EP0921801A1 - Inhibiteurs de la farnesyle-proteine transferase - Google Patents

Inhibiteurs de la farnesyle-proteine transferase

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Publication number
EP0921801A1
EP0921801A1 EP97917116A EP97917116A EP0921801A1 EP 0921801 A1 EP0921801 A1 EP 0921801A1 EP 97917116 A EP97917116 A EP 97917116A EP 97917116 A EP97917116 A EP 97917116A EP 0921801 A1 EP0921801 A1 EP 0921801A1
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European Patent Office
Prior art keywords
substituted
alkyl
aryl
unsubstituted
cycloalkyl
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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EP97917116A
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German (de)
English (en)
Inventor
Dong D. Wei
Theresa M. Williams
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Merck and Co Inc
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Merck and Co Inc
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Priority claimed from GBGB9613460.6A external-priority patent/GB9613460D0/en
Application filed by Merck and Co Inc filed Critical Merck and Co Inc
Publication of EP0921801A1 publication Critical patent/EP0921801A1/fr
Withdrawn legal-status Critical Current

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    • 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
    • C07D401/06—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 carbon chain containing only aliphatic carbon atoms
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/12—Drugs for disorders of the urinary system of the kidneys
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/06—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms

Definitions

  • Ras proteins are part of a signalling pathway that links cell surface growth factor receptors to nuclear signals initiating cellular proliferation.
  • Biological and biochemical studies of Ras action indicate that Ras functions like a G-regulatory protein.
  • Ras In the inactive state, Ras is bound to GDP.
  • Ras Upon growth factor receptor activation Ras is induced to exchange GDP for GTP and undergoes a conformational change.
  • the GTP- bound form of Ras propagates the growth stimulatory signal until the signal is terminated by the intrinsic GTPase activity of Ras, which returns the protein to its inactive GDP bound form (D.R. Lowy and D.M.
  • Mutated ras genes ( a-ras, Ki4a-ra ⁇ , K ⁇ 4b-ras and N-ras) are found in many human cancers, including colorectal carcinoma, exocrine pancreatic carcinoma, and myeloid leukemias. The protein products of these genes are defective in their GTPase activity and constitutively transmit a growth stimulatory signal.
  • Ras must be localized to the plasma membrane for both normal and oncogenic functions. At least 3 post-translational modifications are involved with Ras membrane localization, and all 3 modifications occur at the C-terminus of Ras.
  • the Ras C-terminus contains a sequence motif termed a "CAAX” or "Cys-Aaa ⁇ -Aaa ⁇ -Xaa” box (Cys is cysteine, Aaa is an aliphatic amino acid, the Xaa is any amino acid) (Willumsen et al.. Nature 3/0:583-586 (1984)).
  • this motif serves as a signal sequence for the enzymes farnesyl-protein transferase or geranylgeranyl-protein transferase, which catalyze the alkylation of the cysteine residue of the CAAX motif with a C 1 5 or C20 isoprenoid, respectively.
  • farnesyl-protein transferase or geranylgeranyl-protein transferase which catalyze the alkylation of the cysteine residue of the CAAX motif with a C 1 5 or C20 isoprenoid, respectively.
  • farnesylated proteins include the Ras-related GTP- binding proteins such as Rho, fungal mating factors, the nuclear lamins, and the gamma subunit of transducin. James, et al., J. Biol Chem. 269, 14182 (1994) have identified a peroxisome associated protein Pxf which is also farnesylated. James, et al., have also suggested that there are farnesylated proteins of unknown structure and function in addition to those listed above.
  • Famesyl-protein transferase utilizes farnesyl pyrophosphate to covalently modify the Cys thiol group of the Ras CAAX box with a farnesyl group (Reiss et al, Cell, 62:81 -88 (1990); Schaber et al. J. Biol. Chem., 265: 14701-14704 (1990); Schafer et al, Science, 249: 1133-1139 (1990); Marine et al, Proc. Natl Acad. Sci USA, 87:154 ⁇ -1545 (1990)).
  • Inhibition of farnesyl pyrophosphate biosynthesis by inhibiting HMG-CoA reductase blocks Ras membrane localization in cultured cells.
  • direct inhibition of famesyl- protein transferase would be more specific and attended by fewer side effects than would occur with the required dose of a general inhibitor of isoprene biosynthesis.
  • FPTase famesyl-protein transferase
  • FPP farnesyl diphosphate
  • Ras protein substrates
  • the peptide derived inhibitors that have been described are generally cysteine containing molecules that are related to the CAAX motif that is the signal for protein prenylation.
  • Such inhibitors may inhibit protein prenylation while serving as altemate substrates for the famesyl-protein transferase enzyme, or may be purely competitive inhibitors (U.S. Patent 5,141 ,851 , University of Texas; N.E. Kohl et al, Science, 260: 1934- 1931 (1993); Graham, et al., J. Med. Chem., 37, 725 (1994)).
  • deletion of the thiol from a CAAX derivative has been shown to dramatically reduce the inhibitory potency of the compound.
  • the thiol group potentially places limitations on the therapeutic application of FPTase inhibitors with respect to pharmacokinetics, pharmacodynamics and toxicity. Therefore, a functional replacement for the thiol is desirable.
  • famesyl-protein transferase inhibitors are inhibitors of proliferation of vascular smooth muscle cells and are therefore useful in the prevention and therapy of arteriosclerosis and diabetic disturbance of blood vessels (JP H7- 1 12930).
  • an object of this invention to develop peptidomimetic compounds that do not have a thiol moiety, and that will inhibit famesyl-protein transferase and thus, the post-translational famesylation of proteins. It is a further object of this invention to develop chemotherapeutic compositions containing the compounds of this invention and methods for producing the compounds of this invention.
  • the present invention comprises peptidomimetic piperazine- or piperazinone-containing compounds which inhibit the famesyl-protein transferase.
  • the instant compounds lack a thiol moiety and thus offer unique advantages in terms of improved pharmacokinetic behavior in animals, prevention of thiol-dependent chemical reactions, such as rapid autoxidation and disulfide formation with endogenous thiols, and reduced systemic toxicity.
  • the compounds of this invention are useful in the inhibition of famesyl-protein transferase and the famesylation of the oncogene protein Ras.
