WO2006010077A2 - Prodrugs of lonidamine and lonidamine analogs - Google Patents

Prodrugs of lonidamine and lonidamine analogs Download PDF

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Publication number
WO2006010077A2
WO2006010077A2 PCT/US2005/024434 US2005024434W WO2006010077A2 WO 2006010077 A2 WO2006010077 A2 WO 2006010077A2 US 2005024434 W US2005024434 W US 2005024434W WO 2006010077 A2 WO2006010077 A2 WO 2006010077A2
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Prior art keywords
lonidamine
alkyl
compound
group
cycloalkyl
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PCT/US2005/024434
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French (fr)
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WO2006010077A3 (en
Inventor
Mark Matteucci
Harold E. Selick
Photon Rao
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Threshold Pharmaceuticals, Inc.
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Publication of WO2006010077A2 publication Critical patent/WO2006010077A2/en
Publication of WO2006010077A3 publication Critical patent/WO2006010077A3/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D231/00—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings
    • C07D231/54—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings condensed with carbocyclic rings or ring systems
    • C07D231/56—Benzopyrazoles; Hydrogenated benzopyrazoles

Definitions

  • Lonidamine also known as l-(2,4-dichlorobenzyl)-lH-indazole-3- carboxylic acid
  • LND is an anti-cancer drug approved for the treatment of lung, breast, prostate, and brain cancer in a few European countries.
  • the mechanism of action of lonidamine may involve interference with the energy metabolism of neoplastic cells by disruption of the mitochondrial membrane and by inhibition of hexokinase.
  • Lonidamine also has anti- spermatogenic activity and has been shown to inhibit germ cell respiration. Lonidamine has perhaps been most extensively been studied for use in the treatment of advanced breast cancer. For example, the reference Mansi et al., Sep.
  • Lonidamine has also been studied in lung cancer, particularly non-small cell lung cancer (see the reference Joss et al., Sep. 1984, Cancer Treat. Rev. 11(3):205-36) in combination with radiation or other anti-cancer agents.
  • lung cancer particularly non-small cell lung cancer
  • the references Privitera et al., Dec. 1987, Radiother. Oncol. 10(4):285-90 phase II double-blind randomized study of lonidamine and radiotherapy in epidermoid carcinoma of the lung
  • Cancer 37(3):364-8 which describes a phase II study of paclitaxel, cisplatin and lonidamine); and recurrent papillary carcinomas of the urinary bladder (see the reference Giannotti et al., 1984, Oncology 41 Suppl 1 : 104- 7, which describes treatment results after administration of lonidamine plus adriamycin versus adriamycin alone in adjuvant treatment).
  • BPH benign prostatic hypertrophy
  • PCT Publication No. WO 04/064735 both incorporated herein by reference.
  • BPH benign prostatic hypertrophy
  • U.S. Application Publication 20040167196 see also PCT Publication No. WO 04/064735; both incorporated herein by reference.
  • BPH is a disease in which prostate epithelial cells grow abnormally and block urine flow, and currently afflicts more than 10 million adult males in the United States alone and many millions more throughout the rest of the world.
  • Lonidamine which contains a free carboxylic acid group, is poorly soluble in aqueous or non-aqueous solvents. Many carboxylic acid-containing drugs have reduced therapeutic effectiveness if orally administered due to such physicochemical limitations (for examples of several carboxyl-containing beta-lactam antibiotics, see Durckheimer et. al., Advances in Drug Research, Academic Press, London 17:61-634). There remains a need for lonidamine analogs with improved pharmacokinetic properties for use in the treatment of cancer, BPH and other diseases, particularly for analogs with greater bioavailability.
  • the present invention meets these needs and provides novel prodrugs of lonidamine and its derivatives, described in more detail in the following section.
  • the invention relates to compounds and prodrugs of lonidamine and lonidamine analogs.
  • Compounds according to the present invention include tertiary amine prodrugs, linker prodrugs, polymer conjugates, immunoconjugate prodrugs and other prodrug forms of lonidamine and lonidamine analogs.
  • the prodrugs have high aqueous solubility and extended pharmacokinetics in vivo.
  • the prodrugs have reduced toxicity, as compared with lonidamine or a lonidamine analog.
  • the prodrugs are targeted to specific cells types or cell populations.
  • lonidamine- and lonidamine analog- vehicle combinations have extended pharmacokinetics in vivo.
  • the lonidamine- and lonidamine analog- vehicle combinations have reduced toxicity, as compared with lonidamine or a lonidamine analog.
  • the lonidamine- and lonidamine analog- vehicle combinations can be targeted to specific cell types or cell populations.
  • the present invention provides tertiary amine prodrug compounds of lonidamine and lonidamine analogs.
  • Prodrugs of the present invention have the formula (I):
  • R is an aryl or heteroaryl group, optionally substituted with from one to three R 4 substituents, independently selected from the group consisting of halo and a straight or branched chain (d ⁇ C 8 )alkyl;
  • R 3 is a group of the formula (CR 5 R 6 ) m NR 7 R 8 ; each R 5 and R 6 is independently H, a straight or branched chain (C 1 -C 8 ) alkyl heteroalkyl, heterocyclyl, cycloalkyl, aryl, heteroaryl or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system; each R 7 and R 8 is a straight or branched chain (Ci-C 8 )alkyl, heteroalkyl, heterocyclyl, cycloalkyl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system;
  • X is a straight or branched chain, saturated or unsaturated hydrocarbon linker group
  • Y is CHR 9 ;
  • R 9 is H or a straight or branched chain (C 1 -C 8 )alkyl group; the subscript n is an integer of from 0 to 1 ; the subscript m is an integer of from 1 to 4; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
  • compounds are provided of formula D-Z-M, wherein D is lonidamine or a lonidamine analog; Z is a cleavable linker joined to D; and M is a moiety joined to Z.
  • the D-Z-M compound has a higher Vmax for a transporter expressed in plasma membranes cells than D alone, hi some embodiments, the cells are, for example, epithelial cells lining a human colon, a small intestine, a brain, a kidney, a prostate or a heart. Also provided are an individual isomer or a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate, or a hydrate of a D-Z-M compound.
  • linker prodrugs of the following formulae are provided:
  • D is lonidamine or a lonidamine analog
  • Q 1 is O or CH 2
  • Z 1 and Z 2 are cleavable linkers
  • R' is alpha-OH or hydrogen
  • R" is alpha-OH, beta-OH or hydrogen
  • W is- CH(CH 3 )W 1 , wherein W 1 is a substituted alkyl group containing a moiety which is negatively charged at physiological pH, said moiety is selected from the group consisting Of CO 2 H 5 SO 3 H, SO 2 H, -P(O)(OR)(OH), -OP(O)(OR)(OH), and OSO 3 H, wherein R is C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solv
  • R allc is lower alkyl
  • D is lonidamine or a lonidamine analog
  • R 1 is an aryl group of formula wherein each R 2 is independently halogen or C 1 -C 6 alkyl;
  • X 1 is selected from the group consisting of O, S and NR 3 wherein R 3 is hydrogen or C 1 -C 6 alkyl;
  • Y 1 is -C(R 4 ⁇ - or a sugar moiety, wherein each R 4 is independently hydrogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl;
  • X 2 is selected from the group consisting of halogen, C 1 -C 6 alkoxy, diacylglycerol, amino, C 1 -C 6 alkylamino, C 1 -C 6 dialkylamino, C 1 -C 6 alkylthio, a PEG moiety, a bile acid moiety, a sugar moiety, an amino acid moiety, a di-or tri-peptide, a PEG carboxylic acid, and -U-V wherein
  • U is O or S; and V is selected from the group consisting of C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, CW 2 X 3 , PO(X 3 ) 2 , and SO 2 X 3 ; wherein
  • W 2 is O or NR 5 wherein R 5 is hydrogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl; and each X 3 is independently amino, hydroxyl, mercapto, C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl, C 1 -C 6 alkoxy, C 1 -C 6 alkylamino, C 1 -C 6 dialkylamino, C 1 -C 6 alkylthio, a bile acid based alkoxy group, a sugar moiety, a PEG moiety, and -0-CH 2 -
  • prodrugs of lonidamine and lonidamine analogs are provided that have improve aqueous solubility and extended pharmacokinetics in vivo.
  • the present invention provides a method for prophylaxis of cancer, said method comprising administering a prophylactically effective amount of a compound of the present invention to a human subject in need of such prophylaxis.
  • the present invention provides a method for treatment of cancer, said method comprising administering a therapeutically effective amount of a compound of the present invention to a human subject in need of such treatment.
  • the present invention provides a method for prophylaxis of BPH, said method comprising administering a prophylactically effective amount a compound of the present invention to a human subject.
  • the present invention provides a method for treating BPH, the method comprising administering a therapeutically effective amount of a compound of the present invention to a human subject in need of such treatment.
  • the present invention provides a method for reducing a symptom associated with BPH, said method comprising administering a compound of the present invention to a human subject exhibiting the symptom.
  • the present invention provides a method for reducing prostate size in a human subject, comprising administering a therapeutically effective amount of a compound of the present invention to the subject.
  • Alkyl refers to a linear saturated monovalent hydrocarbon radical or a branched saturated monovalent hydrocarbon radical having the number of carbon atoms indicated in the prefix.
  • (Ci-Cg)alkyl is meant to include methyl, ethyl, n- propyl, 2-propyl, n-butyl, 2-butyl, tert-butyl, pentyl, and the like.
  • alkenyl refers to a linear monovalent hydrocarbon radical or a branched monovalent hydrocarbon radical having the number of carbon atoms indicated in the prefix and containing at least one double bond, but no more than three double bonds.
  • (C2-Cg)alkenyl is meant to include, ethenyl, propenyl, 1,3-butadienyl and the like.
  • Alkoxy refers to a substituted or unsubstituted alkyl group of 1 to 6 carbon atoms covalently attached to O.
  • (d-C 6 )alkoxy group is meant to include, for example, methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentoxy, 2- pentoxy, 3-pentoxy, isopentoxy, neopentoxy, hexoxy, 2-hexoxy, 3-hexoxy and 3- methylpentoxy.
  • Alkylamino refers to a substituted or unsubstituted alkyl group of 1 to 6 carbon atoms covalently attached to -NH-. (CrC ⁇ alkylamino group is meant to include, for example, methylamino, ethylamino, propylamino and butylamino.
  • Alkylthio refers to substituted or unsubstituted alkyl group of 1 to 6 carbon atoms covalently attached to S.
  • (Q-C ⁇ alkylthio is meant to include CH 3 - CH 2 -S-, (CH 3 ) 2 CH-S-, CH 3 -(CH 2 ) S -S-, and the like.
  • Aryl refers to a monovalent monocyclic or bicyclic aromatic hydrocarbon radical of 6 to 10 ring atoms which is substituted independently with one to four substituents.
  • the aryl group has one, two, or three substituents selected from alkyl, cycloalkyl, cycloalkyl-alkyl, halo, nitro, cyano, hydroxy, alkoxy, amino, acylamino, mono-alkylamino, di-alkylamino, haloalkyl, haloalkoxy, heteroalkyl, COR (where R is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), - (CR'R") n -COOR (where n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, and R is hydrogen, alkyl, cycloalkyl, cycloalkyl, cycloalkyl
  • aryl includes, but is not limited to, phenyl, biphenyl, 1-naphthyl, and 2-naphthyl, and the substituted forms thereof.
  • Cycloalkyl refers to a monovalent cyclic hydrocarbon radical of three to seven ring carbons.
  • the cycloalkyl group may have one double bond and may also be optionally substituted independently with one, two, or three substituents selected from alkyl, optionally substituted phenyl, or -C(O)R 2 (where R z is hydrogen, alkyl, haloalkyl, amino, mono-alkylamino, di-alkylamino, hydroxy, alkoxy, or optionally substituted phenyl).
  • cycloalkyl includes, for example, cyclopropyl, cyclohexyl, cyclohexenyl, phenylcyclohexyl, 4-carboxycyclohexyl, 2- carboxamidocyclohexenyl, 2-dimethylaminocarbonyl-cyclohexyl, and the like.
  • Heteroalkyl means an alkyl radical as defined herein with one, two or three substituents independently selected from cyano, -OR W , -NR X RY, and -S(O) n R 2 (where n is an integer from 0 to 2 ), with the understanding that the point of attachment of the heteroalkyl radical is through a carbon atom of the heteroalkyl radical.
  • R w is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, alkoxycarbonyl, aryloxycarbonyl, carboxamido, or mono- or di-alkylcarbamoyl.
  • R x is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl or araalkyl.
  • R y is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, alkoxycarbonyl, aryloxycarbonyl, carboxamido, mono- or di- alkylcarbamoyl or alkylsulfonyl.
  • R z is hydrogen (provided that n is 0), alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, amino, mono-alkylamino, di-alkylamino, or hydroxyalkyl.
  • Representative examples include, for example, 2-hydroxyethyl, 2,3- dihydroxypropyl, 2-methoxyethyl, benzyloxymethyl, 2-cyanoethyl, and 2- methylsulfonyl-ethyl.
  • R w , R x , RY, and R 2 can be further substituted by amino, fluorine, alkylamino, di-alkylamino, OH or alkoxy.
  • the prefix indicating the number of carbon atoms refers to the total number of carbon atoms in the portion of the heteroalkyl group exclusive of the cyano, -OR W , - NR X RY, or -S(O) n R v portions.
  • Heteroaryl means a monovalent monocyclic or bicyclic radical of 5 to 12 ring atoms having at least one aromatic ring containing one, two, or three ring heteroatoms selected from N, O, or S, the remaining ring atoms being C, with the understanding that the attachment point of the heteroaryl radical will be on an aromatic ring.
  • the heteroaryl ring is optionally substituted independently with one to four substituents.
  • the heteroaryl ring is substituted independently with one or two substituents selected from alkyl, cycloalkyl, cycloalkyl-alkyl, halo, nitro, cyano, hydroxy, alkoxy, amino, acylamino, mono-alkylamino, di-alkylamino, haloalkyl, haloalkoxy, heteroalkyl, -COR (where R is hydrogen, alkyl, phenyl or phenylalkyl, -(CR'R") n -COOR
  • n is an integer from 0 to 5
  • R' and R" are independently hydrogen or alkyl, and R is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), or ⁇ (CR'R")n-
  • R X RY (where n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, and R x and RY are, independently of each other, hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl).
  • heteroaryl includes, but is not limited to, pyridyl, furanyl, thienyl, thiazolyl, isothiazolyl, triazolyl, imidazolyl, isoxazolyl, pyrrolyl, pyrazolyl, pyridazinyl, pyrimidinyl, benzofuranyl, tetrahydrobenzofuranyl, isobenzofuranyl, benzothiazolyl, benzoisothiazolyl, benzotriazolyl, indolyl, isoindolyl, benzoxazolyl, quinolyl, tetrahydroquinolinyl, isoquinolyl, benzimidazolyl, benzisoxazolyl or benzothienyl, indazolyl, pyrrolopyrymidinyl, indolizinyl, pyrazolopyridinyl, triazolopyridinyl, imidazolyl, is
  • Heterocyclyl or "cycloheteroalkyl” means a saturated or unsaturated non- aromatic cyclic radical of 3 to 8 ring atoms in which one to four ring atoms are heteroatoms selected from O, NR (where R is independently hydrogen or alkyl), NR Q (where R Q is independently hydrogen or alkyl), or S(O) n (where n is an integer from 0 to
  • the heterocyclyl ring may be optionally substituted independently with one, two, or three substituents selected from alkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, cycloalkyl, cycloalkyl-alkyl, halo, nitro, cyano, hydroxy, alkoxy, amino, mono-alkylamino, di-alkylamino, haloalkyl, haloalkoxy, -COR (where R is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), -COR S (where R s is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), -(CR'R") n
  • heterocyclyl includes, but is not limited to, pyridyl, tetrahydropyranyl, N-methylpiperidin-3-yl, N-methylpyrrolidin-3-yl, 2-pyrrolidon-l-yl, furyl, quinolyl, thienyl, benzothienyl, pyrrolidinyl, piperidinyl, morpholinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydrothiofuranyl, 1 , 1 -dioxo-hexahydro- 1 ⁇ 6 -thiopyran-4-yl, tetrahydroimidazo [4,5-c] pyridinyl, imidazolinyl, piperazinyl, and piperidin-2-onyl, and the derivatives thereof.
  • the prefix indicating the number of carbon atoms e.g., C3-C10) refers to the total number of carbon atoms
  • Alkylene means a linear saturated divalent hydrocarbon radical having from one to twelve carbon atoms or a branched saturated divalent hydrocarbon radical having from one to twelve carbon atoms optionally substituted with substituents including, for example, hydroxy, amino, mono or di(CrC 6 )alkyl amino, halo, C 2 -C 6 alkenyl ether, cyano, nitro, ethenyl, ethynyl, C 1 -C 6 alkoxy, C 1 -C 6 alkylthio, -COOH, -CONH 2 , mono- or di-(C 1 -C 6 )alkyl-carboxamido, -SO 2 NH 2 , -OSO 2 -(C 1 -C 6 ) alkyl, mono or di(C 1 -C 6 ) alkylsulfonamido, aryl and heteroaryl.
  • alkylene includes methylene, ethylene, propylene
  • Heteroalkylene has essentially the meaning given above for alkylene except that one or more heteroatoms (i.e., oxygen, sulfur, nitrogen and/or phosphorous) may be present in the alkylene biradical.
  • heteroalkylene includes, -CH 2 OCH 2 O-, -CH 2 CH 2 OCH 2 CH 2 -, -CH 2 CH 2 N(CH 3 )CH 2 CH 2 -, -CH 2 CH 2 SCH 2 CH 2 -, and the like.
  • sugar moiety refers to a mono-, di-, or trisacharide or a derivative thereof.
  • the sugar moiety is an aldohexose, a ketopentose, an aldopentose, or a ketopentose.
  • the sugar moiety is D-glucose or a derivative thereof, hi one embodiment, the sugar moiety is glucuronic acid.
  • the bonding of a sugar moiety to the rest of a prodrug compound does not involve a peroxo or a -O-O- linkage.
  • Bioisostere of a given molecule, one replaces one or more atoms or groups with atoms or groups having similar size and spatial disposition of electron pair or pairs.
  • Bioisosteres and bioisosterism is a well-known tool for predicting the biological activity of compounds, based upon the premise that compounds with similar size, shape, and electron density can have similar biological activity.
  • a reasonable quantitative prediction of the binding ability of a known molecule can be made based on the spatial arrangement of a small number of atoms or functional groups in the molecule. As used herein, such an arrangement is called a pharmacophore, and once the pharmacophore or pharmacophores in a molecule have been identified, this information can be used to identify other molecules containing the same or similar pharmacophores.
  • Such methods are well known to persons of ordinary skill in the art of medicinal chemistry, and as the structural information described in this application identifies the pharmacophore prodrugs, lonidamine or lonidamine analogs.
  • An example of programs available to perform pharmacophore -related searches is the program 3D Pharmacophore search from the Chemical Computing Group (see Chemical Computing Group internet web site).
  • heterocyclo group optionally mono- or di- substituted with an alkyl group means that the alkyl may but need not be present, and the description includes situations where the heterocyclo group is mono- or disubstituted with an alkyl group and situations where the heterocyclo group is not substituted with the alkyl group.
  • Optionally substituted means a ring or linear radical which optionally may be substituted independently with substituents.
  • a prodrug is a compound that, after administration, is metabolized or otherwise converted to an active or more active form with respect to at least one property.
  • a pharmaceutically active compound can be modified chemically to render it less active or inactive, but the chemical modification is such that an active form of the compound is generated by metabolic or other biological processes.
  • Prodrugs also can be formed by non-covalent modification of a pharmaceutically active compound, such that the compound can be released as an active form or an activatable form.
  • a prodrug may have, relative to the drug, altered metabolic stability or transport characteristics, fewer side effects or lower toxicity, and/or improved flavor, for example (see the reference Nogrady, 1985, Medicinal Chemistry A Biochemical Approach, Oxford University Press, New York, pages 388-392). Prodrugs can also be prepared using compounds that are not drugs.
  • Vmax refers to the maximum initial rate of enzyme or transporter function.
  • Vmax refers to the maximum initial rate of enzyme or transporter function.
  • treating refers to taking steps to obtain beneficial or desired results, including clinical results.
  • beneficial or desired clinical results include, but are not limited to, alleviation or amelioration of one or more symptoms of cancer, BPH or other condition, diminishment of extent of disease, delay or slowing of disease progression, amelioration, palliation or stabilization of the disease state, and other beneficial results described below.
  • preventing or “prophylaxis” mean to reduce the risk of occurrence of disease (e.g., cancer, BPH or other condition) in an individual or in individuals in a population. That is, administration of an agent to a population of individuals "prevents" a condition when, relative to a control population, fewer individuals in the population develop the condition, and/or appearance of the condition is delayed in the administered population relative to a control population. Methods for detecting and quantifying a reduction of risk, including use of clinical trials, are well known in the art. As used herein, "preventing" a condition or disease in a patient refers to taking steps to obtain beneficial or desired results, including clinical results.
  • disease e.g., cancer, BPH or other condition
  • administering or "administration of a drug to a subject include both direct administration, including self-administration, and indirect administration, including the act of prescribing a drug.
  • direct administration including self-administration
  • indirect administration including the act of prescribing a drug.
  • a physician who instructs a patient to self-administer a drug and/or provides a patient with a prescription for a drug is administering the drug to the patient.
  • a "manifestation" of cancer, BPH or other condition refers to a symptom, sign, anatomical state (e.g., prostate size), physiological state (e.g., prostate specific antigen (PSA) level), or report (e.g., American Urological Association Symptom Index score) characteristic of a subject with cancer or BPH.
  • anatomical state e.g., prostate size
  • physiological state e.g., prostate specific antigen (PSA) level
  • report e.g., American Urological Association Symptom Index score
  • a "therapeutically effective amount" of a drug is an amount of a drug that, when administered to a subject with cancer, BPH or other condition, will have the intended therapeutic effect, e.g., alleviation, amelioration, palliation or elimination of one or more manifestations of cancer, BPH or other condition in the subject.
  • the full therapeutic effect does not necessarily occur by administration of one dose, and may occur only after administration of a series of doses.
  • a therapeutically effective amount may be administered in one or more administrations.
  • a prophylactically effective amount of a drug is an amount of a drug that, when administered to a subject, will have the intended prophylactic effect, e.g., preventing or delaying the onset of disease or symptoms, or reducing the likelihood of the onset of disease or symptoms.
  • the full prophylactic effect does not necessarily occur by administration of one dose, and may occur only after administration of a series of doses. Thus, a prophylactically effective amount may be administered in one or more administrations.
  • target cell refers to a cell to be contacted with lonidamine or a lonidamine analog.
  • a target cell population refers to a population having a common characteristic (e.g., a cell surface marker) that is to be contacted with lonidamine or a lonidamine analog.
  • Lonidamine or lonidamine analogs can be used in the prodrugs according to the present invention.
  • Lonidamine is l-(2,4-dichlorobenzyl)-l-H-indazole-3-carboxylic acid.
