Cancer Patent

Compositions and methods for targeting cancer cells

Cancer Patent Abstract



The invention includes compositions and methods useful for treatment of a virus infection in a mammal by double-targeting the virus (i.e. targeting the virus at more than one stage of the virus life cycle) and thereby inhibiting virus replication. The compositions of the invention include compounds which comprise a phosphocholine moiety covalently conjugated with one or more antiviral agents (e.g. nucleoside analogue, protease inhibitor, etc.) to a lipid backbone. The invention also includes pharmaceutical compositions and kits for use in treatment of a virus infection in mammals. The methods of the invention comprise administering a compound of the invention, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition of the invention, in an amount effective to treat the infection, to a mammal infected with a virus. Additionally, the invention includes compositions and methods useful for combating a cancer in a mammal and for facilitating delivery of a therapeutic agent to a mammalian cell. The compositions of the invention include compounds which comprise an alkyl lipid or phospholipid moiety covalently conjugated with an anticancer agent (e.g. a nucleoside analogue). The invention also includes pharmaceutical compositions and kits for combating a cancer and for facilitating delivery of a therapeutic agent to a mammalian cell. The methods of the invention comprise administering a compound of the invention, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition of the invention, in an amount effective to combat a cancer or to facilitate delivery of a therapeutic agent to a mammalian cell.

Cancer Patent Claims
What is claimed is:

1. A compound having the structure of Formula III: ##STR00014## wherein, R.sup.11 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl; R.sup.12 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl; X.sup.11 is O, S, or NHC.dbd.O; X.sup.12 is O, S, or NHC.dbd.O; X.sup.13 is O or S; n is 0, 1 or 2, and R.sup.13 is an anticancer agent, wherein, each and alkynyl of R.sup.11, R.sup.12, and R.sup.13 is, optionally, substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl, and wherein, if n is 1 or 2, the compound is a phospholipase C substrate and is not a phospholipase A substrate, and further wherein, if n is 1 or 2, the compound is converted to an alkyl lipid and a moiety selected from the group consisting of a nucleoside monophosphate and a nucleoside analogue monophosphate intracellularly in a mammal, and is not converted to an alkyl lipid and a moiety selected from the group consisting of a nucleoside monophosphate and a nucleoside analogue monophosphate extracellularly in a mammal.

2. The compound of claim 1, wherein, R.sup.11 is a C.sub.12 alkyl, branched alkyl, alkenyl or alkynyl; R.sup.12 is C.sub.8H.sub.16 alkyl or branched alkyl; n=1, and R.sup.13 is an anticancer agent selected from the group consisting of gemcitabine, 5-azacytidine, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside and 6-mercaptopurine, wherein the phosphorus atom of the phosphate moiety is covalently linked in a phosphate ester linkage to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.13.

3. A compound having the structure of Formula IV: ##STR00015## wherein, R.sup.21 is (C.sub.6 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl; R.sup.22 is (C.sub.1 to C.sub.12) alkyl, branched alkyl, alkenyl, or alkynyl; X.sup.21 is O, S, or NHC.dbd.O; X.sup.22 is O, S, or NHC.dbd.O; X.sup.23 is O or S; n is 1 or 2; R.sup.23 is an anticancer agent, and wherein, each and alkynyl of R.sup.21, R.sup.22, and R.sup.23 is, optionally, substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

4. The compound of claim 3, wherein, R.sup.21 is C.sub.12 alkyl; R.sup.22 is C.sub.10 alkyl; n=1, and R.sup.23 is an anticancer agent selected from the group consisting of gemcitabine, 5-azacytidine, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside and 6-mercaptopurine, wherein the methylene group of the phosphonate moiety is covalently linked to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.23.

5. A compound having the structure of Formula V: ##STR00016## wherein, R.sup.31 is (C.sub.1 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl; R.sup.32 is (C.sub.1 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl; X.sup.31 is O, S, or NHC.dbd.O; X.sup.32 is O, S, or NHC.dbd.O; X.sup.33 is --O, S or amino; R.sup.33 is an anticancer agent, and wherein, each and alkynyl of R.sup.31, R.sup.32, and R.sup.33 is, optionally, substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

6. The compound of claim 5, wherein, R.sup.31 is (C.sub.6-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl; R.sup.32 is (C.sub.1-C.sub.8) alkyl, branched alkyl, alkenyl or alkynyl, and R.sup.33 is an anticancer agent selected from the group consisting of mitoxanthrone, methotrexate and CPT-11, and is covalently linked via an ester, amido or carbamate linkage to the --SH, OH or amino group of X.sup.33.

7. The compound of claim 1, wherein said compound is suspended in a pharmaceutically acceptable carrier and is present in an amount effective to combat a cancer in a mammal.

8. The compound of claim 7, wherein said cancer is a cancer selected from the group consisting of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

9. The compound of claim 1, wherein said compound is present in an amount effective to facilitate delivery of a therapeutic agent to a mammalian cell.

10. The compound of claim 9, wherein the cell is in a mammal.

11. The compound of claim 10, wherein the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

12. The compound of claim 11, wherein the CNS cell is an astrocyte or a glial cell.

13. A pharmaceutically acceptable salt of the compound of claim 1.

14. The pharmaceutically acceptable salt of claim 13, wherein the compound is present in an amount effective to facilitate delivery of an anticancer agent to a mammalian cell.

15. The pharmaceutically acceptable salt of claim 14, wherein the cell is in a mammal.

16. The pharmaceutically acceptable salt of claim 15, wherein the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

17. The pharmaceutically acceptable salt of claim 13, wherein said compound is present in an amount effective to combat a cancer in a mammal.

18. A pharmaceutically acceptable salt of the compound of claim 2.

19. The pharmaceutically acceptable salt of claim 18, wherein said compound is present in an amount effective to facilitate delivery of an anticancer agent to a mammalian cell.

20. The pharmaceutically acceptable salt of claim 19, wherein said cell is in a mammal.

21. The pharmaceutically acceptable salt of claim 19, wherein said cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

22. The pharmaceutically acceptable salt of claim 13, wherein said compound is present in an amount effective to combat a cancer in a mammal.

23. A drug delivery agent comprising a pharmaceutical composition, said composition comprising a compound of claim 1 or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of an anticancer agent to a mammalian cell.

24. The drug delivery agent of claim 23, wherein said cell is in a mammal.

25. The drug delivery agent of claim 23, wherein said cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

26. A drug delivery agent comprising a pharmaceutical composition, said composition comprising a compound of claim 1 or a pharmaceutically acceptable salt thereof, in an amount effective to combat a cancer in a mammal.

27. The drug delivery agent of claim 26, wherein said cancer is a cancer selected from the group consisting of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

28. A drug delivery agent comprising a pharmaceutical composition, the composition comprising a compound of claim 2 or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of an anticancer agent to a mammalian cell.

29. The drug delivery agent of claim 28, wherein the cell is in a mammal.

30. The drug delivery agent of claim 28, wherein said cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

31. A drug delivery agent comprising a pharmaceutical composition, said composition comprising a compound of claim 2 or a pharmaceutically acceptable salt thereof, in an amount effective to combat a cancer in a mammal.

32. The drug delivery agent of claim 31, wherein said cancer is a cancer selected from the group consisting of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

33. A method of facilitating delivery of an anticancer agent to a mammalian cell, said method comprising administering to said cell a pharmaceutical composition comprising a compound of claim 1 or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of said anticancer agent to said cell.

34. The method of claim 33, wherein said cell is in a mammal.

35. The method of claim 33, wherein the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

36. A method of facilitating delivery of an anticancer agent to a cell, said method comprising administering to said cell a pharmaceutical composition comprising a compound of claim 2 or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of said anticancer agent to said cell.

37. The method of claim 36, wherein said cell is in a mammal.

38. The method of claim 36, wherein said cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

39. A method of combating a cancer in a mammal comprising administering to said mammal a pharmaceutical composition comprising a compound of claim 1 or a pharmaceutically acceptable salt thereof, in an amount effective to combat a cancer in the mammal.

40. The method of claim 39, wherein said cancer is a cancer selected from the group consisting of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

41. A kit for combating a cancer in a mammal, said kit comprising a) a composition selected from the group consisting of a compound of claim 1, a pharmaceutically acceptable salt thereof, and a pharmaceutical composition comprising a compound of claim 1, and b) an instructional material.

42. A kit for facilitating delivery of an anticancer agent to a mammalian cell, said kit comprising a) a composition selected from the group consisting of a compound of claim 1, a pharmaceutically acceptable salt thereof, and a pharmaceutical composition comprising a compound of claim 1, and b) an instructional material.

43. The compound of claim 1, wherein, R.sup.11 is a C.sub.12 alkyl, branched alkyl, alkenyl or alkynyl; R.sup.12 is C.sub.8H.sub.16 alkyl or branched alkyl; n=1, and R.sup.13 is an anticancer agent selected from the group consisting of gemcitabine, 5-azacytidine, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside, 6-mercaptopurine, 6-thioguanine, 5-deoxyfluorouridine, ftorafur, capecitabine, 5-deoxy-5-fluorocytidine, 5-aza-cystine arabinoside, troxacitabine, and pentostatin, wherein the phosphorus atom of the phosphate moiety is covalently linked in a phosphate ester linkage to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.13.

44. The compound of claim 3, wherein, R.sup.21 is C.sub.12 alkyl; R.sup.22 is C.sub.10 alkyl; n=1, and R.sup.23 is an anticancer agent selected from the group consisting of gemcitabine, 5-azacytidine, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside, 6-mercaptopurine, 6-thioguanine, 5-deoxyfluorouridine, ftorafur, capecitabine, 5-deoxy-5-fluorocytidine, 5-aza-cytsine arabinoside, troxacitabine, and pentostatin, wherein the methylene group of the phosphonate moiety is covalently linked to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.23.

45. The compound of claim 5, wherein, R.sup.31 is (C.sub.6-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl; R.sup.32 is (C.sub.1-C.sub.8) alkyl, branched alkyl, alkenyl or alkynyl, and R.sup.33 an anticancer agent selected from the group consisting of mitoxanthrone, doxorubicin, idarubicin, epirubicin, daunorubicin, mitomycin, methotrexate, CPT-11, SN-38, camptothecin, topotecan, 9-nitrocamptothecin, and 9-aminocamptothecin, and is covalently linked via an ester, amido or carbamate linkage to the O, S or amino group of X.sup.33.