  • the inhibitors of famesyl-protein transferase are illustrated by the formula A:
  • Rla and R 1D are independently selected from: a) hydrogen, b) aryl, heterocycle, C3-C10 cycloalkyl, C2-C6 alkenyl, C2- C6 alkynyl, RlOO-, Rl lS(0) m -, R 10 C(O)NRl0-, CN(R10)2NC(OX R 10 2N-C(NR 10 )-, CN, N02, RlOc(O)-, N3, -N(RlO)2, orR 1 l ⁇ C(O)NRl0-, c) unsubstituted or substituted C1-C6 alkyl wherein the substitutent on the substituted Cl -C6 alkyl is selected from unsubstituted or substituted aryl, heterocyclic, C3-C10 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, R ⁇ O-, R ⁇ S(O)m-,Rl0C
  • R2 and R3 are independently selected from: H; unsubstituted or substituted Cl-8 alkyl, unsubstituted or substituted C2-8 alkenyl, unsubstituted or substituted C2-8 alkynyl, unsubstituted or substituted aryl,
  • halogen e) CN, f) aryl or heteroaryl, g) perfluoro-C l -4 alkyl, h) SR6 , S(0)R6a, S ⁇ 2R 6a ,
  • R2 and R ⁇ are attached to the same C atom and are combined to form (CH2)u - wherein one of the carbon atoms is optionally replaced by a moiety selected from: O, S(0) m , -NC(O)-, and -N(COR lO)- ;
  • R 4 is selected from H and CH3;
  • R6, R7 and R 7a are independently selected from: H; Cl-4 alkyl, C3-6 cycloalkyl, heterocycle, aryl, aroyl, heteroaroyl, arylsulfonyl, heteroarylsulfonyl, unsubstituted or substituted with: a) Cl-4 alkoxy, b) aryl or heterocycle, c) halogen, d) HO,
  • R6 and R 7 may be joined in a ring; R7 and R ⁇ may be joined in a ring;
  • R oa is selected from: Cl-4 alkyl, C3-6 cycloalkyl, heterocycle, aryl, unsubstituted or substituted with: a) Cl -4 alkoxy, b) aryl or heterocycle, c) halogen, d) HO, pi "1 e » ⁇ O
  • R8 is independently selected from: a) hydrogen, b) aryl, heterocycle, C3-C10 cycloalkyl, C2-C6 alkenyl, C2- C6 alkynyl, perfluoroalkyl, F, Cl, Br, RlOO-, R l l s(0) m - RlOC(0)NRlO-, (RlO) 2 NC(0)-, R!0 2 N-C(NR10)-, CN, N02, R 10 C(O)-, N3, -N(RlO)2, or Rl l ⁇ C(O)NRl0-, and c) Ci-C ⁇ alkyl unsubstituted or substituted by aryl, cyanophenyl, heterocycle, C3-C10 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, perfluoroalkyl, F, Cl, Br, RlOO-,
  • R9 is selected from: a) hydrogen, b) alkenyl, alkynyl, perfluoroalkyl, F, Cl, Br, RlOO-, 1 ⁇ (O)TM-, Rl C(0)NRlO-, (RlO) 2 NC(0)-, RIO 2 N- C(NRlO)-, CN, N02, R 10 C(O)-, N3, -N(RlO) 2 , or RllOC(O)NRl0-, and c) C1-C6 alkyl unsubstituted or substituted by perfluoroalkyl, F, Cl, Br, RlOo-, Rl lS(0) m -, R!0C(O)NR 0-, (Rl )2NC(0)-, Rl ⁇ 2N-C(NRlO)-, CN, RlOC(O)-, N3, -N(RlO)2, or Rl l ⁇ C(O)NRl0-;
  • RlO is independently selected from hydrogen, C1-C6 alkyl, benzyl and aryl;
  • Rl 1 is independently selected from C1-C6 alkyl and aryl
  • G is H2 or O
  • V is selected from: a) hydrogen, b) heterocycle, c) aryl, d) C1 -C20 alkyl wherein from 0 to 4 carbon atoms are replaced with a a heteroatom selected from O, S, and N, and e) C2-C20 alkenyl, provided that V is not hydrogen if Al is S(0)m and V is not hydrogen if A 1 is a bond, n is 0 and A ⁇ is S(0)m;
  • W is a heterocycle
  • Rla and Rib are independently selected from: a) hydrogen, b) aryl, heterocycle, C3-C10 cycloalkyl, C2-C6 alkenyl, C2- C6 alkynyl, RlOO-, RHS(0) m -, R 10 C(O)NRl0-, (Rl0)2NC(O)-, Rl ⁇ 2N-C(NRlO)-, CN, N02, Rl°C(0)-, N3, -N(RlO)2, or Rl l ⁇ C(O)NRl0-, c) unsubstituted or substituted C1-C6 alkyl wherein the substitutent on the substituted C1-C6 alkyl is selected from unsubstituted or substituted aryl, heterocyclic, C3-C10 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, Rl O-, R 1 'S ⁇ m-, Rl0C(O)NR
  • R2 and R3 are independently selected from: H; unsubstituted or substituted Cl-8 alkyl, unsubstituted or substituted C2-8 alkenyl, unsubstituted or substituted C2-8 alkynyl, unsubstituted or substituted aryl,
  • R2 and R3 are attached to the same C atom and are combined to form (CH2)u - wherein one of the carbon atoms is optionally replaced by a moiety selected from: O, S(0) m , -NC(O)-, and -N(CORlO)- ;
  • R4 is selected from H and CH3;
  • R D , R 7 and R 7a are independently selected from: H; Cl-4 alkyl, C3-6 cycloalkyl, heterocycle, aryl, aroyl, heteroaroyl, arylsulfonyl, heteroarylsulfonyl, unsubstituted or substituted with: a) Cl -4 alkoxy, b) aryl or heterocycle, c) halogen, d) HO, i n e) o
  • R6 and R 7 may be joined in a ring; R 7 and R 7a may be joined in a ring;
  • R8 is independently selected from: a) hydrogen, b) aryl, heterocycle, C3-C10 cycloalkyl, C2-C6 alkenyl, C2- C6 alkynyl, perfluoroalkyl, F, Cl, Br, R lOo-, R l l S(0) m -, Rl0C(O)NRl0-, (RlO) 2 NC(0)-, R!0 2 N-C(NR10)-, CN, N02, R 10 C(O)-, N3, -N(RlO) 2 , or RHOC(0)NR10-, and c) C1-C6 alkyl unsubstituted or substituted by aryl, cyanophenyl, heterocycle, C3-C10 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, perfluoroalkyl, F, Cl, Br, RlOO-,
  • R9 is selected from: a) hydrogen, b) C2-C6 alkenyl, C2-C6 alkynyl, perfluoroalkyl, F, Cl, Br, RlOo-, RllS(O) m -,Rl0C(O)NRl0-, (RlO) 2 NC(0)-, Rl0 N-C(NRlO)-, CN, N02, Rl°C(0)-, N3, -N(RlO) 2 , or RllOC(O)NRl0-, and c) C1-C6 alkyl unsubstituted or substituted by perfluoroalkyl, F, Cl, Br, RlOo-, Rl lS(0) m -, Rl°C(O)NRl -, (Rl0)2NC(O)-, Rl0 2 N-C(NRlO)-, CN, RlOC(O)-, N3, -N(RlO)2, or Rl lOC
  • RlO is independently selected from hydrogen, C1-C6 alkyl, benzyl and aryl;
  • Rl 1 is independently selected from Cl-C ⁇ alkyl and aryl
  • V is selected from: a) hydrogen, b) heterocycle, c) aryl, d) C l -C20 alkyl wherein from 0 to 4 carbon atoms are replaced with a a heteroatom selected from O, S, and N, and e) C2-C20 alkenyl, provided that V is not hydrogen if Al is S(0)m and V is not hydrogen if A 1 is a bond, n is 0 and A 2 is S(0)m;
  • W is a heterocycle
  • Z is unsubstituted C1-C alkyl, substituted C1-C6 alkyl, unsubstituted C3-C6 cycloalkyl or substituted C3-C6 cycloalkyl, wherein the substituted C1 -C6 alkyl and substituted C3-C6 cycloalkyl is substituted with one or two of the following: a) Cl -4 alkoxy, c) C3-6 cycloalkyl, d) -NR°C(0)R 7 , e) HO, f) -S(0)mR° , g) halogen, or h) perfluoroalkyl;
  • the inhibitors of famesyl-protein transferase are illustrated by the formula A:
  • R la is independently selected from: hydrogen or C1 -C6 alkyl
  • Rib is independently selected from: a) hydrogen, b) aryl, heterocycle, cycloalkyl, R 10 ⁇ -, -N(R 10) or C2-C6 alkenyl, c) unsubstituted or substituted C1-C6 alkyl wherein the substitutent on the substituted C1 -C6 alkyl is selected from unsubstituted or substituted aryl, heterocycle, cycloalkyl, alkenyl, R lOO- and -N(RlO) ;
  • R3 and R4 are independently selected from H and CH3;
  • R2 is H; O or C ⁇ _5 alkyl, unbranched or branched, unsubstituted or substituted with one or more of: 1 ) aryl, 2) heterocycle,
