Manipulating nitrosative stress to kill pathologic microbes, pathologic helminths and pathologically proliferating cells or to upregulate nitrosative stress defenses

• US 6,057,367 A
• Filed: 05/07/1997
• Issued: 05/02/2000
• Est. Priority Date: 08/30/1996
• Status: Expired due to Fees

• Current Assignee: Duke University, The Medical College of Wisconsin Research Foundation
• Original Assignee: Duke University, The Medical College of Wisconsin Research Foundation
• Inventors: Stamler, Jonathan S., Griffith, Owen W.
• Primary Examiner(s): Weddington, Kevin E.
• Application Number:
  US08/852,490
• Time in Patent Office: 1,091 Days
• Field of Search: 514/561, 514/562
• US Class Current: 514/561
• CPC Class Codes: A61K 31/00 Medicinal preparations containing organic active ingredients
• CPC Class Codes: A61K 31/195 having an amino group
• CPC Class Codes: A61K 31/255 of sulfoxy acids or sulfur analogues thereof
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 33/00 Antiparasitic agents
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07C 381/10 Compounds containing sulfur atoms doubly-bound to nitrogen atoms
• CPC Class Codes: Y02A 50/30 Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

Mammals are treated for infections or for conditions associated with pathologically proliferating mammalian cell growth (for example, certain cancers, restenosis, benign prostatic hypertrophy) by administration of a manipulator of nitrosative stress to selectively kill or reduce the growth of the microbes or helminths causing the infection or of host cells infected with the microbes or of the pathologically proliferating mammalian cells. Novel agents include α-alkyl-S-alkyl-homocysteine sulfoximines wherein the α-alkyl contains 2 to 8 carbon atoms, and the S-alkyl-contains 1 to 10 carbon atoms. In another invention herein, mammals in need of increased nitrosative stress defenses are treated, e.g., humans at risk for a stroke because of having had a transient ischemic attack, are treated. Treatments to increase nitrosative stress defenses include, for example, repeated administrations of low doses of manipulators of nitrosative stress so that the subject treated has increased tolerance to nitrosative stress. In still another invention, mammals are treated for protozoal infections by systemic administration of L-buthionine-S-sulfoximine and agent that increases nitrosative stress.

Claims

1. A method of inhibiting growth of pathologic microbes or pathologic helminths or pathologically proliferating mammalian cells, in a mammal, said method comprising administering to said mammal a pathologic microbe or pathologic helminth or pathologically proliferating mammalian cell antiproliferative effective amount of one or more manipulators of nitrosative stress in said microbes or in host cells infected with said microbes or in said helminths or in said pathologically proliferating mammalian cells whereby nitrosative stress selectively kills or reduces the growth of said microbes or of said helminths or of said mammalian cells or selectively enhances their susceptibility to innate immune defenses or the susceptibility of said microbes to antimicrobial agents or the susceptibility of said helminths to anthelmintic agents or the susceptibility of pathologically proliferating mammalian cells to antiproliferation agents;

provided that where the microbes are protozoa, the manipulator(s) of nitrosative stress administered is (are) one(s) that increase(s) nitrosative stress in the pathologic protozoa, or manipulator of nitrosative stress is administered that selectively inhibits protection against nitrosative stress in the pathologic protozoa but does not increase nitrosative stress in the pathologic protozoa and other manipulator of nitrosative stress is also administered which increases nitrosative stress in the pathologic protozoa, or a manipulator of nitrosative stress is administered which is a selective inhibitor of thiol synthesis by the protozoa; and

provided that when the pathologically proliferating mammalian cells are those that would cause restenosis, the manipulator(s) of nitrosative stress comprise(s) inhibitor of protection against nitrosative stress and is (are) employed to selectively kill or reduce the growth of said cells or to enhance their susceptibility to antiproliferation agents; and

provided that when buthionine sulfoximine or melphalan is locally administered as a manipulator of nitrosative stress, agent that increases nitrosative stress is also administered.

2. The method of claim 1 wherein the manipulator of nitrosative stress is a selective inhibitor of protection against nitrosative stress in the pathologic microbes or in the pathologic helminths or in the pathologically proliferating mammalian cells.

3. The method of claim 2 wherein the manipulator of nitrosative stress is a selective inhibitor of thiol synthesis or selective depleter of thiol in the pathologic microbes or in the pathologic helminths or in the pathologically proliferating mammalian cells.

4. The method of claim 3 wherein the manipulator of nitrosative stress is a selective inhibitor of glutathione synthesis in the pathologic microbes or in the pathologic helminths or in the pathologically proliferating mammalian cells.

