The present invention relates to a drug combination capable of conferring therapeutic benefits in the treatment of both AIDS and malaria. In particular, it relates to a drug combination including at least one quinolinic antimalarial compound such as chloroquine or hydroxychloroquine, and at least one inhibitor of the Human Immunodeficiency Virus (HIV) protease enzyme. This drug combination is capable of inhibiting the replication of both HIV and Plasmodium sp. It also relates to the direct antimalarial effects of the HIV PIs.

Description of the Invention

In addition, the following Sequence Listing material is contained on a disc, and the files are hereby incorporated-by-reference into the present application in their entirety: Savarino Sequence Listing, Patent In Document, 2 KB, created Sep. 21, 2004; Savarino Sequence Listing, Microsoft Word Document, 41 KB, created Sep. 21, 2004; Savarino Sequence Listing, Text Document, 6 KB, created Sep. 21, 2004. Print copies of the Sequence Listings are included as an Appendix to the current Application, and are also incorporated-by-reference herein in their entirety.

FIELD OF THE INVENTION

The present invention relates to a drug combination capable of conferring therapeutic benefits in the treatment of both acquired immunodeficiency syndrome (AIDS) and malaria. In particular, it relates to a drug combination comprising chloroquine or hydroxychloroquine plus an inhibitor of the HIV protease capable of inhibiting the replication of both the human immunodeficiency viruses (HIV) and Plasmodium sp. The present invention also relates to the direct antimalarial effects of the HIV protease inhibitors.

BACKGROUND OF THE INVENTION

Acquired immunodeficiency syndrome (AIDS) and malaria are among the most devastating infectious diseases that have ever affected mankind, causing approximately five million deaths per year in the world. The effects of these diseases are most pronounced in underdeveloped countries in that the diseases are accompanied by financial and living conditions that are already miserable to start with. Several resource-poor countries cannot afford effective therapies that might allow the prevention of many deaths. The difficulties per se in treating both AIDS and malaria, caused in part by the drug-resistance of both their etiological agents, i.e., the human immunodeficiency viruses (HIV) and protozoa belonging to the genus Plasmodium, become exaggerated when the pharmaceutical weapons are extremely limited. In several resource-poor countries with high rates of HIV seroprevalence, the use of highly active antiretroviral therapy (HAART) has encountered major obstacles due to its high costs and the complexities of its prescription. Recently, due to humanitarian considerations, anti-HIV drugs have been offered at reduced prices to some of the least developed countries with a high HIV seroprevalence. The problem is, however, still far from being solved. Compared to antiretrovirals, antimalarials have lower costs, which may in any case weigh heavily on the budgets of several poorer countries. Chloroquine (CQ), recommended for a long time by the World Health Organization (WHO) as a first line treatment of malaria, is still the most affordable and widely adopted antimalarial option in Africa; however, the continuous emergence of drug-resistant Plasmodium strains renders its administration ineffective in a large number of areas in Africa, Latin America and South-Eastern Asia.

As most of the areas heavily stricken by AIDS also exhibit endemic malaria (and frequently individuals are co-infected), it would be useful to develop a treatment effective against both diseases.

In this regard, CQ may be particularly useful in that it has been demonstrated to exhibit in-vitro activity against HIV-1 replication and against several AIDS-related opportunistic microorganisms. It also has well-documented, long-term safety when used in immunocompromised individuals, (including those with HIV/AIDS), when dosed for antimalarial prophylaxis and in the treatment of rheumatic diseases. Although no information is available on the in-vivo effects of CQ on viral load, its hydroxy-analog hydroxychloroquine (HCQ) has proven in-vivo anti-HIV-1 activity. The anti-HIV activity of CQ is due to an impairment of the infectivity of virions produced by cells treated with the drug. Although the present invention is not limited to any particular mechanism, it is believed that the mechanism behind this inhibitory effect is inhibition of gp120 glycosylation. This hypothesis is supported by results showing that CQ impairs the formation of the heavily glycosylated epitope 2G12, which is located on the gp120 envelope glycoprotein surface and is fundamental for virus infectivity. These effects show that CQ inhibits viral replication by a mechanism different than those of currently used antiretroviral drugs, and this new mechanism has led to testing CQ in combination with antiretrovirals in clinical trials.

