mL) and washed successively with 0.1 M citric acid (50 mL, pH 3), water (50 mL) and brine (10 mL), dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 50/50 v/v n-hexane/EtOAc to 25/75 v/v n- hexane/EtOAc) to afford the product as a pale yellow solid (0.68g, 45 %). LC/MS (3.352 min (ES+)), m/z: 501.1 [M+H]+. 1H NMR (400 MHz, CDCI3) δ 7.02 (s, 1 H), 6.66 (m, 1 H), 6.53 (s, 1 H), 6.03 (d, 1 H, J = 15.5 Hz), 5.80 (s, 1 H), 5.63 (d, 1 H, J = 8.9 Hz), 5.55 (m, 1 H), 5.29 (m, 1 H), 4.87 (m, 2H), 4.39 (dd, 1 H, J = 13.5, 4.2 Hz), 4.20 (dd, 1 H, J = 13.2, 5.7 Hz), 3.73 (s, 3H), 3.59 (m, 1 H), 2.81 (dd, 1 H, J = 16.1 , 10.5 Hz), 2.35 (d, 1 H, J = 15.7 Hz), 1 .61 (d, 3H, J = 6.4 Hz), 0.67 (s, 9H), 0.05 (s, 3H), 0.00 (s, 3H). (i) (1 1S, 11aS)-allyl 11 -(tert-butyldimethylsilyloxy)-8-(3-iodopropoxy)-7-methoxy-5-oxo-2- ((E)-prop-1-enyl)- 11, 1 1a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)- carboxylate (49)
Diiodopropane (0.295 g, 1.00 mmol, 5.0 eq) and potassium carbonate (0.028 g, 0.20 mmol, 1.0 eq) were added to a solution of the phenol 12 (0.100 g, 0.020 mmol, 1 .0 eq) in dry acetone (5 mL). The reaction mixture was heated at 60°C for 6 hours when LC/MS showed complete reaction. The reaction mixture was concentrated to dryness under reduced pressure and the residue was purified by flash chromatography (silica gel, 75/25 v/v n- hexane/EtOAc to 50/50 v/v n-hexane/EtOAc) to afford the product as a colourless oil (0.074 g, 56%). LC/MS (3.853 min (ES+)), m/z: 669.0 [M+H]+. 1H NMR (400 MHz, CDCI3) δ 7.26 (s, 1 H), 6.90 (s, 1 H), 6.68 (s, 1 H), 6.24 (d, 1 H, J = 15.3 Hz), 5.87 (d, 1 H, J = 8.9 Hz), 5.78 (m, 1 H), 5.53 (m, 1 H), 5.12 (m, 2H), 4.65 (m, 2H), 4.41 (m, 1 H), 4.1 1 (m, 1 H), 3.93 (s, 3H), 3.81 (m, 1 H), 3.40 (t, 2H, J = 6.7 Hz), 3.05 (dd, 1 H, J = 16.3, 10.1 Hz), 2.57 (m, 1 H), 2.34 (m, 2H), 1 .84 (d, 3H, J = 6.6 Hz), 0.92 (s, 9H), 0.28 (s, 3H), 0.26 (s, 3H).