5. The method of claim 4 wherein the manipulator of nitrosative stress is a selective inhibitor of gamma-glutamylcysteine synthetase in the pathologic microbes or in the pathologic helminths or in the pathologically proliferating mammalian cells.

6. The method of claim 5 wherein the manipulator of nitrosative stress is selected from the group consisting of α

-alkyl-S-alkyl-homocysteine sulfoximines wherein the α

-alkyl contains 2 to 8 carbon atoms and the S-alkyl contains 1 to 10 carbon atoms.

7. The method of claim 6 wherein the manipulator of nitrosative stress is α

-ethyl-L-buthionine-S-sulfoximine.

8. The method of claim 3 wherein the inhibition is carried out on the pathologic microbes and the manipulator of nitrosative stress is an inhibitor of L-homocysteine synthesis or a depleter of L-homocysteine in the pathologic microbes.

9. The method of claim 3 wherein the inhibition is carried out on the pathologic microbes and the manipulator of nitrosative stress is an inhibitor of mycothiol synthesis or a depleter of mycothiol in the pathologic microbes.

10. The method of claim 9 wherein the inhibitor of mycothiol synthesis or depleter of mycothiol is a transportable nitrosylated peptide.

11. The method of claim 3 wherein the inhibition is carried out on the pathologic microbes and the manipulator of nitrosative stress is an inhibitor of ovothiol synthesis or a depleter of ovothiol in the pathologic microbes.

12. The method of claim 3 wherein the inhibition is carried out on the pathologic microbes and the manipulator of nitrosative stress is an inhibitor of δ

-(L-α

-aminoadipoyl)-L-cysteinyl-D-valine synthesis or a depleter of δ

-(L-α

-aminoadipoyl)-L-cysteinyl-D-valine in the pathologic microbes.

13. The method of claim 12 wherein the manipulator of nitrosative stress is δ

-buthylthio-DL-norvaline-SR-sulfoximine.

14. The method of claim 2 wherein the manipulator of nitrosative stress is a selective inhibitor of transcription or translation of an antinitrosative stress gene or a selective inhibitor of an antinitrosative stress gene product.

15. The method of claim 14 wherein the inhibition is carried out on the pathologic microbes and two kinds of manipulators of nitrosative stress in said microbes are administered and one kind is an inhibitor of OxyR transcription or translation of an antinitrosative stress gene and the other kind is a selective inhibitor of thiol synthesis or depleter of thiol in the pathologic microbes.

16. The method of claim 14 wherein the inhibition is carried out on pathologic microbes and a manipulator of nitrosative stress comprises a vector containing an antisense construct to the OxyR promoter or an antisense construct to the OxyR mRNA.

17. The method of claim 1 wherein the manipulator of nitrosative stress is an agent that selectively increases nitrosative stress in the pathologic microbes or the pathologically proliferating cells.

18. The method of claim 17 wherein the manipulator of nitrosative stress is NO-substituted isoniazid or NO-substituted rifampin.

19. The method of claim 1 wherein the inhibition is carried out on the microbes and the agent is one that selectively delivers nitrosating agent to the pathologic microbes.

20. The method of claim 19 wherein the agent is selected from the group consisting of substrates for nitrate reductase, substrates for nitrite reductase and nitrogen oxide microbial sulfite metabolizing enzymes.

21. The method of claim 19 wherein the agent is an organic or inorganic nitrate or nitrite that selectively delivers nitrosating agent to the pathologic microbes.

22. The method of claim 19 wherein the agent is selected from the group consisting of S-nitroso-gamma-methyl-L-homocysteine, S-nitroso-L-homocysteine, S-nitroso-gamma-thio-L-leucine, S-nitroso-δ

-thio-L-leucine, and tripeptides in which two of the residues are selected from the group consisting of serine, glutamine, alanine, leucine and methionine and the other residue is S-nitrosocysteine.

23. The method of claim 19 wherein the manipulator of nitrosative stress is drug that kills or reduces growth of said microbes, substituted to release and transfer NO⁺, NO^- or NO₂⁺ group to the microbes.

24. The method of claim 1 wherein two kinds of manipulator of nitrosative stress in said microbes or host cells or helminths or pathologically proliferating mammalian cells are employed and one kind is a selective inhibitor of protection against nitrosative stress in said microbes or host cells or helminths or pathologically proliferating mammalian cells, and the other kind is an agent that selectively increases nitrosative stress in said microbes or host cells or helminths or pathologically proliferating mammalian cells.