  • R°, R 7 and R 7a are independently selected from:
  • H Cl-4 alkyl, C3-6 cycloalkyl, aryl, heterocycle, unsubstituted or substituted with: a) Cl-4 alkoxy, b) halogen, or c) aryl or heterocycle;
  • R ⁇ i selected from:
  • R8 is independently selected from: a) hydrogen, b) C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 perfluoroalkyl, F, Cl, RlOO-, R!0C(O)NR10-, CN, NO2, (Rl0) 2 N-C(NRl0)-, Rl0c(O)-, -N(RlO) 2 , or Rll ⁇ C(O)NRl0-, and c) C1-C6 alkyl substituted by Cj-C ⁇ perfluoroalkyl, RlOO-,
  • R9 is selected from: a) hydrogen, b) C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 perfluoroalkyl, F, Cl. RlOO-, Rl lS(0) m -, R1°C(0)NR10-, CN, NO2, (RlO) 2 N-C(NRlO)-, RlOC(O)-, -N(RlO)2, or Rll ⁇ C(O)NRl0-, and c) C l -C6 alkyl unsubstituted or substituted by C 1 -C6 perfluoroalkyl, F, Cl, RlOO-, RUS(0) m -, R1°C(0)NR10-, CN, (RlO) N-C(NRlO)-, RIO O)-, -N(R10) 2 , or RllOC(O)NRl0-;
  • RlO is independently selected from hydrogen, Cl-C6 alkyl, benzyl and aryl;
  • R 1 is independently selected from C1-C6 alkyl and aryl
  • V is selected from: a) hydrogen, b) heterocycle selected from pyrrolidinyl, imidazolyl, pyridinyl, thiazolyl, pyridonyl, 2-oxopiperidinyl, indolyl, quinolinyl, isoquinolinyl, and thienyl, c) aryl, d) C1-C20 alkyl wherein from 0 to 4 carbon atoms are replaced with a a heteroatom selected from O, S, and N, and e) C2-C2O alkenyl, and provided that V is not hydrogen if Al is S(0)m and V is not hydrogen if A 1 is a bond, n is 0 and A 2 is S(0)m;
  • G is H2 or O
  • W is a heterocycle selected from pyrrolidinyl, imidazolyl, pyridinyl. thiazolyl, pyridonyl, 2-oxopiperidinyl, indolyl, quinolinyl, or isoquinolinyl;
  • Z is unsubstituted C1-C6 alkyl, substituted Cl -C ⁇ alkyl, unsubstituted C3-C6 cycloalkyl or substituted C3-C6 cycloalkyl,wherein the substituted C1 -C6 alkyl and substituted C3-C6 cycloalkyl is substituted with one or two of the following: a) Cl -4 alkoxy, b) NR6R 7 , c) C3-6 cycloalkyl, d) -NR°C(0)R 7 , e) HO, f) -S(0) m R 6a , g) halogen, or h) perfluoroalkyl;
  • R l a is selected from: hydrogen or C l -C ⁇ alkyl
  • R i b is independently selected from: a) hydrogen, b) aryl, heterocycle, cycloalkyl, R 10 ⁇ -, -N(R 10)2 or C2-C6 alkenyl, c) C ⁇ -C ⁇ alkyl unsubstituted or substituted by aryl, heterocycle, cycloalkyl, alkenyl, R l °0-, or -N(R l°)2;
  • R3 is selected from H and CH3;
  • R2 is selected from H; O or Cl -5 alkyl, unbranched or branched, unsubstituted or substituted with one or more of:
  • R6 and R 7 are independently selected from:
  • R ⁇ is independently selected from: a) hydrogen, b) C l -C ⁇ alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C l -C ⁇ perfluoroalkyl, F, Cl, RlOO-, R1°C(O)NR10-, CN, NO2, (Rl0) 2 N-C(NRl0)-, RlOc(O)-, -N(RlO)2, or
  • RHOC(O)NR10- and c) C1-C6 alkyl substituted by C1-C6 perfluoroalkyl, Rl°0-, Rl0C(O)NRl0-, (R10) 2 N-C(NR1 , RlOC(O)-, -N(Rl0)2, or Rl l ⁇ C(O)NRl0-;
  • R i hydrogen or methyl
  • RlO is independently selected from hydrogen, -C6 alkyl, benzyl and aryl;
  • Rl 1 is independently selected from C1-C6 alkyl and aryl
  • V is selected from: a) hydrogen, b) heterocycle selected from pyrrolidinyl, imidazolyl, pyridinyl, thiazolyl, pyridonyl, 2-oxopiperidinyl, indolyl, quinolinyl, isoquinolinyl, and thienyl, c) aryl, d) C l -C20 alkyl wherein from 0 to 4 carbon atoms are replaced with a a heteroatom selected from O, S, and N, and e) C2-C20 alkenyl, and provided that V is not hydrogen if Al is S(0)m and V is not hydrogen if A 1 is a bond, n is 0 and A is S(0) m ;
  • Z is unsubstituted Cl -C6 alkyl, substituted Cl -C6 alkyl, unsubstituted C3-C6 cycloalkyl or substituted C3-C6 cycloalkyl, wherein the substituted C1 -C6 alkyl and substituted C3-C6 cycloalkyl is substituted with one or two of the following: a) Cl -4 alkoxy, b) NR°R 7 , c) C3-6 cycloalkyl, d) -NR6C(0)R 7 , e) HO, f) -S(0) m R 6a , g) halogen, or h) perfluoroalkyl;
  • n 0, 1 , 2, 3 or 4
  • p 0, 1 , 2, 3 or 4
  • r 0 to 5, provided that r is 0 when V is hydrogen
  • R i b is independently selected from: a) hydrogen, b) aryl, heterocycle, cycloalkyl, R l°0-, -N(R 0)2 or C2-C6 alkenyl, c) Cl-C6 alkyl unsubstituted or substituted by aryl, heterocycle, cycloalkyl, alkenyl, RlOO-, or -N(R 1 °)2;
  • R3 is selected from H and CH3;
  • R2 is selected from H; O or C 1 -5 alkyl, unbranched or branched, unsubstituted or substituted with one or more of:
  • R2 and R ⁇ are optionally attached to the same carbon atom
  • R6 and R 7 are independently selected from: H; Cl-4 alkyl, C3-6 cycloalkyl, aryl, heterocycle, unsubstituted or substituted with: a) Cl-4 alkoxy, b) halogen, or c) aryl or heterocycle;
  • R6 i selected from:
  • R ⁇ is independently selected from: a) hydrogen, b) Cl -C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, Cl -C ⁇ perfluoroalkyl, F, Cl, Rl O-, R!0C(O)NR10-, CN, N02, (Rl0) 2 N-C(NRl0)-, Rl C(O)-, -N(RlO)2, or Rll ⁇ C(O)NRl0-, and c) Cl-C ⁇ alkyl substituted by C1-C6 perfluoroalkyl, RlOO-,
  • RIO is independently selected from hydrogen, C1-C6 alkyl, benzyl and aryl;
  • Rl 1 is independently selected from C1-C6 alkyl and aryl
  • Z is unsubstituted C l -C6 alkyl, substituted C l -C6 alkyl, unsubstituted C3-C6 cycloalkyl or substituted C3-C6 cycloalkyl, wherein the substituted C1-C6 alkyl and substituted C3-C6 cycloalkyl is substituted with one or two of the following: a) Cl -4 alkoxy, b) NR6R 7 , c) C3-6 cycloalkyl, d) -NR°C(0)R 7 , e) HO, f) -S(0) m R6a, g) halogen, or h) perfluoroalkyl;
  • Rib is independently selected from: a) hydrogen, b) aryl. heterocycle, cycloalkyl, R 1 °0-, -N(R 10) 2 or C2-C6 alkenyl. c) Cl-C6 alkyl unsubstituted or substituted by aryl, heterocycle, cycloalkyl, alkenyl, R lOO-, or -N(R l °)2;
  • R2 and R ⁇ are independently selected from: hydrogen or Cl -C6 alkyl
  • Rl i independently selected from hydrogen, Cl -C alkyl, benzyl and aryl;
  • Rl 1 is independently selected from Cl -C6 alkyl and aryl
  • Z is unsubstituted Cl-C6 alkyl, substituted Cl -C6 alkyl, unsubstituted C3-C6 cycloalkyl or substituted C3-C6 cycloalkyl,wherein the substituted C1 -C6 alkyl and substituted C3-C6 cycloalkyl is substituted with one or two of the following: a) Cl -4 alkoxy, b) NR6R7, c) C3-6 cycloalkyl, d) -NR6C(0)R 7 , e) HO, f) -S(0) m R6a, g) halogen, or h) perfluoroalkyl;
  • the preferred compounds of this invention are as follows: 2(S)-n-Butyl-l -[ l -(4-cyanobenzyl)-5-imidazolylmethyl]-4-(2,2,2- trifluoroethyl)piperazin-5-one dihydrochloride
  • the compounds of the present invention may have asymmetric centers and occur as racemates, racemic mixtures, and as individual diastereomers, with all possible isomers, including optical isomers, being included in the present invention.