Example 11
Figure imgf000099_0001
(a) Allyl ((S)- 1 -(((S)-1-((4-((((2-((S)-2-(((ten-butyldimethyl^ 1 -en-
1 -yl)-2,3-dihydro-1 H-pyrrole- 1 -carbonyl)-4-methoxy-5-
((triisopropylsilyl)oxy)phenyl)carbamoyl)oxy)methyl)phenyl)amino)-1-oxopropan-2- yl)amino)-3-methyl- 1 -oxobutan-2-yl)carbamate ( 14)
Triethylamine (0.256 mL, 1 .84 mmol, 2.2 eq) was added to a stirred solution of the amine 7 (0.480 g, 0.835 mmol, 1 .0 eq) and triphosgene (0.089 g, 0.301 mmol, 0.36 eq) in dry THF (15 mL) at 5°C (ice bath). The progress of the isocyanate reaction was monitored by periodically removing aliquots from the reaction mixture and quenching with methanol and performing LCMS analysis. Once the isocyanate reaction was complete a solution of Alloc- Val-Ala-PABOH I5 (0.473 g, 1 .25 mmol, 1.5 eq) and triethylamine (0.174 mL, 1 .25 mmol, 1.5 eq) in dry THF (10 mL) was rapidly added by injection to the freshly prepared isocyanate. The reaction was allowed to stir at 40°C for 4 hours followed by stirring at room temperature overnight. The mixture was concentrated under reduced pressure, and purified by flash chromatography (silica gel, 20/80 v/v n-hexane/EtOAc to 50/50 v/v n- hexane/EtOAc, then 1/99 v/v DCM/MeOH to 5/95 v/v DCM/MeOH) to afford the product as a yellow solid (0.579 g, 71 %). LC/MS (4.468 min (ES+)), m/z: 978.55 [M+H]+. 1H NMR (400 MHz, CDCIs) δ 8.63 (br s, 1 H), 8.42 (s, 1 H), 7.78 (br s, 1 H), 7.53 (d, 2H, J = 8.1 Hz), 7.31 (d, 2H, J = 8.6 Hz), 6.76 (s, 1 H), 6.59 (d, 1 H, J = 7.6 Hz), 6.36 (br s, 1 H), 6.04 (d, 1 H, J = 15.9 Hz), 5.90 (m, 1 H), 5.55 (m, 1 H), 5.33 - 5.21 (m, 3H), 5.10 (s, 2H), 4.66 (m, 2H), 4.57 (dd, 2H, J = 5.6, 1.0 Hz), 3.98 (dd, 1 H, J = 7.3, 6.8 Hz), 3.90 (m, 1 H), 3.81 (m, 1 H), 3.78 (s, 3H), 2.82 (dd, 1 H, J = 15.4, 9.6 Hz), 2.72 (dd, 1 H, J = 15.9, 3.5 Hz), 2.17 (m, 1 H), 1.78 (dd, 3H, J = 6.5, 0.8 Hz), 1 .46 (d, 3H, J = 7.1 Hz), 1 .29 (m, 3H), 1 .1 1 (d, 18H, J = 7.1 Hz), 0.97 (d, 3H, J = 6.8 Hz), 0.92 (d, 3H, J = 6.8 Hz), 0.83 (s, 9H), 0.04 (s, 3H), 0.01 (s, 3H). f Allyl ((S)-1-(((S)-1-((4-((((2-((S)-2-(hydroxymethyl)^
1 H-pyrrole-1 -carbonyl)-4-methoxy-5-
((triisopropylsilyl)oxy)phenyl)carbamoyl)oxy)methyl)phenyl)amino)-1-oxopropan-2- yl)amino)-3-methyl- 1 -oxobutan-2-yl)carbamate ( 15)
The silyl ether 14 (1.49 g, 1 .52 mmol, 1 .0 eq) was dissolved in a 7:1 :1 :2 mixture of acetic acid/ methanol/ tetrahydrofuran/ water (14:2:2:4 mL) and allowed to stir at room
temperature. After 2 hours the reaction was diluted with EtOAc (100 mL), washed sequentially with water, aq. sodium bicarbonate then brine. The organic phase was then dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 100/0 then 99/1 to 92/8 v/v DCM/ MeOH) to afford the product as an orange solid (1.2 g, 92%). LC/MS (3.649 min (ES+)), m/z: 865.44 [M+H]+. 1 H NMR (400 MHz, CDCI3) δ 8.44 (s, 1 H), 8.35 (s, 1 H), 7.69 (br s, 1 H), 7.53 (d, 2H, J = 8.7 Hz), 7.32 (d, 2H, J = 8.3 Hz), 6.78 (s, 1 H), 6.56 (m, 2H), 6.32 (br s, 1 H), 6.05 (d, 1 H, J = 14.9 Hz), 5.90 (m, 1 H), 5.56 (m, 1 H), 5.30 (m, 2H), 5.22 (m, 1 H), 5.10 (d, 2H, J = 3.1 Hz), 4.73 (m, 1 H), 4.64 (m, 1 H), 4.57 (d, 2H, J = 5.8 Hz), 4.01 (m, 1 H), 3.79 (m, 2H), 3.76 (s, 3H), 2.98 (dd, 1 H, J = 16.3, 10.2 Hz), 2.38 (dd, 1 H, J = 16.6, 4.1 Hz), 2.16 (m, 1 H), 1.78 (dd, 3H, J = 6.8, 0.9 Hz), 1.46 (d, 3H, J = 7.1 Hz), 1.29 (m, 3H), 1.1 1 (d, 18H, J = 7.4 Hz), 0.97 (d, 3H, J = 6.7 Hz), 0.92 (d, 3H, J = 6.8 Hz).