  • Lonidamine was first identified as having anti-spermatogenic activity, and subsequently used for treatment of certain cancers. The use of lonidamine and lonidamine analogs for treatment and prevention of benign prostatic hyperplasia has been described (for example, see U.S. patent application number 10/759,337, published as US 2004/0167196).
  • Examples of lonidamine analogs include tolnidamine; AF-2364; and AF-2785 (see Ansari et al, 1998, Contraception 57:271-279; and Corsi et al., 1976, J. Medicinal Chemistry 19:778-83); and compounds described by Silvestrini, 1981, Chemotherapy 27:9-20; Lobl et al., 1981, Chemotherapy 27:61-76; Cheng et al., 2001, Biol. Reprod. 65:449-461; Andreani, F., 1984, Arch. Pharm.
  • lonidamine analogs have the following formula (V):
  • R 1 , R 2 , X, Y, and n are:
  • R 2 is -Cl, -Br, -I, or -CH 3 , monosubstituted phenyl, substituted at the 2, 3, or 4 position; dichloro, dibromo, dimethyl, or chloro and methyl disubstituted phenyl, substituted at the 2 and 3 or 2 and 4 positions; or 2, 4, 5 trichlophenyl;
  • Y is -(CH 2 )-; and n is zero, and R 1 is -COOH, -CONH 2 , -CONHNH 2 , -CONHN(CH 3 ) 2 , -CH 2 CH 2 OH, -CH 2 CH(OH)CH 2 OH, or CH 2 (CH 2 OH) 2 ; or
  • the compounds of the present invention exclude one or more of lonidamine, tolnidamine, AFl 890, AF2364 and/or AF2785.
  • Lonidamine and lonidamine analogs and derivatives can be prepared using by well known synthetic methods. Synthesis of lonidamine is described in U.S. Patent No. 3,895,026 in the presence of an inert base, such as triethyl amine tolnidamine (TND), is described in the art (see, e.g., Corsi et al., 1976, Journal of Medicinal Chemistry 19:778- 83; Cheng et al., 2001, Biol. Reprod. 65:449-61; Silvestrini, 1981, Chemotherapy 27:9- 20; Lobl et al., 1981, Chemotherapy 27:61-76; U.S. Patent Nos. 3,895,026 and 6,001,865; See also PCT/US05/19350).
  • TDD triethyl amine tolnidamine
  • the present invention provides tertiary amine prodrugs of lonidamine and lonidamine analogs having the formula:
  • R 1 represents COOR 3 .
  • R 2 is usually an aryl or heteroaryl group.
  • the aryl or heteroaryl group of R 2 may have substituents on their respective rings, wherein each substituted present can be the same or different from any other substituent. More particularly, R 2 may have from 0 to 3 substituents, more preferably from 1 to 2 substituents, and still more preferably, 2 R 4 substituents.
  • R 4 substituents are independently selected from the group consisting of halo and a straight or branched chain (C 1 -C 8 )alkyl.
  • R 3 represents a group of the formula (CR 5 R 6 ) m NR 7 R 8 wherein each R 5 and R 6 is independently H, a straight or branched chain (C 1 -C 8 )alkyl, heteroalkyl, cycloalkyl, or heterocyclyl or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system.
  • Each R 7 and R 8 is a straight or branched chain (CrC 8 )alkyl, heteroalyl, cycloalkyl, or heterocyclyl or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system.
  • X is a straight or branched chain, saturated or unsaturated hydrocarbon linker group and Y is CHR 9 .
  • R 9 represents H or a straight or branched chain (C 1 -C 8 )alkyl group.
  • the present invention further includes all salts thereof, and particularly, pharmaceutically acceptable salts thereof. Still further, the invention includes compounds that are single isomers of the above formula (e.g., single enantiomers of compounds having a single chiral center), as well as solvate and hydrate forms thereof.
  • the present invention further includes an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
  • R 1 is preferably a COOR 3 moiety.
  • each R 5 and R 6 is preferably independently selected from the group, H, CH 3 , and a member in which R 5 and R 6 are joined together to form a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1,1-dioxo-hexahydro-l ⁇ 6 - thiopyran-4-yl or tetrahydropyran-4-yl group.
  • preferred embodiments are compounds wherein NR 7 R 8 is morpholino.
  • R 2 is preferably phenyl.
  • R 4 is preferably independently selected from the group consisting of Cl, Br, or CH 3 .
  • compounds preferably have the formula:
  • R 4 is, independently Cl or CH 3 ; each R 5 and R 6 is independently H, (C 1 -Cs)alkyl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered ring cycloalkyl or heterocyclyl system;
  • X is an alkenylene linker group; and the subscript n is an integer of from 0 to 1.
  • R 4 is preferably Cl; n is 0; and each R 5 and R 6 is independently, H, CH 3 , or are joined together to form a cyclopropyl, cyclopbutyl, cyclopentyl, cyclohexyl, 1,1-dioxo- hexahydro-l ⁇ 6 -thiopyran-4-yl or tetrahydropyran-4-yl group.
  • each R 5 and R 6 is independently, H, CH 3 , or are joined together to form a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1,1-dioxo-hexahydro-l ⁇ -thiopyran-4- yl or tetrahydropyran-4-yl group.
  • Examples of compounds of Formula (I) include:
  • the present invention further includes an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate of any of these compounds.
  • the lonidamine analog has the formula:
  • R 1 , R 2 , X, Y and n are defined below:
  • R 1 represents COOR 3 .
  • n is O.
  • Y is CH 2 -.
  • R 2 is 2,4-dichlorophenyl.
  • R 3 can be a straight or branched chain (C 1 -C 8 ) alkyl or alkoxy, or a three- to eight-membered cycloalkyl or heterocyclyl ring system.
  • R can be Cr 6 alkoxymethyl, such as methoxymethyl; C 1-6 alkanoyloxymethyl esters such as pivaloyloxymethyl; phthalidyl esters; C ⁇ CgcycloalkoxycarbonyloxyQ-C ⁇ alkyl such as 1- cyclohexylcarbonyloxyethyl; l,3-dioxolen-2-onylmethyl esters, such as 5-methyl-l,3- dioxolen-2-on-ylmethyl; and Q.Qalkoxycarbonyloxyethyl such as 1- methoxycarbonyloxyethyl.
  • the present invention further includes an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate of any of these compounds.
  • Lonidamine and lonidamine analogs and derivatives can be prepared using by well known synthetic methods, as described supra.
  • Ester prodrugs of lonidamine and lonidamine analogs are generally prepared as described in, for example, U.S. Pat. No. 6,146,658, as is administration of polymorphic forms, an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate, a hydrate thereof, and the like.
  • Syntheses of the ester prodrugs start with the free carboxylic acid of lonidamine or a lonidamine analog.
  • the free acid is activated for ester formation in an aprotic solvent and then reacted with a free alcohol group bearing the tertiary amine in the presence of an inert base, such as triethyl amine, to affect ester formation, producing the prodrug.
  • Activating conditions for the carboxylic acid include forming the acid chloride using oxalyl chloride or thionyl chloride in an aprotic solvent, optionally with a catalytic amount of dimethyl formamide, followed by evaporation.
  • aprotic solvents include, but are not limited to methylene chloride, tetrahydrofuran, and the like.
  • activations can be performed in situ by using reagents such as BOP (benzotriazol-l-yloxytris(dimethylamino) phosphonium hexafluorolphosphate, and the like (see Nagy et al., 1993, Proc. Natl. Acad. Sci. USA 90:6373-6376) followed by reaction with the free alcohol.
  • BOP benzotriazol-l-yloxytris(dimethylamino) phosphonium hexafluorolphosphate
  • Isolation of the ester products can be affected by extraction with an organic solvent, such as ethyl acetate or methylene chloride, against a mildly acidic aqueous solution; followed by base treatment of the acidic aqueous phase so as to render it basic; followed by extraction with an organic solvent, for example ethyl acetate or methylene chroride; evaporation of the organic solvent layer; and recrystalization from a solvent, such as ethanol, which has been acidified with an acid, such as HCl or acetic acid.
  • the crude reaction can be passed over an ion exchange column bearing sulfonic acid groups in the protonated form, washed with deionized water, and eluted with aqueous ammonia; followed by evaporation.
  • Suitable free alcohols bearing the tertiary amine are commercially available, such as N-2-hydroxyethylmorpholine, N-2-hydroxyethylimidazole, and the like.
  • Non- commercially available alcohols can be synthesized in straightforward manner via standard literature procedures. Such procedures can be identified via literature search tools such as SciFinder from the American Chemical Society or Beilstein available from MDL Software.
  • the prodrug of the lonidamine or a lonidamine analog is provided in the form of a pharmaceutically acceptable salt.
  • Pharmaceutically acceptable salts include addition salts with acids, as well as the salts with bases.
  • Suitable acids for the formation of acid addition salts are, for example, mineral acids, such as hydrochloric, hydrobromic, sulphuric or phosphoric acid, or organic acids, such as organic sulphonic acids, for example, benzenesulphonic, 4-toluenesulphonic or methanesulphonic acid, and organic carboxylic acids, such as acetic, lactic, palmitic, stearic, malic, maleic, fumaric, tartaric, ascorbic or citric acid. Acid salts of the tertiary amine moiety confer increased aqueous solubility. The use of citric acid salts is especially preferred.
  • the prodrugs of the invention provide for the release of a drug lonidamine and its analogs.
  • the prodrug moiety comprises a tertiary amine having a pKa near the physiological pH of 7.5. Any amines having a pKa within 1 unit of 7.5 are suitable alternatives amines for this purpose.
  • the amine may be provided by the amine of a morpholino group. This pKa range of 6.5 to 8.5 allows for significant concentrations of the basic neutral amine to be present in the mildly alkaline small intestine.
  • the basic, neutral form of the amine prodrug is lipophilic and is absorbed through the wall of the small intestine into the blood.
  • the prodrug moiety is cleaved by esterases which are naturally present in the serum to release lonidamine or the lonidamine analog. More strongly basic amines, such as a trialkyl derivatives with no heteroatom substitutions, will be nearly completely protonated under physiological conditions and will not be as efficiently adsorbed as shown.
  • the serum half live of the prodrug of the lonidamine and lonidamine analogs of the present invention is increased in vivo (compared to the parental form) by the presence of R 5 and R 6 groups.
  • the R 5 and R 6 groups in the prodrug can independently be selected to modulate the rate of cleavage of the prodrug moiety from lonidamine. Increasing the amount of steric hindrance proximal to the ester carbonyl of lonidamine decreases the rate of cleavage of the prodrug moiety. Slowing the rate of cleavage of the prodrug moiety has the effect of increasing serum half life.
  • Hydrogen groups facilitate cleavage of the prodrug moiety and alkyl groups hinder it.
  • the larger and more branched the alkyl group the more cleavage is hindered and the more serum half life is increased.
  • the closer the non-hydrogen substitution is to the lonidamine carbonyl the more cleavage of the prodrug moiety is hindered and the more serum half life of the prodrug form is increased.
  • linkage of the tertiary amine to the lonidamine is stable enough so that the serum half life of the prodrug is from about 8 to about 24 hours.
  • R 4 and R 5 may be joined together to form a cyclic group further comprising heteroatoms. This aspect of the invention further improves upon the aqueous solubility of the compounds of the invention.
  • the present invention provides a prodrug D-Z-M of lonidamine or a lonidamine analog, said prodrug comprising, lonidamine or an analog, D; joined by a cleavable linker Z; to a moiety M.
  • Prodrugs with this structure may be referred to as "linker prodrugs.”
  • the prodrug has a higher Vmax for a transporter expressed in plasma membranes of cells than D alone.
  • the cells are epithelial cells lining a human colon, or small intestine, a prostate, or the like.
  • the transporter is expressed in the plasma membranes of epithelial cell lining in the human gut.
  • the transporter is expressed in the plasma membranes of epithelial cell lining in the prostate. In one embodiment, the transporter is expressed in human kidney, brain, lung, liver and/or heart. In one embodiment, the moiety M is selected from the group consisting of an amino acid, a dipeptide, a tripeptide, a bile acid, and their derivatives.
  • the transporter is selected from the group consisting of ATBO, CAT-I, FATP4, MCTl, MCT4, NADCl, NADC2, OCTN2, PEPTl, PGT, RFC, SAT-I, SAT-6, SMVT, SUT2 and SVCTl (for a description of these transporters see, e.g., Gallop et al., WO02100347).
  • the transporter is PEPT2, which is expressed in human kidney, brain, lung, liver, and heart.
  • the transport system is carrier mediated.
  • the transport system is receptor mediated.
  • the prodrug compound exhibits selective uptake by a subject's prostate as compared to another organ, such as the testis, heart, kidney, brain, lung, and/or liver.
  • the prodrug is selectively taken up by subject's prostate compared to other organs.
  • the prodrug compound exhibits selective uptake by prostate epithelial cells as compared to other epithelial cells of, for example, the testis, heart, kidney, brain, lung, and/or liver.
  • the prodrug is selectively taken up by prostate epithelial cell as compared to other epithelial cells.
  • the M moiety is an androgen, an androgen analog, or a functional androgen analog exhibits selective uptake by a subject's prostate as compared to another organ such as, for example, the testis, heart, kidney, brain, lung, and/or liver.
  • the prodrug D is selectively taken up by subject's prostate.
  • the M moiety is an androgen, an androgen analog, or a functional androgen analog that exhibits selective uptake by prostate epithelial cells as compared to epithelial cells such as, for example, the testis, heart, kidney, brain, lung, and/or liver.
  • the prodrug is selectively taken up by prostate epithelial cells as compared to other epithelial cells.
  • the present invention provides a prodrug of lonidamine or a lonidamine analog comprising a lonidamine- or a lonidamine analog-peptide conjugate, the peptide comprising an amino acid sequence having a cleavage site specific for an enzyme having a proteolytic activity of prostate specific antigen and wherein the peptide is linked to lonidamine or the lonidamine analog to inhibit the therapeutic activity of lonidamine or the lonidamine analog, and wherein lonidamine or- the lonidamine analog is cleaved from the peptide upon proteolysis by an enzyme having a proteolytic activity of prostate specific antigen (PSA).
  • PSA prostate specific antigen
  • the present invention provides a prodrug of lonidamine or a lonidamine analog comprising a lonidamine- or a lonidamine analog-peptide conjugate, the peptide comprising an amino acid sequence having a cleavage site specific for an enzyme having a proteolytic activity of prostate specific antigen, wherein the peptide is 20 or fewer amino acids in length, wherein the sequence comprises the amino acids
  • G 5 is from 0 to 16 amino acids; G 4 is serine, isoleucine, or lysine; G 3 is serine or lysine; G 2 is leucine or lysine; and G 1 is glutamine, asparagine or tyrosine, and wherein the peptide is linked to lonidamine or the lonidamine analog to inhibit the therapeutic activity of the loi ⁇ dam ⁇ ne or the lonidamine analog, and wherein lonidamine or the lonidamine analog is cleaved from the peptide upon proteolysis by an enzyme having a proteolytic activity of prostate specific antigen (PSA).
  • PSA prostate specific antigen
  • the present invention provides a prodrug of lonidamine or an analog comprising a cephalosporin moiety, a dihydronicotinamide moiety, a triglyceride, a long chain fatty acid, or a long chain fatty alcohol.
  • the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is a vitamin or a vitamin precursor. In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is vitamin-D, a vitamin-D analog, or a vitamin-D precursor. In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is vitamin-E, a vitamin-E analog, or a vitamin-E precursor. In a related embodiment, the moiety M is ⁇ -tocopherol.
  • the moiety M is an ⁇ -tocopherol-PEG conjugate. In another related embodiment, the moiety M is an ⁇ -tocopherol- ⁇ , ⁇ -dicarboxylic acid-PEG conjugate. In another related embodiment, the moiety M is an ⁇ -tocopherol-succinic acid-PEG conjugate.
  • Various ⁇ -tocopherol based conjugates employed in the present invention can be adapted from those described in the U.S. Patent Application No. US2005/0142189, to Lambert et al.
  • the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is a hormone or a hormone precursor.
  • the present invention provides a prodrug of lonidamine or a lonidamine analog wherein the prodrug is enzymatically modified to yield lonidamine or the lonidamine analog, wherein the enzyme is carboxypeptidase, aminohydrolase, or glycosidase.
  • the prodrug contains an Aryl-O-CO-N ⁇ moiety which is cleaved by a carboxypeptidase to yield lonidamine or a lonidamine analog from the prodrug.
  • Moiety M and linker Z that can be employed in a D-Z-M prodrug of the present invention is provided for example, in the reference Silverman, January 15, 1992, Organic Chemistry of Drug Design and Drug Action, Academic Press; 1st edition.
  • M moieties including but not limited to a bile acid, an amino acid, and a peptide, and linker Z moieties that can be used in the compounds of the invention are described in the following US Patent Application Nos. 2004/0161424, 2003/0158254, 2003/0158089, and 2003/0017964; and PCT Publication Nos.
  • the moiety can be a targeting peptide, to target lonidamine or a lonidamine analog to a specific cell type. See, e.g., U.S. patent publication No. 2002/0147138.
  • the present invention provides a prodrug D-Z-M of lonidamine or a lonidamine analog, said prodrug comprising lonidamine or an analog, D, joined by a cleavable peptide linker Z, to a stabilizing moiety M.
  • the peptide linker can be any cleavable peptide linker.
  • the linker is cleavable by an endogenous enzyme, hi some embodiments, the linker is a tripeptide, P1-P2-P3, comprising natural or synthetic amino acids.
  • Pl is Leucine, Sarcosine, Tyrosine, Phenylalanine, p-Cl- Phenylalanine, p-Nitrophenylalanine, Valine, Norleucine, Norvaline, Phenylglycine, Tryptophan, tetrahydroisoquinoline-3-carboxylic acid, 3-Pyridylalanine, Alanine, Glycine, or 2-Thienylalanine.
  • P2 can be Alanine, Leucine, Tyrosine, Glycine, Serine, 3-Pyridylalanine, or 2-Thienylalanine.
  • P3 can be Leucine, Phenylalanine, Isoleucine, Alanine, Glycine, Tyrosine, 2- Naphthylalanine, or Serine.
  • the peptide linker can be one of the following: Leu- Ala- Leu, Tyr-Ala-Leu, Met- Ala-Leu, Tyr-Ala-Ile, Phe-Gly-Leu, Met-Gly-Leu, Met-Gly-Ile, Phe-Gly-Ile, Met-Gly-Phe, Leu-Ala-Gly, NIe- Ala-Leu, Phe-Gly-Phe, and Leu-Tyr-Leu. See also U.S. Patent Publication No. 2003/0181359.
  • moiety M is a stabilizing moiety that protects the prodrug from cleavage in circulating blood when it is administered to the patient and allows the prodrug to reach the vicinity of the target cell relatively intact.
  • the stabilizing group typically protects the prodrug from cleavage in blood and blood serum.
  • the stabilizing group is useful in the prodrug when it serves to protect the prodrug from degradation, i.e., inactivation, when tested by storage of the prodrug compound in human blood at 37 0 C. for 2 hours and results in less than 20%, particularly less than 2%, inactivation of the prodrug by the enzymes present in the human blood under the given assay conditions.
  • the stabilizing group can be, for example, an amino acid or an amino acid that is either (i) a non-genetically-encoded amino acid having four or more carbons or (ii) aspartic acid or glutamic acid attached to the N-terminus of the oligopeptide at the beta- carboxyl group of aspartic acid or the gamma-carboxyl group of glutamic acid.
  • dicarboxylic (or a higher order carboxylic) acid or a pharmaceutically acceptable salt thereof may be used as a stabilizing group.
  • the stabilizing group is not an amino acid.
  • linker prodrugs of the following formulae are provided: wherein D is lonidamine or a lonidamine analog; Q 1 is O or CH 2 ; Z 1 and Z 2 are cleavable linkers; R' is alpha-OH or hydrogen; R" is alpha-OH, beta-OH or hydrogen; W is -CH(CH 3 )W 1 , wherein W 1 is a substituted alkyl group containing a moiety which is negatively charged at physiological pH, said moiety is selected from the group consisting OfCO 2 H 5 SO 3 H, SO 2 H, -P(O)(OR)(OH), -OP(O)(OR)(OH), and OSO 3 H wherein R is C 1 - C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a
  • R alk is alkyl (e.g., C 1 -C 6 alkyl); and D is lonidamine or a lonidamine analog, and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof, hi one embodiment, R a i k is lower alkyl.
  • D is covalently attached to a heteroatom in the formula above,
  • R 1 is an aryl group of formula wherein each R 2 is independently halogen or C 1 -C 6 alkyl.
  • R 1 is as defined above.
  • the present invention provides a compound of formula
  • R 1 is an aryl group of formula wherein each R 2 is independently halogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, or heterocyclyl;
  • X 1 is selected from the group consisting of O, S and NR 3 wherein R 3 is hydrogen or C 1 -C 6 alkyl; Y 1 is -C(R 4 ) 2 - or a sugar moiety, wherein each R 4 is independently hydrogen or
  • X 2 is selected from the group consisting of halogen; C 1 -C 6 alkoxy; diacylglycerol; amino; C 1 -C 6 alkylamino; C 1 -C 6 dialkylamino; C 1 -C 6 alkylth ⁇ o; a PEG moiety; a bile acid moiety; a sugar moiety; an amino acid moiety; a di-or tri-peptide; a PEG carboxylic acid; and -U-V wherein
  • U is O or S; and V is selected from the group consisting of C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, CW 2 X 3 , PO(X 3 ) 2 , and SO 2 X 3 wherein W 2 is O or NR 5 wherein R 5 is hydrogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and each X 3 is independently amino, hydroxyl, mercapto, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 alkylamino, C 1 -C 6 dialkylamino, C 1 -C 6 alkylthio; a bile acid based alkoxy group, a sugar moiety, a PEG moiety, and -0-CH 2 -CH(OR 6 )CH 2 X 4 R 6 wherein:
  • V being CO 2 H.
  • the present invention provides the following Y 1 groups:
  • R 31 and each X 3 is defined as in formula (IV);
  • R 34 is independently hydrogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
  • the present invention provides the following compounds:
  • R 31 is defined as in formula (IV);
  • R 34 is independently hydrogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and each X 3 independently is C 1 - C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, C 1 -C 6 alkoxy, C 1 -C 6 alkylamino, C 1 -C 6 dialkylamino, or C 1 -C 6 alkylthio; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
  • compounds of the follow formulae are provided:
  • R 1 is an aryl group of formula wherein each R 2 is independently halogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl,;
  • X 1 is selected from the group consisting of O, S, and NR 3 , wherein R 3 is hydrogen or C 1 -C 6 alkyl; and Y 1 is -C(R 4 ) I - wherein R 4 is independently hydrogen or C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl.
  • the term "cleavable linker”, such as, e.g., Z, refers to a linker which has a short half life in vivo.
  • the breakdown of the linker Z in a compound D-Z-M (supra) releases or generates lonidamine or a lonidamine analog.
  • the cleavable linker has a half life of less than ten hours. In one embodiment, the cleavable linker has a half life of less than an hour. In one embodiment, the half life of the cleavable linker is between one and fifteen minutes.
  • the cleavable linker is hydrolyzed by an esterase enzyme.