Cancer Patent Description
BACKGROUND OF THE INVENTION

Acquired immunodeficiency syndrome (AIDS) is a degenerative disease of the immune system and central nervous system (CNS) resulting from infection of humans by HIV virus. AIDS is responsible for a rapidly growing fatality rate in the world population. At present, no cure has been found, and clinically approved drugs are limited in number. These drugs include nucleoside reverse transcriptase (RT) inhibitors such as 3'-azido-3'-deoxythymidine (AZT, Zidovudine), dideoxyinosine (ddI, Didanosine), dideoxycytidine (ddC, Zalcitabine), 2',3'-dideoxy-3'-thiacytidine (3TC, Lamivudine), and 2',3'-didehydro-3'-deoxythymidine (d4T, Stavudine), a non-nucleoside RT inhibitor (Niverapine), and protease inhibitors such as saquinavir (Inverase), ritonavir (Norvir), indinavir (Crixivan), and nelfinavir (Viracept). Nucleoside RT inhibitors generally have similar structures (2',3'-dideoxynucleosides) and act at an early stage in virus replication to inhibit provirus DNA synthesis (De Clercq, 1995, Journal of Medicinal Chemistry, 38:2491-2517). However, AZT, the recommended initial therapeutic agent, and the other nucleoside analogues have several limitations, including adverse side effects such as bone marrow depression and anemia (Gill et al., 1987, Annals of Internal Medicine, 107:502-505; Richman et al., 1987, New England Journal of Medicine, 317:192-197). Peripheral neuropathy is also a major and common side effect. AZT is rapidly eliminated from the plasma with a half-life of about one hour (Surbone et al., 1988, Annals of Internal Medicine, 108:534-540) and is quickly metabolized in the liver to its corresponding 5'-glucuronide, which is inactive.

Presently, only a small number of antiviral drugs are available for treatment of virus infections. A complication to the development of such drugs is that mutant strains of virus which are resistant to currently available antiviral drugs are developing at an alarming rate. Combinations of new drugs having unique modes of action are urgently needed to replace drugs that have lost their potency against viruses as a result of virus mutations. A further complication to the development of antiviral drugs is that development of viral resistance to available compounds is not the same in different body compartments and fluids. For example, evolution of drug resistance among HIV-1 clinical isolates is often discordant in blood and semen of HIV-1 positive males (Eron et al., 1998, AIDS 12:F181-F189).

Further, currently available drugs useful for antiviral therapy sometimes ineffectively penetrate the genital tract. This is a serious drawback to the use of these drugs to combat viruses which infect the genital tract. If an antiviral drug promotes development of resistance in the genital tract and the virus is commonly transmitted from this body site, the drug will rapidly become ineffective for treatment of the virus infection in the population at risk for transmission. Hence, drug-resistant mutants of certain viruses can be rapidly spread by sexual contact in the human population. It is known that viruses such as HIV, hepatitis B, hepatitis C, herpes simplex virus, cytomegalovirus, papilloma viruses, and many others are transmitted via sexual contact by both males and females. Thus, therapeutic drugs that fully suppress virus infections in the genital tract are a high public health priority.

Another limitation of presently available antiviral drugs is that rapid emergence of drug resistant mutant virus can lead to decreased sensitivity to the drug within a patient or within a patient population (Larder et al., 1989, Science, 243:1731-1734). Thus, the beneficial effects of drugs such as AZT are limited in duration.

The anti-HIV chemotherapy era which started a decade ago has recently made significant progress toward better control of HIV-1 infection by the introduction of protease inhibitors and the use of combinations of nucleoside and non-nucleoside RT inhibitors with protease inhibitors. Monotherapy (e.g. administration of a single drug) using a nucleoside or non-nucleoside RT inhibitor or a protease inhibitor is no longer a recommended form of therapy for treatment of a patient with a virus infection such as HIV-1 infection. Although combinations of AZT, 3TC, and a protease inhibitor have reduced virus load in the plasma of patients to below detectable levels (i.e. fewer than 200 copies of viral RNA per milliliter of plasma) with a concomitant increase in CD4.sup.+ cell count, some drug combinations have been associated with increased toxicity in a person receiving multiple drug therapies. Also, although reduction in virus burden in the plasma of patients to non-detectable levels achieved using some drug combinations is impressive, drug resistance is an escalating problem due to both use and misuse of drug therapy (De Clercq, 1995, Journal of Medicinal Chemistry, 38:2491-2517; Bartlett, 1996, Infectious Diseases in Clinical Practice, 5:172-179) and evolution of resistant mutants in blood and seminal fluids (Eron et al., 1998, AIDS, 12:F181-F189).

The pathogenic events in HIV disease have recently been reviewed by Fauci (1996, Nature {New Biology}, 384:529-534). The current understanding is that entry of HIV into cells varies with the virus strain and cell type. Primary infection of humans is associated with macrophage tropic (M-tropic) virus that utilize the CD4 receptor and a beta-chemokine co-receptor (CCR5) for entry into macrophages. As HIV infection progresses, the initial M-tropic viruses are usually replaced by T-tropic viruses that enter T-lymphocytes via the CD4 receptor and co-receptor CXCR4 (fusin). The viral determinant of cellular tropism maps to the gp 120 subunit of HIV-1 Env protein, particularly the 3rd variable region or V3 loop of gp120. Upon entry into these cells, HIV probably infects dendretic cells, which then carry the virus to CD4+ cells in the lymphoid organs. Infection is then established in the lymphoid organs and a burst of infectious virus seeds itself throughout the body, including the CNS, brain, and lymphoid tissues and sexual organs (e.g. testes). Current drugs used in therapies for HIV infection and AIDS noted above have a limited capacity and half-life for absorption from the stomach to the blood, accumulation into lymphoid organs, crossing the blood-brain barrier into the CNS, or entering the sexual organs (e.g. testes) to attack sanctuaries for HIV replication.

Synthetic phosphocholine lipid (PC lipid) analogues such as, for example, 1-decanamido-2-decyloxypropyl-3-phosphocholine (INK-11) have demonstrated a low incidence of unwanted side effects in mice such as reduction of bone marrow precursor cells and have exhibited high differential selectivity (i.e. the ratio of TC.sub.50 for cytotoxicity to EC.sub.50 for antiviral activity, DS=1342 for INK-11) in human leukocytes in cultured cells. At a dosage of 50 milligrams per kilogram of body weight per day for 21 days, INK-11 inhibited Friend leukemia virus-(FLV-) induced pathogenesis by 42% in infected mice, as indicated by significant activity against splenomegaly. The observation that use of INK-11 resulted in only moderate suppression against RT activity compared with AZT alone (42% vs 98%, respectively) suggests that INK-11 induces production of defective virus, similar to the effect achieved using other lipid compounds alone (Kucera, et al., 1990, AIDS Research & Human Retroviruses 6:491-501).

Other synthetic phospholipids which do not comprise a phosphocholine moiety (non-PC lipids) have been conjugated with antiviral chemotherapeutic agents. For example, thioether lipid-nucleoside conjugates have exhibited improved antineoplastic activity in tumor-bearing mice (Hong et al., 1990, Journal of Medicinal Chemistry 33:1380-1386). Also, natural phospholipids coupled to AZT or to dideoxynucleosides (ddT, ddC) have proven to be markedly active against HIV by inhibiting viral RT activity (Steim et al., 1990, Biochemical & Biophysical Research Communications 171:451-457; Hostetler et al., 1990, Journal of Biological Chemistry 265:6112-6117; Hostetler et al., 1991, Journal of Biological Chemistry 266:11714-11717). Studies of phospholipid antiviral efficacy have also included chemically conjugating AZT or ddI, through a phosphate-ester bond, to selected synthetic phosphatidic acid lipid analogues (Piantadosi et al., 1991, Journal of Medicinal Chemistry 34:1408-1414). Synthetic phosphate-ester linked lipid-nucleoside conjugates were found to be markedly active against infectious HIV-1 production in both acutely- and persistently-infected cells, and were 5- to 10-fold less cytotoxic compared with AZT alone (Piantadosi et al., 1991, Journal of Medicinal Chemistry 34:1408-1414). Results of preliminary studies indicated that synthetic lipid-AZT conjugates block reactivity of HIV-1-induced gp160/gp120 proteins with specific monoclonal antibodies on the surface of infected and treated cells and on the surface of treated HIV-1 particles, as measured by flow cytometry. These conjugate compounds also caused inhibition of HIV-1-induced cell fusion (Kucera et al., 1992, In: Novel Membrane Interactive Ether Lipids With Anti-Human Immunodeficiency Virus Activity, Aloia et al., eds., Membrane Interactions of HIV, pp. 329-350; Krugner-Higby et al., 1995, AIDS Research & Human Retroviruses 11:705-712). However, these phosphate ester-linked lipid-AZT conjugates (non-PC lipid-AZT conjugates) were not very active against AZT-resistant clinical isolates of HIV-1. Moreover, after intracellular metabolism of the conjugate with resulting release of AZT-monophosphate, the lipid moiety exhibited only moderate to non-detectable antiviral activity (Piantadosi et al., 1991, Journal of Medicinal Chemistry 34:1408-1414).

As with the antiviral agents, the development of anticancer agents for treating cancer effectively has also been problematic. Barriers such as cellular mechanisms of anticancer drug resistance, overcoming the blood-brain barrier to provide adequate delivery of drug to the brain and CNS, inadequate uptake of drug by lymphoid and hematopoietic tissues, toxicity, achieving oral bioavailability, overcoming short drug half-life, and preventing extracellular metabolism of the anticancer agent are faced by the skilled artisan.

In order to improve bioavailability to CNS and brain tissue, nucleoside analogues have been encapsulated in liposomes or used with modifying agents to disrupt the blood-brain barrier (Braekman, et al., 1997, Proc. Amer. Soc. for Clinical Oncology, Abstract #810). Implantable devices have been used to provide more sustained drug delivery to increase the pharmacokinetics of anticancer agents (Del Pan, et al., 1997, Proc. Amer. Soc. for Clinical Oncology, Abstract #1384). Additionally, attempts to improve the efficacy of nucleoside analogues in cancer therapy have included the use of multidrug combinations and high-dose nucleoside analogue therapy (Capizzi, 1996, Investigational New Drugs 14:249-256). None of these methods have adequately overcome the problems discussed above with regard to anticancer agents.