(c) (11S, 1 1aS)-4-((S)-2-((S)-2-(allyloxycarbonylamino)-3- methylbutanamido)propanamido)benzyl 11 -hydroxy-7-methoxy-5-oxo-2-((E)-prop-1 -enyl)- 8-(t isopropylsilyloxy)- 1 1, 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)- carboxylate (16)
Dry dimethyl sulfoxide (0.180 g, 2.3 mmol, 3.0 eq) was added dropwise to a solution of oxalyl chloride (0.147 g, 1.1 mmol, 1 .5 eq) in DCM (10 mL) at -78°C under an atmosphere of nitrogen. Maintaining the temperature at -78°C, after 20 minutes, a solution of the primary alcohol 15 (0.666 g, 0.77 mmol, 1.0 eq) in DCM (10 ml.) was added dropwise. After a further 15 minutes, dry triethylamine (0.390 g, 3.85 mmol, 5.0 eq) was added, and the reaction mixture allowed to warm to room temperature. The reaction mixture was washed successively with cold 0.1 N HCI (10 ml_), saturated sodium hydrogen carbonate (10 ml.) and brine (5 ml_). The organic layer was then dried over MgS04, filtered and concentrated under reduced pressure. The residue was then purified by flash
chromatography (silica gel, 50/50 v/v n-hexane/EtOAc to 25/75 v/v n-hexane/EtOAc) to afford the product as a white solid (0.356 g, 54 %). LC/MS (3.487 min (ES+)), m/z: 862.2 [M+H]+. 1 H NMR (400 MHz, CDCI3) δ 8.34 (br s, 1 H), 7.47 (d, 2H, J = 7.6 Hz), 7.17 (s, 1 H), 7.14 (d, 2H, J = 7.5 Hz), 6.86 (br s, 1 H), 6.65 (br s, 1 H), 6.42 (d, 1 H, J = 7.6 Hz), 6.22 (d, 1 H, J = 14.4 Hz), 5.80 (m, 1 H), 5.40 (m, 1 H), 5.53 (m, 1 H), 5.32 (m, 1 H), 5.21 (d, 2H, J = 9.6 Hz), 5.06 (d, 1 H, J = 12.3 Hz), 4.90 (m, 1 H), 4.58 (m, 3H), 3.98 (m, 1 H), 3.84 (m, 1 H), 3.81 (s, 3H), 3.50 (m, 1 H), 3.05 (dd, 1 H, J = 16.0, 10.3 Hz), 2.76 (m, 1 H), 2.15 (m, 1 H), 1.80 (dd, 3H, J = 6.7, 0.8 Hz), 1.44 (d, 3H, J = 7.1 Hz), 1.16 (m, 3H), 1.01 (d, 18H, J = 6.6 Hz), 0.96 (d, 3H, J = 6.8 Hz), 0.92 (d, 3H, J = 6.8 Hz).