  • the linker is a self-immolating linker, such as that disclosed in U.S. patent publication 2002/0147138, to Firestone; PCT Appl. No. US05/08161 and PCT Pub. No. 2004/087075.
  • the linker is a substrate for enzymes. See generally Rooseboom et al., 2004, Pharmacol. Rev. 56:53-102.
  • R C 10 -C 22 alkyl or alkylene
  • R 1 is defined as above.
  • the present invention provides the compounds
  • R 1 , X 1 , Y 1 and X 2 are defined as above.
  • the present invention provides a method for synthesizing prodrugs of lonidamine analogs by reacting
  • X 10 is Cl, Br, I, an alkylsufonyloxy, or arylsulfonyloxy; and R 31 , Y 1 , and X 2 are defined as in formula (IV); in presence of a base such as, for example, a trialkylamine, an alkali metal hydride, or an alkali metal carbonate in a solvent such as dimethylformamide or THF.
  • a base such as, for example, a trialkylamine, an alkali metal hydride, or an alkali metal carbonate in a solvent such as dimethylformamide or THF.
  • Scheme 1 provides a method of synthesizing an acyloxymethyl version of a prodrug of formula (FV) by adapting a method from the reference Sobolev et al., 2002, J. Org. Chem. 67:401-410.
  • V is C 1 -C 6 alkyl
  • Scheme 2 below provides a method for synthesizing a phosphonooxymethyl version of a prodrug of formula (IV) by adapting a method from Mantyla et al., 2004, J. Med. Chem. 47:188-195.
  • V is C 1 -C 6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl.
  • Each Ri in the Schemes is defined as in formula (IV).
  • Lonidamine and lonidamine analogs and derivatives can be prepared using known synthetic methods, as described supra. Methods of synthesizing prodrugs of other carboxylic acids or related compounds are described by Nudelman et al., 2005, J. Med. Chem., 48:1042-1054; Mendes et al., 2002, Bioorg. Med. Chem. 10:809-16; Iley et al., 1999, Eur. J. Pharm. Sci. 9:201-5; Iley et al., 2001, J. Chem. Soc. Perkin Trans. 2 749-53; Kubo et al., 1993, J. Med. Chem.
  • ester prodrugs of lonidamine and lonidamine analogs can be prepared by modifications known to one of skill in the art of the procedure described in U.S. Patent No. 6,146,658, as is administration of a polymorphic forms, enantiomeric forms, tautomeric forms, solvates, hydrates, and the like.
  • the prodrug of the lonidamine or a lonidamine analog can have functionality such as, for example, hydroxyl, amino, alkylamino, mercapto, or a group having an -NH- moiety that is further protected, for example by ester, carbamate, and carbonate moieties, yielding a prodrug of a prodrug.
  • the prodrug of the lonidamine or a lonidamine analog can have functionality that is converted to its acid or base salt.
  • the prodrug of the lonidamine or lonidamine analog is provided in the form of a pharmaceutically acceptable salt.
  • Pharmaceutically acceptable salts include addition salts with acids, as well as the salts with bases.
  • Suitable acids for the formation of acid addition salts are, for example, mineral acids, such as hydrochloric, hydrobromic, sulphuric or phosphoric acid, and organic acids, such as organic sulphonic acids, for example, benzenesulphonic, 4-toluenesulphonic or methanesulphonic acid, and organic carboxylic acids, such as acetic, lactic, palmitic, stearic, malic, maleic, fuinaric, tartaric, ascorbic or citric acid.
  • Acid salts of the tertiary amine moiety confer increased aqueous solubility.
  • citric acid salts is preferred.
  • the invention includes compounds and prodrugs that are single isomers of the formulae described herein (e.g., single enantiomers of compounds having a single chiral center), as well as solvate and hydrate forms thereof.
  • a suitable base for the formulation of base addition salts of prodrugs of lonidamine and lonidamine analogs is a primary amine, a secondary amine, a tertiary amine, an amino acid, or a naturally occurring ⁇ -amino acid.
  • aminoacids include but are limited to glycine, lysine, and arginine.
  • the cation employed in the base addition salt of lonidamine or a lonidamine analog is sodium, potassium, ammonium, or calcium.
  • a base addition salt of a prodrug of lonidamine or a lonidamine analog is formed employing lysine, glycine, or arginine as a base.
  • one equivalent of an amine (wherein amine is as described above) is mixed with one equivalent of a prodrug of lonidamine or a lonidamine analog in water. The mixture is shaken or sonicated to yield a homogenous solution of the base addition salt of the prodrug of lonidamine or the lonidamine analog in water.
  • one equivalent of a prodrug of lonidamine or a lonidamine analog is mixed in water with one equivalent of a metal hydroxide, oxide, bicarbonate, or carbonate wherein the metal comprises sodium, potassium, or calcium resulting in the formation of the metal salt of lonidamine or the lonidamine analog.
  • prodrugs of lonidamine and lonidamine analogs comprise polymer conjugates.
  • lonidamine or a lonidamine analog other compound can be linked directly to a polymer or indirectly via a cleavable linker. See, e.g., Uchegbu, 1999, Pharmaceutical Journal 263:355-358.
  • the prodrug comprises lonidamine or a lonidamine analog covalently linked to a polyethylene glycol.
  • PEG moieties and methods for forming prodrugs with them that can be used in or to make compounds of the invention are described in U.S. Patent Nos. 6,608,076; 6,395,266; 6,194,580; 6,153,655; 6,127,355; 6,111,107; 5,965,566; 5,880,131; 5,840,900; 6,011,042 and 5,681,567. See also Duncan, 2003, Nature Reviews 2:347-360.
  • the prodrug comprises lonidamine or a lonidamine analog covalently linked to another polymer, such as N-(2- hydroxypropyl)methacrylamide), polyglutamate, polysaccharide, proteins, and other polymers. See, e.g., Duncan, 2003 (supra).
  • multi-drug polymer prodrug conjugates can be formed of a water-soluble and non-peptidic polymer covalently attached to lonidamine or a lonidamine analog. See, e.g., U.S. Patent Publication No. 20050112088.
  • the conjugate comprises a plurality of drug molecules attached to the polymer via a hydrolytically degradable linkage, which is released over time following administration of the conjugate to a subject.
  • hydrolytically degradable linkages include, for example, carboxylate ester, carbonate ester, phosphate ester, anhydride, acetal, ketal, acyloxyalkyl ether, imine, orthoester, and oligonucleotides.
  • hydrolytically degradable linkages include esters such as carboxylate and carbonate esters are particularly preferred linkages.
  • the particular linkage and linkage chemistry employed will depend upon the particular drug (e.g., lonidamine or lonidamine analog), the presence of additional functional groups within the active agent, and the like, and can be readily determined by one skilled in the art.
  • a central core organic radical is derived from a molecule that provides a number of polymer attachment sites approximately equal to the desired number of water soluble and non-peptidic polymer arms.
  • the central core molecule of the multi-arm polymer structure can be, for example, a polyol, polythiol, or a polyamine bearing at least three hydroxyl, thiol, or amino groups available for polymer attachment.
  • a "polyol” is a molecule comprising a plurality (greater than 2) of available hydroxyl groups.
  • polythiol is a molecule that possesses a plurality (greater than 2) thiol groups.
  • a "polyamine” is a molecule comprising a plurality (greater than 2) of available amino groups.
  • Illustrative polyols that are preferred for use as the polymer core include aliphatic polyols having from 1 to 10 carbon atoms and from 1 to 10 hydroxyl groups, including for example, ethylene glycol, alkane diols, alkyl glycols, alkylidene alkyl diols, alkyl cycloalkane diols, 1,5-decalindiol, 4,8-bis(hydroxymethyl)tricyclodecane, cycloalkylidene diols, dihydroxyalkanes, trihydroxyalkanes, and the like.
  • Cycloaliphatic polyols include straight chained or closed-ring sugars and sugar alcohols, such as mannitol, sorbitol, inositol, xylitol, quebrachitol, threitol, arabitol, erythritol, adonitol, dulcitol, facose, ribose, arabinose, xylose, lyxose, rhamnose, galactose, glucose, fructose, sorbose, mannose, pyranose, altrose, talose, tagitose, pyranosides, sucrose, lactose, maltose, and the like.
  • sugar alcohols such as mannitol, sorbitol, inositol, xylitol, quebrachitol, threitol, arabitol, erythritol, adonitol,
  • Aromatic polyols include derivatives of glyceraldehyde, glucose, ribose, mannose, galactose, and related stereoisomers.
  • Aromatic polyols may also be used, such as l,l,l-tris(4'-hydroxyphenyl) alkanes, such as 1,1,1- tris(4-hydroxyphenyl)ethane, (l,3-adamantanediyl)diphenol, 2,6- bis(hydroxyalkyl)cresols, 2,2'alkylene-bis(6-t-butyl-4-alkylphenols), 2,2'-alkylene-bis(t- butylphenols), catechol, alkylcatechols, pyrogallol, fluoroglycinol, 1,2,4-benzenetriol, resorcinol, alkylresorcinols, dialkylresorcinols, orcinol monohydrate, olivetol, hydroquinone,
  • polysaccharides examples include crown ethers, cyclodextrins, dextrins and other carbohydrates (e.g., monosaccharides, oligosaccharides, and polysaccharides, starches and amylase).
  • Other polyols include glycerol, trimethylolpropane, reducing sugars such as sorbitol or pentaerythritol, and glycerol oligomers, such as hexaglycerol.
  • Exemplary polyamines include aliphatic polyamines such as diethylene triamine, N,N',N"-trimethyldiethylene triamine, pentamethyl diethylene triamine, Methylene tetramine, tetraethylene pentamine, pentaethylene hexamine, dipropylene triamine, tripropylene tetramine, bis-(3-aminopropyl)-amine, bis-(3-aminopropyl)- methylamine, and N,N-dimethyl-dipropylene-triamine.
  • Naturally occurring polyamines that can be used in the present invention include putrescine, spermidine, and spermine.
  • the polymer conjugate can include a targeting moiety, such as an antibody or ligand, that binds to an antigen, marker or receptor on a target cell or target cell population.
  • a targeting moiety such as an antibody or ligand, that binds to an antigen, marker or receptor on a target cell or target cell population.
  • targeting moieties are further discussed infra.
  • immunoconjugate prodrugs comprising an antibody and a drug are provided.
  • the conjugate includes an antibody conjugated to lonidamine or a lonidamine analog or other compound according to the present invention.
  • the antibody is typically conjugated lonidamine or a lonidamine analog via a linker.
  • the antibody is typically directed to an antigen on the target cell or cell population.
  • the antibody can be an antigen to a tumor-specific antigen, such as a prostate cancer antigen (e.g., prostatic acid phosphatase (PAP); AMACR (x- methylacyl-CoA racemase); CA 15-3 ; prostate-specific membrane antigen (PSMA); Prol09, a human zinc- ⁇ 2-glycoprotein (Freje et al., 1993, Genomics 18(3):575-587); Prol 12, a human cysteine-rich protein with a zinc-finger motif (Liebhaber et al., 1990, Nucleic Acid Research 18(13):3871-79; WO9514772 and WO9845436); Prol 11, a prostate-specific transglutaminase (Dubbink et al., 1998, Genomics 51(3):434-444); Prol 15, a novel serine protease with transmembrane, LDLR
  • PAP
  • Patent No. 5,665,874 and WO9403599 Prol 13, a homeobox gene, HOXB13 (Steinicki et al., 1998, J. Invest. Dermatol. 111 :57-63); Prol 14, a human tetraspan NET-I (WO9839446); and Prol 18, a human JM27 protein (WO9845435). See also U.S. Patent No. 6,902,892.
  • the antibody is directed to another cancer antigen, such as, for example, CA- 125 (epithelial cancer), HER2 (breast cancer), Topoisomerase II alpha (ovarian epithelial cancer), Werner helicase interacting protein (ovarian epithelial cancer), HEXIMl (ovarian epithelial cancer), FLJ20267 (ovarian epithelial cancer),
  • CA- 125 epidermal cancer
  • HER2 breast cancer
  • Topoisomerase II alpha ovarian epithelial cancer
  • Werner helicase interacting protein ovarian epithelial cancer
  • HEXIMl ovarian epithelial cancer
  • FLJ20267 ovarian epithelial cancer
  • Deadbox protein-5 ovarian epithelial cancer
  • Kinesin-like 6 ovarian epithelial cancer
  • p53 ovarian epithelial cancer
  • NY-ESO-I ovarian epithelial cancer
  • linking groups known in the art for making antibody-drug conjugates, including, for example, those disclosed in U.S. patent publication 20020147138.
  • Other suitable linkers include, for example, disulfide groups, thioether groups, acid labile groups, photolabile groups, peptidase labile groups, or esterase labile groups, disulfide and thioether groups being preferred.
  • Conjugates of the antibody and lonidamine or a lonidamine analog can be made using a variety of bifunctional protein coupling agents such as N-succinimidyl (2-pyridyidithio)propionate (SPDP), succinimidyl-fN-maleimidomethyl) cyclohexane-1-carboxylate, iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCL), active esters (such as disuccinimidyl suberate), aldehydes (such as glutaraldehyde), bis-azido compounds (such as his (p-azidobenzoyl) hexanediamine), bis-diazonium derivatives (such as bis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (such as toluene 2,6- diisocyanate), and bis-active fluorine
  • the conjugates can be N-succinimidyl (2- pyridyldithio)pro ⁇ ion- ate (SPDP) (Carlsson et al., 1978, Biochem. J. 173:723-737) and N-succinimidyl (2- ⁇ yridylthio)pentanoate (SPP) to provide for a disulfide linkage.
  • SPDP N-succinimidyl (2- pyridyldithio)pro ⁇ ion- ate
  • SPP N-succinimidyl (2- ⁇ yridylthio)pentanoate
  • the linker may be attached to the lonidamine or lonidamine analog molecule at various positions, depending on the type of the linkage.
  • an ester linkage may be formed by reaction with a hydroxyl group using conventional coupling techniques. The reaction may occur at a position having a hydroxyl group or a hydroxymethyl group
  • a prodrug of lonidamine or a lonidamine analog or other compound according to the present invention can be administered to a target cell or target cell population using Antibody-Dependent Enzyme Mediated Prodrug Therapy (ADEPT).
  • ADEPT Antibody-Dependent Enzyme Mediated Prodrug Therapy
  • an antibody is conjugated to a prodrug-activating enzyme, which converts a prodrug to an active agent (e.g., lonidamine or a lonidamine analog). See, for example, Rooseboom et al., 2004, Pharmacol. Rev. 56:53-102; Uchegbu, 1999, Pharmaceutical Journal 263:355-358.
  • the antibody binds to an antigen or marker on a target cell or target cell population.
  • the prodrug is selected to be a substrate for the enzyme.
  • the prodrug is then administered to the patient, where it is preferentially cleaved (activated) at the target cells or target cell population by the enzyme.
  • the antibody can bind to any suitable antigen marker, such as those discussed supra.
  • the enzyme component of the immunoconjugate useful for ADEPT includes any enzyme capable of acting on a prodrug in such a way so as to covert it into its more active, cytotoxic form.
  • Enzymes that are useful in the method of this invention include, but are not limited to, alkaline phosphatase useful for converting phosphate-containing prodrugs into free drugs; arylsulfatase useful for converting sulfate-containing prodrugs into free drugs; cytosine deaminase useful for converting non-toxic fluorocytosine into the anti-cancer drug, 5-fluorouracil; proteases, such as serratia protease, thermolysin, subtilisin, carboxypeptidases and cathepsins (such as cathepsins B and L), that are useful for converting peptide-containing prodrugs into free drugs; D-alanylcarboxypeptidases, useful for converting prodrugs that contain D-amino acid substituent
  • Antibody-abzyme conjugates can be prepared as described herein for delivery of the abzyme to a target cell population.
  • the enzymes can be covalently bound to the antibodies by techniques well known in the art such as the use of the heterobifunctional crosslinkmg reagents as discussed above.
  • fusion proteins comprising at least the antigen binding region of an antibody linked to at least a functionally active portion of an enzyme can be constructed using recombinant DNA techniques well known in the art (see, e.g., Neuberger et al., 1984, Nature 312:604-608).
  • a prodrug of lonidamine or a lonidamine analog or other compound according to the present invention can be administered to a target cell or target cell population using Gene-Dependent Enzyme Mediated Prodrug Therapy (GDEPT) or Virus-Directed Enzyme Prodrug Therapy (VDEPT).
  • GDEPT Gene-Dependent Enzyme Mediated Prodrug Therapy
  • VDEPT Virus-Directed Enzyme Prodrug Therapy
  • GDEPT Gene-Dependent Enzyme Mediated Prodrug Therapy
  • VDEPT Virus-Directed Enzyme Prodrug Therapy
  • the transgene can be delivered, for example, by any suitable targeting system, including liposomes, viral vectors (e.g., adenovirus, retrovirus, EBV), direct injection or the like.
  • the enzyme can be, for example, the HSV-tk gene, beta-glucuronidase, bacterial nitroreductase, carboxypeptidase, CYP2B1 and p450 reductase, rabbit CYP4B1, carboxylesterase, cytosine deaminase or thymidine kinase. See, e.g., Xu et al., 2001 (supra); Denny, 2003 (supra).
  • a prodrug comprises an emulsion or a micro-emulsion.
  • emulsion refers to a colloidal dispersion of two immiscible liquids in the form of droplets, whose diameter, in general, are between 0.1 and 3.0 microns and which is typically optically opaque, unless the dispersed and continuous phases are refractive index matched.
  • Such systems possess a finite stability, generally defined by the application or relevant reference system, which may be enhanced by the addition of amphiphilic molecules or viscosity enhancers.
  • microemulsion refers to a thermodynamically stable isotropically clear dispersion of two immiscible liquids, such as oil and water, stabilized by an interfacial film of surfactant molecules.
  • the microemulsion has a mean droplet diameter of less than 200 ran, in general between 10- 50 nm.
  • mixtures of oil(s) and non-ionic surfactant(s) form clear and isotropic solutions that are known as self-emulsifying drug delivery systems (SEDDS) and have successfully been used to improve lipophilic drug dissolution and oral absorption.
  • SEDDS self-emulsifying drug delivery systems
  • the emulsion comprises lonidamine or a lonidamine analog, or other compound according to the present invention.
  • the composition of the emulsion according to the solubility of the drug form is soluble.
  • the emulsion is typically stabilized by an emulsifying agent.
  • the emulsifying agent optionally can comprise a targeting agent, such as an antibody or peptide. See generally, Uchegbu, 1999, Pharmaceutical Journal 263:355-358.
  • a prodrug can be administered in a liposome. See generally, e.g., Uchegbu, 1999, Pharmaceutical Journal, 263:309-318.
  • Liposomes generally comprise an "outer membrane” or bulk aqueous phase and "central core” or inner aqueous phase.
  • Liposomal formulations can be prepared, for example, by combining a lonidamine or a lonidamine analog or a prodrug form formulation with a phospholipid such as, for example, dimyristoyl phosphatidyl choline (DMPC), phosphatidyl choline (PC), dipalmitoyl-phosphatidyl choline (DPPC), and distearoylphosphatidyl choline (DSPC).
  • Suitable phosphatidyl glycerols include, for example, dimyristoyl phosphatidylglycerol (DMPG) and egg phosphatidyl-glycerol (EPG).
  • the liposome is a stealthed liposome.
  • “Stealthed” liposomes comprise a biologically effective amount of at least a first stealthing agent in operative association with the outer membrane of the liposome.
  • a “stealthing agent” is a component that increases the biological half life of a liposome or liposome-like composition when operatively associated with the outer membrane of the liposome or liposome-like composition.
  • the outer membrane of the liposome or liposome-like composition is preferably "coated” with the one or more stealthing agents.
  • Stealthing agents can include, for example, a range of biocompatible hydrophilic polymers, such as polyamines, polylactic acid, polyglycolic acid, polylactic-polyglycolic acid (PLGA), polypeptides and related materials.
  • the stealthing agent is a polyethylene glycol (PEG) component, wherein the resulting stealthed liposomes are termed "PEGylated liposomes".
  • Other stealthing agents can include, for example, cholesterol, polyvinyl-pyrrolidone polyacrylamide lipids, glucoronic acid lipids or the high phase transition temperature phospholipid distearoyl phosphatidylcholine.
  • stealthed or PEGylated liposomes further comprise an antibody or antigen-binding fragment thereof, to target the liposome to the target cell type (e.g., a cancer cell).
  • the antibody can bind to, for example, any suitable antigen or marker on a target cell or target cell population, as discussed supra.
  • Liposomes containing the antibody can be prepared by methods known in the art, such as described in Epstein et al., 1985, Proc. Natl. Acad. Sci. USA 82:3688; Hwang et al., 1980, Proc. Natl Acad. Sci. USA77:4030; U.S. Patent Nos.
  • Liposomes with enhanced circulation time are disclosed in, for example, U.S. Patent No. 5,013,556.
  • Fab 1 fragments can be conjugated to the liposomes as described in, for example, Martin et al. (1982, J. Biol. Chem. 257:286-288) via a disulfide interchange reaction.
  • other ligands can be used to target liposomes.
  • the folate receptor overexpressed in ovarian carcinoma
  • the cell adhesion molecules e.g., selectins and integrins
  • Liposomes bearing specific ligands such as folate may be used to target ovarian carcinomas, while specific peptides or carbohydrates may be used to target integrins and selectins.
  • Other ligands can be used according to the target cells.
  • the liposomes are "antibody-coated” stealthed or
  • PEGylated liposomes wherein at least one of lonidamine, a lonidamine analog or other compound according to the present invention is operatively associated with the liposome or dispersed within the liposomal formulation.
  • the lonidamine, a lonidamine analog or other compound according to the present invention is operatively associated with or maintained within the central core of the liposome.
  • liposomal-like compositions can be used.
  • Such compositions can be, for example, non-ionic surfactant vesicles (niosomes) or other synthetic vesicles. See, e.g., Uchegbu, 1999, Pharmaceutical Journal 263:309-318.
  • the compounds of the present invention can be used in any application for which use of lonidamine and/or a lonidamine analogs is contemplated.
  • Applications include, but are not limited to, treatment or prophylaxis of cancer, treatment or phophylaxis of BPH or a symptom of BPH, inhibition of spermatogenesis, treatment or phophylaxis macular degeneration, and antiangiogenic applications. See the references cited herein above, as well as PCT patent publication Nos. WO2004/064734;
  • the prodrugs of lonidamine or lonidamine analogs of the present invention provide improved pharmacokinetic properties compared to lonidamine.
  • the compounds of the present invention have greater bioavailability that lonidamine.
  • these compounds have as much bioavailability as lonidamine.
  • these compounds have 30-80% bioavailability as lonidamine.
  • the present invention provides prodrugs of lonidamine or lonidamine analogs having a higher logP than lonidamine. In another embodiment, the present invention provide prodrugs of lonidamine or lonidamine analogs having logP within ⁇ 0.5 of lonidamine. hi another embodiment, the present invention provides prodrugs of lonidamine or lonidamine analogs having a lower logP than lonidamine.
