Jorge Vila

Absence of viral rebound after treatment of HIV-infected patients with didanosine and hydroxycarbamide

Baseline CD4 count was normal in both patients, except that patient B had a below-normal percentage relative to total lymphocytes. From day 360, both patients remained within normal ranges (table). Plasma RNA was positive at baseline (patient A: 676 copies/mL; patient B: 1120 copies/mL), became undetectable by day 360 and remained undetectable after 12 months’ suspension of treatment, in both patients. Baseline proviral DNA in the PBMC was quantified at 654 copies/10 cells in patient A and 65 copies/10 cells in patient B. At days 360 and 720, proviral DNA was non-quantifiable in patient B, and marginally detectable in patient A. Qualitative evaluation of proviral DNA in PBMC in triplicate experiments at day 720 showed low amplification of the pol region in both patients (one of three samples positive), and of the gag region (one of three samples positive for patient A, and two of three positive for patient B). To assess whether proviral DNA detected was competent for transcription, we tested for intracellular HIV-RNA. Baseline intracellular RNA in the PBMC was quantifiable in both patients but was undetectable at day 360 and remained undetectable despite the absence of treatment. We were unable to isolate any infectious virus from the PBMC. At day 360, lymphnode viral load was undetectable in patient A; in patient B, 26 copies of proviral DNA per 10 cells was found. One year after treatment was stopped, very low levels of proviral DNA were found in lymphnodes of both patients. The presence of provirus was confirmed by qualitative analysis (amplification of gag positive for both patients, and amplification of pol positive for patient B and negative for patient A). However, no extracellular nor intracellular HIV RNA was detectable at day 720 in either

Anti-HIV activity of the combination of didanosine and hydroxyurea in HIV-1-infected individuals.

HIV is known to be present in massive amounts in both resting and actively replicating cells in infected individuals. We tested the combination of didanosine and hydroxyurea, known to suppress viral production in vitro in both of these cell types, in a small number of asymptomatic patients. After 3 months of well tolerated treatment, we observed a large reduction of viral load in the peripheral blood of all 12 patients, down to nonquantifiable levels in 7 of 12 as measured by infectious virus titer, and 6 of 12 as measured by plasma HIV-RNA. In this subgroup of 6 patients, whose baseline HIV-RNA was below 14,000 copies/ml, the median increase in CD4+ count after 90 days of treatment was 244 cells/mm3.

Neurotoxic effect of the anti-HIV drugd-aspartate β-hydroxamate for rat primary neuronal cultures: attenuation by N-methyl-d-aspartate (NMDA) antagonists

The anti-tumor drug D-aspartate beta-hydroxamate (D-A beta H), selectively destroys HIV-1 infected peripheral blood mononuclear cells, but produces anorexia and nausea during prolonged treatment to AIDS patients. Consequently, based on the structural similarity between D-A beta H and the excitotoxins L-aspartate and NMDA, we have investigated the potential neurotoxic action and pharmacology of D-A beta H and of a series of chemically related anti-tumor drugs on rat primary neuronal/glial cultures. In this aim, after a 30 min exposure to D-A beta H (1-2 mM), cortical neurons were selectively destroyed within 24 h. The stereoisomer L-A beta H (0.5-2 mM) was highly neurotoxic for both glial and neuronal cells in mixed cultures but demonstrated no toxicity in glial cell cultures alone. Furthermore, for a series of D-A beta H analogues, VHS.121 and VHS.122 demonstrated a reduced but significant neurotoxicity, whereas VHS.124 and VHS.125 showed no significant neurotoxic effect, and in the case of VHS.125 also prevented D-A beta H and glutamate-mediated neurotoxicity. The related anti-tumor drugs L- or D-glutamate gamma-monohydroxamate or keto-glutamate gamma-monohydroxamate (< or = 2 mM) were not neurotoxic for cortical neurons. The neurotoxic effect of D-A beta H and L-A beta H was attenuated by the NMDA antagonists MK-801, TCP, memantine, ifenprodil, pentamidine and CGS-19755. alpha-Difluoromethylornithine, an inhibitor of polyamine biosynthesis, also protected cultures against the neurotoxicity of L-A beta H and D-A beta H.(ABSTRACT TRUNCATED AT 250 WORDS)

A N-methyl-d-aspartate receptor-mediated neurotoxic effect of aspartate-based hydroxamate compounds in rat primary neuronal cultures

In a previous study (Lockhart et al., Brain Res., 630 (1993) 32-40) we demonstrated a NMDA receptor-mediated neurotoxic effect for the anti-HIV drugs D-aspartate beta-hydroxamate and structurally related analogues in rat primary cortical neurons. Herein, we have examined the neurotoxic action and pharmacology of a novel series of hydroxamate compounds, with potential anti-HIV activity, in a similar paradigm. In this aim, the aspartate-based hydroxamates L-VHS.126 and its stereoisomer D-VHS.126 selectively destroyed (EC50 = 300 microM) rat primary neurons. The D-VHS.126 analogue, VHS.134 was also neurotoxic (EC50 = 450 microM) whereas VHS.129 and VHS.137, or the D-aspartate beta-hydroxamate analogues VHS.128, VHS.135, VHS.132 and VHS.127 or hydroxyurea demonstrated no significant neurotoxicity. The neurotoxic action of L- and D-VHS.126, and VHS.134 was attenuated with MK-801, CGS-19755 but not with CNQX. These observations demonstrate a potent NMDA receptor-mediated excitotoxic action for novel aspartate-based hydroxamate compounds, and further extends this series of potential hydroxamate-based anti-HIV drugs lacking neurotoxicity in vitro.

Binding of the antiretroviral drug, d-aspartate-β-hydroxamate on the NMDA receptor.

d-Aspartate-β-hydroxamate (d-A β H) exhibits antiretroviral properties in vitro and in vivo. It has glutamate agonist properties at the N-methyl-d-aspartate (NMDA) receptor in neuronal cell cultures. This study characterizes its binding properties to the NMDA receptor by measuring its stimulating effect on N-(1-(2-thienyl)[(3)H]cyclohexyl)piperidine ([(3)H]TCP) binding to the ionic channel in rat brain membranes. d-A β H stimulated [(3)H]TCP binding in a dose-dependent manner but to a lower extent than glutamate, suggesting only partial glutamate agonist properties. In the presence of antagonists of the different effector sites of the NMDA receptor the affinity of d-A β H was competitively decreased by CGS-19755 and 7-chlorokynurenate and unaffected by arcaine. Among several d-A β H analogues VHS.125 behaved as a full NMDA agonist, but l- or d-glutamate γ-monohydroxamate (d-GH or l-GH) were without effect. This study shows that d-A β H has potential neurotoxic effects due to its direct interaction with the NMDA receptor and that analogues such as d-GH or l-GH may rather be used in humans.