Regulators of the hedgehog pathway, compositions and uses related thereto

• US 20060020020A1
• Filed: 09/19/2005
• Published: 01/26/2006
• Est. Priority Date: 01/13/1999
• Status: Abandoned Application

• Current Assignee: Curis, Inc.
• Original Assignee: Curis, Inc.
• Inventors: Dudek, Henryk, Ji, Benxiu
• Application Number:
  US11/230,063
• Publication Number:
  US 20060020020A1
• Time in Patent Office: Days
• US Class Current: 514/455
• CPC Class Codes: A61K 31/35 having six-membered rings with one oxygen as the only ring hetero atom
• CPC Class Codes: A61K 31/56 Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
• CPC Class Codes: A61K 31/58 containing heterocyclic rings, e.g. danazol, stanozolol, pancuronium or digitogenin digitoxin A61K31/7048
• CPC Class Codes: A61K 31/585 containing lactone rings, e.g. oxandrolone, bufalin
• CPC Class Codes: A61K 31/70 Carbohydrates; Sugars; Derivatives thereof sorbitol A61K31/047
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 15/16 Masculine contraceptives
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/06 Antipsoriatics
• CPC Class Codes: A61P 19/08 for bone diseases, e.g. rachitism, Paget's disease
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/00 Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system

Abstract

The present invention makes available methods and reagents for inhibiting aberrant growth states resulting from hedgehog gain-of-function, ptc loss-of-function or smoothened gain-of-function comprising contacting a cell with a compound, such as a polypeptide or small molecule in an amount sufficient to control the aberrant growth state, e.g., to agonize a normal ptc pathway or antagonize smoothened or hedgehog activity. The present invention further makes available methods and reagents for ameliorating the consequences of hedgehog loss-of-function, ptc gain-of-function, or smoothened loss-of-function comprising contacting a cell with a compound, such as a polypeptide or small molecule, in an amount sufficient to ameliorate the In certain embodiments, the subject compounds, e.g., a cAMP analog, adenylate cyclase agonist, or cAMP phosphodiesterase inhibitor, regulate cAMP levels, which in turn modulates activity of the hedgehog pathway.

Claims

1. A method for inhibiting an altered growth state of a cell having a ptc loss-of-function phenotype or a smoothened gain-of-function phenotype, comprising contacting the cell with a ptc agonist in a sufficient amount to inhibit the altered growth state, wherein the ptc agonist is a organic molecule having a molecular weight less than about 750 amu.

3. The method of claim 1, wherein the ptc agonist causes repression of smoothened-mediated signal transduction.

4. The method of claim 1, wherein the ptc agonist is a steroidal alkaloid.

5. The method of claim 4, wherein the steroidal alkaloid is represented in the general formulas (I), or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

wherein, as valence and stability permit,

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈;

R₆, R₇, and R₇, are absent or represent, independently, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈, or

R₆ and R₇, or R₇ and R′

₇, taken together form a ring or polycyclic ring, e.g., which is susbstituted or unsubstituted,

with the proviso that at least one of R₆, R₇, or R′

₇ is present and includes a primary or secondary amine;

R₈ represents an aryl, a cycloalkyl, a cycloalkenyl, a heterocycle, or a polycycle; and

m is an integer in the range 0 to 8 inclusive.

6. The method of claim 5, wherein:

R₂ and R₃, for each occurrence, is an —

OH, alkyl, —

O-alkyl, —

C(O)-alkyl, or —

C(O)—

R₈;

R₄, for each occurrence, is an absent, or represents —

OH, ═

O, alkyl, —

O—

alkyl, —

C(O)-alkyl, or —

C(O)—

R₈;

R₆, R₇, and R′

₇ each independently represent, hydrogen, alkyls, alkenyls, alkynyls, amines, imines, amides, carbonyls, carboxyls, carboxamides, ethers, thioethers, esters, or —

(CH₂)_m—

R₈, or

R₇, and R′

₇ taken together form a furanopiperidine, such as perhydrofuro[3,2-b]pyridine, a pyranopiperidine, a quinoline, an indole, a pyranopyrrole, a naphthyridine, a thiofuranopiperidine, or a thiopyranopiperidine

with the proviso that at least one of R₆, R₇, or R′

₇ is present and includes a primary or secondary amine;

R₈ represents an aryl, a cycloalkyl, a cycloalkenyl, a heterocycle, or a polycycle, and preferably R₈ is a piperidine, pyrimidine, morpholine, thiomorpholine, pyridazine,

7. The method of claim 4, wherein the steroidal alkaloid is represented in the general formula (II), or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

wherein

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈;

R₆, R₇, and R′

₇, are absent or represent, independently, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈, or

R₆ and R₇, or R₇ and R′

₇, taken together form a ring or polycyclic ring, e.g., which is susbstituted or unsubstituted,

with the proviso that at least one of R₆, R₇, or R′

₇ is present and includes a primary or secondary amine;

X represents O or S, though preferably O.