  • the lipophilicity of the moieties provided by the present invention such as, for example, R 3 , Z 10 , M, X 1 , Y 1 , and X 2 can modulate the logP of the prodrugs of the present invention.
  • the present invention provides prodrugs of lonidamine or lonidamine analogs having a lower toxicity than lonidamine.
  • the present invention provide prodrugs of lonidamine or lonidamine analogs as much toxicity as lonidamine.
  • the organ specific uptake of the prodrugs of the present invention can lead to altered toxicity of the prodrugs of the present invention.
  • the dose, schedule and duration of administration of the prodrug will depend on a variety of factors, hi some embodiments, the dose, schedule and duration of the prodrug is determined according to the dosage of lonidamine and/or the lonidamine analog to be administered. Factor to be considered include the molecular weight, pharmacokinetic property, pharmacodynamic property, or a combination thereof of the specific prodrug selected. Other important factors include the age, weight and health of the subject, the severity of symptoms, if any, the subject's medical history, co-treatments, goal (e.g., prophylaxis or prevention of relapse), preferred mode of administration, the formulation used, patient response, and the like.
  • Guidance concerning administration is provided by prior experience using lonidamine administered to treat cancer (e.g., 150 mg doses three times a day for a period of about a month), and from new studies in humans and other mammals. Cell culture studies are frequently used in the art to optimize dosages.
  • prodrugs the scientific literature (including, for example, patent and non-patent publications cited herein) provides considerable guidance as to dosages, formulations and dosage forms for specific prodrugs or classes of prodrugs, e.g., dosages known or predicted to result in a biologically effective serum level of the prodrug (or metabolite) in serum.
  • a prodrug can be administered to deliver a dose of lonidamine or a lonidamine analog in the range of about 1 mg to about 100 mg per kg of body weight of the patient to be treated, with more than one dose being administered.
  • a prodrug is administered to deliver a dose of lonidamine or a lonidamine analog in the range of about 1 mg to about 5 mg per kg of body weight of the patient to be treated.
  • a prodrug is administered to deliver a dose of lonidamine or a lonidamine analog in the range of about 1 mg to about 10 mg per kg of body weight of the patient to be treated.
  • a prodrug is administered to deliver a dose of lonidamine or a lonidamine analog of about 5 to about 25 mg per kg of body weight of the patient to be treated.
  • an effective dose is about 5 mg/kg to about 50 mg/kg of lonidamine or the lonidamine analog.
  • dosing regimens contemplated include, for example and not for limitation, "low dosing” (e.g., dosaged in the range of 1-300 mg per day total daily dosage, 5-300 mg/day, 5-70 mg/day, 1-25 mg/day, 20-45 mg/day, 40-65 mg/day, 40-70 mg/day, 50-100 mg/day, 50-200 mg/day, and 50-300 mg/day), "high dosing” (e.g., total daily doses greater than 0.5 g, such as doses in the range 0.5 - 5 g/day, 0.5 - 3 g/day, 0.5 - 1 g/day and 1-3 g/day, or higher doses), and "intermediate dosing” (e.g., doses greater than 300 and less than 500 mg/day, such as doses in the range >300-400 or 400 ⁇ 500, e.g., 450 mg/day).
  • low dosing e.g., dosaged in the range of 1-300 mg
  • the effective dose of a prodrug can be administered daily or once every other day or once a week to the patient.
  • multiple administrations of the prodrug are employed.
  • the entire dose may be administered once daily, or the dose may be administered in multiple smaller doses through the course of a day.
  • the dose may be divided into two smaller doses and administered twice daily, or divided into three smaller doses and administered thrice daily.
  • the dose may be combined and given every other day, or even less frequently, but in any event, the dose is repeatedly administered over a period of time.
  • the administration of the effective dose is continued for multiple days, typically for at least five consecutive days, and often for at least a week and often for several weeks or more.
  • the prodrug is administered once (qday), twice (bid), three times (tid), or four times (qid) a day or once every other day (qod) or once a week (qweek), and treatment is continued for a period ranging from three days to two weeks or longer.
  • the treatment is continued for one to three months.
  • the treatment is continued for a year.
  • a patient may be administered the prodrug for a week, a month, two months, three months, six months, or a year or longer.
  • treatment may continue indefinitely throughout the life of the patient. As is well understood in medicine, treatment may be suspended temporarily if toxicity is observed or for the convenience of the patient without departing from the scope of the invention.
  • the present invention provides a pharmaceutical formulation of a prodrug according to the invention suitable for oral administration (including tablets, capsules, and pills) and contains between 1 and 100 mg of the prodrug, and in another embodiment between 1 and 10 mg of the prodrug.
  • the formulation contains between 200 and 1000 mg of the prodrug, and in another embodiment between 500 and 1000 mg of the prodrug.
  • the present invention provides controlled and sustained release formulations of the prodrugs that allow once a day oral dosing.
  • sustained release formulations can contain between 1 mg and 3 g of the prodrug, with various alternative embodiments, including one that contains between 1 mg and 10 mg of the prodrug; another that contains between 150 and 500 mg of the prodrug; and another that contains between 750 mg and 2 g of the prodrug.
  • preferred dosage forms include pills, tablets, capsules, caplets, and the like, optionally formulated for sustained release.
  • Other suitable forms for oral administration include troches, elixirs, suspensions, syrups, wafers, lozenges, and the like.
  • Other modes of administration are also contemplated, including parenteral, inhalation spray, transdermal, rectal, intraprostetic injection (e.g., of prodrug- containing microparticles) and other routes.
  • dosing schedules are for illustration and not limitation* and that a dosing schedule may change during a course of therapy based on, for example, a patient's response to the drug administered.
  • the HCl salt of Compound A was prepared by dissolving Compound A in ethyl acetate followed by the addition of 1.1 equivalents of HCl in dioxane (4 M) at O 0 C. After evaporating the solvent the HCl salt of Compound A was obtained as a white solid.
  • An aqueous formulation of Compound A is made by dissolving Compound A it or its hydrochloride salt in water.
  • the pharmacokinetic properties of Compound A were determined by measuring plasma levels of Compound A after oral or intravenous administration of a aqueous pharmaceutical formulation of Compound A in rats.
  • the pharmacokinetics of Compound A, including it clearance, distribution and bioavailability were observed to be similar to those of lonidamine.

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Abstract

The invention provides prodrugs of lonidamine and lonidamine analogs as well as methods of using such prodrugs.

Description

PRODRUGSOFLONIDAMINEANDLONIDAMINEANALOGS
CONTINUITY
[0001] This application claims the benefit of U.S. Provisional Patent Application No. 60/586,934 (filed July 8, 2004) and U.S. Provisional Patent Application No. 60/624,505 (filed November 1, 2004), the disclosures of which are incorporated by reference herein.
BACKGROUND OF THE INVENTION [0002] Lonidamine (LND), also known as l-(2,4-dichlorobenzyl)-lH-indazole-3- carboxylic acid, is an anti-cancer drug approved for the treatment of lung, breast, prostate, and brain cancer in a few European countries. The mechanism of action of lonidamine may involve interference with the energy metabolism of neoplastic cells by disruption of the mitochondrial membrane and by inhibition of hexokinase. Lonidamine also has anti- spermatogenic activity and has been shown to inhibit germ cell respiration. Lonidamine has perhaps been most extensively been studied for use in the treatment of advanced breast cancer. For example, the reference Mansi et al., Sep. 1991, Br. J. Cancer 64(3):593-7, reports a phase II study in which lonidamine was administered in a daily divided oral dose of 600 mg. Of the 28 patients evalutable for response, three (11%) achieved a partial response (4-24+ months); three (11%) had a minor response; two had stable disease (greater than 3 months); and 20 progressed. The investigators reported no clear relationship between lonidamine levels and clinical response or toxicity and concluded that lonidamine appeared to be active against advanced breast cancer, and that lonidamine' s low toxicity would allow combination studies.
[0003] Combination studies of lonidamine in advanced breast cancer followed this report, particularly studies in combination with epirubicin or doxorubicin. For examples, see the references Iaffaioli et al., Sep. 1995, Breast Cancer Res. Treat. 35(3):243-8 (phase II trial of high-dose epirubicin, lonidamine, and alpha 2b interferon); Gardin et al., Jan. 1996, Eur. J. Cancer 32A(l):176-7 (phase II trial of lonidamine plus epirubicin and cyclophosphamide); Dogliotti et al., Apr. 1996, J. Clin. Oncol. 14(4): 1165-72 (multicenter prospective randomized trial ~ reports that lonidamine significantly increases the activity of epirubicin); Gebbia et al., Nov. 1997, Anticancer Drugs 8(10):943-8 (phase II trial of cisplatin and epirubicin plus oral lonidamine as first-line treatment for metastatic breast cancer); Amadori et al., Jun. 1998, Breast Cancer Res. Treat. 49(3):209-17 (multicenter prospective randomized trial ~ reports modulating effect of lonidamine on response to doxorubicin in metastatic breast cancer); Dogliotti et al., 1998, Cancer Chemother. Pharmacol. 41(4):333-8 (pilot study of cisplatin, epirubicin, and lonidamine combination regimen as first-line chemotherapy for metastatic breast cancer); Nistico et al., Aug. 1999, Breast Cancer Res. Treat. 56(3):233-7 (study of weekly dosed epirubicin plus lonidamine in advanced breast carcinoma); and Pacini et al., May 2000, Eur J. Cancer 36(8):966-75 (multicentric randomised study of FEC (5-fluorouracil, epidoxorubicin and cyclophosphamide) versus EM (epidoxorubicin and mitomycin-C) with or without lonidamine as first-line treatment). Surprisingly, however, a more recent reference, Berruti et al., 15 Oct. 2002, J. Clin. Oncol. 20(20):4150-9, reports that, in a phase III study with a factorial design, time to progression in metastatic breast cancer patients treated with epirubicin was not improved by the addition of either cisplatin or lonidamine (see also Berruti et al., Jul.-Aug. 1997, Anticancer Res. 17(4A):2763-8).
[0004] Lonidamine has also been studied in lung cancer, particularly non-small cell lung cancer (see the reference Joss et al., Sep. 1984, Cancer Treat. Rev. 11(3):205-36) in combination with radiation or other anti-cancer agents. For examples, see the references Privitera et al., Dec. 1987, Radiother. Oncol. 10(4):285-90 (phase II double-blind randomized study of lonidamine and radiotherapy in epidermoid carcinoma of the lung); Gallo-Curcio et al., Dec. 1988, Semin. Oncol. 15(6 Suppl 7):26-31 (chemotherapy or radiation therapy plus and minus lonidamine); Giaccone et al., 28 Feb. 1989, Tumori 75(l):43-6 (preliminary analysis of lonidamine versus polychemotherapy); Ianniello et al., 1 JuI. 1996, Cancer 78(l):63-9 (multicenter randomized clinical trial of cisplatin, epirubicin, and vindesine with or without lonidamine); Gridelli et al., Mar. -Apr. 1997, Anticancer Res. 17(2B): 1277-9 (phase II trial of VM-26 plus lonidamine in pretreated small cell lung cancer); Cornelia et al., May 1999, J. Clin. Oncol. 17(5): 1526-34 (phase II randomized trial of cisplatin, gemcitabine, and vinorelbine); DeMarinis et al., May. -Jun. 1999, Tumori 85(3):177-82 (phase III randomized trial of vindesine and lonidamine in elderly patients); and Portalone et al., Jul.-Aug. 1999, Tumori 85 (4):239-42 (phase II study with cisplatin, epidoxorubicin, vindesine and lonidamine).
[0005] Lonidamine has been studied as a treatment for other cancers (see the references
Robustelli et al., Apr. 1991, Semin. Oncol. 18(2 Suppl 4):18-22; and Pacilio et al., 1984, Oncology 41 Suppl 1:108-12), including: favorable B-cell neoplasms (see the reference Robins et al., Apr. 1990, Int. J. Radiat. Oncol. Biol. Phys. 18(4):909-20, which describes two pilot clinical trials and laboratory investigations of adjunctive therapy (whole body hyperthermia versus lonidamine) to total body irradiation); advanced colorectal cancer (see the references Passalacqua et al., Jun. 30, 1989, Tumori 75(3):277-9, and Zaniboni et al., Nov. -Dec. 1995, Tumori 81(6):435-7, which describes a phase II study of mitomycin C and lonidamine as second-line therapy); advanced gastric carcinoma (see the reference Barone et al., 15 Apr. 1998, Cancer 82(8): 1460-7, which describes two parallel randomized phase II studies with a 5-fluorouracil-based or a cisplatin-based regimen); malignant glioma (see the references Carapella et al., May 1989, J. Neurooncol. 7(l):103- 8, and Jul.-Dec. 1990, J Neurosurg Sci. 34(3-4):261-4); metastatic cancers (see the references Weinerman, 1990, Cancer Invest. 8(5):505-8, which describes a phase I study of lonidamine and human lymphoblastoid alpha interferon; DeAngelis et al., Sep. 1989, J Neurooncol. 7(3):241-7, and U.S. Patent No. 5,260,327, which describe the combined use of radiation therapy and lonidamine in the treatment of brain metastases; and Weinerman et al., Jun. 1986, Cancer Treat. Rep. 70(6):751-4, which reports a phase II study of lonidamine in patients with metastatic renal cell carcinoma); advanced ovarian cancer (see the references Bottalico et al., Nov.-Dec. 1996, Anticancer Res. 16(6B):3865-9; DeLena et al., Oct. 1997, J. Clin. Oncol. 15(10):3208-13, which reports the revertant and potentiating activity of lonidamine in patients with ovarian cancer previously treated with platinum; and DeLena et al., Feb. 2001, Eur. J. Cancer 37(3):364-8, which describes a phase II study of paclitaxel, cisplatin and lonidamine); and recurrent papillary carcinomas of the urinary bladder (see the reference Giannotti et al., 1984, Oncology 41 Suppl 1 : 104- 7, which describes treatment results after administration of lonidamine plus adriamycin versus adriamycin alone in adjuvant treatment).
[0006] Lonidamine has been studied as a treatment for benign prostatic hypertrophy (BPH), also known as benign prostatic hyperplasia (see U.S. Application Publication 20040167196; see also PCT Publication No. WO 04/064735; both incorporated herein by reference). BPH is a disease in which prostate epithelial cells grow abnormally and block urine flow, and currently afflicts more than 10 million adult males in the United States alone and many millions more throughout the rest of the world.
[0007] Lonidamine, which contains a free carboxylic acid group, is poorly soluble in aqueous or non-aqueous solvents. Many carboxylic acid-containing drugs have reduced therapeutic effectiveness if orally administered due to such physicochemical limitations (for examples of several carboxyl-containing beta-lactam antibiotics, see Durckheimer et. al., Advances in Drug Research, Academic Press, London 17:61-634). There remains a need for lonidamine analogs with improved pharmacokinetic properties for use in the treatment of cancer, BPH and other diseases, particularly for analogs with greater bioavailability. The present invention meets these needs and provides novel prodrugs of lonidamine and its derivatives, described in more detail in the following section.
BRIEF SUMMARY OF THE INVENTION [0008] The invention relates to compounds and prodrugs of lonidamine and lonidamine analogs. Compounds according to the present invention include tertiary amine prodrugs, linker prodrugs, polymer conjugates, immunoconjugate prodrugs and other prodrug forms of lonidamine and lonidamine analogs. In some embodiments, the prodrugs have high aqueous solubility and extended pharmacokinetics in vivo. In some embodiments, the prodrugs have reduced toxicity, as compared with lonidamine or a lonidamine analog. In yet other embodiments, the prodrugs are targeted to specific cells types or cell populations.
[0009] Also provided are vehicles and methods for delivering lonidamine, lonidamine analogs and prodrugs thereof. Such vehicles include, for example, emulsions, liposomes and polymer conjugates. In some embodiments, the lonidamine- and lonidamine analog- vehicle combinations have extended pharmacokinetics in vivo. In some embodiments, the lonidamine- and lonidamine analog- vehicle combinations have reduced toxicity, as compared with lonidamine or a lonidamine analog. In yet other embodiments, the lonidamine- and lonidamine analog- vehicle combinations can be targeted to specific cell types or cell populations.
[0010] In one aspect, the present invention provides tertiary amine prodrug compounds of lonidamine and lonidamine analogs. Prodrugs of the present invention have the formula (I):
Figure imgf000006_0001
(I) wherein R1 is COOR3
R is an aryl or heteroaryl group, optionally substituted with from one to three R4 substituents, independently selected from the group consisting of halo and a straight or branched chain (d~C8)alkyl;
R3 is a group of the formula (CR5R6)mNR7R8; each R5 and R6 is independently H, a straight or branched chain (C1-C8) alkyl heteroalkyl, heterocyclyl, cycloalkyl, aryl, heteroaryl or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system; each R7 and R8 is a straight or branched chain (Ci-C8)alkyl, heteroalkyl, heterocyclyl, cycloalkyl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system;
X is a straight or branched chain, saturated or unsaturated hydrocarbon linker group;
Y is CHR9;
R9 is H or a straight or branched chain (C1-C8)alkyl group; the subscript n is an integer of from 0 to 1 ; the subscript m is an integer of from 1 to 4; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof. [0011] hi another aspect, compounds are provided of formula D-Z-M, wherein D is lonidamine or a lonidamine analog; Z is a cleavable linker joined to D; and M is a moiety joined to Z. In some embodiments, the D-Z-M compound has a higher Vmax for a transporter expressed in plasma membranes cells than D alone, hi some embodiments, the cells are, for example, epithelial cells lining a human colon, a small intestine, a brain, a kidney, a prostate or a heart. Also provided are an individual isomer or a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate, or a hydrate of a D-Z-M compound.
[0012] In another aspect, linker prodrugs of the following formulae are provided:
Figure imgf000007_0001
(II) (III)
wherein D is lonidamine or a lonidamine analog; Q1 is O or CH2; Z1 and Z2 are cleavable linkers; R' is alpha-OH or hydrogen; R" is alpha-OH, beta-OH or hydrogen; W is- CH(CH3)W1, wherein W1 is a substituted alkyl group containing a moiety which is negatively charged at physiological pH, said moiety is selected from the group consisting Of CO2H5 SO3H, SO2H, -P(O)(OR)(OH), -OP(O)(OR)(OH), and OSO3H, wherein R is C1-C6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
[0013] hi another aspect, compounds of the following formulae are provided,
Figure imgf000008_0001
wherein Rallc is lower alkyl; and D is lonidamine or a lonidamine analog; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate, or a hydrate thereof.
[0014] In another aspect, compounds of the following formula are provided:
Figure imgf000008_0002
(IV)
wherein:
R1 is an aryl group of formula
Figure imgf000008_0003
wherein each R2 is independently halogen or C1-C6 alkyl; X1 is selected from the group consisting of O, S and NR3 wherein R3 is hydrogen or C1-C6 alkyl;
Y1 is -C(R4^- or a sugar moiety, wherein each R4 is independently hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl; X2 is selected from the group consisting of halogen, C1-C6 alkoxy, diacylglycerol, amino, C1-C6 alkylamino, C1-C6 dialkylamino, C1-C6 alkylthio, a PEG moiety, a bile acid moiety, a sugar moiety, an amino acid moiety, a di-or tri-peptide, a PEG carboxylic acid, and -U-V wherein
U is O or S; and V is selected from the group consisting of C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, CW2X3, PO(X3)2, and SO2X3; wherein
W2 is O or NR5 wherein R5 is hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl; and each X3 is independently amino, hydroxyl, mercapto, C1-C6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl, C1-C6 alkoxy, C1-C6 alkylamino, C1-C6 dialkylamino, C1-C6 alkylthio, a bile acid based alkoxy group, a sugar moiety, a PEG moiety, and -0-CH2-
CH(OR6)CH2X4R6, wherein:
X4 is selected from the group consisting of O, S, S=O, and SO2; and each R6 is independently C10-C22 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl, alkylene, or heteroalkylene; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof with the proviso that formula (IV) excludes
Figure imgf000010_0001
V being CO2H. [0015] In a related aspect, prodrugs of lonidamine and lonidamine analogs are provided that have improve aqueous solubility and extended pharmacokinetics in vivo.
[0016] In another aspect, the present invention provides a method for prophylaxis of cancer, said method comprising administering a prophylactically effective amount of a compound of the present invention to a human subject in need of such prophylaxis.
[0017] hi another aspect, the present invention provides a method for treatment of cancer, said method comprising administering a therapeutically effective amount of a compound of the present invention to a human subject in need of such treatment.
[0018] hi another aspect, the present invention provides a method for prophylaxis of BPH, said method comprising administering a prophylactically effective amount a compound of the present invention to a human subject.
[0019] hi another aspect, the present invention provides a method for treating BPH, the method comprising administering a therapeutically effective amount of a compound of the present invention to a human subject in need of such treatment. [0020] In another aspect, the present invention provides a method for reducing a symptom associated with BPH, said method comprising administering a compound of the present invention to a human subject exhibiting the symptom.
[0021] In another aspect, the present invention provides a method for reducing prostate size in a human subject, comprising administering a therapeutically effective amount of a compound of the present invention to the subject.
[0022] In another aspect, methods are provided for synthesizing compounds according to the invention and compounds useful as intermediates in such synthetic methods.
[0023] In another aspect, pharmaceutical formulations of the compounds according to the invention are provided.
[0024] These and other aspects and embodiments of the invention are discussed in greater detail below. The description below is organized into sections for convenience only, and disclosure found in any organizational section is applicable to any aspect of the invention.
DETAILED DESCRIPTION OF THE INVENTION [0025] Definitions
[0026] The following definitions are provided to assist the reader. Unless otherwise defined, all terms of art, notations and other scientific or medical terms or terminology used herein are intended to have the meanings commonly understood by those of skill in the chemical and medical arts. In some cases, terms with commonly understood meanings are defined herein for clarity and/or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over the definition of the term as generally understood in the art.
[0027] As used herein, "a" or "an" means "at least one" or "one or more."
[0028] "Alkyl" refers to a linear saturated monovalent hydrocarbon radical or a branched saturated monovalent hydrocarbon radical having the number of carbon atoms indicated in the prefix. For example, (Ci-Cg)alkyl is meant to include methyl, ethyl, n- propyl, 2-propyl, n-butyl, 2-butyl, tert-butyl, pentyl, and the like. For each of the definitions herein (e.g., alkyl, alkenyl, alkoxy, araalkyloxy), when a prefix is not included to indicate the number of main chain carbon atoms in an alkyl portion, the radical or portion thereof will have six or fewer main chain carbon atoms.
[0029] "Alkenyl" refers to a linear monovalent hydrocarbon radical or a branched monovalent hydrocarbon radical having the number of carbon atoms indicated in the prefix and containing at least one double bond, but no more than three double bonds. For example, (C2-Cg)alkenyl is meant to include, ethenyl, propenyl, 1,3-butadienyl and the like.
[0030] "Alkoxy" refers to a substituted or unsubstituted alkyl group of 1 to 6 carbon atoms covalently attached to O. (d-C6)alkoxy group is meant to include, for example, methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentoxy, 2- pentoxy, 3-pentoxy, isopentoxy, neopentoxy, hexoxy, 2-hexoxy, 3-hexoxy and 3- methylpentoxy.