(d) (11S, 11aS)-4-((S)-2-((S)-2-(allyloxycarbonylamino)-3- methylbutanamido )propanamido ) benzyl 1 1 -(tert-butyldimethylsilyloxy) - 7-methoxy-5-oxo -2- ((E)-prop-1 -enyl)-8-(t isopropylsilyloxy)-1 1 , 11 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2- a][1,4]diazepine-10(5H)-carboxylate (17)
7ert-butyldimethylsilyltrifluoromethane sulfonate (0.46 g, 1.74mmol, 3.0 eq) was added to a solution of secondary alcohol 16 (0.5 g, 0.58 mmol, 1 .0 eq) and 2,6-lutidine (0.25 g, 2.32 mmol, 4.0 eq) in dry DCM (10 ml.) at 0°C under an atmosphere of nitrogen. After 10 minutes, the reaction mixture was allowed to warm to room temperature and stirred for a further 120 mins. The organic phase was then washed successively with water (10 ml_), saturated sodium bicarbonate (10 ml.) and brine (5 ml_), dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 50/50 v/v n-hexane/EtOAc) to afford the product as a white solid (0.320 g, 57 %). LC/MS (4.415 min (ES+)), m/z: 976.52 [M+H]+. 1 H NMR (400 MHz, CDCI3) δ 8.31 (br s, 1 H), 7.48 (d, 2H, J = 8.0 Hz), 7.21 (s, 1 H), 7.14 (d, 2H, J = 8.3 Hz), 6.89 (s, 1 H), 6.65 (s, 1 H), 6.38 (d, 1 H, J = 7.3 Hz), 6.25 (d, 1 H, J = 14.6 Hz), 5.93 (m, 1 H), 5.85 (d, 1 H, J = 8.8 Hz), 5.50 (m, 1 H), 5.34 (m, 1 H), 5.24 (m, 2H), 5.15 (d, 1 H, J = 12.5 Hz), 4.86 (d, 1 H, J = 12.2 Hz), 4.62 (m, 3H), 4.01 (m, 1 H), 3.86 (s, 3H), 3.78 (m, 1 H), 3.04 (m, 1 H), 2.56 (m, 1 H), 2.20 (m, 1 H), 1 .84 (dd, 3H, J = 6.6, 0.7 Hz), 1.48 (d, 3H, J = 6.8 Hz), 1 .20 (m, 3H), 1.05 (d, 9H, J = 2.9 Hz), 1 .03 (d, 9H, J = 2.9 Hz), 0.99 (d, 3H, J = 6.8 Hz), 0.95 (d, 3H, J = 6.8 Hz), 0.88 (s, 9H), 0.27 (s, 3H), 0.14 (s, 3H).
(e) (11S, 11aS)-4-((S)-2-((S)-2-(allyloxycarbonylamino)-3- methylbutanamido )propanamido ) benzyl 1 1 -(tert-butyldimethylsilyloxy) -8-hydroxy- 7- methoxy-5-oxo-2-((E)-prop-1 -enyl)-1 1 , 1 1 a-dihydro-1 H-benzo[e]pyrrolo[1 ,2- a][1 ,4]diazepine-10(5H)-carboxylate (18)
Lithium acetate dihydrate (0.010 g, 0.10 mmol, 1.0 eq) was added to a solution of the silyl ether 37 (0.100 g, 0.10 mmol, 1 .0 eq) in wet DMF (2 mL) at 25°C for 3 hours. The reaction mixture was then diluted with ethyl acetate (20 mL) and washed successively with 0.1 M citric acid (20 mL, pH 3), water (20 mL) and brine (5 mL), dried over MgS04, filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (silica gel, 5/95 v/v methanol/DCM) to afford the product as a pale yellow oil (0.070 g, 83 %). LC/MS (3.362 min (ES+)), m/z: 820.2 [M+H]+. 1H NMR (400 MHz, CDCI3) δ 8.39 (s, 1 H), 7.48 (d, 2H, J = 8.2 Hz), 7.25 (s, 1 H), 7.12 (d, 2H, J = 8.1 Hz), 6.88 (s, 1 H), 6.68 (s, 1 H), 6.47 (d, 1 H, J = 7.6 Hz), 6.24 (d, 1 H, J = 15.2 Hz), 6.03 (s, 1 H), 5.92 (m, 1 H), 5.84 (d, 1 H, J = 8.9 Hz), 5.50 (m, 1 H), 5.34 (m, 1 H), 5.26 (m, 2H), 5.18 (d, 1 H, J = 12.3 Hz), 4.80 (d, 1 H, J = 12.4 Hz), 4.66 - 4.60 (m, 3H), 4.02 (m, 1 H), 3.95 (s, 3H), 3.81 (m, 1 H), 3.03 (m, 1 H), 2.57 (m, 1 H), 2.19 (m, 1 H), 1.84 (dd, 3H, J = 6.8, 0.8 Hz), 1.48 (d, 3H, J = 7.1 Hz), 1.00 (d, 3H, J = 6.8 Hz), 0.95 (d, 3H, J = 6.8 Hz), 0.87 (s, 9H), 0.26 (s, 3H), 0.12 (s, 3H).