More detailed information on the anti-HIV effects of CQ can be found in the following two articles, which are hereby incorporated in the present patent application in their entirety: Savarino A, Gennero L, Chen H C, Serrano D, Malavasi F, Boelaert J R, Sperber K. Anti-HIV effects of chloroquine: mechanisms of inhibition and spectrum of activity. AIDS Nov. 23, 2001; 15(17):2221-9. Savarino A, Gennero L, Sperber K, Boelaert J R. The anti-HIV-1 activity of chloroquine. J Clin Virol 2001 February;20(3):131-5.

It is known that CQ may exert additive effects when associated with other anti-HIV drugs such as ddI, hydroxyurea, and AZT. The effects of a combinatorial administration of CQ and inhibitors (PIs) of the HIV protease (SEQ ID NO: 1) have however been totally unknown until the present invention. In view of the future large-scale administration of PI-based regimens in malaria-endemic areas, this interaction may provide the following: 1) CQ/HCQ and PIs are the only drugs tested in humans that inhibit HIV replication at a post-integrational stage; 2) the effects of both CQ and PIs result in an impairment of the infectivity of newly produced virions; 3) both CQ and PIs are substrates of and, at varying levels, inhibit important cell surface drug transporters, ie., the P-glycoprotein (P-gp) and the multi-drug resistance-associated proteins (MRP), which belong to the ATP-binding cassette family and modulate the intracellular concentrations of antiretroviral drugs. Of note, recent data indicate that CQ is capable of increasing the level of inhibition of P-gp- and MRP-mediated efflux exerted by PIs in CD4.sup.+ lymphocytes (Savarino et al., JAIDS 2004, in press).

The inhibitory effects of PIs on cell surface drug transporters may make the combination of CQ and a PI particularly useful in treatment of malaria.

Drug transport on the cell surface has been hypothesized to be involved in plasmodial drug-resistance. This theory is supported by several pieces of evidence.

First, a glycoprotein of P. falciparum, namely Pf-MDR, presents a high degree of homology with human P-gp and may be in some ways related to CQ-resistance. Ward S A, Bray P G. Definitive proof for a role of pfmdr 1 in quinoline resistance in Plasmodium falciparum. Drug Resist Updat 2000 April;3(2):80-81

Second, CQ-resistance in vitro is characteristically reverted by verapamil, a known inhibitor of the ATP-binding cassette in human cells. Sidhu A B, Verdier-Pinard D, Fidock D A. Chloroquine resistance in Plasmodium falciparum malaria parasites conferred by pfcrt mutations. Science Oct. 4, 2002;298(5591):210-3

Third, erythrocytes parasited by CQ-resistant P. falciparum strains accumulate more limited intracellular CQ pools than those parasited by CQ-sensitive strains. The capacity of a P. falciparum strain to decrease CQ accumulation within erythrocytes is strictly associated with mutations in a gene (Pf-crt) that encodes the so-called CQ-resistance transport (CRT) protein. The precise mechanisms by which P. falciparum CRT intervenes in these phenomena have not been elucidated yet. Of note, these mutations are present in the vast majority of the CQ-resistant field isolates of P. falciparum coming from different areas of the world and are not present in CQ-sensitive isolates. Sidhu A B, Verdier-Pinard D, Fidock D A. Chloroquine resistance in Plasmodium falciparum malaria parasites conferred by pfcrt mutations. Science Oct. 4, 2002;298(5591):210-3.

It would be beneficial to have compositions and treatments using a combination of CQ and a PI that inhibits both HIV and Plasmodium sp.

SUMMARY OF THE INVENTION

The present invention relates to a drug combination capable of conferring therapeutic benefits in the treatment of both AIDS and malaria. In particular, it relates to a drug combination including an inhibitor of the HIV protease (SEQ ID NO: 1) plus CQ or HCQ or another antimalarial with similar characteristics. This drug combination is capable of inhibiting the replication of both HIV and Plasmodium sp. It also relates to the direct antimalarial effects of the HIV PIs.

The combination claimed in the present patent application unexpectedly demonstrated enhanced capability of conferring a more sustained inhibition of both HIV and Plasmodium sp. than the single agents administered alone, that is, CQ can reinforce the antiretroviral activity of a PI and a PI can strengthen the antimalarial activity of CQ.

The combination of a PI plus CQ may thus be used for the purpose of inhibiting HIV replication, for the purpose of inhibiting Plasmodium sp. growth, or for the purpose of inhibiting both agents.

From a clinical perspective, the combination of a PI plus CQ/HCQ may be capable of treating AIDS and malaria. Therefore, it can be utilized in the treatment of individuals infected with HIV, in individuals affected by or at risk for contracting malaria or in people with HIV/malaria coinfection.