[0031] "Alkylamino" refers to a substituted or unsubstituted alkyl group of 1 to 6 carbon atoms covalently attached to -NH-. (CrC^alkylamino group is meant to include, for example, methylamino, ethylamino, propylamino and butylamino.
[0032] "Alkylthio" refers to substituted or unsubstituted alkyl group of 1 to 6 carbon atoms covalently attached to S. For example, (Q-C^alkylthio is meant to include CH3- CH2-S-, (CH3)2CH-S-, CH3-(CH2)S-S-, and the like.
[0033] "Aryl" refers to a monovalent monocyclic or bicyclic aromatic hydrocarbon radical of 6 to 10 ring atoms which is substituted independently with one to four substituents. In some embodiments, the aryl group has one, two, or three substituents selected from alkyl, cycloalkyl, cycloalkyl-alkyl, halo, nitro, cyano, hydroxy, alkoxy, amino, acylamino, mono-alkylamino, di-alkylamino, haloalkyl, haloalkoxy, heteroalkyl, COR (where R is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), - (CR'R")n-COOR (where n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, and R is hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, phenyl or phenylalkyl) or -(CR'R")n-CONRxRy (where n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, and Rx and RY are independently selected from hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, phenyl or phenylalkyl). In some embodiments, the term aryl includes, but is not limited to, phenyl, biphenyl, 1-naphthyl, and 2-naphthyl, and the substituted forms thereof. [0034] "Cycloalkyl" refers to a monovalent cyclic hydrocarbon radical of three to seven ring carbons. The cycloalkyl group may have one double bond and may also be optionally substituted independently with one, two, or three substituents selected from alkyl, optionally substituted phenyl, or -C(O)R2 (where Rz is hydrogen, alkyl, haloalkyl, amino, mono-alkylamino, di-alkylamino, hydroxy, alkoxy, or optionally substituted phenyl). More specifically, the term cycloalkyl includes, for example, cyclopropyl, cyclohexyl, cyclohexenyl, phenylcyclohexyl, 4-carboxycyclohexyl, 2- carboxamidocyclohexenyl, 2-dimethylaminocarbonyl-cyclohexyl, and the like.
[0035] "Heteroalkyl" means an alkyl radical as defined herein with one, two or three substituents independently selected from cyano, -ORW, -NRXRY, and -S(O)nR2 (where n is an integer from 0 to 2 ), with the understanding that the point of attachment of the heteroalkyl radical is through a carbon atom of the heteroalkyl radical. Rw is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, alkoxycarbonyl, aryloxycarbonyl, carboxamido, or mono- or di-alkylcarbamoyl. Rx is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl or araalkyl. Ry is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, alkoxycarbonyl, aryloxycarbonyl, carboxamido, mono- or di- alkylcarbamoyl or alkylsulfonyl. Rz is hydrogen (provided that n is 0), alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, amino, mono-alkylamino, di-alkylamino, or hydroxyalkyl. Representative examples include, for example, 2-hydroxyethyl, 2,3- dihydroxypropyl, 2-methoxyethyl, benzyloxymethyl, 2-cyanoethyl, and 2- methylsulfonyl-ethyl. For each of the above, Rw, Rx, RY, and R2 can be further substituted by amino, fluorine, alkylamino, di-alkylamino, OH or alkoxy. Additionally, the prefix indicating the number of carbon atoms (e.g., CJ-CJO) refers to the total number of carbon atoms in the portion of the heteroalkyl group exclusive of the cyano, -ORW, - NR XRY, or -S(O)nRv portions.
[0036] "Heteroaryl" means a monovalent monocyclic or bicyclic radical of 5 to 12 ring atoms having at least one aromatic ring containing one, two, or three ring heteroatoms selected from N, O, or S, the remaining ring atoms being C, with the understanding that the attachment point of the heteroaryl radical will be on an aromatic ring. The heteroaryl ring is optionally substituted independently with one to four substituents. In some embodiments, the heteroaryl ring is substituted independently with one or two substituents selected from alkyl, cycloalkyl, cycloalkyl-alkyl, halo, nitro, cyano, hydroxy, alkoxy, amino, acylamino, mono-alkylamino, di-alkylamino, haloalkyl, haloalkoxy, heteroalkyl, -COR (where R is hydrogen, alkyl, phenyl or phenylalkyl, -(CR'R")n-COOR
(where n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, and R is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), or ~(CR'R")n-
CONRXRY (where n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, and Rx and RY are, independently of each other, hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl). More specifically, in some embodiments the term heteroaryl includes, but is not limited to, pyridyl, furanyl, thienyl, thiazolyl, isothiazolyl, triazolyl, imidazolyl, isoxazolyl, pyrrolyl, pyrazolyl, pyridazinyl, pyrimidinyl, benzofuranyl, tetrahydrobenzofuranyl, isobenzofuranyl, benzothiazolyl, benzoisothiazolyl, benzotriazolyl, indolyl, isoindolyl, benzoxazolyl, quinolyl, tetrahydroquinolinyl, isoquinolyl, benzimidazolyl, benzisoxazolyl or benzothienyl, indazolyl, pyrrolopyrymidinyl, indolizinyl, pyrazolopyridinyl, triazolopyridinyl, pyrazolopyrimidinyl, triazolopyrimidinyl, pyrrolotriazinyl, pyrazolotriazinyl, triazolotriazinyl, pyrazolotetrazinyl, hexaaza-indenly, and heptaaza-indenyl and the derivatives thereof Unless indicated otherwise, the arrangement of the hetero atoms within the ring may be any arrangement allowed by the bonding characteristics of the constituent ring atoms.
[0037] "Heterocyclyl" or "cycloheteroalkyl" means a saturated or unsaturated non- aromatic cyclic radical of 3 to 8 ring atoms in which one to four ring atoms are heteroatoms selected from O, NR (where R is independently hydrogen or alkyl), NRQ (where RQ is independently hydrogen or alkyl), or S(O)n (where n is an integer from 0 to
2), the remaining ring atoms being C, where one or two C atoms may optionally be replaced by a carbonyl group. The heterocyclyl ring may be optionally substituted independently with one, two, or three substituents selected from alkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, cycloalkyl, cycloalkyl-alkyl, halo, nitro, cyano, hydroxy, alkoxy, amino, mono-alkylamino, di-alkylamino, haloalkyl, haloalkoxy, -COR (where R is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), -CORS (where Rs is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), -(CR'R")n-
COOR (n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, and R is hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl), -(CRpRL)m- COORS (m is an integer from O to 5, Rp and RL are independently hydrogen or alkyl), - (CR'R")n-CONRxRy (where n is an integer from 0 to 5, R' and R" are independently hydrogen or alkyl, Rx and RY are, independently of each other, hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, phenyl or phenylalkyl), or -(CRpRL)m-CONR(S)2 (where m is an integer from 0 to 5, Rp and RL are independently hydrogen or alkyl, and each Rs is independently, hydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, phenyl or phenylalkyl). More specifically the term heterocyclyl includes, but is not limited to, pyridyl, tetrahydropyranyl, N-methylpiperidin-3-yl, N-methylpyrrolidin-3-yl, 2-pyrrolidon-l-yl, furyl, quinolyl, thienyl, benzothienyl, pyrrolidinyl, piperidinyl, morpholinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydrothiofuranyl, 1 , 1 -dioxo-hexahydro- 1 Δ6-thiopyran-4-yl, tetrahydroimidazo [4,5-c] pyridinyl, imidazolinyl, piperazinyl, and piperidin-2-onyl, and the derivatives thereof. The prefix indicating the number of carbon atoms (e.g., C3-C10) refers to the total number of carbon atoms in the portion of the cyclohetero alkyl or heterocyclyl group exclusive of the number of heteroatoms.
[0038] " Alkylene" means a linear saturated divalent hydrocarbon radical having from one to twelve carbon atoms or a branched saturated divalent hydrocarbon radical having from one to twelve carbon atoms optionally substituted with substituents including, for example, hydroxy, amino, mono or di(CrC6)alkyl amino, halo, C2-C6 alkenyl ether, cyano, nitro, ethenyl, ethynyl, C1-C6 alkoxy, C1-C6 alkylthio, -COOH, -CONH2, mono- or di-(C1-C6)alkyl-carboxamido, -SO2NH2, -OSO2-(C1-C6) alkyl, mono or di(C1-C6) alkylsulfonamido, aryl and heteroaryl. For example alkylene includes methylene, ethylene, propylene, 2-methyl-proρylene, pentylene, hexylene, and the like. In one embodiment, an alkylene radical has from one to 25 carbon atoms.
[0039] "Heteroalkylene" has essentially the meaning given above for alkylene except that one or more heteroatoms (i.e., oxygen, sulfur, nitrogen and/or phosphorous) may be present in the alkylene biradical. For example, heteroalkylene includes, -CH2OCH2O-, -CH2CH2OCH2CH2-, -CH2CH2N(CH3)CH2CH2-, -CH2CH2SCH2CH2-, and the like.
[0040] As used herein, "sugar moiety" refers to a mono-, di-, or trisacharide or a derivative thereof. Li one embodiment, the sugar moiety is an aldohexose, a ketopentose, an aldopentose, or a ketopentose. hi one embodiment, the sugar moiety is D-glucose or a derivative thereof, hi one embodiment, the sugar moiety is glucuronic acid. In some embodiments, the bonding of a sugar moiety to the rest of a prodrug compound does not involve a peroxo or a -O-O- linkage.
[0041] As used herein, in a "bioisostere" of a given molecule, one replaces one or more atoms or groups with atoms or groups having similar size and spatial disposition of electron pair or pairs. Bioisosteres and bioisosterism is a well-known tool for predicting the biological activity of compounds, based upon the premise that compounds with similar size, shape, and electron density can have similar biological activity. Known bioisosteric replacements include, for example, the interchangeability of -F, -OH, -NH2, - Cl, and -CH3; the interchangeability of -Br and -i-C3H7; the interchangeability of -I and - t-GjHp; the interchangeability of -O-, -S-, -NH-, -CH2, and -Se-; the interchangeability of -N=, -CH=, and -P= (in cyclic or noncyclic moieties); the interchangeability of phenyl and pyridyl groups; the interchangeability of -C=C- and -S- (for example, benzene and thiophene); the interchangeability of an aromatic nitrogen (Rar-N(Rar)-Rar) for an unsaturated carbon (Rar-C(=Rar)-Rar); and the interchangeability of -CO-, -SO-, and - SO2-. These examples are not limiting on the range of bioisosteric equivalents and one of skill in the art will be able to identify other bioisosteric replacements known in the art. See, for example, Patani et al, 1996, Chem. Rev. 96:3147-76; and Burger, 1991, A. Prog. Drug Res. 37:287-371.
[0042] Often a reasonable quantitative prediction of the binding ability of a known molecule can be made based on the spatial arrangement of a small number of atoms or functional groups in the molecule. As used herein, such an arrangement is called a pharmacophore, and once the pharmacophore or pharmacophores in a molecule have been identified, this information can be used to identify other molecules containing the same or similar pharmacophores. Such methods are well known to persons of ordinary skill in the art of medicinal chemistry, and as the structural information described in this application identifies the pharmacophore prodrugs, lonidamine or lonidamine analogs. An example of programs available to perform pharmacophore -related searches is the program 3D Pharmacophore search from the Chemical Computing Group (see Chemical Computing Group internet web site).
[0043] The terms "optional" or "optionally" as used throughout the specification mean that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs and instances in which it does not. For example, "heterocyclo group optionally mono- or di- substituted with an alkyl group" means that the alkyl may but need not be present, and the description includes situations where the heterocyclo group is mono- or disubstituted with an alkyl group and situations where the heterocyclo group is not substituted with the alkyl group.
[0044] "Optionally substituted" means a ring or linear radical which optionally may be substituted independently with substituents.
[0045] As used herein, a "prodrug" is a compound that, after administration, is metabolized or otherwise converted to an active or more active form with respect to at least one property. To produce a prodrug, a pharmaceutically active compound can be modified chemically to render it less active or inactive, but the chemical modification is such that an active form of the compound is generated by metabolic or other biological processes. Prodrugs also can be formed by non-covalent modification of a pharmaceutically active compound, such that the compound can be released as an active form or an activatable form. A prodrug may have, relative to the drug, altered metabolic stability or transport characteristics, fewer side effects or lower toxicity, and/or improved flavor, for example (see the reference Nogrady, 1985, Medicinal Chemistry A Biochemical Approach, Oxford University Press, New York, pages 388-392). Prodrugs can also be prepared using compounds that are not drugs.
[0046] As used herein, "Vmax" refers to the maximum initial rate of enzyme or transporter function. For a detailed description of Vmax, see Nelson et al., Lehninger Principles of Biochemistry, 2000, Third Edition, W. H. Freeman.
[0047] As used herein, "treating" a condition or patient refers to taking steps to obtain beneficial or desired results, including clinical results. For purposes of this invention, beneficial or desired clinical results include, but are not limited to, alleviation or amelioration of one or more symptoms of cancer, BPH or other condition, diminishment of extent of disease, delay or slowing of disease progression, amelioration, palliation or stabilization of the disease state, and other beneficial results described below.
[0048] As used herein, "preventing" or "prophylaxis" mean to reduce the risk of occurrence of disease (e.g., cancer, BPH or other condition) in an individual or in individuals in a population. That is, administration of an agent to a population of individuals "prevents" a condition when, relative to a control population, fewer individuals in the population develop the condition, and/or appearance of the condition is delayed in the administered population relative to a control population. Methods for detecting and quantifying a reduction of risk, including use of clinical trials, are well known in the art. As used herein, "preventing" a condition or disease in a patient refers to taking steps to obtain beneficial or desired results, including clinical results.
[0049] As used herein, "reduction" of a symptom or symptoms means decreasing of the severity or frequency of the symptom(s), or elimination of the symptom(s).
[0050] As used herein, "administering" or "administration of a drug to a subject include both direct administration, including self-administration, and indirect administration, including the act of prescribing a drug. For example, as used herein, a physician who instructs a patient to self-administer a drug and/or provides a patient with a prescription for a drug is administering the drug to the patient.
[0051] As used herein, a "manifestation" of cancer, BPH or other condition refers to a symptom, sign, anatomical state (e.g., prostate size), physiological state (e.g., prostate specific antigen (PSA) level), or report (e.g., American Urological Association Symptom Index score) characteristic of a subject with cancer or BPH.
[0052] As used herein, a "therapeutically effective amount" of a drug is an amount of a drug that, when administered to a subject with cancer, BPH or other condition, will have the intended therapeutic effect, e.g., alleviation, amelioration, palliation or elimination of one or more manifestations of cancer, BPH or other condition in the subject. The full therapeutic effect does not necessarily occur by administration of one dose, and may occur only after administration of a series of doses. Thus, a therapeutically effective amount may be administered in one or more administrations.
[0053] As used herein, a "prophylactically effective amount" of a drug is an amount of a drug that, when administered to a subject, will have the intended prophylactic effect, e.g., preventing or delaying the onset of disease or symptoms, or reducing the likelihood of the onset of disease or symptoms. The full prophylactic effect does not necessarily occur by administration of one dose, and may occur only after administration of a series of doses. Thus, a prophylactically effective amount may be administered in one or more administrations. [0054] As used herein, "target cell" refers to a cell to be contacted with lonidamine or a lonidamine analog. As used herein, a "target cell population" refers to a population having a common characteristic (e.g., a cell surface marker) that is to be contacted with lonidamine or a lonidamine analog.
A. Lonidamine and Lonidamine Analogs
[0055] Lonidamine or lonidamine analogs can be used in the prodrugs according to the present invention. Lonidamine is l-(2,4-dichlorobenzyl)-l-H-indazole-3-carboxylic acid. Lonidamine was first identified as having anti-spermatogenic activity, and subsequently used for treatment of certain cancers. The use of lonidamine and lonidamine analogs for treatment and prevention of benign prostatic hyperplasia has been described (for example, see U.S. patent application number 10/759,337, published as US 2004/0167196).
[0056] Examples of lonidamine analogs include tolnidamine; AF-2364; and AF-2785 (see Ansari et al, 1998, Contraception 57:271-279; and Corsi et al., 1976, J. Medicinal Chemistry 19:778-83); and compounds described by Silvestrini, 1981, Chemotherapy 27:9-20; Lobl et al., 1981, Chemotherapy 27:61-76; Cheng et al., 2001, Biol. Reprod. 65:449-461; Andreani, F., 1984, Arch. Pharm. 317:847-851; Besner et al., 1997, Drug Metabolism Reviews 29(l&2):219-234; Caputo, A., 1981, Chemotherapy 27(Suppl. 2):107-120; Hagishita et al., 1996, J. Med. Chem. 39:3636-3658; Kakehi et al., 1978, Bulletin of Chemical Society of Japan, 51(l):251-256; Milanesio et al., 2000, J. Org. Chem. 65:3416-3425; Palacios et al., 1995, Tetrahedron 51(12):3683-3690; Silvestrini et al., 1984, Prog. Med. Chem. 21, G.P. Ellis and G.B. West, eds., Elsevier Science Publishers, Amsterdam, p.l 11-135; Tapia et al., 1999, J. Med. Chem. 42:2870-2880; Wright and Collins, 1956, 78:221-224; U.S. Patent Nos. 3,895,026; 6,001,865; 5,621,002; 4,002,749; 3,470,194; 3,625,971; 5,034,398; U.S. patent publication No. 2004/0167196; and WO 96/03383. Synthesis of exemplary lonidamine analogs, including tolnidamine (TND), is described in the art including the aforelisted references.
[0057] hi one embodiment, lonidamine analogs have the following formula (V):
Figure imgf000020_0001
where R1, R2, X, Y, and n are:
R2 is -Cl, -Br, -I, or -CH3, monosubstituted phenyl, substituted at the 2, 3, or 4 position; dichloro, dibromo, dimethyl, or chloro and methyl disubstituted phenyl, substituted at the 2 and 3 or 2 and 4 positions; or 2, 4, 5 trichlophenyl;
Y is -(CH2)-; and n is zero, and R1 is -COOH, -CONH2, -CONHNH2, -CONHN(CH3)2, -CH2CH2OH, -CH2CH(OH)CH2OH, or CH2(CH2OH)2; or
n is one, R1 is -COOH, and X is -CH=CH-; and
an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
[0058] In one embodiment, the compounds of the present invention exclude one or more of lonidamine, tolnidamine, AFl 890, AF2364 and/or AF2785.
[0059] Lonidamine and lonidamine analogs and derivatives can be prepared using by well known synthetic methods. Synthesis of lonidamine is described in U.S. Patent No. 3,895,026 in the presence of an inert base, such as triethyl amine tolnidamine (TND), is described in the art (see, e.g., Corsi et al., 1976, Journal of Medicinal Chemistry 19:778- 83; Cheng et al., 2001, Biol. Reprod. 65:449-61; Silvestrini, 1981, Chemotherapy 27:9- 20; Lobl et al., 1981, Chemotherapy 27:61-76; U.S. Patent Nos. 3,895,026 and 6,001,865; See also PCT/US05/19350). B. Tertiary Amine Prodrugs of Lonidamine and Lonidamine Analogs
1. Compounds
[0060] In one aspect, the present invention provides tertiary amine prodrugs of lonidamine and lonidamine analogs having the formula:
Figure imgf000021_0001
(I) wherein R1, R2, X, Y and n are defined below: R1 represents COOR3. R2 is usually an aryl or heteroaryl group. The aryl or heteroaryl group of R2 may have substituents on their respective rings, wherein each substituted present can be the same or different from any other substituent. More particularly, R2 may have from 0 to 3 substituents, more preferably from 1 to 2 substituents, and still more preferably, 2 R4 substituents. R4 substituents are independently selected from the group consisting of halo and a straight or branched chain (C1-C8)alkyl.
[0061] R3 represents a group of the formula (CR5R6)mNR7R8 wherein each R5 and R6 is independently H, a straight or branched chain (C1-C8)alkyl, heteroalkyl, cycloalkyl, or heterocyclyl or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system.
[0062] Each R7 and R8 is a straight or branched chain (CrC8)alkyl, heteroalyl, cycloalkyl, or heterocyclyl or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system.
[0063] Returning to formula (I), X is a straight or branched chain, saturated or unsaturated hydrocarbon linker group and Y is CHR9. The symbol R9 represents H or a straight or branched chain (C1-C8)alkyl group.
[0064] Again returning to formula (I), the subscript n is an integer of from 0 to 1 and the subscript m is an integer of from 1 to 4. [0065] In addition to compounds having formula I above, the present invention further includes all salts thereof, and particularly, pharmaceutically acceptable salts thereof. Still further, the invention includes compounds that are single isomers of the above formula (e.g., single enantiomers of compounds having a single chiral center), as well as solvate and hydrate forms thereof.
[0066] In additional embodiments, in addition to compounds having Formula I above, the present invention further includes an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
[0067] A number of other groups of embodiments are preferred and are set forth below.
[0068] In a first group of embodiments, R1 is preferably a COOR3 moiety.
[0069] The subscript m is preferably 2 and each R5 and R6 is preferably independently selected from the group, H, CH3, and a member in which R5 and R6 are joined together to form a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1,1-dioxo-hexahydro-lΔ6- thiopyran-4-yl or tetrahydropyran-4-yl group.
[0070] With regard to the prodrug group, preferred embodiments are compounds wherein NR7R8 is morpholino.
[0071] R2 is preferably phenyl. Within this embodiment R4 is preferably independently selected from the group consisting of Cl, Br, or CH3.
[0072] In another preferred embodiment n is 1 and X is -CH=CH-. In yet another preferred embodiment n is 0. In another embodiment, Y is preferably CH2.
[0073] In one embodiment, compounds preferably have the formula:
Figure imgf000022_0001
wherein R4 is, independently Cl or CH3; each R5 and R6 is independently H, (C1-Cs)alkyl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered ring cycloalkyl or heterocyclyl system; X is an alkenylene linker group; and the subscript n is an integer of from 0 to 1. Within this embodiment R4 is preferably Cl; n is 0; and each R5 and R6 is independently, H, CH3, or are joined together to form a cyclopropyl, cyclopbutyl, cyclopentyl, cyclohexyl, 1,1-dioxo- hexahydro-lλ6-thiopyran-4-yl or tetrahydropyran-4-yl group.
[0074] Even further preferred are those embodiments in which the compound has the formula:
Figure imgf000023_0001
wherein each R5 and R6 is independently, H, CH3, or are joined together to form a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1,1-dioxo-hexahydro-lλ -thiopyran-4- yl or tetrahydropyran-4-yl group.
[0075] Still other preferred groups of embodiments are provided in the Examples below.
