MIXTURES OF AT LEAST TWO REVERSE TRANSCRIPTASE INHIBITORS AND HYDROXYCARBAMIDE, IN PARTICULAR FOR INHIBITING RETROVIRAL SPREAD

Abstract: Composition as a medicament, in particular for use in inhibiting the spread of a retrovirus, comprising at least two different reverse transcriptase inhibitors and hydroxycarbamide.

Description: Mixtures of at least two reverse transcriptase inhibitors and hydroxycarbamide, in particular for inhibiting retroviral spread FIELD OF THE INVENTION The present invention relates to a combination or an association of at least two reverse transcriptase inhibitors and hydroxycarbamide wherein the combination or association is useful in inhibiting retroviral spread. BACKGROUND OF THE INVENTION The expression "Acquired Immuno-Deficiency Syndrome" (AIDS) was first used in 1981 to describe a state of cellular immune deficiency in homosexuals, characterized by the appearance of opportunistic infections and Kaposi's Sarcoma evolving very aggressively (CDC (Center for Disease Control) , MM R, 30 : 305-308. DC (1981)). In 1983 a retrovirus since called HIV (Human Immunodeficiency Virus type 1) was isolated among AIDS patients (Barre-Sinoussi F. et al Science, 220 : 868-870 (1983)). Over the past several years, researchers and clinicians have gained considerable experience in studying and caring for individuals infected with HIV throughout the often prolonged course of HIV disease and AIDS. On the basis of this experience, it has become clear that the pathogenic mechanisms underlying HIV infection and disease are not unidimensional, but rather are extremely complex (Fauci AS., Science, 239, 617 (1988)). Any attempt to design a comprehensive therapeutic strategy for HIV disease must take this fact into account. (Fauci, 1993, Science, 262:1011- 1018) . After entry of the HIV virus into cells and uncoating of the HIV particle, reverse transcription of the viral RNA genome into DNA replicas occurs. Among several forms of unintegrated viral DNA (now containing the long repeats [LTRs], at both the 5' and the 3* ends) only the two-LTR linear forms can integrate into the host genome. Such a process appears strictly dependent upon cell activation/replication of T lymphocytes, although "resting" T cells are clearly susceptible to HIV infection. (Zack J.A. et al, Cell; 61, 213-222, (1990)). Furthermore, part of the reverse transcription process also can occur in unactivated T cells, a process that results in the accumulation of incomplete DNA molecules, which may persist for several hours and remain capable of being integrated into the host genome if the cell undergoes sufficient activation (Zack J.A. et al, Cell ; 61, 213-222, (1990)). Therefore, infected "resting" CD4 + T lymphocytes can be considered a transient viral reservoir in infected individuals (Bokrinsky M.I. et al; Science, 254, 423-427, (1991)). These observations are of particular importance in anatomic compartments such as the peripheral blood and lymphoid organs, where the CD4 + T cell subset represents the predominant infected cell type (Schmittman S M. et al, science, 245, 305-308, (1989)); (Fox CH. et al J . Infect . Diε . , 164, 1051-1057, (1991)). The above discussion provides a sound scientific basis for blocking the initial burst of virus replication and dissemination as well as the persistent replication throughout the course of disease by treating HIV-infected individuals with anti-retroviral agents from the earliest time that HIV infection is recognized through the entire course of infection. Unfortunately, currently available agents used alone are only partially effective in suppressing virus replication and spread, and this effect is transient (Hirsch MS, et al, New Engl . J . Med . 328 1686, (1993)). A number of reverse transcriptase inhibitors have been found to be useful for the treatment or prophylaxis of retroviral infections and especially HIV and AIDS. Examples of such materials include: 3 1 - azido-2 • , 3 • -dideoxythymidine or azidothymidine (AZT) ; 2 ' , 3 '-dideoxycytosine (ddC) ; 2 ' , 3 ' -dideoxy-adenosine (ddA) ; 2 ' , 3 ' -dideoxy-guanosine (ddG) ; and 2', 3'- dideoxy-inosine (ddl) ; 2 • , 3 • -dideoxy-thymidine (ddT) ; 2 ' ,3 '-didehydro-3 ' -deoxythy idine (d4T) ; 2'-deoxy-3'- thiocytidine (3TC) . See European patent application 0206497 and published PCT application number WO 87/01284. Non nucleoside reverse transcriptase inhibitors (in abreviation NNRTI) have been also proposed, such as nevirapine (Boeringher Ingelheim) , pyridinone (Merck) , piperazine or atevidine, imidazo- benzodiazepine (Tibo) , delavirdine (Upjohn) , loviridine. Hydroxycarbamide (HC) was initially synthesized over 120 years ago and has been found to demonstrate activity against a broad spectrum of tumors. (Donehower, Seminars in Oncology, Vol. 19, No. 3, Suppl. 