  • any variable e.g. aryl, heterocycle, Rl , R2 etc.
  • its definition on each occurence is independent at every other occurence.
  • combinations of substituents/or variables are permissible only if such combinations result in stable compounds.
  • alkyl is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms;
  • alkoxy represents an alkyl group of indicated number of carbon atoms attached through an oxygen bridge.
  • Halogen or “halo” as used herein means fluoro, chloro, bromo and iodo.
  • aryl is intended to mean any stable monocyclic or bicyclic carbon ring of up to 7 members in each ring, wherein at least one ring is aromatic.
  • aryl elements include phenyl, naphthyl, tetrahydronaphthyl, indanyl, biphenyl, phenanthryl, anthryl or acenaphthyl.
  • heterocycle or heterocyclic represents a stable 5- to 7-membered monocyclic or stable 8- to 1 1 - membered bicyclic heterocyclic ring which is either saturated or unsaturated, and which consists of carbon atoms and from one to four heteroatoms selected from the group consisting of N, O, and S, and including any bicyclic group in which any of the above-defined heterocyclic rings is fused to a benzene ring.
  • the heterocyclic ring may be attached at any heteroatom or carbon atom which results in the creation of a stable structure.
  • heterocyclic elements include, but are not limited to, azepinyl, benzimidazolyl, benzisoxazolyl, benzofurazanyl, benzopyranyl, benzothiopyranyl, benzofuryl, benzothiazolyl, benzothienyl, benzoxazolyl, chromanyl, cinnolinyl, dihydrobenzofuryl, dihydrobenzothienyl, dihydrobenzothiopyranyl, dihydrobenzothiopyranyl sulfone, furyl, imidazolidinyl, imidazolinyl, imidazolyl, indolinyl, indolyl, isochromanyl, isoindolinyl, isoquinolinyl, isothiazolidinyl, isothiazolyl, isothiazolidinyl, morpholinyl, naphthyridinyl, oxadiazolyl,
  • heteroaryl is intended to mean any stable monocyclic or bicyclic carbon ring of up to 7 members in each ring, wherein at least one ring is aromatic and wherein from one to four carbon atoms are replaced by heteroatoms selected from the group consisting of N, O, and S.
  • heterocyclic elements include, but are not limited to, benzimidazolyl, benzisoxazolyl, benzofurazanyl, benzopyranyl, benzothiopyranyl, benzofuryl, benzothiazolyl, benzothienyl, benzoxazolyl, chromanyl, cinnolinyl, dihydrobenzofuryl, dihydrobenzothienyl, dihydrobenzothiopyranyl, dihydrobenzothiopyranyl sulfone, furyl, imidazolyl, indolinyl, indolyl, isochromanyl, isoindolinyl, isoquinolinyl, isothiazolyl, naphthyridinyl, oxadiazolyl, pyridyl, pyrazinyl, pyrazolyl, pyridazinyl, pyrimidinyl, pyrrolyl, quinazolin
  • the substituted group intended to mean a substituted Cl-8 alkyl, substituted C2-8 alkenyl, substituted C2-8 alkynyl, substituted aryl or substituted heterocycle from which the substitutent(s) R2 and R ⁇ are selected.
  • the substituted Cl -8 alkyl, substituted C3-6 cycloalkyl, substituted aroyl, substituted aryl, substituted heteroaroyl, substituted arylsulfonyl, substituted heteroarylsulfonyl and substituted heterocycle include moieties containing from 1 to 3 substitutents in addition to the point of attachment to the rest of the compound.
  • cyclic moieties When R and R3 are combined to form - (CH2)u -, cyclic moieties are formed. Examples of such cyclic moieties include, but are not limited to:
  • Such cyclic moieties may optionally include a heteroatom(s).
  • heteroatom-containing cyclic moieties include, but are not limited to:
  • Lines drawn into the ring systems from substituents indicate that the indicated bond may be attached to any of the substitutable ring carbon atoms.
  • R l a and R ib are independently selected from: hydrogen, -N(Rl )2, R ! 0C(O)NR 10- or unsubstituted or substituted Cl -C6 alkyl wherein the substituent on the substituted Cj -C6 alkyl is selected from unsubstituted or substituted phenyl, -N(R l )2, R lOo- and R l0C(O)NR l0-.
  • R2 is selected from: H,
  • R is selected from: hydrogen and C l -C6 alkyl.
  • R4 and R$ are hydrogen.
  • R6, R7 and R a is selected from: hydrogen, unsubstituted or substituted Cl -C6 alkyl, unsubstituted or substituted aryl and unsubstituted or substituted cycloalkyl.
  • R6a i unsubstituted or substituted Cl -C6 alkyl, unsubstituted or substituted aryl and unsubstituted or substituted cycloalkyl.
  • R is hydrogen or methyl. Most preferably, R a is hydrogen.
  • R lO is selected from H, Cl -C6 alkyl and benzyl.
  • a l and A2 are independently selected from: a bond, -C(O)NRl0-, -NR IOC(O)-, O, -N(R 10)-, -S(0)2N(R 10). and-
  • V is selected from hydrogen, heterocycle and aryl. More preferably, V is phenyl.
  • Z is unsubstituted or substituted Cl -C6 alkyl.
  • W is selected from imidazolinyl, imidazolyl, oxazolyl, pyrazolyl, pyyrolidinyl, thiazolyl and pyridyl. More preferably, W is selected from imidazolyl and pyridyl.
  • n and r are independently 0, 1 , or 2.
  • p is 1 , 2 or 3.
  • s is 0.
  • t is 1. It is intended that the definition of any substituent or variable (e.g., Rl a, R 5 n , etc.) at a particular location in a molecule be independent of its definitions elsewhere in that molecule.
  • -N(R 10)2 represents -NHH, -NHCH3, -NHC2H5, etc. It is understood that substituents and substitution patterns on the compounds of the instant invention can be selected by one of ordinary skill in the art to provide compounds that are chemically stable and that can be readily synthesized by techniques known in the art, as well as those methods set forth below, from readily available starting materials.