[0076] Examples of compounds of Formula (I) include:
[0077] 1 -(2,4-dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid 2-morpholm-4-yl-ethyl ester:
Figure imgf000023_0002
[0078] l-(2,4-dichloro-benzyl)-lH-indazole-3-carboxylic acid 2-methyl-2-morpholin-4- yl-propyl ester:
Figure imgf000024_0001
[0079] l-(2,4-Dichloro-benzyl)-lH-indazole-3-carboxylic acid l-morpholin-4- ylmethyl-cyclopropyl ester:
Figure imgf000024_0002
[0080] 1 -(2,4-Dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid 1 -morpholin-4- ylmethyl-cyclobutyl ester:
Figure imgf000024_0003
[0081] 1 -(2,4-Dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid l-morpholin-4- ylmethyl-cyclopentyl ester:
Figure imgf000025_0001
[0082] 1 -(2,4-Dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid 1 -morpholin-4- ylmethyl-cyclohexyl ester:
Figure imgf000025_0002
[0083] 1 -(2,4-dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid 4-morpholin-4-ylmethyl- 1 , 1 -dioxo-hexahydro- 1 λ6-tbiopyran-4-yl ester:
'
Figure imgf000025_0003
[0084] 1 -(2,4-dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid 4-morpholin-4-ylmethyl- tetrahydro-pyran-4-yl ester:
Figure imgf000025_0004
[0053] l-(2,4-dichloro-benzyl)-lH-indazole-3-carboxylic acid 2,2-dimethyl-2- morpholin-4 yl-butyl ester:
Figure imgf000026_0001
[0054] 1 -(2,4-dichloro-benzyl)- lH-indazole-3-carboxylic acid 1 , 1 -dimethyl-2- morpholin-4 -yl butyl ester:
Figure imgf000026_0002
[0085] In additional embodiments, the present invention further includes an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate of any of these compounds.
[0086] In a related aspect, the lonidamine analog has the formula:
Figure imgf000026_0003
(XII)
wherein R1, R2, X, Y and n are defined below: R1 represents COOR3. n is O. Y is CH2-. R2 is 2,4-dichlorophenyl. [0087] R3 can be a straight or branched chain (C1-C8) alkyl or alkoxy, or a three- to eight-membered cycloalkyl or heterocyclyl ring system. For example, R can be Cr6 alkoxymethyl, such as methoxymethyl; C1-6 alkanoyloxymethyl esters such as pivaloyloxymethyl; phthalidyl esters; C^CgcycloalkoxycarbonyloxyQ-Cβalkyl such as 1- cyclohexylcarbonyloxyethyl; l,3-dioxolen-2-onylmethyl esters, such as 5-methyl-l,3- dioxolen-2-on-ylmethyl; and Q.Qalkoxycarbonyloxyethyl such as 1- methoxycarbonyloxyethyl. In additional embodiments, the present invention further includes an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate of any of these compounds.
2. Synthesis of Tertiary Amine Prodrugs of Lonidamine and Lonidamine Analogs
[0088] Lonidamine and lonidamine analogs and derivatives can be prepared using by well known synthetic methods, as described supra. Ester prodrugs of lonidamine and lonidamine analogs are generally prepared as described in, for example, U.S. Pat. No. 6,146,658, as is administration of polymorphic forms, an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate, a hydrate thereof, and the like.
[0089] Syntheses of the ester prodrugs start with the free carboxylic acid of lonidamine or a lonidamine analog. The free acid is activated for ester formation in an aprotic solvent and then reacted with a free alcohol group bearing the tertiary amine in the presence of an inert base, such as triethyl amine, to affect ester formation, producing the prodrug. Activating conditions for the carboxylic acid include forming the acid chloride using oxalyl chloride or thionyl chloride in an aprotic solvent, optionally with a catalytic amount of dimethyl formamide, followed by evaporation. Examples of aprotic solvents, include, but are not limited to methylene chloride, tetrahydrofuran, and the like. Alternatively, activations can be performed in situ by using reagents such as BOP (benzotriazol-l-yloxytris(dimethylamino) phosphonium hexafluorolphosphate, and the like (see Nagy et al., 1993, Proc. Natl. Acad. Sci. USA 90:6373-6376) followed by reaction with the free alcohol. Isolation of the ester products can be affected by extraction with an organic solvent, such as ethyl acetate or methylene chloride, against a mildly acidic aqueous solution; followed by base treatment of the acidic aqueous phase so as to render it basic; followed by extraction with an organic solvent, for example ethyl acetate or methylene chroride; evaporation of the organic solvent layer; and recrystalization from a solvent, such as ethanol, which has been acidified with an acid, such as HCl or acetic acid. Alternatively the crude reaction can be passed over an ion exchange column bearing sulfonic acid groups in the protonated form, washed with deionized water, and eluted with aqueous ammonia; followed by evaporation.
[0090] Suitable free alcohols bearing the tertiary amine are commercially available, such as N-2-hydroxyethylmorpholine, N-2-hydroxyethylimidazole, and the like. Non- commercially available alcohols can be synthesized in straightforward manner via standard literature procedures. Such procedures can be identified via literature search tools such as SciFinder from the American Chemical Society or Beilstein available from MDL Software.
[0091] In certain embodiments, the prodrug of the lonidamine or a lonidamine analog is provided in the form of a pharmaceutically acceptable salt. Pharmaceutically acceptable salts include addition salts with acids, as well as the salts with bases. Suitable acids for the formation of acid addition salts are, for example, mineral acids, such as hydrochloric, hydrobromic, sulphuric or phosphoric acid, or organic acids, such as organic sulphonic acids, for example, benzenesulphonic, 4-toluenesulphonic or methanesulphonic acid, and organic carboxylic acids, such as acetic, lactic, palmitic, stearic, malic, maleic, fumaric, tartaric, ascorbic or citric acid. Acid salts of the tertiary amine moiety confer increased aqueous solubility. The use of citric acid salts is especially preferred.
[0092] The prodrugs of the invention provide for the release of a drug lonidamine and its analogs. An illustrative example discussed below illustrated how a prodrug of the invention can be designed to exhibit increased aqueous solubility and extended pharmacokinetics in vivo.
[0093] In an embodiment of the invention, the prodrug moiety comprises a tertiary amine having a pKa near the physiological pH of 7.5. Any amines having a pKa within 1 unit of 7.5 are suitable alternatives amines for this purpose. The amine may be provided by the amine of a morpholino group. This pKa range of 6.5 to 8.5 allows for significant concentrations of the basic neutral amine to be present in the mildly alkaline small intestine. The basic, neutral form of the amine prodrug is lipophilic and is absorbed through the wall of the small intestine into the blood. Following absorption into the bloodstream, the prodrug moiety is cleaved by esterases which are naturally present in the serum to release lonidamine or the lonidamine analog. More strongly basic amines, such as a trialkyl derivatives with no heteroatom substitutions, will be nearly completely protonated under physiological conditions and will not be as efficiently adsorbed as shown.
[0094] In one aspect of the invention, the serum half live of the prodrug of the lonidamine and lonidamine analogs of the present invention is increased in vivo (compared to the parental form) by the presence of R5 and R6 groups. The R5 and R6 groups in the prodrug, as shown in the structure above, can independently be selected to modulate the rate of cleavage of the prodrug moiety from lonidamine. Increasing the amount of steric hindrance proximal to the ester carbonyl of lonidamine decreases the rate of cleavage of the prodrug moiety. Slowing the rate of cleavage of the prodrug moiety has the effect of increasing serum half life. Hydrogen groups facilitate cleavage of the prodrug moiety and alkyl groups hinder it. The larger and more branched the alkyl group, the more cleavage is hindered and the more serum half life is increased. Similarly, the closer the non-hydrogen substitution is to the lonidamine carbonyl, the more cleavage of the prodrug moiety is hindered and the more serum half life of the prodrug form is increased.
[0095] In a preferred embodiment, linkage of the tertiary amine to the lonidamine is stable enough so that the serum half life of the prodrug is from about 8 to about 24 hours.
[0096] In another aspect of the invention, R4 and R5 may be joined together to form a cyclic group further comprising heteroatoms. This aspect of the invention further improves upon the aqueous solubility of the compounds of the invention.
C. Linker Prodrugs of Lonidamine and Lonidamine Analogs
1. Compounds
[0097] In one aspect, the present invention provides a prodrug D-Z-M of lonidamine or a lonidamine analog, said prodrug comprising, lonidamine or an analog, D; joined by a cleavable linker Z; to a moiety M. Prodrugs with this structure may be referred to as "linker prodrugs." In some embodiments, the prodrug has a higher Vmax for a transporter expressed in plasma membranes of cells than D alone. In some embodiments, the cells are epithelial cells lining a human colon, or small intestine, a prostate, or the like. In one embodiment, the transporter is expressed in the plasma membranes of epithelial cell lining in the human gut. In another embodiment, the transporter is expressed in the plasma membranes of epithelial cell lining in the prostate. In one embodiment, the transporter is expressed in human kidney, brain, lung, liver and/or heart. In one embodiment, the moiety M is selected from the group consisting of an amino acid, a dipeptide, a tripeptide, a bile acid, and their derivatives. In one embodiment, the transporter is selected from the group consisting of ATBO, CAT-I, FATP4, MCTl, MCT4, NADCl, NADC2, OCTN2, PEPTl, PGT, RFC, SAT-I, SAT-6, SMVT, SUT2 and SVCTl (for a description of these transporters see, e.g., Gallop et al., WO02100347). In one embodiment, the transporter is PEPT2, which is expressed in human kidney, brain, lung, liver, and heart. In one embodiment, the transport system is carrier mediated. In a related embodiment, the transport system is receptor mediated.
[0098] In one embodiment, the prodrug compound exhibits selective uptake by a subject's prostate as compared to another organ, such as the testis, heart, kidney, brain, lung, and/or liver. In a related embodiment, the prodrug is selectively taken up by subject's prostate compared to other organs. In another embodiment, the prodrug compound exhibits selective uptake by prostate epithelial cells as compared to other epithelial cells of, for example, the testis, heart, kidney, brain, lung, and/or liver. In a related embodiment, the prodrug is selectively taken up by prostate epithelial cell as compared to other epithelial cells.
[0099] In one embodiment, the M moiety is an androgen, an androgen analog, or a functional androgen analog exhibits selective uptake by a subject's prostate as compared to another organ such as, for example, the testis, heart, kidney, brain, lung, and/or liver. In a related embodiment, the prodrug D is selectively taken up by subject's prostate. In another embodiment, the M moiety is an androgen, an androgen analog, or a functional androgen analog that exhibits selective uptake by prostate epithelial cells as compared to epithelial cells such as, for example, the testis, heart, kidney, brain, lung, and/or liver. In a related embodiment, the prodrug is selectively taken up by prostate epithelial cells as compared to other epithelial cells.
[0100] In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog comprising a lonidamine- or a lonidamine analog-peptide conjugate, the peptide comprising an amino acid sequence having a cleavage site specific for an enzyme having a proteolytic activity of prostate specific antigen and wherein the peptide is linked to lonidamine or the lonidamine analog to inhibit the therapeutic activity of lonidamine or the lonidamine analog, and wherein lonidamine or- the lonidamine analog is cleaved from the peptide upon proteolysis by an enzyme having a proteolytic activity of prostate specific antigen (PSA).
[0101] In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog comprising a lonidamine- or a lonidamine analog-peptide conjugate, the peptide comprising an amino acid sequence having a cleavage site specific for an enzyme having a proteolytic activity of prostate specific antigen, wherein the peptide is 20 or fewer amino acids in length, wherein the sequence comprises the amino acids
G5-G4-G3-G2-G1,
wherein G5 is from 0 to 16 amino acids; G4 is serine, isoleucine, or lysine; G3 is serine or lysine; G2 is leucine or lysine; and G1 is glutamine, asparagine or tyrosine, and wherein the peptide is linked to lonidamine or the lonidamine analog to inhibit the therapeutic activity of the loiύdamϊne or the lonidamine analog, and wherein lonidamine or the lonidamine analog is cleaved from the peptide upon proteolysis by an enzyme having a proteolytic activity of prostate specific antigen (PSA).
[0102] In one embodiment, the present invention provides a prodrug of lonidamine or an analog comprising a cephalosporin moiety, a dihydronicotinamide moiety, a triglyceride, a long chain fatty acid, or a long chain fatty alcohol.
[0103] In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is a vitamin or a vitamin precursor. In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is vitamin-D, a vitamin-D analog, or a vitamin-D precursor. In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is vitamin-E, a vitamin-E analog, or a vitamin-E precursor. In a related embodiment, the moiety M is α-tocopherol. In another related embodiment, the moiety M is an α-tocopherol-PEG conjugate. In another related embodiment, the moiety M is an α-tocopherol- α,ω-dicarboxylic acid-PEG conjugate. In another related embodiment, the moiety M is an α-tocopherol-succinic acid-PEG conjugate. Various α-tocopherol based conjugates employed in the present invention can be adapted from those described in the U.S. Patent Application No. US2005/0142189, to Lambert et al.
[0104] In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog, wherein the moiety M is a hormone or a hormone precursor.
[0105] In one embodiment, the present invention provides a prodrug of lonidamine or a lonidamine analog wherein the prodrug is enzymatically modified to yield lonidamine or the lonidamine analog, wherein the enzyme is carboxypeptidase, aminohydrolase, or glycosidase. hi one embodiment, the prodrug contains an Aryl-O-CO-N< moiety which is cleaved by a carboxypeptidase to yield lonidamine or a lonidamine analog from the prodrug.
[0106] Moiety M and linker Z that can be employed in a D-Z-M prodrug of the present invention is provided for example, in the reference Silverman, January 15, 1992, Organic Chemistry of Drug Design and Drug Action, Academic Press; 1st edition.
[0107] Other M moieties including but not limited to a bile acid, an amino acid, and a peptide, and linker Z moieties that can be used in the compounds of the invention are described in the following US Patent Application Nos. 2004/0161424, 2003/0158254, 2003/0158089, and 2003/0017964; and PCT Publication Nos. WO 04/053192, WO 04/052844, WO 04/052841, WO 04/052360, WO 04/041203, WO 04/033655, WO 03/104184, WO 03/099338, WO 03/080588, WO 03/077902, WO 03/065982, WO 03/020214, WO 02/100392, WO 02/100347, WO 02/100344, WO 02/100172, WO 02/44324, WO 02/42414, WO 02/32376, WO 02/28883, WO 02/28882, WO 02/28881, and WO 02/28411. hi a related embodiment, the moiety can be a targeting peptide, to target lonidamine or a lonidamine analog to a specific cell type. See, e.g., U.S. patent publication No. 2002/0147138.
[0108] In another aspect, the present invention provides a prodrug D-Z-M of lonidamine or a lonidamine analog, said prodrug comprising lonidamine or an analog, D, joined by a cleavable peptide linker Z, to a stabilizing moiety M. The peptide linker can be any cleavable peptide linker. In some embodiments, the linker is cleavable by an endogenous enzyme, hi some embodiments, the linker is a tripeptide, P1-P2-P3, comprising natural or synthetic amino acids. [0109] In some embodiments, Pl is Leucine, Sarcosine, Tyrosine, Phenylalanine, p-Cl- Phenylalanine, p-Nitrophenylalanine, Valine, Norleucine, Norvaline, Phenylglycine, Tryptophan, tetrahydroisoquinoline-3-carboxylic acid, 3-Pyridylalanine, Alanine, Glycine, or 2-Thienylalanine. In some embodiments, P2 can be Alanine, Leucine, Tyrosine, Glycine, Serine, 3-Pyridylalanine, or 2-Thienylalanine. In some embodiments, P3 can be Leucine, Phenylalanine, Isoleucine, Alanine, Glycine, Tyrosine, 2- Naphthylalanine, or Serine.
[0110] In some embodiments, the peptide linker can be one of the following: Leu- Ala- Leu, Tyr-Ala-Leu, Met- Ala-Leu, Tyr-Ala-Ile, Phe-Gly-Leu, Met-Gly-Leu, Met-Gly-Ile, Phe-Gly-Ile, Met-Gly-Phe, Leu-Ala-Gly, NIe- Ala-Leu, Phe-Gly-Phe, and Leu-Tyr-Leu. See also U.S. Patent Publication No. 2003/0181359.
[0111] In some embodiments, moiety M is a stabilizing moiety that protects the prodrug from cleavage in circulating blood when it is administered to the patient and allows the prodrug to reach the vicinity of the target cell relatively intact. The stabilizing group typically protects the prodrug from cleavage in blood and blood serum. In some embodiments, the stabilizing group is useful in the prodrug when it serves to protect the prodrug from degradation, i.e., inactivation, when tested by storage of the prodrug compound in human blood at 370C. for 2 hours and results in less than 20%, particularly less than 2%, inactivation of the prodrug by the enzymes present in the human blood under the given assay conditions.
[0112] The stabilizing group can be, for example, an amino acid or an amino acid that is either (i) a non-genetically-encoded amino acid having four or more carbons or (ii) aspartic acid or glutamic acid attached to the N-terminus of the oligopeptide at the beta- carboxyl group of aspartic acid or the gamma-carboxyl group of glutamic acid. For example, dicarboxylic (or a higher order carboxylic) acid or a pharmaceutically acceptable salt thereof may be used as a stabilizing group. In other embodiments, the stabilizing group is not an amino acid.
[0113] In another aspect, linker prodrugs of the following formulae are provided:
Figure imgf000034_0001
wherein D is lonidamine or a lonidamine analog; Q1 is O or CH2; Z1 and Z2 are cleavable linkers; R' is alpha-OH or hydrogen; R" is alpha-OH, beta-OH or hydrogen; W is -CH(CH3)W1, wherein W1 is a substituted alkyl group containing a moiety which is negatively charged at physiological pH, said moiety is selected from the group consisting OfCO2H5 SO3H, SO2H, -P(O)(OR)(OH), -OP(O)(OR)(OH), and OSO3H wherein R is C1- C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof. .
[0114] In another aspect, compounds of the following formulae, and enantiomers and diastereomers thereof, are provided:
Figure imgf000035_0001
wherein Ralk is alkyl (e.g., C1-C6 alkyl); and D is lonidamine or a lonidamine analog, and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof, hi one embodiment, Raik is lower alkyl. In one embodiment, when D is covalently attached to a heteroatom in the formula above,
Figure imgf000035_0002
then D is Ri
wherein R1 is an aryl group of formula
Figure imgf000035_0003
wherein each R2 is independently halogen or C1-C6 alkyl. In one embodiment, when D is covalently attached to a carbonyl (-C=O)- moiety, then D
IS
Figure imgf000036_0001
wherein
R1 is as defined above.
[0115] In another aspect, the present invention provides a compound of formula
Figure imgf000036_0002
wherein:
R1 is an aryl group of formula
Figure imgf000036_0003
wherein each R2 is independently halogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, or heterocyclyl;
X1 is selected from the group consisting of O, S and NR3 wherein R3 is hydrogen or C1-C6 alkyl; Y1 is -C(R4)2- or a sugar moiety, wherein each R4 is independently hydrogen or
C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl;
X2 is selected from the group consisting of halogen; C1-C6 alkoxy; diacylglycerol; amino; C1-C6 alkylamino; C1-C6 dialkylamino; C1-C6 alkylthϊo; a PEG moiety; a bile acid moiety; a sugar moiety; an amino acid moiety; a di-or tri-peptide; a PEG carboxylic acid; and -U-V wherein
U is O or S; and V is selected from the group consisting of C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, CW2X3, PO(X3 )2, and SO2X3 wherein W2 is O or NR5 wherein R5 is hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and each X3 is independently amino, hydroxyl, mercapto, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, C1-C6 dialkylamino, C1-C6 alkylthio; a bile acid based alkoxy group, a sugar moiety, a PEG moiety, and -0-CH2-CH(OR6)CH2X4R6 wherein:
X4 is selected from the group consisting of O, S, S=O, and SO2; and each R6 is independently C10-C22 alkyl, heteroalkyl, heteroalkylene or alkylene; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof; with the proviso that formula (IV) excludes
Figure imgf000037_0001
V being CO2H.
[0116] hi one embodiment, the present invention provides the following Y1 groups:
CH2, CHMe, CH(isopropyl), CH(tertiarybutyl), C(Me)2, C(Et)2, C(isopropyl)2, and
C(ρroρyl)2. [0117] In one embodiment, the present invention provides the following X2 groups:
Figure imgf000038_0001
OMe, OEt, -O-isopropyl, O-isobutyl, O-tertiarybutyl, -O-COMe, -O-C(=O)(isoρropyl), -O-C(=O)(isobutyl), -O-C(=O)(tertiarybutyl), -O-C(=O)-NMe2, -O-C(=O)-NHMe, -O-C(=O)-NH2, -O-C(=O)-N(H)-C(Ra)-CO2Et wherein Ra is a side chain alkyl, heteroalkyl, or heterocyclyl group selected from the side chain groups present in essential amino acids, -O-P(=O)(OMe)2, -O-P(=O)(O-isopropyl)2, and -O-P(=O)(O-isobutyl)2. [0118] In one embodiment, the present invention provides the following compounds:
Figure imgf000038_0002
Figure imgf000038_0003
wherein R31 and each X3 is defined as in formula (IV); R34 is independently hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
[0119] In one embodiment, the present invention provides the following compounds:
and
Figure imgf000038_0004
Figure imgf000039_0001
wherein R31 is defined as in formula (IV); R34 is independently hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and each X3 independently is C1- C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, C1-C6 alkoxy, C1-C6 alkylamino, C1-C6 dialkylamino, or C1-C6 alkylthio; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof. [0120] In one embodiment, compounds of the follow formulae are provided:
Figure imgf000039_0002
Figure imgf000039_0003
(X), wherein
R1 is an aryl group of formula
Figure imgf000039_0004
wherein each R2 is independently halogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl,;
X1 is selected from the group consisting of O, S, and NR3, wherein R3 is hydrogen or C1-C6 alkyl; and Y1 is -C(R4)I- wherein R4 is independently hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl.
[0121] Various polyethylene glycol (PEG) moieties and methods for forming prodrugs with them that can be used in or to make compounds of the invention are described in U.S. Patent Nos. 6,608,076; 6,395,266; 6,194,580; 6,153,655; 6,127,355; 6,111,107; 5,965,566; 5,880,131; 5,840,900; 6,011,042 and 5,681,567.
[0122] Various protecting groups and methods for forming prodrugs with them that can be used in or to make compounds of the invention can be adapted from the references Testa et al, Hydrolysis in Drug and Prodrug Metabolism, June 2003, Wiley- VCH, Zurich, 419-534 and Beaumont et al., Curr. Drug Metab. 2003, 4:461-85.
[0123] In another embodiment, the term "cleavable linker", such as, e.g., Z, refers to a linker which has a short half life in vivo. The breakdown of the linker Z in a compound D-Z-M (supra) releases or generates lonidamine or a lonidamine analog. In one embodiment, the cleavable linker has a half life of less than ten hours. In one embodiment, the cleavable linker has a half life of less than an hour. In one embodiment, the half life of the cleavable linker is between one and fifteen minutes. In one embodiment, the cleavable linker has at least one connection with the structure: C*- C(=X*)X*-C* wherein C* is a substituted or unsubstituted methylene group, and X* is S or O. In one embodiment, the cleavable linker has at least one C*-C(=O)O-C* connection. In one embodiment, the cleavable linker has at least one C*-C(=O)S-C* connection. In one embodiment, the cleavable linker has at least one -C(=O)N*-C*-SC>2- N*-connection, wherein N* is -NH- or C1-C6 alkylamino. In one embodiment, the cleavable linker is hydrolyzed by an esterase enzyme.