Also, the combination of CQ and PIs can be used to restore the sensitivity of drug-resistant isolates of HIV and P. falciparum to the PIs and to CQ, respectively.

In another embodiment, the present invention relates to the intrinsic antimalarial effects of PIs. The present inventor found that PIs clinically used in the treatment of HIV exert direct antimalarial effects. These direct effects are observable in vitro at therapeutically achievable concentrations (See example III).

Although this invention is not related to any particular mechanism, bioinformatic analysis suggests that the target of PIs may be plasmepsin II (SEQ ID NO: 2), a member of the plasmepsins family, a potential target for new antimalarials.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to a drug combination effective against both of the etiological agents of the two major infectious diseases in the world, ie., AIDS and malaria. In particular, it relates to a drug combination including an inhibitor of the HIV protease (SEQ ID NO: 1) plus an antimalarial such as, for example, CQ or HCQ, capable of inhibiting the replication of both HIV and Plasmodium sp.

The combination claimed in the present patent application may be capable of conferring a more sustained inhibition of both HIV and Plasmodium sp. than the single agents alone, that is, CQ can reinforce the antiretroviral activity of a PI and a PI can strengthen the antimalarial activity of CQ.

As the therapeutic benefit of the above-described combination can be seen on both HIV and Plasmodium sp., the combination may be used for the purpose of inhibiting HIV replication, for the purpose of inhibiting Plasmodium sp. growth, or for the purpose of inhibiting both HIV replication and Plasmodium sp. growth.

From a clinical perspective, the combination of a PI plus an antimalarial such as CQ/HCQ can be used for treatment of AIDS and malaria. Therefore, it could be utilized in the treatment of individuals infected with HIV, in individuals affected by or at risk for contracting malaria or in people with HIV/malaria coinfection. The two agents used in combination may increase the inhibition level of drug-sensitive HIV and Plasmodium strains, but also that the combination PI+CQ restores the sensitivity of drug-resistant isolates of HIV and P. falciparum to the PIs and to CQ, respectively.

Regarding the treatment of HIV, it is important to point out that the effects of CQ in combination with protease inhibitors are synergistic. When administered to acutely infected cells in combination with a PI, CQ decreases the concentration of PIs necessary to produce a certain level of HIV inhibition [see EXAMPLE 1].

In addition, CQ partially restores sensitivity to PIs in PI-resistant strains, as exemplified below [see EXAMPLE 1].

Although the invention is not limited to any particular mechanism, it is believed that the use of a P-gp and MRP blocking agent such as CQ may increase the intracellular concentrations of PIs.

In one embodiment, the present invention allows a treatment strategy whereby the co-administration of an antimalarial, such as CQ/HCQ or another quinolinic agent, to HIV positive individuals allows the effective dose of PIs to be decreased, lessening cost and possibly toxicity. Also, the ability of CQ to overcome resistance to PIs could be of greatest importance for the treatment of drug-experienced HIV positive subjects who have developed multiple resistance to antiretroviral drugs and thus have limited therapeutic options.

Embodiments of the present invention may also be used in the treatment of drug-resistant malaria. Indeed, in several areas of the world with endemic malaria, P. falciparum strains with a multi-drug resistant phenotype are becoming prevalent, and the use of a PI may restore sensitivity to CQ. The availability of one such drug may therefore be expected to save enormous numbers of lives. Cost-related problems in Third World areas where PIs are currently not affordable are expected to be resolved--at least partially--in the near future when PIs will become available on a large scale for the treatment of HIV infection. Considering that AIDS and malaria often co-exist in the same areas, PIs may become more commonly available in those areas than they are today, and therefore it will be possible to postulate a more cost effective use of these drugs in the treatment of malaria. Similarly, HIV-infected individuals living in areas with endemic drug-resistant malaria and treated with the PI+CQ combination may become protected from the occurrence of malarial episodes.

Furthermore, said effects of PIs in combination with a quinolinic agent may contribute to a revival of drugs such as CQ and first generation PIs (RTV, SVQ, IDV), which otherwise would be doomed to be replaced by newer drugs in the near future.

To sum up, the present invention involves administration of a drug combination that may be effective against HIV and malaria. Embodiments of the combination may include: 1) chloroquine (CQ) or hydroxychloroquine (HCQ) or another quinolinic agent such as mefloquine (MQ) and quinine (Q) combined with 2) one or more inhibitors of the HIV protease (PIs). PIs may Include: Indinavir (IDV), ritonavir (RTV), saquinavir (SQV), nelfinavir (NFV), lopinavir (LPV), the combination RTV plus LPV, amprenavir (APV), fosamprenavir (FPV), tipranavir (TPV), atazanavir (ATZ), TMC-114.