  • the pharmaceutically acceptable salts of the compounds of this invention include the conventional non-toxic salts of the compounds of this invention as formed, e.g., from non-toxic inorganic or organic acids.
  • such conventional non-toxic salts include those derived from inorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric and the like: and the salts prepared from organic acids such as acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic, maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic, sulfanilic, 2-acetoxy-benzoic, fumaric, toluenesulfonic, methanesulf onic, ethane disulfonic, oxalic, isethionic, trifluoroacetic and the like.
  • the pharmaceutically acceptable salts of the compounds of this invention can be synthesized from the compounds of this invention which contain a basic moiety by conventional chemical methods.
  • the salts are prepared either by ion exchange chromatography or by reacting the free base with stoichiometric amounts or with an excess of the desired salt-forming inorganic or organic acid in a suitable solvent or various combinations of solvents.
  • Reactions used to generate the compounds of this invention are prepared by employing reactions as shown in the Schemes 1 -21 , in addition to other standard manipulations such as ester hydrolysis, cleavage of protecting groups, etc., as may be known in the literature or exemplified in the experimental procedures.
  • Substituents R, R a and Rb represent the substituents R2, R 5 R4 a nd R ⁇ however their point of attachment to the ring is illustrative only and is not meant to be limiting.
  • Substituent Z' represents an alkyl moiety or a substitutent on an alkyl moiety such that Z'CH2- is the substiutent Z as defined hereinabove.
  • Boc-protected amino acids I available commercially or by procedures known to those skilled in the art, can be coupled to N-benzyl amino acid esters using a variety of dehydrating agents such as DCC (dicyclohexycarbodiimide) or EDC-HC1 ( l -ethyl-3- (3-dimethylaminopropyl)carbodiimide hydrochloride) in a solvent such as methylene chloride , chloroform, dichloroethane, or in dimethylformamide.
  • dehydrating agents such as DCC (dicyclohexycarbodiimide) or EDC-HC1 ( l -ethyl-3- (3-dimethylaminopropyl)carbodiimide hydrochloride) in a solvent such as methylene chloride , chloroform, dichloroethane, or in dimethylformamide.
  • the product II is then deprotected with acid, for example hydrogen chloride in chloroform or ethyl acetate, or trifluoroacetic acid in methylene chloride, and cyclized under weakly basic conditions to give the diketopiperazine III.
  • acid for example hydrogen chloride in chloroform or ethyl acetate, or trifluoroacetic acid in methylene chloride, and cyclized under weakly basic conditions to give the diketopiperazine III.
  • Reduction of III with lithium aluminum hydride in refluxing ether gives the piperazine IV, which is protected as the Boc derivative V.
  • the N-benzyl group can be cleaved under standard conditions of hydrogenation, e.g., 10% palladium on carbon at 60 psi hydrogen on a Parr apparatus for 24-48 h.
  • the product VI can be reductively alkylated with a suitably substituted aldehyde to provide the protected piperazine VII; a final acid deprotection as previously described gives the intermediate VIII (Scheme 2).
  • the intermediate VIII can itself be reductively alkylated with a variety of aldehydes, such as IX.
  • the aldehydes can be prepared by standard procedures, such as that described by O. P. Goel, U. Krolls, M. Stier and S. Kesten in Organic Syntheses. 1988, 67, 69-75, from the appropriate amino acid (Scheme 3).
  • the reductive alkylation can be accomplished at pH 5-7 with a variety of reducing agents, such as sodium triacetoxyborohydride or sodium cyanoborohydride in a solvent such as dichloroethane, methanol or dimethylformamide.
  • the product X can be deprotected to give the final compounds XI with trifluoroacetic acid in methylene chloride.
  • the final product XI is isolated in the salt form, for example, as a trifluoroacetate, hydrochloride or acetate salt, among others.
  • the product diamine XI can further be selectively protected to obtain XII, which can subsequently be reductively alkylated with a second aldehyde to obtain XIII. Removal of the protecting group, and conversion to cyclized products such as the dihydroimidazole XV can be accomplished by literature procedures.
  • the piperazine intermediate VIII can be reductively alkylated with other aldehydes such as l -trityl-4-imidazolyl- carboxaldehyde or l-trityl-4-imidazolylacetaldehyde, to give products such as XVI (Scheme 4).
  • the trityl protecting group can be removed from XVI to give XVII, or altematively, XVI can first be treated with an alkyl halide then subsequently deprotected to give the alkylated imidazole XVIII.
  • the intermediate VIII can be acylated or sulfonylated by standard techniques.
  • the imidazole acetic acid XIX can be converted to the acetate XXI by standard procedures, and XXI can be first reacted with an alkyl halide, then treated with refluxing methanol to provide the regiospecifically alkylated imidazole acetic acid ester XXII.
  • Hydrolysis and reaction with piperazine VIII in the presence of condensing reagents such as l-(3-dimethylaminopropyl)-3- ethylcarbodiimide (EDC) leads to acylated products such as XXIV.
  • the piperazine VIII is reductively alkylated with an aldehyde which also has a protected hydroxyl group, such as XXV in Scheme 6, the protecting groups can be subsequently removed to unmask the hydroxyl group (Schemes 6, 7).
  • the alcohol can be oxidized under standard conditions to e.g. an aldehyde, which can then be reacted with a variety of organometallic reagents such as Grignard reagents, to obtain secondary alcohols such as XXIX.
  • the fully deprotected amino alcohol XXX can be reductively alkylated (under conditions described previously) with a variety of aldehydes to obtain secondary amines, such as XXXI (Scheme 7), or tertiary amines.
  • the Boc protected amino alcohol XXVII can also be utilized to synthesize 2-aziridinylmethylpiperazines such as XXXII (Scheme 8).
  • the aziridine reacted in the presence of a nucleophile, such as a thiol, in the presence of base to yield the protected ring-opened product XXXIII.
  • piperazine VIII can be reacted with aldehydes derived from amino acids such as O-alkylated tyrosines, according to standard procedures, to obtain compounds such as XXXIX.
  • R' is an aryl group
  • XXXIX can first be hydrogenated to unmask the phenol, and the amine group deprotected with acid to produce XL. Altematively, the amine protecting group in XXXIX can be removed, and O-alkylated phenolic amines such as XLI produced.
  • various side chains can be incorporated into the piperazine.
  • the isomeric piperazin-3-ones can be prepared as described in Scheme 13.
  • the imine formed from arylcarboxamides LII and 2- aminoglycinal diethyl acetal (LIII) can be reduced under a variety of conditions, including sodium triacetoxyborohydride in dichloroethane, to give the amine LIV.
  • Amino acids I can be coupled to amines LIV under standard conditions, and the resulting amide LV when treated with aqueous acid in tetrahydrofuran can cyclize to the unsaturated LVI.
  • Catalytic hydrogenation under standard conditions gives the requisite intermediate LVII, which is elaborated to final products as described in Schemes 3-9.
  • Reaction Scheme 14 provides an illustrative example the synthesis of compounds of the instant invention wherein the substituents R2 and R ⁇ are combined to form - (CH2)u -•
  • substituents R2 and R ⁇ are combined to form - (CH2)u -•
  • 1- aminocyclohexane- 1 -carboxylic acid LVIII can be converted to the spiropiperazine LXVI essentially according to the procedures outlined in Schemes 1 and 2.
  • the piperazine intermediate LXVI can be deprotected as before, and carried on to final products as described in Schemes 3-9. It is understood that reagents utilized to provide the imidazolylalkyl substituent may be readily replaced by other reagents well known in the art and readily available to provide other N- substituents on the piperazine.
  • the aldehyde XLIX from Scheme 12 can also be reductively alkylated with an alkyl amine as shown in Scheme 15.