[0124] In one embodiment, the linker is a self-immolating linker, such as that disclosed in U.S. patent publication 2002/0147138, to Firestone; PCT Appl. No. US05/08161 and PCT Pub. No. 2004/087075. hi another embodiment, the linker is a substrate for enzymes. See generally Rooseboom et al., 2004, Pharmacol. Rev. 56:53-102.
[0125] Ih one embodiment the present invention, compounds of the following formulae are provided:
Figure imgf000041_0001
Figure imgf000041_0002
R = C10-C22 alkyl or alkylene
In the following examples Rf H or C1^C6 alk>
Figure imgf000041_0003
Figure imgf000041_0004
Figure imgf000042_0001
Figure imgf000042_0002
Figure imgf000042_0003
chain alkyl groups present in naturally alpha amino acids
Figure imgf000042_0004
Figure imgf000042_0005
X = (CH2)n,N = 2-4 X = (CH2)n,N = 2-4 CO-Y-NH2 = aminoacyl Y = (CH2)n,N=l-3 Z = O, NMe
Figure imgf000042_0006
Figure imgf000043_0001
wherein R1 is defined as above.
[0126] In one embodiment, the present invention provides the compounds
Figure imgf000043_0003
wherein R1, X1, Y1 and X2 are defined as above.
2. Synthesis of Linker Prodrugs of Lonidamine and Analogs
[0127] In one embodiment, the present invention provides a method for synthesizing prodrugs of lonidamine analogs by reacting
Figure imgf000043_0004
with X10-Y1-X2 wherein X10 is Cl, Br, I, an alkylsufonyloxy, or arylsulfonyloxy; and R31, Y1, and X2 are defined as in formula (IV); in presence of a base such as, for example, a trialkylamine, an alkali metal hydride, or an alkali metal carbonate in a solvent such as dimethylformamide or THF. Exemplery methods for synthesizing prodrugs of Lonidamine are schematically provided below:
[0128] Scheme 1 below provides a method of synthesizing an acyloxymethyl version of a prodrug of formula (FV) by adapting a method from the reference Sobolev et al., 2002, J. Org. Chem. 67:401-410.
Figure imgf000044_0001
wherein V is C1-C6 alkyl.
Scheme 1
[0129] Scheme 2 below provides a method for synthesizing a phosphonooxymethyl version of a prodrug of formula (IV) by adapting a method from Mantyla et al., 2004, J. Med. Chem. 47:188-195.
bromide
Figure imgf000044_0002
Figure imgf000044_0003
Scheme 2
[0130] Scheme 3 below provides a method of synthesizing an alkyloxymethyl version of a prodrug of formula (IV).
Figure imgf000045_0001
Scheme 3
wherein V is C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl. Each Ri in the Schemes is defined as in formula (IV).
[0131] Lonidamine and lonidamine analogs and derivatives can be prepared using known synthetic methods, as described supra. Methods of synthesizing prodrugs of other carboxylic acids or related compounds are described by Nudelman et al., 2005, J. Med. Chem., 48:1042-1054; Mendes et al., 2002, Bioorg. Med. Chem. 10:809-16; Iley et al., 1999, Eur. J. Pharm. Sci. 9:201-5; Iley et al., 2001, J. Chem. Soc. Perkin Trans. 2 749-53; Kubo et al., 1993, J. Med. Chem. 36:2343-9; Popovski et al., 2000, Molecules 5:927-36; and Bundgaard et al., 1987, J. Med. Chem. 30:451-454. One skilled in the art will know how to adapt these methods to synthesize the prodrugs provided in the present invention using lonidamine or a lonidamine analog as a starting material. Some linker moieties useful in the synthesis of prodrugs of the present invention are described in the PCT Publication Nos. WO 02/028411 , pages 4 and 5, and WO 02/100347, pages 5-7. In one embodiment, such synthetic procedures can be identified via search tools such as SciFinder from the American Chemical Society and Beilstein from MDL Software.
[0132] In some embodiments, ester prodrugs of lonidamine and lonidamine analogs can be prepared by modifications known to one of skill in the art of the procedure described in U.S. Patent No. 6,146,658, as is administration of a polymorphic forms, enantiomeric forms, tautomeric forms, solvates, hydrates, and the like.
[0133] In certain embodiments, the prodrug of the lonidamine or a lonidamine analog can have functionality such as, for example, hydroxyl, amino, alkylamino, mercapto, or a group having an -NH- moiety that is further protected, for example by ester, carbamate, and carbonate moieties, yielding a prodrug of a prodrug. In certain embodiments, the prodrug of the lonidamine or a lonidamine analog can have functionality that is converted to its acid or base salt. [0134] In certain embodiments, the prodrug of the lonidamine or lonidamine analog is provided in the form of a pharmaceutically acceptable salt. Pharmaceutically acceptable salts include addition salts with acids, as well as the salts with bases. Suitable acids for the formation of acid addition salts are, for example, mineral acids, such as hydrochloric, hydrobromic, sulphuric or phosphoric acid, and organic acids, such as organic sulphonic acids, for example, benzenesulphonic, 4-toluenesulphonic or methanesulphonic acid, and organic carboxylic acids, such as acetic, lactic, palmitic, stearic, malic, maleic, fuinaric, tartaric, ascorbic or citric acid. Acid salts of the tertiary amine moiety confer increased aqueous solubility. The use of citric acid salts is preferred. Still further, the invention includes compounds and prodrugs that are single isomers of the formulae described herein (e.g., single enantiomers of compounds having a single chiral center), as well as solvate and hydrate forms thereof.
[0135] In another embodiment, a suitable base for the formulation of base addition salts of prodrugs of lonidamine and lonidamine analogs is a primary amine, a secondary amine, a tertiary amine, an amino acid, or a naturally occurring α-amino acid. Examples of aminoacids include but are limited to glycine, lysine, and arginine. In one embodiment, the cation employed in the base addition salt of lonidamine or a lonidamine analog is sodium, potassium, ammonium, or calcium. In one embodiment, a base addition salt of a prodrug of lonidamine or a lonidamine analog is formed employing lysine, glycine, or arginine as a base. In one embodiment, one equivalent of an amine (wherein amine is as described above) is mixed with one equivalent of a prodrug of lonidamine or a lonidamine analog in water. The mixture is shaken or sonicated to yield a homogenous solution of the base addition salt of the prodrug of lonidamine or the lonidamine analog in water. In another embodiment, one equivalent of a prodrug of lonidamine or a lonidamine analog is mixed in water with one equivalent of a metal hydroxide, oxide, bicarbonate, or carbonate wherein the metal comprises sodium, potassium, or calcium resulting in the formation of the metal salt of lonidamine or the lonidamine analog.
D. Polymer Conjugate Prodrugs
[0136] In another aspect, prodrugs of lonidamine and lonidamine analogs comprise polymer conjugates. For example, lonidamine or a lonidamine analog other compound can be linked directly to a polymer or indirectly via a cleavable linker. See, e.g., Uchegbu, 1999, Pharmaceutical Journal 263:355-358.
[0137] In some embodiments, the prodrug comprises lonidamine or a lonidamine analog covalently linked to a polyethylene glycol. PEG moieties and methods for forming prodrugs with them that can be used in or to make compounds of the invention are described in U.S. Patent Nos. 6,608,076; 6,395,266; 6,194,580; 6,153,655; 6,127,355; 6,111,107; 5,965,566; 5,880,131; 5,840,900; 6,011,042 and 5,681,567. See also Duncan, 2003, Nature Reviews 2:347-360.
[0138] In other embodiments, the prodrug comprises lonidamine or a lonidamine analog covalently linked to another polymer, such as N-(2- hydroxypropyl)methacrylamide), polyglutamate, polysaccharide, proteins, and other polymers. See, e.g., Duncan, 2003 (supra).
[0139] In other embodiments, multi-drug polymer prodrug conjugates can be formed of a water-soluble and non-peptidic polymer covalently attached to lonidamine or a lonidamine analog. See, e.g., U.S. Patent Publication No. 20050112088. In some embodiments, the conjugate comprises a plurality of drug molecules attached to the polymer via a hydrolytically degradable linkage, which is released over time following administration of the conjugate to a subject. Representative hydrolytically degradable linkages include, for example, carboxylate ester, carbonate ester, phosphate ester, anhydride, acetal, ketal, acyloxyalkyl ether, imine, orthoester, and oligonucleotides. In some embodiments, hydrolytically degradable linkages include esters such as carboxylate and carbonate esters are particularly preferred linkages. The particular linkage and linkage chemistry employed will depend upon the particular drug (e.g., lonidamine or lonidamine analog), the presence of additional functional groups within the active agent, and the like, and can be readily determined by one skilled in the art.
[0140] A central core organic radical is derived from a molecule that provides a number of polymer attachment sites approximately equal to the desired number of water soluble and non-peptidic polymer arms. The central core molecule of the multi-arm polymer structure can be, for example, a polyol, polythiol, or a polyamine bearing at least three hydroxyl, thiol, or amino groups available for polymer attachment. A "polyol" is a molecule comprising a plurality (greater than 2) of available hydroxyl groups. A
"polythiol" is a molecule that possesses a plurality (greater than 2) thiol groups. A "polyamine" is a molecule comprising a plurality (greater than 2) of available amino groups.
[0141] Illustrative polyols that are preferred for use as the polymer core include aliphatic polyols having from 1 to 10 carbon atoms and from 1 to 10 hydroxyl groups, including for example, ethylene glycol, alkane diols, alkyl glycols, alkylidene alkyl diols, alkyl cycloalkane diols, 1,5-decalindiol, 4,8-bis(hydroxymethyl)tricyclodecane, cycloalkylidene diols, dihydroxyalkanes, trihydroxyalkanes, and the like. Cycloaliphatic polyols include straight chained or closed-ring sugars and sugar alcohols, such as mannitol, sorbitol, inositol, xylitol, quebrachitol, threitol, arabitol, erythritol, adonitol, dulcitol, facose, ribose, arabinose, xylose, lyxose, rhamnose, galactose, glucose, fructose, sorbose, mannose, pyranose, altrose, talose, tagitose, pyranosides, sucrose, lactose, maltose, and the like. Additional examples of aliphatic polyols include derivatives of glyceraldehyde, glucose, ribose, mannose, galactose, and related stereoisomers. Aromatic polyols may also be used, such as l,l,l-tris(4'-hydroxyphenyl) alkanes, such as 1,1,1- tris(4-hydroxyphenyl)ethane, (l,3-adamantanediyl)diphenol, 2,6- bis(hydroxyalkyl)cresols, 2,2'alkylene-bis(6-t-butyl-4-alkylphenols), 2,2'-alkylene-bis(t- butylphenols), catechol, alkylcatechols, pyrogallol, fluoroglycinol, 1,2,4-benzenetriol, resorcinol, alkylresorcinols, dialkylresorcinols, orcinol monohydrate, olivetol, hydroquinone, alkylhydroquinones, l,l-bi-2-naphthol, phenyl hydroquinones, dihydroxynaphthalenes, 4,4'-(9-fluorenylidene)-diphenol, anthrarobin, dithranol, bis (hydroxyphenyl) methane biphenols, dialkylstilbesterols, bis(hydroxyphenyl)alkanes, bisphenol-A and derivatives thereof, meso-hexesterol, nordihydroguaiaretic acid, calixarenes and derivatives thereof, tannic acid, and the like. Other polyols that may be used include crown ethers, cyclodextrins, dextrins and other carbohydrates (e.g., monosaccharides, oligosaccharides, and polysaccharides, starches and amylase). Other polyols include glycerol, trimethylolpropane, reducing sugars such as sorbitol or pentaerythritol, and glycerol oligomers, such as hexaglycerol.
[0142] Exemplary polyamines include aliphatic polyamines such as diethylene triamine, N,N',N"-trimethyldiethylene triamine, pentamethyl diethylene triamine, Methylene tetramine, tetraethylene pentamine, pentaethylene hexamine, dipropylene triamine, tripropylene tetramine, bis-(3-aminopropyl)-amine, bis-(3-aminopropyl)- methylamine, and N,N-dimethyl-dipropylene-triamine. Naturally occurring polyamines that can be used in the present invention include putrescine, spermidine, and spermine. Numerous suitable pentamines, tetramines, oligoamines, and pentamidine analogs suitable for use in the present invention are described in Bacchi et al., January 2002, Antimicrobial Agents and Chemotherapy 46(1):55-61.
[0143] In related embodiments, the polymer conjugate can include a targeting moiety, such as an antibody or ligand, that binds to an antigen, marker or receptor on a target cell or target cell population. Such targeting moieties are further discussed infra.
E. Immunoconjugate Prodrugs
[0144] In another aspect, immunoconjugate prodrugs comprising an antibody and a drug are provided. The conjugate includes an antibody conjugated to lonidamine or a lonidamine analog or other compound according to the present invention. The antibody is typically conjugated lonidamine or a lonidamine analog via a linker.
[0145] The antibody is typically directed to an antigen on the target cell or cell population. For example, the antibody can be an antigen to a tumor-specific antigen, such as a prostate cancer antigen (e.g., prostatic acid phosphatase (PAP); AMACR (x- methylacyl-CoA racemase); CA 15-3 ; prostate-specific membrane antigen (PSMA); Prol09, a human zinc-α 2-glycoprotein (Freje et al., 1993, Genomics 18(3):575-587); Prol 12, a human cysteine-rich protein with a zinc-finger motif (Liebhaber et al., 1990, Nucleic Acid Research 18(13):3871-79; WO9514772 and WO9845436); Prol 11, a prostate-specific transglutaminase (Dubbink et al., 1998, Genomics 51(3):434-444); Prol 15, a novel serine protease with transmembrane, LDLR, and SRCR domains and maps to 21q22.3 (Paoloni-Giacobino et al., 1997, Genomics 1997 44(3):309-320; WO9837418 and WO987093); Prol 10, a human breast carcinoma fatty acid synthase (U.S. Patent No. 5,665,874 and WO9403599); Prol 13, a homeobox gene, HOXB13 (Steinicki et al., 1998, J. Invest. Dermatol. 111 :57-63); Prol 14, a human tetraspan NET-I (WO9839446); and Prol 18, a human JM27 protein (WO9845435). See also U.S. Patent No. 6,902,892.
[0146] In other embodiments, the antibody is directed to another cancer antigen, such as, for example, CA- 125 (epithelial cancer), HER2 (breast cancer), Topoisomerase II alpha (ovarian epithelial cancer), Werner helicase interacting protein (ovarian epithelial cancer), HEXIMl (ovarian epithelial cancer), FLJ20267 (ovarian epithelial cancer),
Deadbox protein-5 (ovarian epithelial cancer), Kinesin-like 6 (ovarian epithelial cancer), p53 (ovarian epithelial cancer) and NY-ESO-I (ovarian epithelial cancer). See, e.g., Menard et al., 2004, Cell MoI. Life Sci. 61(23):2965-78; WO 03/064593.
[0147] There are many linking groups known in the art for making antibody-drug conjugates, including, for example, those disclosed in U.S. patent publication 20020147138. Other suitable linkers include, for example, disulfide groups, thioether groups, acid labile groups, photolabile groups, peptidase labile groups, or esterase labile groups, disulfide and thioether groups being preferred. Conjugates of the antibody and lonidamine or a lonidamine analog can be made using a variety of bifunctional protein coupling agents such as N-succinimidyl (2-pyridyidithio)propionate (SPDP), succinimidyl-fN-maleimidomethyl) cyclohexane-1-carboxylate, iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCL), active esters (such as disuccinimidyl suberate), aldehydes (such as glutaraldehyde), bis-azido compounds (such as his (p-azidobenzoyl) hexanediamine), bis-diazonium derivatives (such as bis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (such as toluene 2,6- diisocyanate), and bis-active fluorine compounds (such as l,5-difluoro-2,4- dinitrobenzene). In some embodiments, the conjugates can be N-succinimidyl (2- pyridyldithio)proρion- ate (SPDP) (Carlsson et al., 1978, Biochem. J. 173:723-737) and N-succinimidyl (2-ρyridylthio)pentanoate (SPP) to provide for a disulfide linkage.
[0148] The linker may be attached to the lonidamine or lonidamine analog molecule at various positions, depending on the type of the linkage. For example, an ester linkage may be formed by reaction with a hydroxyl group using conventional coupling techniques. The reaction may occur at a position having a hydroxyl group or a hydroxymethyl group
F. ADEPT
[0149] In another aspect, a prodrug of lonidamine or a lonidamine analog or other compound according to the present invention can be administered to a target cell or target cell population using Antibody-Dependent Enzyme Mediated Prodrug Therapy (ADEPT). In ADEPT, an antibody is conjugated to a prodrug-activating enzyme, which converts a prodrug to an active agent (e.g., lonidamine or a lonidamine analog). See, for example, Rooseboom et al., 2004, Pharmacol. Rev. 56:53-102; Uchegbu, 1999, Pharmaceutical Journal 263:355-358. The antibody binds to an antigen or marker on a target cell or target cell population. The prodrug is selected to be a substrate for the enzyme. The prodrug is then administered to the patient, where it is preferentially cleaved (activated) at the target cells or target cell population by the enzyme. The antibody can bind to any suitable antigen marker, such as those discussed supra.
[0150] The enzyme component of the immunoconjugate useful for ADEPT includes any enzyme capable of acting on a prodrug in such a way so as to covert it into its more active, cytotoxic form. Enzymes that are useful in the method of this invention include, but are not limited to, alkaline phosphatase useful for converting phosphate-containing prodrugs into free drugs; arylsulfatase useful for converting sulfate-containing prodrugs into free drugs; cytosine deaminase useful for converting non-toxic fluorocytosine into the anti-cancer drug, 5-fluorouracil; proteases, such as serratia protease, thermolysin, subtilisin, carboxypeptidases and cathepsins (such as cathepsins B and L), that are useful for converting peptide-containing prodrugs into free drugs; D-alanylcarboxypeptidases, useful for converting prodrugs that contain D-amino acid substituents; carbohydrate- cleaving enzymes such as O-galactosidase and neuramimidase useful for converting glycosylated prodrugs into free drugs; P-lactamase useful for converting drugs derivatized with P-lactams into free drugs; and penicillin amidases, such as penicillin V amidase or penicillin G amidase, useful for converting drugs derivatized at their amine nitrogens with phenoxyacetyl or phenylacetyl groups, respectively, into free drugs. Alternatively, antibodies with enzymatic activity, also known in the art as "abzymes", can be used to convert the prodrugs of the invention into free active drugs (see, e.g., Massey, 1987,
Nature 328: 457-458). Antibody-abzyme conjugates can be prepared as described herein for delivery of the abzyme to a target cell population. The enzymes can be covalently bound to the antibodies by techniques well known in the art such as the use of the heterobifunctional crosslinkmg reagents as discussed above.
[0151] Alternatively, fusion proteins comprising at least the antigen binding region of an antibody linked to at least a functionally active portion of an enzyme can be constructed using recombinant DNA techniques well known in the art (see, e.g., Neuberger et al., 1984, Nature 312:604-608).
G. GDEPT and VDEPT
[0152] In another aspect, a prodrug of lonidamine or a lonidamine analog or other compound according to the present invention can be administered to a target cell or target cell population using Gene-Dependent Enzyme Mediated Prodrug Therapy (GDEPT) or Virus-Directed Enzyme Prodrug Therapy (VDEPT). See, e.g., Xu et al., 2001, Clinical Cancer Research 7:3314-24; Denny, 2003, J. Biomed. Biotech. 1:48-40. In GDEPT and VDEPT, a transgene encoding a heterologous activating enzyme is administered to the target cell or target cell population. The activating enzyme is expressed by the cell. The prodrug is selected to be a substrate for the enzyme. The prodrug is then administered to the patient, where it is preferentially cleaved (activated) in the target cells or target cell population by the enzyme.
[0153] The transgene can be delivered, for example, by any suitable targeting system, including liposomes, viral vectors (e.g., adenovirus, retrovirus, EBV), direct injection or the like. The enzyme can be, for example, the HSV-tk gene, beta-glucuronidase, bacterial nitroreductase, carboxypeptidase, CYP2B1 and p450 reductase, rabbit CYP4B1, carboxylesterase, cytosine deaminase or thymidine kinase. See, e.g., Xu et al., 2001 (supra); Denny, 2003 (supra).
H. Emulsions and Micromulsions
[0154] In another embodiment, a prodrug comprises an emulsion or a micro-emulsion. The term "emulsion" refers to a colloidal dispersion of two immiscible liquids in the form of droplets, whose diameter, in general, are between 0.1 and 3.0 microns and which is typically optically opaque, unless the dispersed and continuous phases are refractive index matched. Such systems possess a finite stability, generally defined by the application or relevant reference system, which may be enhanced by the addition of amphiphilic molecules or viscosity enhancers. The term "microemulsion" refers to a thermodynamically stable isotropically clear dispersion of two immiscible liquids, such as oil and water, stabilized by an interfacial film of surfactant molecules. The microemulsion has a mean droplet diameter of less than 200 ran, in general between 10- 50 nm. In the absence of water, mixtures of oil(s) and non-ionic surfactant(s) form clear and isotropic solutions that are known as self-emulsifying drug delivery systems (SEDDS) and have successfully been used to improve lipophilic drug dissolution and oral absorption.
[0155] The emulsion comprises lonidamine or a lonidamine analog, or other compound according to the present invention. The composition of the emulsion according to the solubility of the drug form is soluble. The emulsion is typically stabilized by an emulsifying agent. The emulsifying agent optionally can comprise a targeting agent, such as an antibody or peptide. See generally, Uchegbu, 1999, Pharmaceutical Journal 263:355-358.
I. Liposomes
[0156] In a related embodiment, a prodrug can be administered in a liposome. See generally, e.g., Uchegbu, 1999, Pharmaceutical Journal, 263:309-318. Liposomes generally comprise an "outer membrane" or bulk aqueous phase and "central core" or inner aqueous phase. Liposomal formulations can be prepared, for example, by combining a lonidamine or a lonidamine analog or a prodrug form formulation with a phospholipid such as, for example, dimyristoyl phosphatidyl choline (DMPC), phosphatidyl choline (PC), dipalmitoyl-phosphatidyl choline (DPPC), and distearoylphosphatidyl choline (DSPC). Suitable phosphatidyl glycerols include, for example, dimyristoyl phosphatidylglycerol (DMPG) and egg phosphatidyl-glycerol (EPG).
[0157] In some embodiments, the liposome is a stealthed liposome. "Stealthed" liposomes comprise a biologically effective amount of at least a first stealthing agent in operative association with the outer membrane of the liposome. A "stealthing agent" is a component that increases the biological half life of a liposome or liposome-like composition when operatively associated with the outer membrane of the liposome or liposome-like composition. In "operative association", the outer membrane of the liposome or liposome-like composition is preferably "coated" with the one or more stealthing agents.
[0158] Stealthing agents can include, for example, a range of biocompatible hydrophilic polymers, such as polyamines, polylactic acid, polyglycolic acid, polylactic-polyglycolic acid (PLGA), polypeptides and related materials. In some embodiments, the stealthing agent is a polyethylene glycol (PEG) component, wherein the resulting stealthed liposomes are termed "PEGylated liposomes". Other stealthing agents can include, for example, cholesterol, polyvinyl-pyrrolidone polyacrylamide lipids, glucoronic acid lipids or the high phase transition temperature phospholipid distearoyl phosphatidylcholine.
