No abstract

Combination antiretroviral therapy has become the standard of care for patients with human immunodeficiency virus (HIV) infection in the United States (19,20,23,25). Antiretroviral treatment regimens usually employing combinations of drugs from at least two of the three classes of antiretroviral therapy, namely, nucleoside analog reverse transcriptase (RT) inhibitors (NRTI), nonnucleoside analog RT inhibitors (NNRTI), and protease inhibitors, exhibit a potent and sustained antiviral effect and confer consistent long-term viral suppression in patients with HIV infection (19,20,23,25). However, each of these drugs can select for drug-resistant viruses, and the emergence of antiviral drug resistance limits their clinical benefit (3,4,6,8,10,11,14,15,17,18,21,22,24,26,31). The transmission of drug-resistant HIV is a serious problem that merits further attention by public health officials as well as virologists and clinicians. There is an urgent need for new anti-HIV agents capable of inhibiting the replication of NRTIas well as NNRTI-resistant HIV.Stavudine (STV) (also known as "d4T") is a pyrimidine nucleoside analogue used in the treatment of HIV infection. It inhibits HIV RT, as do zidovudine (ZDV), didanosine, zalcitabine, and lamivudine (3TC), which comprise the family of NRTI. The 5Ј-triphosphates of these NRTI, which are generated intracellularly by the action of nucleoside and nucleotide kinases, are potent inhibitors of HIV type 1 (HIV-1) RT (20,27). The rate-limiting step for the generation of the bioactive STV metabolite STV-triphosphate is the conversion of STV to its monophosphate derivative (27). In an attempt to overcome the dependence of STV on intracellular nucleoside kinase activation, we prepared STV-5Ј-(p-bromophenyl methoxyalaninyl phosphate(stampidine [STAMP])/HI-113, STV-5Ј-[p-bromophenyl methoxyalaninyl phosphate], a novel aryl phosphate derivative of STV (27,30). In preliminary studies, we found that STAMP is substantially more potent than STV in inhibiting HIV-1 replication in thymidine kinase-deficient T cells (27,30,32).Recently, we completed a detailed analysis of the in vitro activity profile of STAMP against HIV-1 isolates with B and non-B envelope subtypes (28). STAMP exhibited potent anti-HIV activity against the HIV-1 strain HTLV IIIB in human peripheral blood mononuclear cells with nanomolar 50% inhibitory concentration (IC 50 ) and IC 90 values and a selectivity index of Ͼ100,000. The IC 50 (mean Ϯ standard error of the mean [SEM]) of STAMP (0.001 Ϯ 0.000 M) against HTLV-IIIB was 20 times lower than the IC 50 of STV (0.023 Ϯ 0.008 M; P Ͻ 0.001), 3 times lower than the IC 50 of ZDV (0.003 Ϯ 0.001 M; P Ͻ 0.001), 40 times lower than the IC 50 of 3TC (0.040 Ϯ 0.025 M; P Ͻ 0.001), 6 times lower than the IC 50 of nelfinavir (0.006 Ϯ 0.003 M; P Ͻ 0.001), and 20 times lower than the IC 50 of nevirapine (0.024 Ϯ 0.007 M; P Ͻ 0.001) (28) (all values are means Ϯ SEMs). Similarly, in a side-by-side comparison against 10 ZDV-sensitive primary clinical HIV-1 isolates, 9 of which had a non-B en...

Here we provide experimental evidence that identifies JAK3 as one of the regulators of platelet function. Treatment of platelets with thrombin induced tyrosine phosphorylation of the JAK3 target substrates STAT1 and STAT3. Platelets from JAK3-deficient mice displayed a decrease in tyrosine phosphorylation of STAT1 and STAT3. In accordance with these data, pretreatment of human platelets with the JAK3 inhibitor WHI-P131 markedly decreased the base-line enzymatic activity of constitutively active JAK3 and abolished the thrombin-induced tyrosine phosphorylation of STAT1 and STAT3. Following thrombin stimulation, WHI-P131-treated platelets did not undergo shape changes indicative of activation such as pseudopod formation. WHI-P131 inhibited thrombin-induced degranulation/serotonin release as well as platelet aggregation. Highly effective platelet inhibitory plasma concentrations of WHI-P131 were achieved in mice without toxicity. WHI-P131 prolonged the bleeding time of mice in a dose-dependent manner and improved event-free survival in a mouse model of thromboplastin-induced generalized and invariably fatal thromboembolism. To our knowledge, WHI-P131 is the first anti-thrombotic agent that prevents platelet aggregation by inhibiting JAK3.