Another attempt to circumvent the problems associated with conventional nucleoside analogue cancer therapy has been the conjugation of these molecules to phospholipids. Thus far, the conjugation of nucleoside analogues to phospholipid molecules has focused on ara-C and a limited number of diacyl, alkylacyl and thioether phospholipids (Hong, 1990, Cancer Res. 50:4401-4406). Although these conjugates have shown efficacy in the treatment of hematologic malignancies, these drugs must be administered intraperitoneally or intravenously and do not overcome the problems discussed above regarding anticancer agents. These conjugates are degraded by phospholipase A and phospholipase B extracellularly and do not provide the option of oral administration.

Despite the promising attributes of compounds such as PC lipids, and non-PC lipid-nucleoside analogue conjugates, currently available antiviral and anticancer agents such as nucleoside analogues and anti-HIV nucleoside drugs have severe inherent limitations. Although such drugs are capable of delaying the onset of symptoms of virus infection and extending survival time for patients, new compounds having the attributes of increased tolerability, potency, and selectivity against specific viruses, differential mechanisms of action, ability to cross the blood-brain barrier, and freedom from myelosuppressive side effects are urgently needed for improved treatment of virus infections. Also, new antiviral and anticancer compounds are needed which more effectively combat cancers or target multiple aspects of the virus life cycle, which facilitate delivery of an anticancer agent to cells and tissues not normally accessible to anticancer agents (e.g. CNS and lymphoid tissues), which combine lipophilic (e.g. phospholipid) and antiretroviral or anticancer agents within the same molecule (e.g. conjugate compounds) in order to yield a drug with a more sustained antiviral or anticancer effect, which decrease the rate of emergence of drug-resistant virus strains, and which inhibit virus replication in a wider range of cellular or tissue reservoirs of virus infection. The present invention satisfies these needs.

BRIEF SUMMARY OF THE INVENTION

The invention includes a compound having the structure of Formula I:

##STR00001## wherein,

n and m are each independently 0 or 1, but n and m are not both 0;

R.sup.1 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl if n is 0 and (C.sub.1-C.sub.16) alkylene, branched alkyl, alkenyl or alkynyl if n is 1;

R.sup.2 is (C.sub.1-C.sub.16) alkyl branched alkyl alkenyl or alkynyl if m is 0 and (C.sub.1-C.sub.16) alkylene, branched alkyl, alkenyl or alkynyl if m is 1;

R.sup.3, R.sup.4 and R.sup.5 are each independently (C.sub.1-C.sub.8) alkylene;

R.sup.6, R.sup.7 and R.sup.8 are each independently (C.sub.1-C.sub.8) alkyl;

X.sup.1 and X.sup.2 are each independently S, O, NHC.dbd.O, OC.dbd.O or NH;

X.sup.3 is O or S;

E.sup.1 is H, S, halo or N.sub.3;

Z.sup.1 is H, S, or halo; or E.sup.1 and Z.sup.1 together are a covalent bond;

E.sup.2 is H, S, halo, or N.sub.3;

Z.sup.2 is H, S, or halo; or E.sup.2 and Z.sup.2 together are a covalent bond;

D.sup.1 and D.sup.2 are each independently selected from the group consisting of purine, pyrimidine, adenine, thymine, cytosine, guanine, hypoxanthine, inosine, uracil and ring modifications thereof, including O, N, and S substitutions, and

wherein, each alkyl, alkylene, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5, R.sup.6, R.sup.7, R.sup.8, D.sup.1, and D.sup.2 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In one aspect, the compound is present in an amount effective to inhibit virus replication in a mammal.

In another aspect, R.sup.1 is (C.sub.6-C.sub.16) alkyl if n is 0 or --CH.dbd.CH-- if n is 1.

In yet another aspect, R.sup.2 is (C.sub.6-C.sub.16) alkyl if m is 0 or --CH.dbd.CH-- if m is 1.

In a further aspect, R.sup.3 is --CH.sub.2CH.sub.2--.

In one embodiment, R.sup.4 is --CH.sub.2--.

In another embodiment, R.sup.5 is --CH.sub.2--.

In yet another embodiment, R.sup.6, R.sup.7 and R.sup.8 are each --CH.sub.3.

In one aspect, X.sup.1 is S, NHC.dbd.O, --NH-- or O.

In another aspect, X.sup.2 is S, NHC.dbd.O or O.

In a further aspect, X.sup.3 is O or S.

In another aspect, E.sup.1 is N.sub.3, S or H.

In one embodiment, Z.sup.1 is H or S.

In another embodiment, E.sup.2 is N.sub.3, S or H.

In a further embodiment, Z.sup.2 is H or S.

In one aspect, n is 0 and m is 1.

In another aspect, n is 1 and m is 0.

In yet another aspect, D.sup.1 is selected from the group consisting of cytosine, guanine, inosine and thymine.

In a further aspect, D.sup.2 is selected from the group consisting of cytosine, guanine, inosine and thymine.

In another aspect, the compound is in the form of a pharmaceutically acceptable salt.

In one embodiment, the compound is present in an amount effective to inhibit virus replication in a mammal.

In a preferred embodiment, R.sup.1 is (C.sub.6-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl; R.sup.2 is (C.sub.4-C.sub.12) alkylene; R.sup.3 is --CH.sub.2CH.sub.2--; R.sup.5 is --CH.sub.2--; R.sup.6, R.sup.7 and R.sup.8 are each CH.sub.3; X.sup.1 and X.sup.2 are each independently S, O or NHC.dbd.O; E.sup.2 is H or N.sub.3; D.sup.2 is selected from the group consisting of thymine, cytosine, guanine and inosine, and wherein each alkyl, branched alkyl, alkylene, alkenyl, alkynyl, thymine, cytosine, guanine, and inosine of R.sup.1, R.sup.2, R.sup.3, R.sup.5, R.sup.6, R.sup.7, R.sup.8, and D.sup.2 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b), wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

The invention also includes a method of treating a virus infection in a mammal. The method comprises administering to the mammal, in an amount effective to treat the infection, a compound having the structure of Formula I:

##STR00002## wherein,

n and m are each independently 0 or 1, but n and m are not both 0;

R.sup.1 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl if n is 0 and (C.sub.1-C.sub.16) alkylene, branched alkyl, alkenyl or alkynyl if n is 1;

R.sup.2 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl if m is 0 and (C.sub.1-C.sub.16) alkylene, branched alkyl, alkenyl or alkynyl if m is 1;

R.sup.3, R.sup.4 and R.sup.5 are each independently (C.sub.1-C.sub.8) alkylene;

R.sup.6, R.sup.7 and R.sup.8 are each independently (C.sub.1-C.sub.8) alkyl;

X.sup.1 and X.sup.2 are each independently S, O, NHC.dbd.O, OC.dbd.O or NH;

X.sup.3 is O or S;

E.sup.1 is H, S, halo or N.sub.3;

Z.sup.1 is H, S, or halo; or E.sup.1 and Z.sup.1 together are a covalent bond;

E.sup.2 is H, S, halo, or N.sub.3;

Z.sup.2 is H, S, or halo; or E.sup.2 and Z.sup.2 together are a covalent bond;

D.sup.1 and D.sup.2 are each independently selected from the group consisting of purine, pyrimidine, adenine, thymine, cytosine, guanine, hypoxanthine, inosine, uracil and ring modifications thereof, including O, N, and S substitutions, and

wherein, each alkyl, alkylene, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5, R.sup.6, R.sup.7, R.sup.8, D.sup.1, and D.sup.2 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In one aspect, R.sup.1 is (C.sub.8-C.sub.12) alkyl if n is 0 and --CH.sub.2CH.sub.2-- if n is 1.

In another aspect, R.sup.2 is (C.sub.8-C.sub.12) alkyl if m is 0 and --CH.sub.2CH.sub.2-- if m is 1.

In yet another aspect, R.sup.3 is --CH.sub.2CH.sub.2--.

In one embodiment, R.sup.4 is --CH.sub.2--.

In another embodiment, R.sup.5 is --CH.sub.2--.

In a further embodiment, R.sup.6, R.sup.7 and R.sup.8 are each --CH.sub.3.

In another embodiment, X.sup.1 is S or O.

In one aspect, X.sup.2 is S or O.

In another aspect, X.sup.3 is O.

In a further aspect, E.sup.1 is N.sub.3 or H.

In a still further aspect, Z.sup.1 is H.

In one embodiment, E.sup.2 is N.sub.3 or H.

In another embodiment, Z.sup.2 is H.

In yet another embodiment, n is 0 and m is 1.

In one aspect, n is 1 and m is 0.

In another aspect, D.sup.1 is selected from the group consisting of cytosine, guanine, inosine, and thymine.

In a further aspect, D.sup.2 is selected from the group consisting of cytosine, guanine, inosine, and thymine.

In a still further aspect, the virus infection is an infection by a virus selected from the group consisting of HIV, hepatitis virus, and a herpes virus.

In one embodiment, the HIV is selected from the group consisting of HIV-1 and HIV-2.

Preferably, the hepatitis virus is selected from the group consisting of hepatitis A, hepatitis B, hepatitis C, hepatitis D, and hepatitis E viruses.

Preferably, the herpes virus is selected from the group consisting of herpes simplex virus type 1, herpes simplex virus type 2, varicella-zoster virus, cytomegalovirus, Epstein Barr virus, human herpes virus type 6, human herpes virus type 7, and human herpes virus type 8.

In a preferred aspect, R.sup.1 is (C.sub.6-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl; R.sup.2 is (C.sub.4-C.sub.12) alkylene; R.sup.3 is --CH.sub.2CH.sub.2--; R.sup.5 is --CH.sub.2--; R.sup.6, R.sup.7 and R.sup.8 are each CH.sub.3; X.sup.1 and X.sup.2 are each independently S, O or NHC.dbd.O; E.sup.2 is H or N.sub.3; D.sup.2 is selected from the group consisting of thymine, cytosine, guanine and inosine, and wherein each alkyl, branched alkyl, alkylene, alkenyl, alkynyl, thymine, cytosine, guanine, and inosine of R.sup.1, R.sup.2, R.sup.3, R.sup.5, R.sup.6, R.sup.7, R.sup.8, and D.sup.2 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b), wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In one aspect, a pharmaceutically acceptable salt of the compound is administered to the mammal.

Preferably, the mammal is a human.

The invention also includes a method of inhibiting virus replication in a cell. The method comprises administering to the cell a compound of Formula I in an amount effective to inhibit virus replication in the cell.