9 (June) 1992: pp 11-19). Additionally, hydroxycarbamide has been used as a virucide. In published PCT application number WO 93/09718, hydroxycarbamide is taught to be useful in a hydrogel polymer coating of a blood bag in order to inhibit viral and HIV infectivity. Gao et al (PNAS, USA, Vol. 900 pp. 8925-8928, October 1993) disclose that hydroxyurea (hydroxycarbamide) treatment of peripheral blood lymphocytes (PBLs) decreases dNTP levels and the DNA synthesis rate to levels comparable to quiescent PBLs. The article alleges a possible use of hydroxyurea in AIDS therapy. However, there still remains a need for more effective treatments for the suppression of retroviruses and, in particular, the prevention and/or inhibition of HIV and viral spread. By viral spread, it is intended to include the inhibition of viral replication, and also may include the ability of inhibiting the virus to infect a further host cells. Objectives of the present invention in the search for new antiretroviral agents include: 1) the identification of treatments with less toxicity and antiviral activity greater than a combination of two reverse-transcriptase inhibitors alone, 2) the development of drug combinations or associations which potentiate the synergistic (if any) activity of at least two reverse-transcriptase inhibitors co-administered in a same treatment. BRIEF SUMMARY OF THE INVENTION The present invention relates to a combination or association of at least two different reverse transcriptase inhibitors and hydroxycarbamide wherein the combination or association is capable of preventing and/or inhibiting the spread of retroviruses including HIV. More specifically, the present invention relates to a method of preventing and/or inhibiting the spread of retroviruses, including HIV (HIV-1 and HIV-2) , HTLV-1, HTLV-2 , SIV, by exposing a cell population, including cells infected by a retrovirus such as, for example, HIV, to a combination or association of at least two reverse transcriptase inhibitors and hydroxycarbamide. Additionally, the present invention encompasses the treatment of HIV-infected and AIDS patients with a combination or association of at least two different reverse transcriptase inhibitors and hydroxycarbamide in order to prevent and/or inhibit the spread of HIV in these patients. In a preferred embodiment of the present invention, each reverse transcriptase inhibitor is a dideoxynucleoside, in particular at least one of AZT, ddC, ddl, ddA, ddG, ddT, and for example ddl. In another preferred embodiment of the invention, each reverse transcriptase inhibitor is a didehydrodeoxynucleoside, in particular d4T. In another preferred embodiment of the invention, each reverse transcriptase inhibitor is a non nucleoside reverse transcriptase inhibitor, in particular at least one of hevirapine, pyridinone or atevidine, imidazo-benzodiazepine, delavirdine and loviridine. In particular and in the preferred combination or association of the present invention, it has been found that the co-administration of hydroxycarbamide (HC) , 2 • ,3 '-dideoxy-inosine (ddl), and didehydrodeoxy- thymidine (d4T) is especially effective in preventing and/or inhibiting HIV spread. The preferred embodiment of the invention encompasses a composition including a pharmaceutical composition comprising a combination of ddl, d4T and HC. The pharmaceutical composition can optionally contain a pharmaceutically acceptable carrier and/or excipient and/or vehicle. The preferred method of the instant invention comprises preventing and/or inhibiting retroviral or HIV spread by treating a cell population, including cells infected with HIV, with a combination or association of ddl, d4T and HC. Additionally, the preferred method comprises treating an HIV infected or AIDS patient with an association or combination of ddl, d4T and HC so as to prevent and/or inhibit HIV spread in the patient. DETAILED DESCRIPTION OF THE INVENTION The following example of a specific embodiment of the present invention is offered for illustrative purposes only and is not limiting with respect to the scope of the disclosure or claim coverage. Clinical evaluation of the use of hydroxycarbamide, ddl and d4T in HIV infection We evaluated 6 months' treatment of HIV-infected patients with the combination therapy hydroxycarbamide (hydroxyurea) , didanosine (ddl) and didehydrodeoxy- thymidine (d4T) . The combination or association was evaluated in asymptomatic patients. The objective was to evaluate the tolerance and the anti-viral response. A significant decline in viral load was observed after 6 months' treatment with this combination therapy. 27 volunteers, ddl- and d4T-naive individuals classified by CDC at entry as Al or A2 (without symptoms and CD4 cell count >200 cells/μl) , seropositive for HIV-1 confirmed by ELISA and Western Blot, were treated for 6 months, ddl was given at the dose of 200 mg twice daily, hydroxycarbamide at the dose of 15 mg/kg per day in two divided doses and d4T at the dose of 40 mg twice daily. Plasma viral load was evaluated for HIV-RNA by PCR using the ultrasensitive PCR Amplicor HIV Monitor (Roche Diagnostic Systems, Branchburg, NJ, USA) . Treatment was well tolerated; no patient interrupted treatment due to side-effects. A few minor transient symptoms such as digestive disorders were noted in some patients, thrombopenia in one patient, neuropathy in one patient and an increase in mean corpuscular volume was seen in all patients. After 180 days, leucopenia was observed in three patients. No changes is haemaglobin, amylases, lipases, or transaminases were recorded. Average plasma HIV-RNA at baseline was 172 113 copies/ml (see Table) . All patients showed a very significant drop in plasma viral load and in 23 out of 27 patients evaluated at 6 months, viraemia became undetectable. 