xample 12
Figure imgf000103_0001
(11S, 11aS)-allyl 8-(3-((11S, 11aS)-10-((4-((R)-2-((R)-2-(allyloxycarbonylamino)-3- methylbutanamido )propanamido )benzyloxy)carbonyl) - 1 1 -(tert-butyldimethylsilyloxy) - 7- methoxy-5-oxo-2-((E)-prop-1-enyl)-5, 10, 1 1, 1 1 a-tetrahydro-1 H-benzo[e]pyrrolo[1 ,2- a][1 ,4]diazepin-8-yloxy)propoxy)-11 -(tert-butyldimethylsilyloxy)-7-methoxy-5-oxo-2-((E)- prop-1-enyl)- 11, 1 1 a-dihydro- 1 H-benzo[e]pyrrolo[1 ,2-a][1 ,4]diazepine-10(5H)-carboxylate (50)
Potassium carbonate (0.030 g, 0.21 mmol, 1.0 eq) was added to a solution of the phenol 18 (0.175 g, 0.21 mmol, 1.0 eq) and the iodo linker 49 (0.214 g, 0.32 mmol, 1 .5 eq) in acetone (10 mL). The reaction mixture was heated under a nitrogen atmosphere at 75°C in a sealed flask for 17 hours. The reaction mixture was concentrated to dryness under reduced pressure and purified by flash chromatography (silica gel, 2/98 v/v methanol/DCM to 5/95 v/v methanol/DCM) to afford the product as a pale yellow solid (0.100 g, 35%). LC/MS (4.293 min (ES+)), m/z: 1359.13 [M]+.

Claims

1. A compound of formula I:
Figure imgf000104_0001
wherein:
R7 is selected from: ORA, where RA is selected from C1-4 saturated alkyl, optionally substituted by phenyl, which may bear a chloro substituent, pyridyl and furanyl; chloro; NH2; -CH2-0-C(=0)Me;
R8 is OProt°, where Prot° is a silicon-based oxygen protecting group orthogonal to Rc; R9 is selected from H, methyl and methoxy;
Rs is selected from CF3, (CF2)3CF3, CH3 and
Figure imgf000104_0002
;
and
Rc is selected from:
(i) -C(=0)-ORc1 , where RC1 is a saturated Ci-4 alkyl group;
(ii) -CH2-0-C(=0)RC2, where RC2 is methyl or phenyl;
(iii) -CH2-0-Si-(RSi1)(RSi2)(RSi3), where RSi1 , RSi2, RSi3 are independently selected from d-6 saturated alkyl group and a phenyl group; and
(iv) -C(-YRC3)(-YRC4) where each Y is independently O or S, and where RC3 and RC4 are independently a saturated Ci-4 alkyl group, or together form a C2-3 alkylene.
2. The compound according to claim 1 , which is is substantially free of any of the corresponding compound of formula IB:
Figure imgf000104_0003
3. A compound according to either claim 1 or claim 2, wherein R7 is ORA, where RA is selected from C1-4 saturated alkyl, optionally substituted by phenyl, pyridyl and furanyl.
4. A compound according to any one of claims 1 to 3, wherein RA is unsubstituted C1-4 saturated alkyl.
5. A compound according to claim 4, wherein RA is methyl.
6. A compound according to any one of claims 1 to 5, wherein Rs is CF3.
7. A compound according to any one of claims 1 to 6, wherein RC1 is methyl.
8. A compound according to any one of claims 1 to 7, wherein -Si-(RSl1)(RSl2)(RSl3) is TBDMS.
9. A method of synthesising a compound of formula I :
as defined in any one of rmula II:
Figure imgf000105_0001
comprising treating II with the appropriate anhydride and anhydrous 2,6-lutidine or anhydrous 2,6-fBu-pyridine at a temperature of -35°C or lower in a dry organic solvent under a inert atmosphere.
10. The method according to claim 9, wherein the reaction is carried out at a temperature of -40°C or lower. A compound of formul
Figure imgf000106_0001
wherein Rc, R7, R8 and R9 are as defined in any one of claims 1 to 8;
R2 is selected from:
(i) an optionally substituted C5-20 aryl group;
(ii) an optionally substituted Ci-5 alkyl group; and
(iii) H.