8. The method of claim 4, wherein the steroidal alkaloid is represented in the general formula (III), or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

wherein

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈;

R₈ represents an aryl, a cycloalkyl, a cycloalkenyl, a heterocycle, or a polycycle; and

A and B represent monocyclic or polycyclic groups;

T represent an alkyl, an aminoalkyl, a carboxyl, an ester, an amide, ether or amine linkage of 1-10 bond lengths;

T′

is absent, or represents an alkyl, an aminoalkyl, a carboxyl, an ester, an amide, ether or amine linkage of 1-3 bond lengths, wherein if T and T′

are present together, than T and T′

taken together with the ring A or B form a covelently closed ring of 5-8 ring atoms;

R₉ represent one or more substitutions to the ring A or B, which for each occurrence, independently represent halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈; and

n and m are, independently, zero, 1 or 2;

with the proviso that A and R₉, or T, T′

B and R₉, taken together include at least one primary or secondary amine.

9. The method of claim 4, wherein the steroidal alkaloid is represented in the general formula (IV), or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

wherein

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈;

R₆ is absent or represent, independently, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈;

R₉ represent one or more substitutions to the ring A or B, which for each occurrence, independently represent halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈; and

R₂₂ is absent or represents an alkyl, an alkoxyl or —

OH.

10. The method of claim 4, wherein the steroidal alkaloid is represented in the general formula (V) or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈;

R₆ is absent or represent, independently, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈; and

R₉ represent one or more substitutions to the ring A or B, which for each occurrence, independently represent halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxarnides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈.

11. The method of claim 4, wherein the steroidal alkaloid is represented in the general formula (VI), or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

wherein

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈; and

R₉ represent one or more substitutions to the ring A or B, which for each occurrence, independently represent halogens;

alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈.

12. The method of claim 4, wherein the steroidal alkaloid is represented in the general formula (VII) or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

wherein

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈; and

R₉ represent one or more substitutions to the ring A or B, which for each occurrence, independently represent halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈.

13. The method of claim 4, wherein the steroidal alkaloid does not substantially interfere with the biological activity of such steroids as aldosterone, androstane, androstene, androstenedione, androsterone, cholecalciferol, cholestane, cholic acid, corticosterone, cortisol, cortisol acetate, cortisone, cortisone acetate, deoxycorticosterone, digitoxigenin, ergocalciferol, ergosterol, estradiol-1 7-α

, estradiol-17-β

, estriol, estrane, estrone, hydrocortisone, lanosterol, lithocholic acid, mestranol, β

-methasone, prednisone, pregnane, pregnenolone, progesterone, spironolactone, testosterone, triamcinolone and their derivatives.

14. The method of claim 4, wherein the steroidal alkaloid does not specifically bind a nuclear hormone receptor.

15. The method of claim 4, wherein the steroidal alkaloid does not specfically bind estrogen or testerone receptors.

16. The method of claim 4, wherein the steroidal alkaloid has no estrogenic activity at therapeutic concentrations.

17. The method of claim 1, wherein the ptc agonist inhibits ptc loss-of-function or smoothened gain-of-function mediated signal transduction with an ED₅₀ of 1 mM or less.

18. The method of claim 1, wherein the ptc agonist inhibits ptc loss-of-function or smoothened gain-of-function mediated signal transduction with an ED₅₀ of 1 μ

M or less.

19. The method of claim 1, wherein the ptc agonist inhibits ptc loss-of-function or smoothened gain-of-function mediated signal transduction with an ED₅₀ of 1 nM or less.