  • the product LXVII1 can be converted to a piperazinone by acylation with chloroacetyl chloride to give LXIX, followed by base-induced cyclization to LXX.
  • Deprotection, followed by reductive alkylation with a protected imidazole carboxaldehyde leads to LXXII, which can be alkylation with an arylmethylhalide to give the imidazolium salt LXXIII.
  • Final removal of protecting groups by either solvolysis with a lower alkyl alcohol, such as methanol, or treatment with triethylsilane in methylene chloride in the presence of trifluoroacetic acid gives the final product LXXIV.
  • Scheme 16 illustrates the use of an optionally substituted homoserine lactone LXXV to prepare a Boc-protected piperazinone LXXVIII.
  • Intermediate LXXVIII may be deprotected and reductively alkylated or acylated as illustrated in the previous Schemes. Altematively, the hydroxyl moiety of intermediate LXXVIII may be mesylated and displaced by a suitable nucleophile, such as the sodium salt of ethane thiol, to provide an intermediate LXXIX.
  • Intermediate LXXVIII may also be oxidized to provide the carboxylic acid on intermediate LXXXX, which can be utilized form an ester or amide moiety.
  • Amino acids of the general formula LXXXI which have a sidechain not found in natural amino acids may be prepared by the reactions illustrated in Scheme 17 starting with the readily prepared imine LXXXII.
  • the instant compounds are useful as pharmaceutical agents for mammals, especially for humans. These compounds may be administered to patients for use in the treatment of cancer.
  • Examples of the type of cancer which may be treated with the compounds of this invention include, but are not limited to, colorectal carcinoma, exocrine pancreatic carcinoma, myeloid leukemias and neurological tumors. Such tumors may arise by mutations in the ras genes themselves, mutations in the proteins that can regulate Ras activity (i.e., neurofibromin (NF-1), neu, ser, abl , lck, fyn) or by other mechanisms.
  • the compounds of the instant invention inhibit famesyl- protein transferase and the famesylation of the oncogene protein Ras.
  • the instant compounds may also inhibit tumor angiogenesis, thereby affecting the growth of tumors (J. Rak et al. Cancer Research, 55:4575- 4580 (1995)). Such anti-angiogenesis properties of the instant compounds may also be useful in the treatment of certain forms of blindness related to retinal vascularization.
  • the compounds of this invention are also useful for inhibiting other proliferative diseases, both benign and malignant, wherein Ras proteins are aberrantly activated as a result of oncogenic mutation in other genes (i.e., the Ras gene itself is not activated by mutation to an oncogenic form) with said inhibition being accomplished by the administration of an effective amount of the compounds of the invention to a mammal in need of such treatment.
  • a component of NF-1 is a benign proliferative disorder.
  • the instant compounds may also be useful in the treatment of certain viral infections, in particular in the treatment of hepatitis delta and related viruses (J.S. Glenn et al. Science, 256:1331 -1333 (1992).
  • the compounds of the instant invention are also useful in the prevention of restenosis after percutaneous transluminal coronary angioplasty by inhibiting neointimal formation (C. Indolfi et al. Nature medicine, 1 :541 -545( 1995).
  • the instant compounds may also be useful in the treatment and prevention of polycystic kidney disease (D.L. Schaffner et al. American Journal of Pathology, 142: 1051 - 1060 (1993) and B. Cowley, Jr. et ⁇ X.FASEB Journal, 2:A3160 (1988)).
  • the instant compounds may also be useful for the treatment of fungal infections.
  • the compounds of this invention may be administered to mammals, preferably humans, either alone or, preferably, in combination with pharmaceutically acceptable carriers or diluents, optionally with known adjuvants, such as alum, in a pharmaceutical composition, accordmg to standard pharmaceutical practice.
  • the compounds can be administered orally or parenterally, including the intravenous, intramuscular, intraperitoneal, subcutaneous, rectal and topical routes of administration.
  • the selected compound may be administered, for example, in the form of tablets or capsules, or as an aqueous solution or suspension.
  • carriers which are commonly used include lactose and corn starch, and lubricating agents, such as magnesium stearate, are commonly added.
  • useful diluents include lactose and dried com starch.
  • aqueous suspensions are required for oral use, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening and/or flavoring agents may be added.
  • sterile solutions of the active ingredient are usually prepared, and the pH of the solutions should be suitably adjusted and buffered.
  • the total concentration of solutes should be controlled in order to render the preparation isotonic.
  • the compounds of the instant invention may also be co- administered with other well known therapeutic agents that are selected for their particular usefulness against the condition that is being treated.
  • the instant compounds may be useful in combination with known anti-cancer and cytotoxic agents.
  • the instant compounds may be useful in combination with agents that are effective in the treatment and prevention of NF- 1 , restinosis, polycystic kidney disease, infections of hepatitis delta and related viruses and fungal infections.
  • Such combination products employ the compounds of this invention within the dosage range described below and the other pharmaceutically active agent(s) within its approved dosage range.
  • Compounds of the instant invention may altematively be used sequentially with known pharmaceutically acceptable agent(s) when a combination formulation is inappropriate.
  • the present invention also encompasses a pharmaceutical composition useful in the treatment of cancer, comprising the administration of a therapeutically effective amount of the compounds of this invention, with or without pharmaceutically acceptable carriers or diluents.
  • suitable compositions of this invention include aqueous solutions comprising compounds of this invention and pharmacolo- gically acceptable carriers, e.g., saline, at a pH level, e.g., 7.4. The solutions may be introduced into a patient's blood-stream by local bolus injection.
  • the daily dosage will normally be determined by the prescribing physician with the dosage generally varying according to the age, weight, and response of the individual patient, as well as the severity of the patient's symptoms.
  • a suitable amount of compound is administered to a mammal undergoing treatment for cancer.
  • Administration occurs in an amount between about 0.1 mg/kg of body weight to about 60 mg/kg of body weight per day, preferably of between 0.5 mg/kg of body weight to about 40 mg/kg of body weight per day.
  • the compounds of the instant invention are also useful as a component in an assay to rapidly determine the presence and quantity of farnesyl-protein transferase (FPTase) in a composition.
  • FPTase farnesyl-protein transferase
  • the composition to be tested may be divided and the two portions contacted with mixtures which comprise a known substrate of FPTase (for example a tetrapeptide having a cysteine at the amine terminus) and farnesyl pyrophosphate and, in one of the mixtures, a compound of the instant invention.
  • the chemical content of the assay mixtures may be determined by well known immunological, radiochemical or chromatographic techniques. Because the compounds of the instant invention are selective inhibitors of FPTase, absence or quantitative reduction of the amount of substrate in the assay mixture without the compound of the instant invention relative to the presence of the unchanged substrate in the assay containing the instant compound is indicative of the presence of FPTase in the composition to be tested.
  • potent inhibitor compounds of the instant invention may be used in an active site titration assay to determine the quantity of enzyme in the sample.
  • a series of samples composed of aliquots of a tissue extract containing an unknown amount of famesyl-protein transferase, an excess amount of a known substrate of FPTase (for example a tetrapeptide having a cysteine at the amine terminus) and farnesyl pyrophosphate are incubated for an appropriate period of time in the presence of varying concentrations of a compound of the instant invention.
  • concentration of a sufficiently potent inhibitor i.e., one that has a Ki substantially smaller than the concentration of enzyme in the assay vessel
  • concentration of a sufficiently potent inhibitor i.e., one that has a Ki substantially smaller than the concentration of enzyme in the assay vessel
  • Step A N-Methoxy-N-methyl 2(S)-(/c/7-butoxycarbonylamino)- hexanamide 2(S)-Butoxycarbonylaminohexanoic acid (24.6 g, 0.106 mol), N,0-dimethylhydroxylamine hydrochloride (15.5 g, 0.15 mol), EDC hydrochloride ( 22.3 g, 0.1 17 mol) and HOBT (14.3 g, 0.106 mol) were stirred in dry, degassed DMF (300 mL) at 20°C under nitrogen. N-Methylmorpholine was added to obtain pH 7.