25. The method of claim 1 wherein two kinds of manipulator of nitrosative stress in said microbes or host cells or helminths or pathologically proliferating mammalian cells are employed and one kind is inhibitor of protection against nitrosative stress and the other kind is an increaser of nitrosative stress in the pathologic microbes or in the host cells infected with said microbes or in the pathologic helminths or in the pathologically proliferating mammalian cells and either the inhibition of protection or the increasing of nitrosative stress or both are selective in said microbes or host cells or helminths or pathologically proliferating mammalian cells.

26. The method of claim 1 wherein an agent which increases oxidative stress is administered concurrently with the manipulator of nitrosative stress and the combination of said agent and said manipulator selectively kills or reduces the growth of said microbes or enhances their susceptibility to innate immune defenses or the susceptibility of said microbes to antimicrobial agents or selectively kills or reduces the growth of said helminths or enhances their susceptibility to innate immune defenses or the susceptibility of said helminths to anthelmintic agents or selectively kills or reduces the growth of said pathologically proliferating cells or enhances their susceptibility to innate immune defenses or enhances their susceptibility to antiproliferation agents.

27. The method of claim 1 wherein the inhibition is carried out on the pathologic microbes.

28. The method of claim 27 wherein the microbes are bacteria.

29. The method of claim 28 wherein the bacteria are E. coli.

30. The method of claim 1 wherein the inhibition is carried out on pathologic helminths.

31. The method of claim 1 wherein the inhibition is carried out on the pathologically proliferating mammalian cells and the cells are cancer cells.

32. The method of claim 1 where the mammal is a human.

33. The method of claim 31, wherein the pathologically proliferating mammalian cells are those in Hodgkin'"'"'s disease, in small cell lung cancer, in cancer of the breast, in testicular cancer or in prostate cancer.

34. The method of claim 31, wherein the pathologically proliferating mammalian cells are those in Hodgkin'"'"'s disease.

35. The method of claim 31, wherein the pathologically proliferating mammalian cells are those in small cell lung cancer.

36. The method of claim 31 wherein two kinds of manipulators of nitrosative stress are administered and one kind is an inhibitor of protection against nitrosative stress and the other kind is an agent that increases nitrosative stress and one or both are selective for this(these) effect(s) in the cancer cells.

37. The method of claim 31 wherein the cancer cells are of a type normally treated with an alkylating agent and the manipulator of nitrosative stress is an alkylating agent that releases and transfers NO⁺, NO^- or NO₂⁺ group to the cancer cells.

38. The method of claim 31 wherein the manipulator of nitrosative stress is a chemotherapeutic agent derivatized to release and transfer NO⁺, NO^- or NO₂⁺ group to said cells.

39. The method of claim 1 wherein the inhibition is carried out on the pathologically proliferating cells and the cells are those causing restenosis.

40. The method of claim 39 wherein said manipulator of nitrosative stress is associated with a stent.

41. The method of claim 40 wherein the manipulator of nitrosative stress associated with a stent is agent which inhibits the synthesis of glutathione.

42. The method of claim 39 wherein the manipulator of nitrosative stress is an inhibitor of glutathione synthesis attached to a stent placed in an occluded artery during angioplasty and beta-irradiation, gamma-irradiation or X-ray irradiation is administered to the occluded artery.

43. The method of claim 42 wherein the inhibitor of glutathione synthesis is L-buthionine-S-sulfoximine.

44. The method of claim 40 wherein the manipulator of nitrosative stress associated with a stent is agent which depletes glutathione.

45. The method of claim 39 wherein the manipulator of nitrosative stress comprises agent causing overexpression of transaldolase.

46. The method of claim 39 wherein two kinds of manipulators of nitrosative stress are administered and one is L-buthionine-S-sulfoximine administered in association with a stent emplaced during angioplasty and the other is agent that increases nitrosative stress administered by intracoronary infusion or intravenously.

47. The method of claim 39 wherein two kinds of manipulators of nitrosative stress are administered and either or both kinds are associated with a porous polymer coated on a stent whereby the two kinds of manipulators of nitrosative stress leach out of the porous polymer and contact the pathologically proliferating cells or one kind is associated with a stent and the other kind is administered systemically and the one kind of manipulator of nitrosative stress is L-buthionine-S-sulfoximine and the other kind of manipulator of nitrosative stress is an agent that increases nitrosative stress, or wherein the manipulator of nitrosative stress is NO-substituted L-buthionine-S-sulfoximine and it is associated with a porous polymer coated on a stent.