[0159] In some embodiments, stealthed or PEGylated liposomes further comprise an antibody or antigen-binding fragment thereof, to target the liposome to the target cell type (e.g., a cancer cell). The antibody can bind to, for example, any suitable antigen or marker on a target cell or target cell population, as discussed supra. Liposomes containing the antibody can be prepared by methods known in the art, such as described in Epstein et al., 1985, Proc. Natl. Acad. Sci. USA 82:3688; Hwang et al., 1980, Proc. Natl Acad. Sci. USA77:4030; U.S. Patent Nos. 4,485,045 and 4,544,545; and WO97/38731 , published Oct. 23 , 1997. Liposomes with enhanced circulation time are disclosed in, for example, U.S. Patent No. 5,013,556. Fab1 fragments can be conjugated to the liposomes as described in, for example, Martin et al. (1982, J. Biol. Chem. 257:286-288) via a disulfide interchange reaction.
[0160] In additional embodiments, other ligands can be used to target liposomes. For example, the folate receptor, overexpressed in ovarian carcinoma, and the cell adhesion molecules (e.g., selectins and integrins), which are implicated in metastatic events can be targeted. Liposomes bearing specific ligands such as folate may be used to target ovarian carcinomas, while specific peptides or carbohydrates may be used to target integrins and selectins. Other ligands can be used according to the target cells.
[0161] In some embodiments, the liposomes are "antibody-coated" stealthed or
PEGylated liposomes wherein at least one of lonidamine, a lonidamine analog or other compound according to the present invention is operatively associated with the liposome or dispersed within the liposomal formulation. Typically, the lonidamine, a lonidamine analog or other compound according to the present invention is operatively associated with or maintained within the central core of the liposome.
[0162] hi other embodiments, liposomal-like compositions can be used. Such compositions can be, for example, non-ionic surfactant vesicles (niosomes) or other synthetic vesicles. See, e.g., Uchegbu, 1999, Pharmaceutical Journal 263:309-318.
J. Administration of Prodrugs
[0163] The compounds of the present invention can be used in any application for which use of lonidamine and/or a lonidamine analogs is contemplated. Applications include, but are not limited to, treatment or prophylaxis of cancer, treatment or phophylaxis of BPH or a symptom of BPH, inhibition of spermatogenesis, treatment or phophylaxis macular degeneration, and antiangiogenic applications. See the references cited herein above, as well as PCT patent publication Nos. WO2004/064734;
2004/064735; WO2004/064734; 2004/064736; U.S. Provisional application Nos. 60/256,234; 60/491,039; 60/121,205; 60/385,649; and Del Bufalo et al., 2004, Neoplasia 6:513-22.
[0164] The prodrugs of lonidamine or lonidamine analogs of the present invention provide improved pharmacokinetic properties compared to lonidamine. In one embodiment, the compounds of the present invention have greater bioavailability that lonidamine. In another embodiment these compounds have as much bioavailability as lonidamine. hi another embodiment these compounds have 30-80% bioavailability as lonidamine.
[0165] In another embodiment, the present invention provides prodrugs of lonidamine or lonidamine analogs having a higher logP than lonidamine. In another embodiment, the present invention provide prodrugs of lonidamine or lonidamine analogs having logP within ±0.5 of lonidamine. hi another embodiment, the present invention provides prodrugs of lonidamine or lonidamine analogs having a lower logP than lonidamine. The lipophilicity of the moieties provided by the present invention, such as, for example, R3, Z10, M, X1, Y1, and X2 can modulate the logP of the prodrugs of the present invention.
[0166] In another embodiment, the present invention provides prodrugs of lonidamine or lonidamine analogs having a lower toxicity than lonidamine. hi another embodiment, the present invention provide prodrugs of lonidamine or lonidamine analogs as much toxicity as lonidamine. The organ specific uptake of the prodrugs of the present invention can lead to altered toxicity of the prodrugs of the present invention.
[0167] A variety of routes, dosage schedules, and dosage forms are appropriate for administration of the prodrugs according to the invention. Appropriate dosage schedules and modes of administration will be apparent to the ordinarily skilled practitioner reading the present disclosure and/or can be determined using routine pharmacological methods.
[0168] The dose, schedule and duration of administration of the prodrug will depend on a variety of factors, hi some embodiments, the dose, schedule and duration of the prodrug is determined according to the dosage of lonidamine and/or the lonidamine analog to be administered. Factor to be considered include the molecular weight, pharmacokinetic property, pharmacodynamic property, or a combination thereof of the specific prodrug selected. Other important factors include the age, weight and health of the subject, the severity of symptoms, if any, the subject's medical history, co-treatments, goal (e.g., prophylaxis or prevention of relapse), preferred mode of administration, the formulation used, patient response, and the like. Guidance concerning administration is provided by prior experience using lonidamine administered to treat cancer (e.g., 150 mg doses three times a day for a period of about a month), and from new studies in humans and other mammals. Cell culture studies are frequently used in the art to optimize dosages. For particular prodrugs, the scientific literature (including, for example, patent and non-patent publications cited herein) provides considerable guidance as to dosages, formulations and dosage forms for specific prodrugs or classes of prodrugs, e.g., dosages known or predicted to result in a biologically effective serum level of the prodrug (or metabolite) in serum.
[0169] For example, a prodrug can be administered to deliver a dose of lonidamine or a lonidamine analog in the range of about 1 mg to about 100 mg per kg of body weight of the patient to be treated, with more than one dose being administered. In one embodiment, a prodrug is administered to deliver a dose of lonidamine or a lonidamine analog in the range of about 1 mg to about 5 mg per kg of body weight of the patient to be treated. In another embodiment, a prodrug is administered to deliver a dose of lonidamine or a lonidamine analog in the range of about 1 mg to about 10 mg per kg of body weight of the patient to be treated. In certain other embodiments, a prodrug is administered to deliver a dose of lonidamine or a lonidamine analog of about 5 to about 25 mg per kg of body weight of the patient to be treated. In another embodiment, an effective dose is about 5 mg/kg to about 50 mg/kg of lonidamine or the lonidamine analog. For reference exemplary dosage schedules for lonidamine are described in co- pending U.S. patent application No. 10/759,337, filed January 16, 2004, now U.S. patent publication No. US 2004-0167196 Al, which is incorporated herein by reference.
[0170] Other dosing regimens contemplated include, for example and not for limitation, "low dosing" (e.g., dosaged in the range of 1-300 mg per day total daily dosage, 5-300 mg/day, 5-70 mg/day, 1-25 mg/day, 20-45 mg/day, 40-65 mg/day, 40-70 mg/day, 50-100 mg/day, 50-200 mg/day, and 50-300 mg/day), "high dosing" (e.g., total daily doses greater than 0.5 g, such as doses in the range 0.5 - 5 g/day, 0.5 - 3 g/day, 0.5 - 1 g/day and 1-3 g/day, or higher doses), and "intermediate dosing" (e.g., doses greater than 300 and less than 500 mg/day, such as doses in the range >300-400 or 400<500, e.g., 450 mg/day). [0171] For illustration, the effective dose of a prodrug can be administered daily or once every other day or once a week to the patient. Generally, multiple administrations of the prodrug are employed. Depending on the dose selected by the practitioner and the convenience of the patient, the entire dose may be administered once daily, or the dose may be administered in multiple smaller doses through the course of a day. For example, the dose may be divided into two smaller doses and administered twice daily, or divided into three smaller doses and administered thrice daily. Alternatively, the dose may be combined and given every other day, or even less frequently, but in any event, the dose is repeatedly administered over a period of time. For optimum treatment benefit, the administration of the effective dose is continued for multiple days, typically for at least five consecutive days, and often for at least a week and often for several weeks or more. In one embodiment, the prodrug is administered once (qday), twice (bid), three times (tid), or four times (qid) a day or once every other day (qod) or once a week (qweek), and treatment is continued for a period ranging from three days to two weeks or longer. In one embodiment, the treatment is continued for one to three months. In another embodiment, the treatment is continued for a year. Thus, a patient may be administered the prodrug for a week, a month, two months, three months, six months, or a year or longer. For some applications, treatment may continue indefinitely throughout the life of the patient. As is well understood in medicine, treatment may be suspended temporarily if toxicity is observed or for the convenience of the patient without departing from the scope of the invention.
[0172] For illustration and not limitation, the present invention provides a pharmaceutical formulation of a prodrug according to the invention suitable for oral administration (including tablets, capsules, and pills) and contains between 1 and 100 mg of the prodrug, and in another embodiment between 1 and 10 mg of the prodrug. In another embodiment, the formulation contains between 200 and 1000 mg of the prodrug, and in another embodiment between 500 and 1000 mg of the prodrug.
[0173] In addition, the present invention provides controlled and sustained release formulations of the prodrugs that allow once a day oral dosing. Such sustained release formulations (including tablets, capsules, and pills) can contain between 1 mg and 3 g of the prodrug, with various alternative embodiments, including one that contains between 1 mg and 10 mg of the prodrug; another that contains between 150 and 500 mg of the prodrug; and another that contains between 750 mg and 2 g of the prodrug. [0174] When formulated for oral delivery, preferred dosage forms include pills, tablets, capsules, caplets, and the like, optionally formulated for sustained release. Other suitable forms for oral administration include troches, elixirs, suspensions, syrups, wafers, lozenges, and the like. Other modes of administration are also contemplated, including parenteral, inhalation spray, transdermal, rectal, intraprostetic injection (e.g., of prodrug- containing microparticles) and other routes.
[0175] It will be appreciated that these dosing schedules are for illustration and not limitation* and that a dosing schedule may change during a course of therapy based on, for example, a patient's response to the drug administered.
EXAMPLES
[0176] Synthesis of
Figure imgf000058_0001
(Compound A)
[0177] To a solution of 2-morpholinoethanol and pyridine in THF was added l-(2,4- dichloro-benzyl)-3-chlorocarbonyl-l-H-indazole and stirred for 30 min at room temperature. Volatiles were removed in vacuo and the crude reaction mixture was separated by column chromatography on silica gel employing 0-100% ethyl acetate/hexane as an eluent to yield
Figure imgf000058_0002
Compound A
[0178] Formulation of Compound A
[0179] The HCl salt of Compound A was prepared by dissolving Compound A in ethyl acetate followed by the addition of 1.1 equivalents of HCl in dioxane (4 M) at O0C. After evaporating the solvent the HCl salt of Compound A was obtained as a white solid. An aqueous formulation of Compound A is made by dissolving Compound A it or its hydrochloride salt in water.
[0180] The pharmacokinetic properties of Compound A were determined by measuring plasma levels of Compound A after oral or intravenous administration of a aqueous pharmaceutical formulation of Compound A in rats. The pharmacokinetics of Compound A, including it clearance, distribution and bioavailability were observed to be similar to those of lonidamine.
[0181] Although the present invention has been described in detail with reference to specific embodiments, those of skill in the art will recognize that modifications and improvements are within the scope and spirit of the invention, as set forth in the claims which follow. All publications and patent documents (patents, published patent applications, and unpublished patent applications) cited herein are incorporated herein by reference as if each such publication or document was specifically and individually indicated to be incorporated herein by reference. Citation of publications and patent documents is not intended as an admission that any such document is pertinent prior art, nor does it constitute any admission as to the contents or date of the same. The invention having now been described by way of written description and example, those of skill in the art will recognize that the invention can be practiced in a variety of embodiments and that the foregoing description and examples are for purposes of illustration and not limitation of the following claims.

Claims

WHAT IS CLAIMED IS:
1. A compound of the formula (I) :
Figure imgf000060_0001
(I) wherein R1 is COOR3; R2 is an aryl or heteroaryl group, optionally substituted with from one to three R4 substituents, independently selected from the group consisting of halo and a straight or branched chain (C1-C8)alkyl, heteroalkyl, cycloalkyl, and heterocyclyl; R3 is a group of the formula (CR5R6)mNR7R8; each R5 and R6 is independently H, a straight or branched chain (C1-Cs)alkyl, heteroalyl, cycloalkyl, or heterocyclyl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system; each R7 and R8 is a straight or branched chain (C1-C8)alkyl, heteroalkyl, cycloalkyl, or heterocyclyl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered cycloalkyl or heterocyclyl ring system; X is a straight or branched chain, saturated or unsaturated hydrocarbon linker group; Y is CHR9; R9 is H or a straight or branched chain (CrC8)alkyl, heteroalkyl, cycloalkyl, or heterocyclyl group; the subscript n is an integer of from 0 to 1 ; the subscript m is an integer of from 1 to 4; and pharmaceutically acceptable salts, solvates, and hydrates thereof.
2. A compound of claim 1 wherein m is 2 and each R5 and R6 is independently, H, CH3, or are joined together to form a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1 , 1 -dioxo-hexahydro- 1 λ6-thiopyran-4-yl or tetrahydropyran-4-yl group .
3. A compound of claim 1 wherein NR7R8 is morpholino.
4. A compound of claim 1 wherein R2 is phenyl.
5. A compound of claim 4 wherein R is independently selected from the group consisting of Cl, Br, or CH3.
6. A compound of claim 1 wherein n is 1 and X is -CH=CH- or n is 0.
7. A compound of claim 1 wherein Y is CH2.
8. A compound of claim 1 having the following formula:
Figure imgf000061_0001
wherein R4 is, independently Cl or CH3; each R5 and R6 is independently H, (C1-C8)alkyl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered ring cycloalkyl or heterocyclyl system; X is an alkenylene linker group; the subscript n is an integer of from 0 to 1 ; and pharmaceutically acceptable salts, solvates, and hydrates thereof.
9. The compound of claim 8 wherein each R4 is CL
10. The compound of claim 8 wherein n is 0.
11. The compound of claim 8 wherein each R5 and R6 is independently, H, CH3, or are joined together to form a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 1,1- dioxo-hexahydro- 1 λ6-thiopyran-4-yl or tetrahydropyran-4-yl group.
12. The compound of claim 1 having the following formula:
Figure imgf000062_0001
wherein each R5 and R6 is independently, H, CH3, or are joined together to form a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, I,l-dioxo-hexahydro-lλ6-thiopyran- 4-yl or tetrahydropyran-4-yl group.
13. The compound of claim 12 that is selected from the group consisting of: l-(2,4-dichloro-benzyl)-lH-indazole-3-carboxylic acid 2-morpholin-4-yl-ethyl ester:
Figure imgf000062_0002
l-(2,4-dichloro-benzyl)-lH-indazole-3-carboxylic acid 2-methyl-2-morpholin-4-yl-propyl ester:
Figure imgf000062_0003
l-(2,4-Dichloro-benzyl)-lH-indazole-3-carboxylic acid l-morpholin-4-ylmethyl-cyclopropyl ester:
Figure imgf000063_0001
1 -(2,4-Dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid 1 -morpholin-4-ylmethyl-cyclobutyl ester:
Figure imgf000063_0002
1 -(2,4-Dichloro-benzyl)- 1 H-indazole-3 -carboxylic acid 1 -morpholin-4-ylmethyl-cyclopentyl ester:
Figure imgf000063_0003
l-(2,4-Dichloro-benzyl)-lH-indazole-3-carboxylic acid l-morpholin-4-ylmethyl-cyclohexyl ester:
Figure imgf000064_0001
l-(2,4-dichloro-benzyl)-lH-indazole-3-carboxylic acid 4-morpholin-4-ylmethyl-l,l-dioxo- hexahydro-lλ6-thiopyran-4-yl ester:
Figure imgf000064_0002
1 -(2,4-dichloro-benzyl)- 1 H-indazole-3-carboxylic acid 4-morpholin-4-ylmethyl-tetrahydro- pyran-4-yl ester:
Figure imgf000064_0003
1 -(2,4-dichloro-benzyl)- 1 H-indazole-3-carboxylic acid 2,2-dimethyl-2-morpholin-4 yl-butyl ester:
Figure imgf000064_0004
l-(2,4-dichloro-benzyl)-lH-indazole-3-carboxylic acid l,l-dimethyl-2-morpholin-4 -yl butyl ester:
Figure imgf000065_0001
14. A compound of formula D-Z-M, wherein D is lonidamine or a lonidamine analog; Z is a cleavable linker joined to D; and M is a moiety joined to Z wherein the D-Z-M has a higher Vmax for a transporter expressed in plasma membranes of epithelial cells lining a human colon, a small intestine, a brain, a kidney, a prostate, or a heart, than D alone.
15. A compound of the following formula
Figure imgf000065_0002
wherein D is lonidamine or a lonidamine analog; Q1 is O or CH2; Z1 and Z2 are cleavable linkers; R' is alpha-OH or hydrogen; R" is alpha-OH, beta-OH or hydrogen; W is - CH(CH3)W1, wherein W1 is a substituted alkyl group containing a moiety which is negatively charged at physiological pH, said moiety is selected from the group consisting of CO2H, SO3H, SO2H, -P(O)(OR)(OH), -OP(O)(OR)(OH), and OSO3H wherein R is C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and pharmaceutically acceptable salts thereof.
16. A compound selected from the group consisting of
Figure imgf000066_0001
wherein Ralk is lower alkyl; and D is lonidamine or a lonidamine analog; and an individual isomer, a racemic or non-racemic mixture of isomers, a bioisostere, a pharmacophore, a pharmaceutically acceptable salt, a solvate or a hydrate thereof.
17. A compound of the following formula
Figure imgf000066_0002
R1 is an aryl group of formula
Figure imgf000067_0001
wherein each R2 is independently halogen or C1-C6 alkyl; X1 is selected from the group consisting of O, S and NR3 wherein R3 is hydrogen or C1-C6 alkyl; Y1 is -C(R4)2- or a sugar moiety, wherein each R4 is independently hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl; X2 is selected from the group consisting of halogen, C1-C6 alkoxy; diacylglycerol; amino; C1-C6 alkylamino; C1-C6 dialkylamino; C1-C6 alkylthio, a PEG moiety; a bile acid moiety; a sugar moiety, an amino acid moiety; a di-or tri-peptide; a PEG carboxylic acid; and -U-V wherein U is O or S; and V is selected from the group consisting Of C1-C6 alkyl, heteroalkyl, cycloalkyl, hetrocyclyl, aryl, or heteroaryl, CW2X3, PO(X3)2, and SO2X3 wherein W2 is O or NR5 wherein R5 is hydrogen or C1-C6 alkyl; and each X3 is independently amino, hydroxyl, mercapto, C1-C6 alkyl, C1- C6 alkoxy, C1-C6 alkylamino, C1-C6 dialkylamino, C1-C6 alkylthio; a bile acid based alkoxy group, a PEG moiety, and -0-CH2-CH(OR6)CH2X4R6 wherein: X4 is selected from the group consisting of O, S, S=O, and SO2; and each R6 is independently C10-C22 alkyl, heteroalkyl, heteroalkylene, or alkylene; with the proviso that formula (IV) excludes
Figure imgf000068_0001
V being CO2H.
18. The compound of claim 17 wherein X2 is : .
Figure imgf000068_0002
OMe, OEt, -O-isopropyl, O-isobutyl, O-tertiarybutyl, -0-COMe, -O-C(=O)(isopropyl), -O-C(=O)(isobutyl), -O-C(=O)(tertiarybutyl), -0-C(O)-NMe2, -0-C(O)-NHMe, -O- C(O)-NH2, -O-C(=O)-N(H)-C(Ra)-CO2Et, -O-P(=O)(OMe)2, -O-P(=O)(O-isopropyl)2, or ■ O-P(=O)(O-isobutyl)2.
19. The compound of claim 17, wherein Y1 is: CH2, CHMe, CH(isoproρyl), CH(tertiarybutyl), CMe2, CEt2, C(isoρropyl)2, or C(propyl)2.
20 A compound of the formula:
Figure imgf000068_0003
Figure imgf000068_0004
61 wherein R1 is an aryl group of formula
Figure imgf000069_0001
wherein each R2 is independently halogen or C1-C6 alkyl; R4 is independently hydrogen or C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; and each X3 independently is C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, C1-C6 alkoxy, C1-C6 alkylamino, C1-C6 dialkylamino, or C1-C6 alkylthio.
21. A compound selected from the group consisting of:
Figure imgf000070_0001
Figure imgf000070_0002
Figure imgf000071_0001
Figure imgf000071_0002
Figure imgf000072_0001
Figure imgf000072_0002
wherein R1 is an aryl group of formula
Figure imgf000072_0003
each R2 is independently halogen, C1-C6 alkyl; R is C1-C6 alkyl, heteroalkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl; Ra is a side chain alkyl group present in naturally occurring alpha amino acids or side chain alkyl, heteroalkyl, or heterocyclyl group selected from the side chain groups present in essential amino acids; -(O-CH2-CH2)n- is an oligoethyleneoxy moiety wherein n is an integer from 100- 1000.
22. A compound of formula:
Figure imgf000072_0004
wherein R is Ci0 - C22 alkyl, heteroalkyl, alkenyl, alkylene, or heteroalkylene;and
R1 is an aryl group of formula wherein each R2 is independently halogen or C1-C6 alkyl.
23. A compound of formula:
Figure imgf000073_0002
wherein X is (CH2)N wherein N is 2-4; -CO-Y-NH2 together is aminoacyl or Y is (CH2)N wherein N is 1 -3; Z is NMe or O; and
R1 is an aryl group
Figure imgf000073_0003
wherein each R2 is independently halogen or C1-C6 alkyl.
24. A method for prophylactically or therapeutically treating a patient . comprising administering a therapeutically effective amount of a compound of any one of claims 1, 14-17 or 20-22 to a human subject in need of such treatment.
25. A method for treating cancer, said method comprising administering to a mammal a therapeutically effective amount of a compound of any one of claims 1 , 14- 17 or 20-22 to a human subject in need of such treatment.
26. A method for treating benign prostatic hypertrophy (BPH) comprising administering a therapeutically effective amount of a compound of any one of claims 1, 14- 17 or 20-22 to a human subject in need of such treatment.
27. A method for reducing a symptom associated with BPH comprising administering a compound of any one of claims 1, 14-17 or 20-22 to a human subject exhibiting the symptom.
28. A method of reducing prostate size in a human subject, comprising administering a therapeutically effective amount of a compound of any one of claims 1, 14- 17 or 20-22 to the subject.
29. A method for prophylaxis of BPH comprising administering a prophylactically effective amount a compound of any one of claims 1, 14-17 or 20-22 to a human subject.
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EP3564214A1 (en) 2018-05-04 2019-11-06 Universita' Degli Studi G. D Annunzio Chieti - Pescara Indazole derivatives as modulators of the cannabinoid system

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US5260327A (en) * 1985-10-02 1993-11-09 Sloan-Kettering Institute For Cancer Research Methods for inhibiting the proliferation of brain and hepatic metastases by using lonidamine

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WO2015002996A1 (en) * 2013-07-01 2015-01-08 University Of Georgia Research Foundation, Inc. Precise delivery of therapeutic agents to cell mitochondria for anti-cancer therapy
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EP3564214A1 (en) 2018-05-04 2019-11-06 Universita' Degli Studi G. D Annunzio Chieti - Pescara Indazole derivatives as modulators of the cannabinoid system

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