  • Step B 2(S)-(fc?; -Butoxycarbonylamino)hexanal
  • 2,2,2-Trifluoroethylamine hydrochloride (0.407 g, 3.0 mmol) was dissolved in dichloroethane under nitrogen. N-Methyl mo ⁇ holine (0.330 mL, 3.0 mmol) was added to obtain pH 5-6, and sodium triacetoxyborohydride (0.795 g, 3.75 mmol) was added. A solution of the product from Step B (0.573 g, 2.5 mmol) in dichloroethane (80 mL) was added slowly dropwise at 20°C. The reaction was stirred overnight, then quenched with saturated sodium bicarbonate solution. The aqueous layer was removed, the organic phase washed with saturated brine and dried over magnesium sulfate. The title compound was obtained as an oil.
  • Step D 1 -fc/t-Butoxycarbonyl-2(S)-/?-butyl-4-(2,2,2- trifluoroethyl)piperazin-5-one
  • Step E l-Triphenylmethyl-4-(hydroxymethyl)imidazole
  • Step F l -Triphenylmethyl-4-(acetoxymethyl)-imidazole
  • Step G 1 -(4-Cyanobenzyl)-5-(acetoxymethyl)-imidazole hydrobromide
  • a solution of the product from Step F (85.8 g, 225 mmol) and oc-bromo-/?-tolunitrile (50.1 g, 232 mmol) in 500 mL of EtOAc was stirred at 60 °C for 20 hours, during which a pale yellow precipitate formed.
  • the reaction was cooled to room temperature and filtered to provide the solid imidazolium bromide salt.
  • the filtrate was concentrated in vacuo to a volume 200 mL, reheated at 60 °C for two hours, cooled to room temperature, and filtered again.
  • the filtrate was concentrated in vacuo to a volume 100 mL, reheated at 60 °C for another two hours, cooled to room temperature, and concentrated in vacuo to provide a pale yellow solid. All of the solid material was combined, dissolved in 500 mL of methanol, and warmed to 60 °C. After two hours, the solution was reconcentrated in vacuo to provide a white solid which was triturated with hexane to remove soluble materials. Removal of residual solvents in vacuo provided the titled product hydrobromide as a white solid which was used in the next step without further purification.
  • Step H l -(4-Cyanobenzyl)-5-(hydroxymethyl)-imidazole
  • Step I l-(4-Cyanobenzyl)-5-imidazolecarboxaldehyde
  • the reaction was stirred overnight at 20°C then poured into saturated sodium bicarbonate solution. The organic phase was washed with saturated brine and dried over magnesium sulfate.
  • the cmde product was purified by preparative HPLC on a 40 X 100 mm Waters PrepPak® reverse phase HPLC column (Delta-PakTM Cl R 15 ⁇ m, 100 ⁇ ) using a gradient elution of 25% (0.1 % TFA in acetonitrile), 75% (0.1 % TFA in water) progressing to 45% (0.1 % TFA in acetonitrile), 55% (0.1 % TFA in water) over 50 min.
  • Step A N- 1 -(3,3,3-Trifluoropropyl)-2(S)-( r.'- butoxycarbonylamino)-hexanamine
  • the title compound is prepared according to the procedure described in Example 1, Step C, except using 1 -(3,3,3- trifluoropropyl)amine hydrochloride in place of 2,2,2- trifluoroethylamine hydrochloride.
  • Step B 1 -te/7-Butoxycarbonyl-2(S)-/7-butyl-4-[ 1 -(3,3,3- trifluoropropyI)lpiperazin-5-one
  • the title compound is prepared according to the procedure described in Example 1 , Step D, except using N- 1 -(3,3,3- trifluoropropyl)-2(S)-(ferr-butoxycarbonylamino)hexanamine in place of N-(2,2,2,-trifluoroethyI)-2(S)-( /7- butoxycarbonylamino)hexanamine.
  • Step C 2(S)- «-Butyl- 1 -[ 1 -(4-cyanobenzyl)-5-imidazolylmethyl]-4- T 1 -(3.3.3-trifluoropropyl)lpiperazin-5-one dihydrochloride
  • the title compound is prepared according to the procedure described in Example 1 , Step J, except using 1 -fer -butoxy carbonyl - 2(S)-/7-butyl-4-[ l-(3,3,3-trifluoropropyl)]piperazin-5-one in place of 1 - f ⁇ ?/ -butoxycarbonyl-2(S)-/7-butyl-4-(2,2,2-trifluoroethyl)piperazin-5- one.
  • the purified product is converted to the hydrochloride salt with
  • Step A N-(Cyclopropylmethyl)-2(S)-(rcrr-butoxycarbonylamino)- hexanamine
  • the title compound is prepared according to the procedure described in Example 1 , Step C, except using cyclopropylmethylamine hydrochloride in place of 2,2,2-trifluoroethylamine hydrochloride.
  • Step B 1 - ⁇ ?/- -Butoxycarbonyl-2(S)-/7-butyl-4-
  • Step C 2(S)- «-Butyl-l -[ l- ⁇ -cyanobenzy -S-imidazolylmethylM- teyclopropylmethyPpiperazin ⁇ -one dihydrochloride
  • the title compound is prepared according to the procedure described in Example 1 , Step J, except using l- / -butoxycarbonyl- 2(S)-/7-butyl-4-(cyclopropylmethyI)piperazin-5-one in place of ⁇ -tert- butoxycarbonyI-2(S)- 7-butyl-4-(2,2,2-trifluoroethyI)piperazin-5-one to obtain the title compound.
  • the purified product is converted to the dihydrochloride salt with HCl in dichloromethane.
  • Step A 3-(4-Cvanobenzyl)pyridin-4-carboxylic acid methyl ester
  • Step B 3-(4-Cvanobenzyl)-4-(hvdroxymethyl)pyridine
  • the title compound was obtained by sodium borohydride (300 mg) reduction of the ester from Step A (0.415 g) in methanol (5 mL) at room temperature. After stirring for 4 h the solution was evaporated and the product was purified on silica gel, eluting with 2% methanol in chloroform to give the title compound.
  • the title compound was obtained by activated manganese dioxide (1.0 g) oxidation of the alcohol from Step B (0.240 g, 1.07 mmol) in dioxane ( 10 mL) at reflux for 30 min. Filtration and evaporation of the solvent provided title compound, mp 80-83°C.
  • Step D 3(S)-n-Butyl- 1 -[3-(4-cyanobenzyl)pyridin-4-yl]-4-(2,2,2- trifluoroethyl)piperazin-5-one dihydrochloride
  • Bovine FPTase was assayed in a volume of 100 ⁇ l containing 100 mM N-(2- hydroxy ethyl) piperazine-N'-(2-ethane sulfonic acid) (HEPES), pH 7.4, 5 mM MgCl2, 5 mM dithiothreitol (DTT), 100 mM H] -farnesyl diphosphate ([ 3 HJ-FPP; 740 CBq/mmol, New England Nuclear), 650 nM Ras-CVLS and 10 ⁇ g/ml FPTase at 31 °C for 60 min. Reactions were initiated with FPTase and stopped with 1 ml of 1.0 M HCL in ethanol.
  • Precipitates were collected onto filter-mats using a TomTec Mach II cell harvestor, washed with 100% ethanol, dried and counted in an LKB ⁇ - plate counter.