48. The method of claim 1 wherein the inhibition is carried out on pathologically proliferating cells and the manipulator of nitrosative stress comprises depleter of glutathione which is locally administered.

49. The method of claim 48 wherein the depleter of glutathione is melphalan.

50. The method of claim 1 wherein the inhibition is carried out on the pathologically proliferating cells and the cells are those causing benign prostatic hypertrophy.

51. The method of claim 50 wherein the manipulator of nitrosative stress is administered systemically with coadministration of drug which does not manipulate nitrosative stress which inhibits the cell proliferation or enlargement of the benign prostatic hypertrophy.

52. A method for treating microbial infections of the oral cavity in a mammal comprising topically administering to the site of the infection an infection reducing effective amount of a manipulator of nitrosative stress in the microbes causing the microbial infection.

53. The method of claim 52 wherein the manipulator of nitrosative stress is a S-nitrosothiol.

54. The method of claim 52 wherein the infection treated is oral candidiasis in an immunocompromised mammal.

55. A method for treating microbe caused infection of skin lesions in a mammal comprising topically administering to the skin lesions an infection reducing effective amount of an S-nitrosothiol.

56. The method of claim 55 where the S-nitrosothiol is formed from administration of a mixture comprising inorganic nitrite, pharmacologically acceptable acid and thiol.

57. A method for inhibiting growth of pathologic bacteria in a mammal, said method comprising administering to said mammal a pathologic bacteria antiproliferative effective amount of a chelating agent and of an agent that increases nitrosative stress.

58. A method for inhibiting growth of pathologic microbes or pathologic helminths or pathologically proliferating mammalian cells, in a mammal, comprising administering to said mammal a therapeutic amount of agent that converts endogenously produced NO to nitric-oxide related compound or NO⁺ or NO^-.

59. The method of claim 58 wherein the agent that converts endogenously produced NO to nitricoxide-related compound or NO⁺ or NO^- is administered in conjunction with agent that causes increased NO production.

60. The method of claim 59 wherein the agent that converts endogenously produced NO to nitricoxide-related compound or NO⁺ or NO^- is a redox active metal catalyst and the agent that causes increased NO production is a cytokine.

61. A method for inhibiting growth of pathologically proliferating mammalian cells or for killing or inhibiting the growth of mammalian host cells containing pathologic microbes, in a mammal, comprising administering to said mammal a pathologically proliferating mammalian cell or pathologic microbe antiproliferative effective amount of a blocker of the ability of the mammalian cells to export nitrosant and of an agent that increases nitrosative stress.

62. The method of claim 61 wherein said blocker is a substrate for P-glycoprotein or an inhibitor of P-glycoprotein.

63. The method of claim 62 where a single agent is administered which both blocks the ability of the mammalian cells to export nitrosative stress and increases nitrosative stress in the mammalian cells and the single agent is NO-substituted verpamil.

64. A method of inhibiting growth of pathologic microbes or pathologic helminths or pathologically proliferating mammalian cells, in a mammal, said method comprising systemically administering to said mammal a pathologic microbe or pathologically proliferating cell antiproliferative effective amount of agent that increases nitrosative stress in said microbes or in host cells infected with said microbes or in the pathologic helminths or in said pathologically proliferating cells whereby nitrosative stress selectively kills or reduces the growth of said microbes or host cells or helminths or pathologically proliferating mammalian cells or selectively enhances their susceptibility to innate immune defenses or the susceptibility of said microbes to antimicrobial agents or the susceptibility of said helminths to anthelmintic agents or the susceptibility of the pathologically proliferating mammalian cells to antiproliferative agents, and also administering to said mammal a hypotensive response preventing amount of an inhibitor of guanylyl cyclase.

65. A method of inhibiting growth of non-viral pathologic microbes in a mammal, said method comprising administering to said mammal a pathologic microbe antiproliferative effective amount of a selective inhibitor of thiol synthesis in the microbe to mediate selective killing or growth reduction of the microbe by agent which is selected from the group consisting of antimicrobials, immune cells in the mammal and products of mammalian antimicrobial response.

66. A method for inhibiting the growth of pathologic viruses infecting a mammal, said method comprising administering to the mammal a therapeutic amount of a non-selective inhibitor of glutathione synthesis in the host cells infected by the virus to thereby sensitize the virus to selective killing by antiviral agent, immune cells in the mammal and products of the mammalian antiviral response.