The antimalarial and PI combination may be administered with the contemporary co-administration of nucleosidic inhibitors of the HIV reverse transcriptase (NRTIs).

NRTIs may Include:

Zidovudine (AZT or ZDV), lamivudine (3TC), abacavir (ABC), zalcitabine (ddC), didanosine (ddI), stavudine (d4T), tenofovir (TDF), emitricitabine (FTC), amdoxovir (DAPD).

The invention is not limited in this regard, and any appropriate quinolinic agent, PI and/or NRTI may be used.

The antimalarial and PI combination may also be administered with the contemporary co-administration of other antimalarial drugs, or with the contemporary co-administration of antibiotics against concomitant infections, or any drug against co-existing or related diseases.

The present invention also relates to the direct antimalarial effects of the HIV PIs. Not only can PIs revert CQ resistance, but PIs also are endowed with intrinsic antimalarial effects. These direct effects are observable in vitro at therapeutically achievable concentrations (See example II) and in vivo in a murine malaria model (See example III).

The mechanism for the direct antimalarial effects of PIs has not been elucidated yet. Interesting insights however come from the observation that the HIV-1 protease (i.e., the target against which these drugs were designed) shares a significant sequence- and structure-similarity with proteases which are members of the plasmepsins family of Plasmodium sp.(FIGS. 1 and 2; SEQ ID NO: 1 AND 2, see Original Patent). Similarly to the HIV-1 protease, plasmepsins are aspartyl-proteases and have a fundamental role in the intracellular growth of P. falciparum. They intervene in the first steps of the degradation of hemoglobin, which constitutes the principal nutrient for the intraerythrocytic stages of the parasite. Given the structural similarity between the HIV-1 protease and plasmepsins, it is possible to hypothesize that PIs impair plasmodial growth by targeting these enzymes. This hypothesis is sustained by the fact that the regions of maximal similarity between the two proteins is their catalytic site, which, in the HIV-1 protease, is non-covalently bound to and inhibited by PIs. If this mechanism is confirmed by experimental data, the HIV PIs will become the first drugs subjected to safety tests in humans to inhibit a member of the plasmepsins family, recently indicated by WHO as a potential target for the development of new antimalarials. In a time in which drug-resistant Plasmodium strains are continuously emerging, the availability in the pharmaceutical arsenal of drugs directed to a new target will increase the therapeutic options.

Other potential ground for the antimalarial effect of PIs is the recently described down-modulation of CD36 (a receptor for P. falciparum) induced by these drugs in human erythrocytes. Nathoo S, Serghides L, Kain K C. Effect of HIV-1 antiretroviral drugs on cytoadherence and phagocytic clearance of Plasmodium falciparum-parasitised erythrocytes. Lancet. Sep. 27, 2003;362(9389):1039-41.

The description of the mechanisms reported above has been done only for explanatory purposes: the present invention relates to the effects of PIs on Plasmodium sp. growth in vitro and in vivo and is not limited to any particular mechanism.

The direct antimalarial effects of PIs corroborate their use in combination with CQ, as described above. The direct antimalarial effect of PIs also indicates that HIV-infected individuals living in areas with endemic malaria and treated with an antiretroviral cocktail including a PI may become protected, at least partially, from the occurrence of malarial episodes. Protection from malarial episodes is an advantage for treatment of HIV in view of the limited budgets of several resource-poor countries. Indeed, in Sub-Saharian Africa, there are malaria-endemic areas where the levels of HIV seroprevalence can reach 30%. As HIV-infected people are at higher risk for complicated malaria, one can imagine that the direct anti-plasmodial effects of a PI could save a huge amount of human and financial resources.

Claim 1 of 2 Claims

1. A method of treating malaria in humans comprising administering to a patient in need thereof a therapeutically effective amount of a composition comprising: at least one inhibitor of the HIV protease, or pharmaceutically acceptable salt thereof, selected from the group consisting of indinavir (IDV), ritonavir (RTV), saquinavir (SQV), nelfinavir (NFV), lopinavir (LPV), amprenavir (APV), fosamprenavir, tipranavir, atazanavir, TMC-114, and combinations thereof, in an amount that is therapeutically effective to inhibit the growth of Plasmodium sp.
 

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