  • the assay was linear with respect to both substrates, FPTase levels and time; less than 10% of the [ 3 H]-FPP was utilized during the reaction period.
  • Purified compounds were dissolved in 100% dimethyl sulfoxide (DMSO) and were diluted 20-fold into the assay. Percentage inhibition is measured by the amount of inco ⁇ oration of radioactivity in the presence of the test compound when compared to the amount of inco ⁇ oration in the absence of the test compound.
  • DMSO dimethyl sulfoxide
  • Human FPTase was prepared as described by Omer et al-, Biochemistry 32:5167-5176 (1993). Human FPTase activity was assayed as described above with the exception that 0.1 % (w/v) polyethylene glycol 20,000, 10 ⁇ M ZnCl 2 and 100 ⁇ Ras-CVIM were added to the reaction mixture. Reactions were performed for 30 min., stopped with 100 ⁇ l of 30% (v/v) trichloroacetic acid (TCA) in ethanol and processed as described above for the bovine enzyme.
  • TCA trichloroacetic acid
  • the cell line used in this assay is a v-ras line derived from either Ratl or NIH3T3 cells, which expressed viral Ha-ras p21.
  • the assay is performed essentially as described in DeClue, J.E. et al., Cancer Research 51:712-717. (1991). Cells in 10 cm dishes at 50-75% confluency are treated with the test compound (final concentration of solvent, methanol or dimethyl sulfoxide, is 0.1%).
  • the cells After 4 hours at 37°C, the cells are labelled in 3 ml methionine-free DMEM supple- meted with 10% regular DMEM, 2% fetal bovine serum and 400 mCi[35S]methionine (1000 Ci/mmol). After an additional 20 hours, the cells are lysed in 1 ml lysis buffer (1 % NP40/20 mM HEPES, pH 7.5/5 mM MgCl2/lmM DTT/ 10 mg/ml aprotinen/2 mg/ml leupeptin/2 mg/ml antipain/0.5 mM PMSF) and the lysates cleared by centrifugation at 100,000 x g for 45 min.
  • 1 ml lysis buffer (1 % NP40/20 mM HEPES, pH 7.5/5 mM MgCl2/lmM DTT/ 10 mg/ml aprotinen/2 mg/ml leupeptin/2 mg/ml antipain/0.5 mM PM
  • the immunoprecipitates are washed four times with IP buffer (20 nM HEPES, pH 7.5/1 mM EDTA/1% Triton X- 100.0.5% deoxycholate/0.1%/SDS/0.1 M NaCl) boiled in SDS-PAGE sample buffer and loaded on 13% acrylamide gels. When the dye front reached the bottom, the gel is fixed, soaked in Enlightening, dried and autoradiographed. The intensities of the bands corresponding to farnesylated and nonfarnesylated ras proteins are compared to determine the percent inhibition of farnesyl transfer to protein.
  • IP buffer 20 nM HEPES, pH 7.5/1 mM EDTA/1% Triton X- 100.0.5% deoxycholate/0.1%/SDS/0.1 M NaCl
  • Rat 1 cells transformed with either v-ras, v-raf, or v-mos are seeded at a density of 1 x IO 4 cells per plate (35 mm in diameter) in a 0.3% top agarose layer in medium A (Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine semm) over a bottom agarose layer (0.6%). Both layers contain 0.1 % methanol or an appropriate concentration of the instant compound (dissolved in methanol at 1000 times the final concentration used in the assay).
  • the cells are fed twice weekly with 0.5 ml of medium A containing 0.1 % methanol or the concentration of the instant compound. Photomicrographs are taken 16 days after the cultures are seeded and comparisons are made.

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Abstract

La présente invention concerne des composés inhibiteurs de la farnésyle-protéine transférase (FTase) et de la farnésylation de la protéine oncogène Ras. L'invention concerne également, non seulement des compositions chimiothérapiques contenant les composés de l'invention, mais aussi des procédés d'inhibition de la farnésyle-protéine transférase et de la farnésylation de la protéine oncogène Ras.
EP97917116A 1996-04-03 1997-03-27 Inhibiteurs de la farnesyle-proteine transferase Withdrawn EP0921801A1 (fr)

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US1459396P 1996-04-03 1996-04-03
US14593P 1996-04-03
GBGB9613460.6A GB9613460D0 (en) 1996-06-27 1996-06-27 Inhibitors of farnesyl-protein transferase
GB9613460 1996-06-27
PCT/US1997/005144 WO1997036593A1 (fr) 1996-04-03 1997-03-27 Inhibiteurs de la farnesyle-proteine transferase

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WO1999009985A1 (fr) * 1997-08-27 1999-03-04 Merck & Co., Inc. Inhibiteurs de prenyl-proteine-transferases
US6387903B1 (en) 1997-08-27 2002-05-14 Merck & Co., Inc. Inhibitors of prenyl-protein transferase
AU4562799A (en) * 1998-06-15 2000-01-05 Merck & Co., Inc. Inhibitors of prenyl-protein transferase
JP2002519428A (ja) 1998-07-02 2002-07-02 メルク エンド カムパニー インコーポレーテッド プレニル−タンパク質トランスフェラーゼの阻害剤
JP2002538120A (ja) 1999-03-03 2002-11-12 メルク エンド カムパニー インコーポレーテッド プレニルタンパク質トランスフェラーゼの阻害剤
CA2363169A1 (fr) 1999-03-03 2000-09-08 Merck & Co., Inc. Inhibiteurs de la prenyle-proteine transferase
AU4005300A (en) * 1999-03-03 2000-09-21 Merck & Co., Inc. Inhibitors of prenyl-protein transferase
GB9923748D0 (en) * 1999-10-07 1999-12-08 Glaxo Group Ltd Chemical compounds
USRE39921E1 (en) 1999-10-07 2007-11-13 Smithkline Beecham Corporation Chemical compounds
GB0025354D0 (en) 2000-10-17 2000-11-29 Glaxo Group Ltd Chemical compounds
JP2005522436A (ja) 2002-02-08 2005-07-28 グラクソ　グループ　リミテッド ピペリジルカルボキシアミド誘導体またはそのタキキニン介在疾患の治療における使用
GB0203020D0 (en) 2002-02-08 2002-03-27 Glaxo Group Ltd Chemical compounds
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US3915981A (en) * 1973-03-16 1975-10-28 Yoshitomi Pharmaceutical 1-{8 2-(2-Chlorobenzoyl)-4-nitrophenyl{9 -2-(diethylaminomethyl)-imidazole
US5138069A (en) * 1986-07-11 1992-08-11 E. I. Du Pont De Nemours And Company Angiotensin II receptor blocking imidazoles
US4835154A (en) * 1987-06-01 1989-05-30 Smithkline Beckman Corporation 1-aralykyl-5-piperazinylmethyl-2-mercaptoimidazoles and 2-alkylthioimidazoles and their use as dopamine-βhydroxylase inhibitors
US5219856A (en) * 1992-04-06 1993-06-15 E. I. Du Pont De Nemours And Company Angiotensin-II receptor blocking, heterocycle substituted imidazoles
US5478934A (en) * 1994-11-23 1995-12-26 Yuan; Jun Certain 1-substituted aminomethyl imidazole and pyrrole derivatives: novel dopamine receptor subtype specific ligands
WO1996016057A1 (fr) * 1994-11-23 1996-05-30 Neurogen Corporation Certains derives d'aminoethyl imidazole et pyrrole substitues en 1; nouveaux ligands specifiques d'un sous-type de recepteur dopaminergique

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JP2000507582A (ja) 2000-06-20
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AU706495B2 (en) 1999-06-17
WO1997036593A1 (fr) 1997-10-09

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