The invention includes a pharmaceutical composition comprising a compound of Formula I in combination with a pharmaceutically acceptable carrier.

In one aspect, the compound is present in an amount effective to inhibit virus replication in a mammal.

In another aspect, the compound is present in an amount effective to inhibit virus replication in a mammal.

The invention also includes a kit for treatment of a viral infection in a mammal. The kit comprises a) a composition selected from the group consisting of a compound of Formula I, a pharmaceutically acceptable salt thereof, and a pharmaceutical composition comprising a compound of Formula I, and b) an instructional material.

The invention also includes a kit for inhibition of virus replication in a cell. The kit comprises a) a composition selected from the group consisting of a compound of Formula I, a pharmaceutically acceptable salt thereof, and a pharmaceutical composition comprising a compound of Formula I, and b) an instructional material.

The invention also includes a compound having the structure of Formula III:

##STR00003## wherein,

R.sup.11 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl;

R.sup.12 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl;

X.sup.11 is O, S, or NHC.dbd.O;

X.sup.12 is O, S, or NHC.dbd.O;

X.sup.13 is O or S;

n is 0, 1 or 2, and

R.sup.13 is a therapeutic agent,

wherein, each alkyl, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.11, R.sup.12, and R.sup.13 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl, and

wherein, if n is 1 or 2, the compound is a phospholipase C substrate and is not a phospholipase A substrate, and

further wherein, if n is 1 or 2, the compound is converted to an alkyl lipid and a moiety selected from the group consisting of a nucleoside monophosphate and a nucleoside analogue monophosphate intracellularly in a mammal, and is not converted to an alkyl lipid and a moiety selected from the group consisting of a nucleoside monophosphate and a nucleoside analogue monophosphate extracellularly in a mammal.

In a preferred embodiment, R.sup.11 is a C.sub.12 alkyl, branched alkyl, alkenyl or alkynyl; R.sup.12 is C.sub.8H.sub.16 alkyl or branched alkyl; n=1, and R.sup.13 is an anticancer agent selected from the group consisting of gemcitabine, ara-C, 5-azacytidine, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside, 6-mercaptopurine, 6-thioguanine 5-deoxyfluorouridine, ftorafur, capecitabine, 5-deoxy-5-fluorocytidine, 5-aza-cytsine arabinoside, troxacitabine, and pentostatin, wherein the phosphorus atom of the phosphate moiety is covalently linked in a phosphate ester linkage to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.13.

The invention also includes a compound having the structure of Formula IV:

##STR00004## wherein,

R.sup.21 is (C.sub.6 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl;

R.sup.22 is (C.sub.1 to C.sub.12) alkyl, branched alkyl, alkenyl, or alkynyl;

X.sup.21 is O, S, or NHC.dbd.O;

X.sup.22 is O, S, or NHC.dbd.O;

X.sup.23 is O or S;

n is 1 or 2;

R.sup.23 is a therapeutic agent, and

wherein, each alkyl, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.21, R.sup.22, and R.sup.23 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In a preferred aspect, R.sup.21 is C.sub.12 alkyl; R.sup.22 is C.sub.10 alkyl; n=1, and R.sup.23 is an anticancer agent selected from the group consisting of gemcitabine, ara-C, 5-azacytidme, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside, 6-mercaptopurine, 6-thioguanine, 5-deoxyfluorouridine, ftorafur, capecitabine, 5-deoxy-5-fluorocytidine, 5-aza-cytsine arabinoside, troxacitabine, and pentostatin, wherein the methylene group of the phosphonate moiety is covalently linked to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.23.

The invention also includes a compound having the structure of Formula V:

##STR00005## wherein,

R.sup.31 is (C.sub.1 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl;

R.sup.32 is (C.sub.1 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl;

X.sup.31 is O, S, or NHC.dbd.O;

X.sup.32 is O, S, or NHC.dbd.O;

X.sup.33 is --OH, --SH, or amino;

R.sup.33 is a therapeutic agent, and

wherein, each alkyl, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.31, R.sup.32, and R.sup.33 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In a preferred embodiment, R.sup.31 is (C.sub.6-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl; R.sup.32 is (C.sub.1-C.sub.8) alkyl, branched alkyl, alkenyl or alkynyl, and R.sup.32 is an anticancer agent selected from the group consisting of mitoxanthrone, doxorubicin, idarubicin, epirubicin, daunorubicin, mitomycin, methotrexate, CPT-11, SN-38, camptothecin, topotecan, 9-nitrocamptothecin, and 9-aminocamptothecin, and is covalently linked via an ester, amido or carbamate linkage to the --SH, OH or amino group of X.sup.33.

In one aspect, the compound is suspended in a pharmaceutically acceptable carrier and is present in an amount effective to combat a cancer in a mammal.

Preferably, the cancer is a cancer selected from the group consisting of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

In one aspect, the compound is present in an amount effective to facilitate delivery of a therapeutic agent to a mammalian cell.

Preferably, the therapeutic agent is an anticancer agent.

Preferably, the cell is in a mammal.

Preferably, the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

In one aspect, the CNS cell is an astrocyte or a glial cell.

In one embodiment, the compound is in the form of a pharmaceutically acceptable salt.

In one aspect, the compound is present in an amount effective to facilitate delivery of a therapeutic agent to a mammalian cell.

In one embodiment, the cell is in a mammal.

Preferably, the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

In one aspect, the compound is present in an amount effective to combat a cancer in a mammal.

In one embodiment, the compound in the pharmaceutically acceptable salt is present in an amount effective to facilitate delivery of a therapeutic agent to a mammalian cell.

Preferably, the therapeutic agent is an anticancer agent.

In one aspect, the cell is in a mammal.

In one embodiment, the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

In one aspect, the compound is present in an amount effective to combat a cancer in a mammal.

The invention also includes a drug delivery agent comprising a pharmaceutical composition. The composition comprises a compound of Formula III or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of a therapeutic agent to a mammalian cell.

In one aspect, the therapeutic agent is an anticancer agent.

In another aspect, the cell is in a mammal.

Preferably, the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

The invention also includes a drug delivery agent comprising a pharmaceutical composition. The composition comprises a compound of Formula III or a pharmaceutically acceptable salt thereof, in an amount effective to combat a cancer in a mammal.

Preferably, the cancer is a cancer selected from the group consisting of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

The invention also includes a method of facilitating delivery of a therapeutic agent to a mammalian cell. The method comprises administering to the cell a pharmaceutical composition comprising a compound of Formula III or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of the therapeutic agent to the cell.

In one aspect, the therapeutic agent is an anticancer agent.

In another aspect, the cell is in a mammal.

Preferably, the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

The invention also includes a method of facilitating delivery of a therapeutic agent to a cell. The method comprises administering to the cell a pharmaceutical composition comprising a compound of Formula III or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of the therapeutic agent to the cell.

In one aspect, the cell is in a mammal.

In another aspect, the cell is a cell selected from the group consisting of a CNS cell and a lymphoid cell.

The invention also includes a method of combating a cancer in a mammal. The method comprises administering to the mammal a pharmaceutical composition comprising a compound of Formula III or a pharmaceutically acceptable salt thereof, in an amount effective to combat a cancer in the mammal.

In one aspect, the cancer is a cancer selected from the group consisting of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

The invention also includes a method of treating a disease in a mammal. The method comprises administering to the mammal a pharmaceutical composition comprising a compound of Formula III, or a pharmaceutically acceptable salt thereof, in an amount effective to facilitate delivery of a therapeutic agent to a cell in the mammal, thereby treating the disease.

In one aspect, the disease is a disease selected from the group consisting of a brain disease, a CNS disease, a lymphatic system disease, a reproductive system disease, a cardiovascular disease, a kidney disease and a liver disease.

The invention also includes a kit for combating a cancer in a mammal. The kit comprises a) a composition selected from the group consisting of a compound of Formula III, a pharmaceutically acceptable salt thereof, and a pharmaceutical composition comprising a compound of Formula III, and b) an instructional material.

The invention also includes a kit for facilitating delivery of a therapeutic agent to a mammalian cell. The kit comprises a) a composition selected from the group consisting of a compound of Formula III, a pharmaceutically acceptable salt thereof, and a pharmaceutical composition comprising a compound of Formula III, and b) an instructional material.

Preferably, the therapeutic agent is an anticancer agent.

BRIEF DESCRIPTION OF THE DRAWINGS

The foregoing summary, as well as the following detailed description of the invention, will be better understood when read in conjunction with the appended drawings.

FIG. 1, comprising FIGS. 1A, 1B, and 1C is a series of formulae depicting the chemical structures of several anticancer agents. FIG. 1A depicts the chemical structure of gemcitabine. FIG. 1B depicts the chemical structure of ara-C. FIG. 1C depicts the chemical structure of 5-azacytidine.

FIG. 2 is a reaction scheme depicting the synthesis method for preparing a lipid backbone (i.e. an alkyl lipid) for the compounds of the invention.

FIG. 3 is a reaction scheme depicting the synthesis method for preparing an AZT-malonic acid (AZT-MA) compound, which is an intermediate compound in the synthesis of the double targeting PC lipid-AZT conjugate compounds of the invention.

FIG. 4 is a reaction scheme depicting the synthesis method for preparing a double targeting PC lipid-AZT conjugate compound of the invention (INK-20).

FIG. 5, comprising FIGS. 5A and 5B, is a pair of formulae depicting the chemical structures of exemplary compounds of Formula III.

FIG. 6, comprising FIGS. 6A and 6B, is a pair of formulae depicting the chemical structures of exemplary compounds of Formula IV.

FIG. 7 is a formula depicting the chemical structure of an exemplary compound of Formula V.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to methods and compositions useful in drug delivery for treatment of a virus infection in a mammal by targeting the virus at two or more stages of the virus life cycle and thereby inhibiting virus replication. This mode of use of antiviral compositions is referred to herein as double-targeting a virus infection. The compositions of the invention include compounds comprising at least two chemically combined (e.g. covalently conjugated) antiviral agents which have different modes of action. Because the antiviral agents have different modes of action, they target the virus life cycle at two or more different stages. By way of example and not by limitation, the compositions of the invention include compounds having a nucleoside analogue or protease inhibitor moiety conjugated with a phosphocholine lipid (PC lipid) moiety. Also by way of example and not by limitation, the targets in the viral life cycle of the compounds of the invention may include stages involving reverse transcription, protease activity, and virus assembly. The methods and compositions of the invention are particularly useful in combating drug-resistant mutants of viruses because viruses resistant to nucleoside analogues and protease inhibitors are still sensitive to inhibition by phospholipids.