12. A compound according to claim 1 1 , which is substantially free of any of the corresponding compound of formula 1MB:
Figure imgf000106_0002
A method of synthesising a compound of formula
Figure imgf000106_0003
as defined in any one of claims 1 to 8 by:
(i) a Suzuki coupling between a compound of formula I and an appropriate aryl boron derivative; (ii) a Heck coupling with an appropriate alkene, acrylamide or acrylate;
(iii) a Stille coupling with an appropriate organotin reagent;
(iv) a Sonagishira coupling with an appropriate alkyne; or
(v) a hydride transfer using a triethylsilane.
14. A compound of formula IV:
Figure imgf000107_0001
wherein R7, R8 and R9 are as defined in any one of claims 1 to 8, R2 is as defined in claim 1 1 ;
RD is selected from:
(ii) -CH2-0-C(=0)RC2, where RC2 is methyl or phenyl;
(iii) -CH2-0-Si-(RSi1)(RSi2)(RSi3), where RSi1 , RSi2, RSi3 are independently selected from C1-6 saturated alkyl group and a phenyl group; and
(iv) -C(-YRC3)(-YRC4) where each Y is independently O or S, and where RC3 and RC4 are independently a saturated Ci-4 alkyl group, or together form a C2-3 alkylene.
15. A compound according to claim 14,
which is substantially free of any of the corresponding compound of formula IVB:
Figure imgf000107_0002
16. A compound according to either claim 14 or claim 15, wherein -Si-(RSi1)(RSi2)(RSi3) is TBDMS.
17. A compound according to either claim 14 or claim 15, wherein R is methyl.
18. A method of synthesising a compound of formula IV:
Figure imgf000108_0001
defined in any one of claims 14 to 17, from a compound of formula
Figure imgf000108_0002
as defined in claim 12, by:
(i) if Rc is -C(=0)-ORc1 , conversion of -C(=0)-ORc1 to -CH2-0-C(=0)RC2 or -CH2-0-Si- (RSi1)(RSi2)(RSi3), followed by:
(ii) if Rc is -CH2-0-C(=0)RC2, reduction of the nitro group by zinc in acetic acid or if Rc is -CH2-0-Si-(RSi1)(RSi2)(RSi3), reduction of the nitro group using zinc in 5% formic acid in ethanol;
(iii) if Rc is -C(-YRC3)(-YRC4), reduction of the nitro group using Cd/Pb couple, sodium dithionite or tin II chloride.
A compound of formula V
Figure imgf000108_0003
wherein R7, R8 and R9 are as defined in any one of claims 1 to 8, R2 is as defined in claim 1 1 and RD is as defined in any one of claims 14 to 17,
R10 is a carbamate-based nitrogen protecting group, which is palladium-labile and/or labile under the conditions used to add the group:
Figure imgf000108_0004
or contains a moiety which is palladium-labile and/or is labile under the conditions used to add the group:
Figure imgf000109_0001
20. A compound according to claim 19, which is substantially free of any of the corresponding compound of formula VB:
Figure imgf000109_0002
A compound according to either claim 19 or 20, wherein R10 is selected from one of:
Figure imgf000109_0003
Figure imgf000110_0001
22. A compound according to eit 20, wherein R10 is of the formula:
Figure imgf000110_0002
where the asterisk indicates the point of attachment to the N10 position, G1 is a functional group to form a connection to a cell binding agent, L1 is a cleavable linker, L2 is a covalent bond or together with -OC(=0)- forms a self-immolative linker.
23. A method of synthesising
Figure imgf000110_0003
as defined in any one of claims 19 to 22, from a compound of formula IV:
Figure imgf000111_0001
as defined in any one of claims 14 to 17, by reaction of IV with triphosgene to obtain isocyanate followed by reaction with R10-OH.
24. A compound of formula
Figure imgf000111_0002
wherein R7, R8 and R9 are as defined in any one of claims 1 to 8, R2 is as defined in claim 1 1 , RD is as defined in any one of claims 14 to 17, and
R10 is as defined in any one of claims 19 to 22.