Here we report the antiretroviral activity of the experimental nucleoside reverse transcriptase inhibitor (NRTI) compound stampidine in cats chronically infected with feline immunodeficiency virus (FIV). Notably, a single oral bolus dose of 50 or 100 mg of stampidine per kg resulted in a transient >1-log decrease in the FIV load of circulating peripheral blood mononuclear cells in five of six FIV-infected cats and no side effects. A 4-week stampidine treatment course with twice-daily administration of hard gelatin capsules containing 25 to 100 mg of stampidine per kg was also very well tolerated by cats at cumulative dose levels as high as 8.4 g/kg and exhibited a dose-dependent antiretroviral effect. One of three cats treated at the 25-mg/kg dose level, three of three cats treated at the 50-mg/kg dose level, and three of three cats treated at the 100-mg/kg dose level (but none of three control cats treated with placebo pills) showed a therapeutic response, as evidenced by a >1-log reduction in the FIV load in peripheral blood mononuclear cells within 2 weeks. The previously documented in vitro and in vivo antiretroviral activity of stampidine against primary clinical human immunodeficiency virus type 1 isolates with genotypic and/or phenotypic NRTI resistance, together with its favorable animal toxicity profile, pharmacokinetics, and in vivo antiretroviral activity in FIV-infected cats, warrants further development of this promising new NRTI compound.Stavudine (STV) is a pyrimidine nucleoside analogue used in the treatment of human immunodeficiency virus (HIV) infection. It inhibits viral reverse transcriptase (RT), as do zidovudine (ZDV), didanosine, zalcitabine, and lamivudine, which make up the family of nucleoside RT inhibitors (NRTIs). The 5Ј triphosphates of these NRTI, which are generated intracellularly by the action of nucleoside and nucleotide kinases, are potent inhibitors of HIV type 1 (HIV-1) RT (13). The rate-limiting step in the generation of the bioactive STV metabolite STV triphosphate is conversion of STV to its monophosphate derivative (3,13,23). To overcome the dependence of STV on intracellular nucleoside kinase activation, we prepared stampidine (STAMP) HI-113, STV-5Ј-(p-bromophenyl methoxyalaninyl phosphate), a novel aryl phosphate derivative of STV (18,23,25). In preliminary studies, we found that STAMP is substantially more potent than STV at inhibiting HIV-1 replication in thymidine kinase-deficient T cells (23). STAMP was a much more potent anti-HIV agent than STV, and it was active against phenotypically and/or genotypically NRTI-resistant HIV strains for which the 50% inhibitory concentrations (IC 50 s) are in the low nanomolar-to-subnanomolar range (21). Similarly, STAMP inhibited the replication of laboratory HIV-1 strains and primary clinical HIV-1 isolates with non-NRTI (NNRTI) (1) binding site mutations and NNRTI-resistant phenotypes for which the IC 50 s are in the low nanomolar-to-subnanomolar range (21).

The purpose of the present study was to evaluate the effects of graft-versus-host disease (GVHD) prophylaxis with the Janus kinase 3 (JAK3) inhibitor WHI-P131/ JANEX-1 on the graft-versus-leukemic (GVL) function of marrow allografts in mice undergoing bone marrow transplantation (BMT) after being challenged with an otherwise invariably fatal dose of BCL-1 leukemia cells. GVHD prophylaxis using WHI-P131 markedly improved the survival outcome after BMT. The probability of survival at 30 days after BMT was 11% ؎ 6% for vehicle-treated recipients