As used herein, the term "conjugated with" means covalently attached to the same molecule.

The targeted virus may be any type of virus, and non-limiting exemplary viruses include HIV-1, HIV-2, hepatitis virus (e.g. hepatitis A, hepatitis B, hepatitis C, hepatitis D, and hepatitis E viruses), and herpesviruses (e.g. herpes simplex virus types 1 and 2, varicella-zoster virus, cytomegalovirus, Epstein Barr virus, and human herpes viruses types 6, 7, and 8).

The compounds of the invention exhibit biological properties which are superior to currently available antiviral drugs, including (i) reduced cytotoxicity accompanied by the ability of the mammal to tolerate a higher dose of the drug compared with nucleoside analogues or protease inhibitors alone, (ii) ability to target multiple distinct stages of virus replication (e.g. reverse transcription, protease processing of viral proteins and virus assembly, leading to non-replication or to production of defective progeny virus) (iii) ability to simultaneously deliver constant amounts of multiple antiviral agents (e.g. phosphocholine lipid and a nucleoside analogue or phosphocholine lipid and a protease inhibitor) to virus-infected cells with preferential uptake into the CNS, lymphoid tissues, and male and female genital tracts, (iv) intracellular metabolism of the conjugate compound and simultaneous release of two antiviral agents in cells in which virus is multiplying, (v) increased half-life of the compound in vivo compared with nucleoside analogues or protease inhibitors alone, (vi) prolonged duration of biological effect, presumably owing to protection of nucleoside analogue from rapid glucuronide formation in the liver of intact animals, and (vii) capacity to conjugate other small molecular weight compounds to the phosphocholine (PC) lipid backbone for treatment of other diseases of the central nervous system (e.g. Alzheimer's, cancer), in addition to diseases such as AIDS, resulting from virus infection.

Previous studies have established that a PC moiety is an essential component for a phospholipid to exhibit optimal antiviral activity (Piantadosi et al., 1991, J. Med. Chem. 34:1408-1414; Krugner-Higby et al., 1995, AIDS Res. & Human Retrovir. 11:705-712). Compounds comprising phosphatidic acid, phosphoethanolamine, phosphoalkylpyridine, alcohol, or quaternary amine salt moieties were less active, more toxic, exhibited much lower differential selectivities, or some combination of these, relative to the corresponding PC lipids. In certain preferred compounds of the invention, a PC moiety is incorporated into the lipid backbone to result in compounds which exhibit optimal antiviral activity, can accumulate into lymphoid tissues, testes, and vaginal secretions, and can pass the blood-brain barrier into the CNS. These anatomical sites serve as important reservoirs of virus during infection by viruses such as HIV-1, and also serve as sources of transmission of drug-resistant mutants.

The invention also includes methods of treating a virus infection in a cell or in a mammal, such as a human, comprising administering to the cell or mammal a compound of the invention in an amount effective to alleviate or eliminate the virus infection or to alleviate a symptom associated with the infection.

The present invention also includes methods and compositions useful in drug delivery for facilitating delivery of a therapeutic agent to a mammalian cell. As used herein, the term "facilitating delivery" or "to facilitate delivery" of a therapeutic agent to a mammalian cell means enhancing the uptake of a therapeutic agent in a mammalian cell to a level higher than the level of uptake of the therapeutic agent in an otherwise identical mammalian cell which is not administered a compound or composition of the invention. The uptake of a therapeutic agent can be enhanced, by way of example and not by limitation, by any one or more of the following means: by bypassing the requirement for a cellular active transport mechanism for uptake of the therapeutic agent into a cell; by providing the therapeutic agent (i.e. a drug) intracellularly in an activated form, (i.e. the monophosphorylated form in the case of a nucleoside analogue anticancer drug) thereby bypassing the requirement for intracellular activation of the therapeutic agent by an enzyme such as an intracellular kinase; by overcoming a physiological barrier to uptake of the therapeutic agent in a desired cell, such as low solubility, poor absorption from the stomach or small intestine, or impermeability to the blood-brain barrier, to enable delivery of the therapeutic agent to sites not normally accessible thereto (i.e. CNS and lymphoid tissues).

The present invention also includes methods and compositions useful in drug delivery for combating a cancer in a mammal or for treating or alleviating a disease in a mammal.

As used herein, the term "combating a cancer" or "to combat a cancer" in a mammal means, for example, any one or more of the following: to increase survival of a mammal, to decrease or arrest tumor size in a mammal, or to increase the time period of remission of cancer regrowth in a mammal, relative to an otherwise identical mammal which was not administered a composition or compound of the invention.

As used herein, the term "therapeutic agent" means any compound or composition, which, upon entering a mammalian cell, is capable of being of benefit in alleviating or treating a disease in a mammal. By way of example and not by limitation, such compounds and compositions include small organic molecules, peptides, nucleoside analogues, anticancer agents, antiviral agents, ribozymes, antisense oligonucleotides and other drugs. The disease may be any disease experienced by a mammal. By way of example and not by limitation, such diseases include diseases of the brain, CNS, lymphatic system, reproductive system, cardiovascular system, renal system and liver, among others.

As used herein, "alleviating a disease" means reducing the severity of a symptom of the disease.

As used herein, "treating a disease" means reducing the frequency with which a symptom of the disease is experienced by a mammal.

As used herein, the term "anticancer agent" means a therapeutic agent which is capable of exhibiting efficacy at combating a cancer in a mammal or in a mammalian cell, or any compound which is capable of being converted intracellularly to a compound which is capable of exhibiting efficacy at combating a cancer in a mammal or in a mammalian cell.

The mammalian cell can be any type of mammalian cell, including both cancerous and non-cancerous cells. Examples of preferred cells include, but are not limited to, CNS and lymphoid cells. Preferred lymphoid cells include lymphoma, spleen and thymus cells. Preferred CNS cells include brain cells, astrocytes, and glial cells. The cancer can be any type of cancer in a mammal. Preferably, the cancer is one or more of a carcinoma, a sarcoma, a neuroblastoma, a leukemia, a lymphoma and a solid tumor.

The compositions of the invention include compounds which comprise an alkyl lipid or a phospholipid moiety covalently conjugated with a therapeutic agent. As used herein, the term "alkyl lipid" means that portion of any of the compounds of Formulas III, IV and V as described herein without the therapeutic agent moiety.

The invention also includes pharmaceutical compositions and kits for combating a cancer and/or for facilitating delivery of a therapeutic agent to a mammalian cell.

The invention also includes methods which comprise administering a compound of the invention, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition of the invention, in an amount effective to combat a cancer or in an amount effective to facilitate delivery of a therapeutic agent to a mammalian cell.

As used herein, the following terms are defined as follows, unless otherwise described: halo is fluoro, chloro, bromo, or iodo. Alkyl, alkoxy, alkylene, etc. denote both straight and branched groups; but reference to an individual radical such as "propyl" embraces only the straight chain radical, a branched chain isomer such as "isopropyl" being specifically referred to.

The articles "a" and "an" are used herein to refer to one or to more than one (i.e. to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

Compounds of the invention having a chiral center can exist in and be isolated in distinct optically active or racemic forms. The present invention encompasses any racemic, optically-active, polymorphic, or stereoisomeric form, or mixtures of such forms of a compound of the invention. Preparation of optically active forms of a compound is well known in the art (for example, by resolution of the racemic form by recrystallization techniques, by synthesis from optically-active starting materials, by chiral synthesis, or by chromatographic separation using a chiral stationary phase). Determination or assessment of antiviral activity can be performed using standard tests described herein or other tests known in the art.

Specific and preferred definitions listed below for radicals and substituents are for illustration only; they do not exclude other defined values or other values within defined ranges for the radicals and substituents described herein.

C.sub.1-C.sub.8 alkyl moieties include, for example, methyl, ethyl, propyl, isopropyl, butyl, iso-butyl, sec-butyl, pentyl, sec-pentyl, iso-pentyl, hexyl, sec-hexyl, iso-hexyl, heptyl, sec-heptyl, iso-heptyl, and octyl moieties. C.sub.1-C.sub.8 alkoxy moieties include, for example, methoxy, ethoxy, propoxy, isopropoxy, butoxy, iso-butoxy, sec-butoxy, pentoxy, sec-pentoxy, iso-pentoxy, hexyloxy, sec-hexyloxy, heptoxy, sec-heptoxy, iso-heptoxy, and octyloxy moieties. C.sub.1-C.sub.8 alkylene moieties include, for example, methylene, ethylene, propylene, isopropylene, butylene, iso-butylene, sec-butylene, pentylene, sec-pentylene, iso-pentylene, hexylene, sec-hexylene, iso-hexylene, heptylene, sec-heptylene, iso-heptylene, and octylene moieties. C.sub.6-C.sub.15 alkyl moieties include, for example, hexyl, heptyl, sec-heptyl, iso-heptyl, octyl, sec-octyl, iso-octyl, nonyl, sec-nonyl, iso-nonyl, decyl, sec-decyl, iso-decyl, undecyl, sec-undecyl, iso-undecyl, dodecyl, sec-dodecyl, iso-dodecyl, tridecyl, sec-tridecyl, iso-tridecyl, tetradecyl, sec-tetradecyl, iso-tetradecyl, and pentadecyl moieties. C.sub.6-C.sub.15 alkylene moieties include, for example, hexylene, heptylene, sec-heptylene, iso-heptylene, octylene, sec-octylene, iso-octylene, nonylene, sec-nonylene, iso-nonylene, decylene, sec-decylene, iso-decylene, undecylene, sec-undecylene, iso-undecylene, dodecylene, sec-dodecylene, iso-dodecylene, tridecylene, sec-tridecylene, iso-tridecylene, tetradecylene, sec-tetradecylene, iso-tetradecylene, and pentadecylene moieties. C.sub.8-C.sub.12 alkyl moieties include, for example, octyl, sec-octyl, iso-octyl, nonyl, sec-nonyl, iso-nonyl, decyl, sec-decyl, iso-decyl, undecyl, sec-undecyl, iso-undecyl, and dodecyl moieties. C.sub.8-C.sub.12 alkylene moieties include, for example, octylene, sec-octylene, iso-octylene, nonylene, sec-nonylene, iso-nonylene, decylene, sec-decylene, iso-decylene, undecylene, sec-undecylene, iso-undecylene, and dodecylene moieties.