25. A compound according to claim 24, which is substantially free of any of the corresponding compound of formula VIB:
Figure imgf000111_0003
A method of synthesising a compound of formula VI
Figure imgf000111_0004
as defined in either claim 24 or claim 25, from a compound of formula V:
Figure imgf000112_0001
as defined in any one of claims 19 to 22, by:
(a) (i) where RD is -CH2-0-C(=0)Me, removal of the acetate protecting group, with potassium carbonate in aqueous methanol, or with lithium triethylborohydride;
(ii) where RD is -CH2-0-Si-(RSi1)(RSi2)(RSi3), removal of the silyl ether protecting group using: TBAF in THF; acetic acid in aqueous THF; CsF in DMF; or HF in pyridine; and
(b) oxidation of the product of step (a); or
(c) where RD is -C(-YRC3)(-YRC4), removal of the acetal or thioacetal protecting groups, respectively with acid or reaction with Hg (II) salts.
27. A method according to claim 26, wherein the oxidation is carried out with
Dess-Martin periodinane.
28. A compound of formula
Figure imgf000112_0002
vii
wherein R7, R8 and R9 are as defined in any one of claims 1 to 8, R2 is as defined in claim 1 1 , R10 is as defined in any one of claims 19 to 22, and
RE is selected from the group consisting of: 0-C1-2 alkyl; 0-Si-(RSi1)(RSi2)(RSi3), where RSi1 , RSi2, RSi3 are as defined above; 0-CH2OCH3 (OMOM), 0-CH2OC2H4OCH3 (OMEM), OC5H10 (THP).
29. A compound according to claim 28, which is substantially free of any of the corresponding compound of formula VIIB:
Figure imgf000113_0001
VIIB
A compound according to either claim 28 or claim 29, wherein RE is O-TBS
A method of synthesising a compound of formula VII
Figure imgf000113_0002
VII
as defined in any one of claims d of formula VI:
Figure imgf000113_0003
as defined in either claims 24 to 25, by:
by adding a protecting group to OH to yield RE.
A compound of formula
Figure imgf000113_0004
VIII
wherein R7 and R9 are as defined in any one of claims 1 to 8, R2 is as defined in claim 1 1 , R10 is as defined in any one of claims 19 to 22, and RE is as defined in any one of claims 28 to 30.
33. A compound according to claim 32, which is substantially free of any of the corresponding compound of formula VIIIB:
Figure imgf000114_0001
VIIIB
A method of synthesisin II
Figure imgf000114_0002
VIII
as defined in either claim 32 or claim 33, from a compound of formula VII:
Figure imgf000114_0003
VII
as defined in any one of claims 28 to 30, by:
removing the protecting group in R8 to yield OH.
35. A compound of formula IX:
Figure imgf000114_0004
IX wherein R7 and R9 are as defined in any one of claims 1 to 8, R2 is as defined in claim 1 1 , R10 is as defined in any one of claims 19 to 22, RE is as defined in any one of claims x to y, n is from 1 to 12 and Hal is selected from I, CI and Br.
36. A compound according to claim 35, which is substantially free of any of the corresponding compound of formula IXB:
Figure imgf000115_0001
IXB
A compound according to either claim 35 or 36, wherein Hal is I.
A method of synthesising a compound of formula IX:
Figure imgf000115_0002
IX
as defined in any one of claims of formula VIII:
Figure imgf000115_0003
VIII
as defined in either claim 32 or claim 33, by reacting with a compound of formula X:
Hal-^ wherein Q is selected from I, CI and Br.
39. A method according to claim 38, wherein Q is I.
A compound of formula XI
Figure imgf000116_0001
XI
wherein R7 and R9 are as defined in any one of claims 1 to 8, R2 is as defined in claim 1 1 , R10 is as defined in any one of claims 19 to 22, RE is as defined in any one of claims 28 to 30, and where R2 , R7 , R9 , R10 and RE are independently selected from the same groups as R2, R7, R9, R10 and RE respectively.
A method of synthesising a compound of formula XI
Figure imgf000116_0002
XI
As defined in claim 40, comprising coupling compounds of formulae VIII and IX:
Figure imgf000116_0003
VIE ix
as defined in either claim 32 or claim 33 and any one of claims 35 to 37 respectively.
42. The method according to claim 41 , wherein the reaction is carried out in refluxing acetone in the presence of K2C03.
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EA029046B1 (en) 2018-02-28
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CN104011018A (en) 2014-08-27
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