4 out of 27 patients showed an average reduction in plasma viremia of 3.17 log (from 464 312 to 318 copies/ml) . After 180 days of therapy, average CD4+ lymphocytes relative to total lymphocytes remained unchanged (28%) . The subject of the present invention is also a new composition or association for the treatment of a cell population in the presence of a retrovirus. Additionally, the invention includes a pharmaceutical composition intended, in particular, for the treatment and prevention of retroviral infections, especially those linked to HIV and AIDS wherein the composition contains hydroxycarbamide (HC) and at least two different reverse transcriptase inhibitors, in particular at least two different dideoxynucleosides and hydroxycarbamide, most preferably a combination of dideoxyinosine, didehydrodeoxy-thymidine, and hydroxycarbamide as active principle, in a pharmaceutically acceptable vehicle. The composition of the invention can also contain inert or pharmacodynamically active additives, carriers and/or excipients. The pharmaceutical composition of the invention can take the form of a lyophilized powder of the active combination, to be dissolved immediately before use in a physiological solution for the purpose of injection. The medicament can then be administered parenterally, for example intravenously, intraperitoneally, in the cerebrospinal fluid, and the like. For injection, the active principle is dissolved in a physiological solution until the desired concentration for administration is obtained. TABLE Vlrologtcal data at baseline and after 6 months of treatment of HIV-lnf β ot β d Individuals with a combination of hydroxycarbamiπe, dldanαsinβ and d4T. FCR RNA RNA cαplθs/ml plasma Patient Day 0 Day 180 1 non dot non det 2 33S7S 1 non det 3 5737 non det 4 43680 πoπ dot 5 1 820 πoπ det 6 10141 1 " non det 7 88S28 <2Q0 8 S 13S non det 9 160806 non det 1 0 s 17 πoπ det 1 1 1 64S946 625 1 2 98539 non dot 3 33394 non det 1 4 216S42 non det 5 1 17540 non det 1 6 1 60 non det 1 7 76395 non det 1 8 1 987 non det 1 9 52407 non det 20 23093 256 21 1 382 non det 22 721 2 non det 23 1 18986 nan det 24 822913 non det 25 99682 21 8 26 non det non det 27 241862 non det on dot: non detectable (sensitivity threshold = 200 RNA copies/ml) The pharmaceutical composition according to the invention can also take a form which is suitable for oral administration. For example, suitable forms are tablets, hard gelatin capsules, dragees, powders and granules. The formation of such oral forms is well- known to those skilled in the art. Any of the known formulations are useful in preparing the instant oral pharmaceutical compositions. As regards the dosage of the medicament according to the invention, it will be clear to the artisan that the doses to be administered are variable according to the treatment period, and frequency of administration, the host and the nature and severity of the disease and that the dosages can be easily determined without any undue amount of experimentation. The compositions of the invention are administered in substantially non-toxic dosage concentrations sufficient to ensure the release of a sufficient dosage unit of the present combination or association into the patient to provide the desired inhibition of the spread of the retrovirus. The actual dosage administered will be determined by physical and physiological factors such as age, body weight, severity of condition, and/or clinical history of the patient. With these considerations in mind, the dosage of the instant combination or association for a particular subject can be readily determined by the physician. It might be noted that in extreme cases a dosage approaching the toxic level may be the acceptable treatment protocol. For example, in the treatment of HIV-infected and AIDS patients, the composition can comprise from about 1 to 66 mg/Kg/day of ddl, from 0.1 to 2 mg/Kg/day of d4T, and from about greater than 1 mg/Kg/day to about 20 mg/Kg/day of HC. The invention also covers the use of hydroxycarbamide (HC) , didehydrodeoxythymidine and dideoxyinosine in combination with other medicinal compositions intended for the treatment of retroviral infections and tumors, in particular HIV protease inhibitors, such as for example indinavir, ritonavir, saquinavir, nelfinavir, palinavir, VX-478 (Vertex) , and AG1343 (Agouron) , immunostimulants and immunomodulators such as for example cytokines, including IL-2, IL-12 and interferon molecules can be used in combination with the present invention. By association of compounds is meant a co- administration of these compounds, whether they are administered in a same composition comprising said compounds, or they are separately administered, so as to be available in the blood plasma almost at a same time. All of the references cited hereinabove are expressly incorporated herein, in toto, by reference thereto. The invention has been described with reference to specific and preferred embodiments. It will be recognized by those skilled in the art that numerous changes and substitutions may be made without departing from the spirit and scope of the invention.