The present invention includes compounds which exhibit antiviral activity and are particularly useful because they exhibit antiviral activity against drug-resistant viruses. Accordingly, the invention includes a compound having the chemical structure of Formula I or a pharmaceutically acceptable salt thereof.

Formula I is

##STR00006## wherein

n and m are each independently 0 or 1, but n and m are not both 0;

R.sup.1 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl if n is 0 and (C.sub.1-C.sub.16) alkylene, alkenyl or alkynyl if n is 1;

R.sup.2 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl if m is 0 and (C.sub.1-C.sub.16) alkylene, alkenyl or alkynyl if m is 1;

R.sup.3, R.sup.4 and R.sup.5 are each independently (C.sub.1-C.sub.8) alkylene;

R.sup.6, R.sup.7 and R.sup.8 are each independently (C.sub.1-C.sub.8) alkyl;

X.sup.1 and X.sup.2 are each independently S, O, NHC.dbd.O, OC.dbd.O or NH;

X.sup.3 is O or S;

E.sup.1 is H, S, halo or N.sub.3;

Z.sup.1 is H, S, or halo; or E.sup.1 and Z.sup.1 together are a covalent bond;

E.sup.2 is H, S, halo, or N.sub.3;

Z.sup.2 is H, S, or halo; or E.sup.2 and Z.sup.2 together are a covalent bond, and

D.sup.1 and D.sup.2 are each independently selected from the group consisting of purine, pyrimidine, adenine, thymine, cytosine, guanine, hypoxanthine, inosine, uracil and ring modifications thereof, including O, N, and S substitutions.

In Formula I, each alkyl, alkylene, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5, R.sup.6, R.sup.7, R.sup.8, D.sup.1, and D.sup.2 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoro, trifluoromethyl, trifluoromethoxy, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

The following are examples of definitions for radicals and substituents of Formula I in preferred embodiments. These examples are not limiting, but are instead provided as examples of several preferred embodiments which are included in the invention.

In preferred embodiments, R.sup.1 can be one of (C.sub.2-C.sub.16) alkylene, --(CH.sub.2).sub.12--, and --CH.dbd.CH--. In these embodiments, R.sup.1 is optionally substituted with 1, 2, 3, or 4 substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In preferred embodiments, R.sup.2 can be one of (C.sub.2-C.sub.16) alkylene, --(CH.sub.2).sub.8--, --(CH.sub.2).sub.9--, --(CH.sub.2).sub.10--, --(CH.sub.2).sub.11--, --(CH.sub.2).sub.12--, and --CH.dbd.CH--. In these embodiments, R.sup.2 is optionally substituted with 1, 2, 3 or 4 substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

R.sup.3 is preferably --CH.sub.2CH.sub.2--, optionally substituted with 1, 2, 3, or 4 substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

R.sup.4 is preferably --CH.sub.2--, optionally substituted with 1 or 2, substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

R.sup.5 is preferably --CH.sub.2--, optionally substituted with 1 or 2 substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In one preferred embodiment, each of R.sup.6, R.sup.7 and R.sup.8 is --CH.sub.3, each optionally substituted with 1 or 2 substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

X.sup.1 is preferably S, O or NHC.dbd.O.

X.sup.2 is preferably S, O or NHC.dbd.O.

X.sup.3 is preferably O or S.

E.sup.1 is preferably N.sub.3, S, or H.

Z.sup.1 is preferably H.

E.sup.2 is preferably N.sub.3, S or H.

Z.sup.2 is preferably H.

D.sup.1 is preferably cytosine, guanine, inosine or thymine, wherein D.sub.1 is optionally substituted with 1, 2, 3, or 4 substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

D.sup.2 is preferably cytosine, guanine, inosine or thymine, wherein D.sub.2 is optionally substituted with 1, 2, 3 or 4 substituents selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

The invention also includes a compound which exhibits antiviral activity having the chemical structure of Formula II or a pharmaceutically acceptable salt thereof.

Formula II is

##STR00007## wherein,

R.sup.1 is (C.sub.6-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl;

R.sup.2 is (C.sub.4-C.sub.12) alkylene;

R.sup.3 is --CH.sub.2CH.sub.2--;

R.sup.5 is --CH.sub.2--;

R.sup.6, R.sup.7 and R.sup.8 are each CH.sub.3;

X.sup.1 and X.sup.2 are each independently S, O or NHC.dbd.O;

E.sup.2 is H or N.sub.3, and

D.sup.2 is selected from the group consisting of thymine, cytosine, guanine and inosine.

In Formula II, each alkyl, branched alkyl, alkenyl, alkynyl, thymine, cytosine, guanine, and inosine of R.sup.1, R.sup.2, R.sup.3, R.sup.5, R.sup.6, R.sup.7, R.sup.8, and D.sup.2 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b), wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

The present invention also includes compounds which are useful in drug delivery for treating or alleviating a disease or combating a cancer in a mammal. The compounds are also useful for facilitating delivery of a therapeutic agent to a mammalian cell. Accordingly, the invention includes a compound having the chemical structure of Formula III or a pharmaceutically acceptable salt thereof.

Formula III is

##STR00008## wherein,

R.sup.11 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl;

R.sup.12 is (C.sub.1-C.sub.16) alkyl, branched alkyl, alkenyl or alkynyl;

X.sup.11 is O, S, or NHC.dbd.O;

X.sup.12 is O, S, or NHC.dbd.O;

X.sup.13 is O or S;

n is 0, 1 or 2, and

R.sup.13 is a therapeutic agent.

In Formula III, each alkyl, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.11, R.sup.12, and R.sup.13 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In Formula III, if n is 1 or 2, the compound is a phospholipase C substrate and is not a phospholipase A substrate. Also, if n is 1 or 2, the compound is converted to an alkyl lipid and a moiety selected from the group consisting of a nucleoside monophosphate and a nucleoside analogue monophosphate intracellularly in a mammal, and is not converted to an alkyl lipid and a moiety selected from the group consisting of a nucleoside monophosphate and a nucleoside analogue monophosphate extracellularly in a mammal.

The conjugate compounds of Formula III can be formulated in pharmaceutical compositions as described herein, which have the advantageous properties of being suitable for oral administration, can be readily absorbed from the gastrointestinal tract, can cross the blood-brain barrier and be of value in the treatment of CNS diseases and cancers. These conjugates can be used in a number of different cell lines including, by way of example and not by limitation, brain tumor cells, lymphoid cells and pancreatic tumor cells.

In compounds of Formula III which have at least one phosphate group (i.e., n=1 or 2) the phosphate ester linkage is cleaved intracellularly in a mammal by the action of a phospholipase C-like activity to release intracellularly a phospholipid and an anticancer agent. These compounds are substrates of phospholipase C, but not substrates of phospholipase A. Because the phospholipase C activity is intracellular, the conjugates are only converted to a phospholipid and a nucleoside monophosphate intracellularly, and not extracellularly. The metabolism of these compounds by an intracellular phospholipase C-like activity enables the compounds to be used in methods which circumvent the rate limiting step for the activation of nucleoside analogue prodrugs, namely, the conversion of nucleoside analogue to nucleoside analog monophosphate. Because they are metabolized intracellularly to release a nucleoside analogue monophosphate, the administration of these compounds results in the ability to provide an anticancer agent which can be effective in cancer cells which lack a kinase enzyme such as, for example, deoxycytidine kinase, as a mechanism of cellular anticancer drug resistance. Additionally, the phospholipid moiety can affect signal transduction pathways involving protein kinase C and MAP kinase signaling cascades.

The released nucleoside monophosphate serves two purposes. First, it bypasses the rate limiting step in the activation of several nucleoside prodrugs, namely, deoxycytidine kinase. Second, the polar phosphate group "locks" the nucleoside within the cell. The phospholipid conjugate also serves as a reservoir for the drug, increasing the drugs half-life. The capacity to conjugate other small molecular weight compounds to the phospholipid backbone for the treatment of other diseases of the central nervous system (i.e. Alzheimer's) is also of great utility. For example, an ether-lipid moiety can be used as a backbone for conjugation to a variety of therapeutic agents including nucleoside analogues, anticancer and antiviral agents, ribozymes and antisense oligonucleotides. Since the ether-lipid backbone is lipophilic, these conjugates can cross the blood-brain barrier and be used as prodrugs in the treatment of CNS diseases, such as Alzheimer's and neurologic degenerative diseases. The lipophilic property of the conjugates enables them to cross the blood-brain barrier, and thus bypass the requirement for an active transport system in the cell in which uptake of the drug is desired.

In preferred compounds of Formula III,

R.sup.11 is a C.sub.12 alkyl, branched alkyl, alkenyl or alkynyl;

R.sup.12 is C.sub.8H.sub.16 alkyl or branched alkyl;

n=1,

and R.sup.13 is an anticancer agent.

Preferably, the anticancer agent is selected from the group consisting of gemcitabine, ara-C, 5-azacytidine, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside 6-mercaptopurine, 6-thioguanine, 5-deoxyfluorouridine, ftorafur, capecitabine, 5-deoxy-5-fluorocytidine, 5-aza-cytsine arabinoside, troxacitabine, and pentostatin, wherein the phosphorus atom of the phosphate moiety is covalently linked in a phosphate ester linkage to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.13.

The invention includes additional compounds which are useful in drug delivery for treating or alleviating a disease or combating a cancer in a mammal. The compounds are also useful for facilitating delivery of a therapeutic agent to a mammalian cell. Accordingly, the invention includes a compound having the chemical structure of Formula IV or a pharmaceutically acceptable salt thereof.

Formula IV is

##STR00009## wherein,

R.sup.21 is (C.sub.6 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl;

R.sup.22 is (C.sub.1 to C.sub.12) alkyl, branched alkyl, alkenyl, or alkynyl;

X.sup.21 is O, S, or NHC.dbd.O;

X.sup.22 is O, S, or NHC.dbd.O;

X.sup.23 is O or S;

n is 1 or 2, and

R.sup.23 is a therapeutic agent.

In Formula IV, each alkyl, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.21, R.sup.22, and R.sup.23 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In preferred compounds of Formula IV,

R.sup.21 is C.sub.12 alkyl;

R.sup.22 is C.sub.10 alkyl;

n=1, and

R.sup.23 is an anticancer agent.