20. The method of claim 1, wherein the cell is contacted with the ptc agonist in vitro.

21. The method of claim 1, wherein the cell is contacted with the ptc agonist in vivo.

22. The method of claim 1, wherein the ptc agonist is administered as part of a therapeutic or cosmetic application.

23. The method of claim 22, wherein the therapeutic or cosmetic application is selected from the group consisting of regulation of neural tissues, bone and cartilage formation and repair, regulation of spermatogenesis, regulation of smooth muscle, regulation of lung, liver and other organs arising from the primative gut, regulation of hematopoietic function, regulation of skin and hair growth, etc.

2. A method for inhibiting aberrant proliferation of a cell having a ptc loss-of-function phenotype or a smoothened gain-of-function phenotype comprising contacting the cell with a ptc agonist in a sufficient amount to inhibit proliferation of the cell.

24. A pharmaceutical preparation comprising a steroidal alkaloid represented in the general formulas (I), or unsaturated forms thereof and/or seco-, nor- or homo-derivatives thereof:

wherein, as valence and stability permit,

R₂, R₃, R₄, and R₅, represent one or more substitutions to the ring to which each is attached, for each occurrence, independently represent hydrogen, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈;

R₆, R₇, and R′

₇, are absent or represent, independently, halogens, alkyls, alkenyls, alkynyls, aryls, hydroxyl, ═

O, ═

S, alkoxyl, silyloxy, amino, nitro, thiol, amines, imines, amides, phosphoryls, phosphonates, phosphines, carbonyls, carboxyls, carboxamides, anhydrides, silyls, ethers, thioethers, alkylsulfonyls, arylsulfonyls, selenoethers, ketones, aldehydes, esters, or —

(CH₂)_m—

R₈, or

R₆ and R₇, or R₇ and R′

₇, taken together form a ring or polycyclic ring, e.g., which is susbstituted or unsubstituted,

with the proviso that at least one of R₆, R₇, or R′

₇ is present and includes a primary or secondary amine;

R₈ represents an aryl, a cycloalkyl, a cycloalkenyl, a heterocycle, or a polycycle; and

m is an integer in the range 0 to 8 inclusive.

25. A method for inhibiting an altered growth state of a cell having a ptc loss-of-function phenotype, hedgehog gain-of-function phenotype, or a smoothened gain-of-function phenotype, comprising contacting the cell with a composition including at least one cAMP agonist.

26. The method of claim 25, wherein at least one cAMP agonist activates adenylate cyclase.

27. The method of claim 25, wherein at least one cAMP agonist is a cAMP analog.

28. The method of claim 25, wherein at least one cAMP agonist is a cAMP phosphodiesterase inhibitor.

29. The method of claim 25, wherein the composition inhibits ptc loss-of-function, hedgehog gain-of-function, or smoothened gain-of-function mediated signal transduction with an ED₅₀ of 1 mM or less.

30. The method of claim 25, wherein the composition inhibits ptc loss-of-function, hedgehog gain-of-function, or smoothened gain-of-function mediated signal transduction with an ED₅₀ of 1 μ

M or less.

31. The method of claim 25, wherein the composition inhibits ptc loss-of-function, hedgehog gain-of-function, or smoothened gain-of-function mediated signal transduction with an ED₅₀ of 1 nM or less.

32. The method of claim 25, wherein the cell is contacted with the composition in vitro.

33. The method of claim 25, wherein the cell is contacted with the composition in vivo.

34. The method of claim 25, wherein the composition is administered as part of a therapeutic or cosmetic application.

35. The method of claim 34, wherein the therapeutic or cosmetic application is selected from the group consisting of regulation of neural tissues, bone and cartilage formation and repair, regulation of spermatogenesis, regulation of smooth muscle, regulation of lung, liver and other organs arising from the primative gut, regulation of hematopoietic function, regulation of skin and hair growth, etc.

36. The method of claim 25, wherein the composition includes forskolin or a derivative thereof.

37. A method for treating or preventing basal cell carcinoma, comprising administering a composition including a cAMP agonist to a patient in an amount sufficient to inhibit progression of basal cell carcinoma.

38. A method for inhibiting an altered growth state of a cell having a ptc loss-of-function phenotype, hedgehog gain-of-function phenotype, or a smoothened gain-of-function phenotype, comprising

determining the phenotype of the cell;

and

if the phenotype is a ptc loss-of-function, hedgehog gain-of-function, or a smoothened gain-of-function phenotype, treating the cell with a cAMP agonist in an amount sufficient to inhibit the altered growth state of the cell.