Claims: We Claim : 1. A composition as a medicament, in particular for use in inhibiting the spread of a retrovirus, comprising at least two different reverse transcriptase inhibitors and hydroxycarbamide. 2. A composition according to claim 1, wherein each reverse transcriptase inhibitor is a dideoxynucleoside, in particular at least one of AZT, ddC, ddl, ddA, ddG, ddT, and for example ddl. 3. A composition according to claim 1, wherein each reverse transcriptase inhibitor is a didehydrodeoxynucleoside, in particular d4T. 4. A composition according to claim 1, wherein each reverse transcriptase inhibitor is a non nucleoside reverse transcriptase inhibitor, in particular at least one of hevirapine, pyridinone or atevidine, imidazo-benzodiazepine, delavirdine and loviridine. 5. A composition according to claim 1, wherein two reverse transcriptase inhibitors are respectively ddl and d4T. 6. A composition according to any one of claims 1 to 5, wherein the retrovirus is HIV. 7. A composition according to any one of claims 1 to 5, wherein the retrovirus is HIV-1. 8. A method of inhibiting spread of a retrovirus in an infected human cell population, comprising administering to the human cell population an effective spreading inhibiting amount of a composition according to any one of claims 1 to 5. 9. Use in assocation of at least two different reverse transcriptase inhibitors and hydroxycarbamide for inhibiting the replication of a retrovirus. 10. Use according to claim 9, wherein each reverse transcriptase inhibitor is a dideoxynucleoside, in particular at least one of AZT, ddC, ddl, ddA, ddG, ddT, and for example ddl. 11. Use according to claim 9, wherein each reverse transcriptase inhibitor is a didehydrodeoxynucleoside, in particular d4T. 12. Use according to claim 9, wherein each reverse transcriptase inhibitor is a non nucleoside reverse transcriptase inhibitor, in particular at least one of hevirapine, pyridinone or atevidine, imidazo-benzodiazepine, delavirdine and loviridine. 13. Use according to claim 9 , wherein two reverse transcriptase inhibitors are respectively ddl and d4T. 14. An association of at least two different reverse transcriptase inhibitors and hydroxycarbamide, for use in the treatment or prophylaxis of a human disease in relation to the spread or replication of a retrovirus, in particular HIV, in a cell population. 15. An association according to claim 14, wherein each reverse transcriptase inhibitor is a dideoxynucleoside, in particular at least one of AZT, ddC, ddl, ddA, ddG, ddT, and for example ddl. 16. An association according to claim 14, wherein each reverse transcriptase inhibitor is a didehydrodeoxynucleoside, in particular d4T. 17. An association according to claim 14, wherein each reverse transcriptase inhibitor is a non nucleoside reverse transcriptase inhibitor, in particular at least one of hevirapine, pyridinone or atevidine, imidazo-benzodiazepine, delavirdine and loviridine. 18. An association according to claim 14, wherein two reverse transcriptase inhibitors are respectively ddl and d4T.

Inventors: VILA JORGE, VILA, JORGE
Applicants: AGUETTANT LAB, VILA JORGE, COMPAGNIE DE DEVELOPPEMENT AGUETTANT, VILA, JORGE
Application year: 1997
Main classification: A61K31
IPC codes: A61K31, A61K45, A61P31, A61P37