Preferably, the anticancer agent is selected from the group consisting of gemcitabine, ara-C, 5-azacytidine, cladribine, fludarabine, fluorodeoxyuridine, cytosine arabinoside 6-mercaptopurine, 6-thioguanine, 5-deoxyfluorouridine, ftorafur, capecitabine, 5-deoxy-5 -fluorocytidine, 5-aza-cytsine arabinoside, troxacitabine, and pentostatin, wherein the methylene group of the phosphonate moiety is covalently linked to the oxygen atom of the 5' hydroxyl group of a sugar moiety of R.sup.23.

The invention includes additional compounds which are useful in drug delivery for treating or alleviating a disease or combating a cancer in a mammal. The compounds are also useful for facilitating delivery of a therapeutic agent to a mammalian cell. Accordingly, the invention includes a compound having the chemical structure of Formula V or a pharmaceutically acceptable salt thereof.

Formula V is

##STR00010## wherein,

R.sup.31 is (C.sub.1 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl;

R.sup.32 is (C.sub.1 to C.sub.16) alkyl, branched alkyl, alkenyl, or alkynyl;

X.sup.31 is O, S, or NHC.dbd.O;

X.sup.32 is O, S, or NHC.dbd.O;

X.sup.33 is --OH, --SH, or amino, and

R.sup.33 is a therapeutic agent.

In Formula V, each alkyl, branched alkyl, alkenyl, alkynyl, adenine, thymine, cytosine, guanine, pyrimidine, purine, hypoxanthine, inosine and uracil of R.sup.31, R.sup.32, and R.sup.33 can, optionally, be substituted with 1, 2, 3, or 4 substituents independently selected from the group consisting of halo, nitro, trifluoromethyl, (C.sub.1-C.sub.8) alkyl, (C.sub.1-C.sub.8) alkoxy, aryl, and N(R.sup.a)(R.sup.b) wherein R.sup.a and R.sup.b are each independently selected from the group consisting of H and (C.sub.1-C.sub.8) alkyl.

In preferred compounds of Formula V,

R.sup.31 is (C.sub.6-C.sub.16) alkyl branched alkyl alkenyl or alkynyl;

R.sup.32 is (C.sub.1-C.sub.8) alkyl, branched alkyl, alkenyl or alkynyl, and

R.sup.33 is an anticancer agent.

Preferably, the anticancer agent is selected from the group consisting of mitoxanthrone, doxorubicin, idarubicin, epirubicin, daunorubicin, mitomycin, methotrexate, CPT-11, SN-38, camptothecin, topotecan, 9-nitrocamptothecin, and 9-aminocamptothecin, and is covalently linked via an ester, amido or carbamate linkage to the --SH, OH or amino group of X.sup.33.

Compounds of Formula I and Formula II can be prepared according to procedures known to the skilled artisan (See, for example, Marx et al., 1988, Journal of Medicinal Chemistry 31:858-863; Meyer et al., 1991, Journal of Medicinal Chemistry 34:1377-1383; Morris-Natschke et al., 1986, Journal of Medicinal Chemistry 29:2114-2117; Piantadosi et al., 1991, Journal of Medicinal Chemistry 34:1408-1414; and Surles et al., 1993, Lipids 28:55-57).

An example of such a procedure is illustrated in FIGS. 2-4. The structures presented in the reaction schemes of FIGS. 2-4 are representative and not meant to limit the compounds of the invention. Modifications to the reactions in FIGS. 2-4 using different compounds are apparent to the skilled artisan. Briefly, a compound of Formula I or Formula II is prepared by reacting a lipid backbone moiety, prepared as shown in FIG. 2, for example, with an AZT-malonic acid (AZT-MA) moiety, for example, prepared as shown in FIG. 3. Synthetic methods for the preparation of a lipid backbone as described in FIG. 2 are known in the art. For example, the synthesis method for preparing a lipid backbone for a thiophosphocholine is described in Morris-Natschke et al., 1986, Journal of Medicinal Chemistry 29(10):2114-2117, except one would substitute the benzyloxy alkyl bromide for the C-2 alkyl chain described in the reference. To prepare an amidophosphocholine, for example, one would follow the synthesis method described in Kucera et al., 1998, Antiviral Chemistry and Chemotherapy, 9:157-165. However, one would substitute C.sub.6H.sub.5CH.sub.2O(CH.sub.2).sub.8Br (8-benzyloxyoctyl bromide) for CH.sub.3(CH.sub.2).sub.7Br (octyl bromide) described in the reference. To prepare a lipid backbone for various other phosphocholine syntheses, one would follow the synthesis procedures described in Meyer et al., 1991, Journal of Medicinal Chemistry 34(4):1377-1383 and Morris-Natschke et al., 1993, Journal of Medicinal Chemistry 36(14) 2018-2023. Again, one would substitute the benzyloxy alkyl bromide for the C-2 alkyl chain described in the references.

A preferred compound of the invention (e.g. INK-20, a PC lipid-AZT conjugate) can be prepared as described in the Examples herein and depicted in FIG. 4 by reacting the lipid backbone moiety generated as shown in FIG. 2 with the AZT-malonic acid (AZT-MA) moiety generated as shown in FIG. 3. The AZT-MA moiety can be prepared, for example, as described in the Examples herein. FIGS. 2-4 together illustrate the reaction scheme for preparation of certain preferred compounds of the invention, wherein AZT is linked to a PC lipid at the terminal functionality of position-2 on a modified thioglycerol backbone. The intermediate thiophosphocholine in FIG. 4 has a terminal hydroxyl group on the position-2 side chain which is used as a site for conjugating AZT to the PC lipid. An antiviral agent such as, for example, AZT or a protease inhibitor can be linked to the PC lipid via a malonic ester. This synthetic pathway allows manipulation of the rate of esterase-catalyzed hydrolysis of the AZT moiety in the cell by incorporation of substituted malonic linking groups. While not wishing to be bound by any particular theory, it is expected that, as with accepted prodrug strategy, the ester bond linking the PC lipid with the AZT moiety is cleaved by the action of esterase-like activity in vivo, thereby releasing both active antiviral agents (e.g. nucleoside or protease inhibitor and PC lipid) inside treated cells (See Chapter 47, "Chemotherapy of Microbial Agents," pp. 1130 and 1141, respectively, in Goodman and Gilman, 1996, "The Pharmacological Basis of Therapeutics", Ninth Ed.).

The following compounds are illustrative of compounds having structures according to one or both of Formula I and Formula II, as described above. These compounds can be prepared by the procedures described herein, or by variations thereof which are apparent to those skilled in the art in view of the instant disclosure. Exemplary compounds include INK-20, INK-25 and INK-26. The chemical structures of these compounds are depicted in Table 1 herein.

Compounds of Formulae III, IV and V can be prepared according to procedures known to the skilled artisan. An example of such a procedure is described, for example, in Piantadosi et al., 1991, J. Med. Chem. 34:1408-1414. The synthesis of a compound of Formula V involves direct esterification of the lipid portion with the therapeutic agent rather than conjugation of the therapeutic agent with the phosphatidic acid portion of Formulae III and IV.

Exemplary compounds having structures according to Formulae III, IV and V, are described herein in the Figures. These compounds can be prepared by the procedures described herein, or by variations thereof which are apparent to those skilled in the art in view of the instant disclosure. Structural formulae of exemplary compounds are shown in FIG. 7 (Formula III), FIG. 8 (Formula IV), and FIG. 9 (Formula V).

The compounds of the present invention can be prepared in the form of a pharmaceutically acceptable salt or a non-pharmaceutically acceptable salt. Non-pharmaceutically acceptable salts are useful, for example, as intermediates for preparation of a pharmaceutically acceptable salt. When the compounds are sufficiently basic or acidic to form stable non-toxic acid or base salts, the compounds may be prepared as a pharmaceutically acceptable salt. Pharmaceutically acceptable salts are salts that retain the desired biological activity of the parent compound and do not impart undesirable toxicological effects.

Examples of such salts are acid addition salts formed with inorganic acids, for example, hydrochloric, hydrobromic, sulfuric, phosphoric, and nitric acids and the like; salts formed with organic acids such as acetic, oxalic, tartaric, succinic, maleic, fumaric, gluconic, citric, malic, methanesulfonic, p-toluenesulfonic, napthalenesulfonic, and polygalacturonic acids, and the like; salts formed from elemental anions such as chlorine, bromine, and iodine; salts formed from metal hydroxides, for example, sodium hydroxide, potassium hydroxide, calcium hydroxide, lithium hydroxide, and magnesium hydroxide; salts formed from metal carbonates, for example, sodium carbonate, potassium carbonate, calcium carbonate, and magnesium carbonate; salts formed from metal bicarbonates, for example, sodium bicarbonate and potassium bicarbonate; salts formed from metal sulfates, for example, sodium sufate and potassium sulfate; and salts formed from metal nitrates, for example, sodium nitrate and potassium nitrate.

Pharmaceutically acceptable and non-pharmaceutically acceptable salts may be prepared using procedures well known in the art, for example by reacting a sufficiently basic compound such as an amine with a suitable acid comprising a physiologically acceptable anion. Alkali metal (for example, sodium, potassium, or lithium) or alkaline earth metal (for example, calcium) salts of carboxylic acids can also be made.

The compounds of Formulae I-V can be formulated as pharmaceutical compositions and administered to a mammal, such as a human patient by a chosen route of administration. Pharmaceutical compositions that are useful in the methods of the invention can be prepared, packaged, or sold in a variety of formulations which can be suitable for one or more routes of administration such as, for example, oral, intravenous, intramuscular, topical, subcutaneous, rectal, vaginal, parenteral, pulmonary, intranasal, buccal, ophthalmic, or another route of administration. Other contemplated formulations include projected nanoparticles, liposomal preparations, resealed erythrocytes containing the active ingredient, and immunologically-based formulations.

Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions which are suitable for ethical administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and perform such modification with merely ordinary, if any, experimentation. Subjects to which administration of the pharmaceutical compositions of the invention is contemplated include, but are not limited to, humans and other primates and mammals including commercially relevant mammals such as cattle, pigs, horses, sheep, cats, and dogs.

Thus, the present compounds can be systemically administered (e.g. orally) in combination with a pharmaceutically acceptable vehicle such as an inert diluent or an assimilable edible carrier. They can be enclosed in hard or soft shell gelatin capsules, compressed into tablets, or incorporated directly into the food of the patient's diet. For oral therapeutic administration, the active compound can be combined with one or more excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like. Such compositions and preparations should contain at least 0.1% (w/w) of active compound. The percentage of the compositions and preparations can, of course, be varied, for example from about 0.1% to nearly 100% of the weight of a given unit dosage form. The amount of active compound in such therapeutically useful compositions is such that an effective dosage level will be obtained upon administration.

The tablets, troches, pills, capsules, and the like can also contain one or more of the following: binders such as gum tragacanth, acacia, corn starch, or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid, and the like; a lubricant such as magnesium stearate; a sweetening agent such as sucrose, fructose, lactose, or aspartame; and a flavoring agent such as peppermint, oil of wintergreen, or cherry flavoring. When the unit dosage form is a capsule, it can contain, in addition to materials of the above type, a liquid carrier, such as a vegetable oil or a polyethylene glycol. Various other materials can be present as coatings or to otherwise modify the physical form of the solid unit dosage form. For instance, tablets, pills, or capsules can be coated with gelatin, wax, shellac, sugar, and the like. A syrup or elixir can contain the active compound, sucrose or fructose as a sweetening agent, methyl and propylparabens as preservatives, a dye, and flavoring such as cherry or orange flavor. Of course, any material used in preparing a unit dosage form should be pharmaceutically acceptable and substantially non-toxic in the amounts employed. In addition, the active compound can be incorporated into sustained-release preparations and devices.

The active compound can be administered intravenously or intraperitoneally by infusion or injection. Solutions of the active compound or its salts can be prepared in water, optionally mixed with a non-toxic surfactant. Dispersions can be prepared in glycerol, liquid polyethylene glycols, triacetin, mixtures thereof, and in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent growth of microorganisms.

Pharmaceutical dosage forms suitable for injection or infusion can include sterile aqueous solutions or dispersions or sterile powders comprising the active ingredient which are adapted for the extemporaneous preparation of sterile injectable or infusible solutions or dispersions, optionally encapsulated in liposomes. The ultimate dosage form should be sterile, fluid, and stable under conditions of manufacture and storage. The liquid carrier or vehicle can be a solvent or liquid dispersion medium comprising, for example, water, ethanol, a polyol (for example, glycerol, propylene glycol, liquid polyethylene glycols, and the like), vegetable oils, nontoxic glyceryl esters, and suitable mixtures thereof. The proper fluidity can be maintained, for example, by formation of liposomes, by the maintenance of the required particle size (in the case of dispersions) or by use of one or more surfactants. Microbial growth can be prevented using various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, buffers, or sodium chloride. Prolonged absorption of the injectable compositions can be achieved using agents which delay absorption, for example, aluminum monostearate and gelatin.

Sterile injectable solutions are prepared by incorporating the active compound in the required amount in an appropriate solvent, optionally with one or more of the other ingredients enumerated above, followed by filter sterilization. In the case of sterile powders for preparation of sterile injectable solutions, preferred methods of preparation include vacuum drying and the freeze drying techniques, which yield a powder of the active ingredient and any additional desired ingredient present in the previously sterile-filtered solution(s).

A pharmaceutical composition of the invention may be prepared, packaged, or sold in a formulation suitable for rectal administration. Such a composition may be in the form of, for example, a suppository, a retention enema preparation, and a solution for rectal or colonic irrigation.

Suppository formulations may be made by combining the active ingredient with a non-irritating pharmaceutically acceptable excipient which is solid at ordinary room temperature (i.e. about 20.degree. C.) and which is liquid at the rectal temperature of the subject (i.e. about 37.degree. C. in a healthy human). Suitable pharmaceutically acceptable excipients include, but are not limited to, cocoa butter, polyethylene glycols, and various glycerides. Suppository formulations may further comprise various additional ingredients including, but not limited to, antioxidants and preservatives.

Retention enema preparations or solutions for rectal or colonic irrigation may be made by combining the active ingredient with a pharmaceutically acceptable liquid carrier. As is well known in the art, enema preparations may be administered using, and may be packaged within, a delivery device adapted to the rectal anatomy of the subject. Enema preparations may further comprise various additional ingredients including, but not limited to, antioxidants and preservatives.

A pharmaceutical composition of the invention may be prepared, packaged, or sold in a formulation suitable for vaginal administration. Such a composition may be in the form of, for example, a suppository, an impregnated or coated vaginally-insertable material such as a tampon, a douche preparation, or a solution for vaginal irrigation.

Methods for impregnating or coating a material with a chemical composition are known in the art, and include, but are not limited to methods of depositing or binding a chemical composition onto a surface, methods of incorporating a chemical composition into the structure of a material during the synthesis of the material (i.e. such as with a physiologically degradable material), and methods of absorbing an aqueous or oily solution or suspension into an absorbent material, with or without subsequent drying.

Douche preparations or solutions for vaginal irrigation may be made by combining the active ingredient with a pharmaceutically acceptable liquid carrier. As is well known in the art, douche preparations may be administered using, and may be packaged within, a delivery device adapted to the vaginal anatomy of the subject. Douche preparations may further comprise various additional ingredients including, but not limited to, antioxidants, antibiotics, antifungal agents, and preservatives.

A pharmaceutical composition of the invention may be prepared, packaged, or sold in a formulation suitable for pulmonary administration via the buccal cavity. Such a formulation may comprise dry particles which comprise the active ingredient and which have a diameter in the range from about 0.5 to about 7 nanometers, and preferably from about 1 to about 6 nanometers. Such compositions are conveniently in the form of dry powders for administration using a device comprising a dry powder reservoir to which a stream of propellant may be directed to disperse the powder or using a self-propelling solvent/powder-dispensing container such as a device comprising the active ingredient dissolved or suspended in a low-boiling propellant in a sealed container. Preferably, such powders comprise particles wherein at least 98% of the particles by weight have a diameter greater than 0.5 nanometers and at least 95% of the particles by number have a diameter less than 7 nanometers. More preferably, at least 95% of the particles by weight have a diameter greater than 1 nanometer and at least 90% of the particles by number have a diameter less than 6 nanometers. Dry powder compositions preferably include a solid fine powder diluent such as sugar and are conveniently provided in a unit dose form.

Low boiling propellants generally include liquid propellants having a boiling point of below 65.degree. F. at atmospheric pressure. Generally the propellant may constitute 50 to 99.9% (w/w) of the composition, and the active ingredient may constitute 0.1 to 20% (w/w) of the composition. The propellant may further comprise additional ingredients such as a liquid non-ionic or solid anionic surfactant or a solid diluent (preferably having a particle size of the same order as particles comprising the active ingredient).

Pharmaceutical compositions of the invention formulated for pulmonary delivery may also provide the active ingredient in the form of droplets of a solution or suspension. Such formulations may be prepared, packaged, or sold as aqueous or dilute alcoholic solutions or suspensions, optionally sterile, comprising the active ingredient, and may conveniently be administered using any nebulization or atomization device. Such formulations may further comprise one or more additional ingredients including, but not limited to, a flavoring agent such as saccharin sodium, a volatile oil, a buffering agent, a surface active agent, or a preservative such as methylhydroxybenzoate. The droplets provided by this route of administration preferably have an average diameter in the range from about 0.1 to about 200 nanometers.

The formulations described herein as being useful for pulmonary delivery are also useful for intranasal delivery of a pharmaceutical composition of the invention.

Another formulation suitable for intranasal administration is a coarse powder comprising the active ingredient and having an average particle from about 0.2 to 500 micrometers. Such a formulation is administered in the manner in which snuff is taken i.e. by rapid inhalation through the nasal passage from a container of the powder held close to the nares.

Formulations suitable for nasal administration may, for example, comprise from about as little as 0.1% (w/w) and as much as 100% (w/w) of the active ingredient, and may further comprise one or more of the additional ingredients described herein.

A pharmaceutical composition of the invention may be prepared, packaged, or sold in a formulation suitable for ophthalmic administration. Such formulations may, for example, be in the form of eye drops including, for example, a 0.1-1.0% (w/w) solution or suspension of the active ingredient in an aqueous or oily liquid carrier. Such drops may further comprise buffering agents, salts, or one or more other of the additional ingredients described herein. Other ophthalmalmically-administrable formulations which are useful include those which comprise the active ingredient in microcrystalline form or in a liposomal preparation.

For topical administration, the present compounds can be applied in pure form, i.e., as a liquid. However, it will generally be desirable to administer the compounds to the skin as compositions or formulations, in combination with a dermatologically acceptable carrier, which may be a solid or a liquid.

Useful solid carriers include finely divided solids such as talc, clay, microcrystalline cellulose, silica, alumina, and the like. Useful liquid carriers include water, alcohols, glycols, and blends of two or more of these, in which the present compounds can be dissolved or dispersed at effective levels, optionally with the aid of non-toxic surfactants. Adjuvants such as fragrances and additional antimicrobial agents can be added to optimize properties for a given use. The resulting liquid compositions can be applied using absorbent pads, used to impregnate bandages or other dressings, or sprayed onto the affected area using pump-type or aerosol sprayers.

Thickeners such as synthetic polymers, fatty acids, fatty acid salts and esters, fatty alcohols, modified celluloses, or modified mineral materials can also be employed with liquid carriers to form spreadable pastes, gels, ointments, soaps, and the like, for application directly to the skin of the user.

Examples of useful dermatological compositions which can be used to deliver the compounds of the invention to the skin are disclosed in Jacquet et al. (U.S. Pat. No. 4,608,392), Geria (U.S. Pat. No. 4,992,478), Smith et al. (U.S. Pat. No. 4,559,157) and Wortzman (U.S. Pat. No. 4,820,508).

Accordingly, the invention includes pharmaceutical compositions comprising one or more compounds of Formula I, Formula II, Formula III, Formula IV or Formula V, or any combination thereof, or a pharmaceutically acceptable salt thereof, in combination with a pharmaceutically acceptable carrier.

In a preferred embodiment, the pharmaceutical composition is adapted for oral, topical, or parenteral administration to a mammal such as a human, and comprises one or more compounds of Formula I or Formula II, or any combination thereof, or a pharmaceutically acceptable salt thereof, in an amount effective to treat a virus infection in a mammal or in a cell, particularly wherein the virus is HIV, hepatitis virus, or herpes simplex virus.

As used herein, "treatmen