Safety, pharmacokinetics and neutralization of the broadly neutralizing HIV-1 human monoclonal antibody VRC01 in healthy adults.

VRC-HIVMAB060-00-AB (VRC01) is a broadly neutralizing HIV-1 monoclonal antibody (mAb) isolated from the B cells of an HIV-infected patient. It is directed against the HIV-1 CD4 binding site and is capable of potently neutralizing the majority of diverse HIV-1 strains. This Phase I dose-escalation study in healthy adults was conducted at the National Institutes of Health (NIH) Clinical Center (Bethesda, MD, USA). Primary objectives were the safety, tolerability and pharmacokinetics (PK) of VRC01 intravenous (i.v.) infusion at 5, 20 or 40 mg/kg, given either once (20 mg/kg) or twice 28 days apart (all doses), and of subcutaneous (s.c.) delivery at 5 mg/kg compared to s.c. placebo given twice, 28 days apart. Cumulatively, 28 subjects received 43 VRC01 and nine received placebo administrations. There were no serious adverse events or dose-limiting toxicities. Mean 28-day serum trough concentrations after the first infusion were 35 and 57 µg/ml for groups infused with 20 mg/kg (n = 8) and 40 mg/kg (n = 5) doses, respectively. Mean 28-day trough concentrations after the second infusion were 56 and 89 µg/ml for the same two doses. Over the 5-40 mg/kg i.v. dose range (n = 18), the clearance was 0.016 l/h and terminal half-life was 15 days. After infusion VRC01 retained expected neutralizing activity in serum, and anti-VRC01 antibody responses were not detected. The human monoclonal antibody (mAb) VRC01 was well tolerated when delivered i.v. or s.c. The mAb demonstrated expected half-life and pharmacokinetics for a human immunoglobulin G. The safety and PK results support and inform VRC01 dosing schedules for planning HIV-1 prevention efficacy studies.

Glutamate stimulates ascorbate transport by astrocytes.

The concentrations of glutamate and ascorbate in brain extracellular fluid increase following seizure activity, trauma and ischemia. Extracellular ascorbate concentration also rises following intracerebral glutamate injection. We hypothesized that glutamate triggers the release of ascorbate from astrocytes. We observed in primary cultures of rat cerebral astrocytes that glutamate increased ascorbate efflux significantly within 30 min. The half-maximal effective concentration of glutamate was 180+/-30 microM. Glutamate-stimulated efflux of ascorbate was attenuated by hypertonic media. 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid inhibited both Na(+)-dependent glutamate uptake and ascorbate efflux. Two other inhibitors of volume-sensitive organic anion channels (1, 9-dideoxyforskolin and 5-nitro-2-(3-phenylpropylamino) benzoic acid) did not slow glutamate uptake but prevented stimulation of ascorbate efflux. Glutamate also stimulated the uptake of ascorbate by ascorbate-depleted astrocytes. In contrast, glutamate uptake was not affected by intracellular ascorbate, thus ruling out a putative glutamate-ascorbate heteroexchange mechanism. These results are consistent with activation by glutamate of ascorbate-permeant channels in astrocytes.

Propofol prevents peroxide-induced inhibition of glutamate transport in cultured astrocytes.

BACKGROUND: Glutamate transporters located in the plasma membrane of cerebral astrocytes take up excitatory neurotransmitters from the synaptic cleft. In diseases characterized by oxidative stress, the extracellular glutamate concentration increases and contributes to neuronal death. The authors wanted to determine whether propofol defends brain cells against oxidant-induced changes in their transport of glutamate. METHODS: Primary cultures of rat cerebral astrocytes were exposed to tert-butyl hydroperoxide (1 mM) to serve as an in vitro model of oxidative stress. Astrocytes were incubated with propofol for 2 h and tert-butyl hydroperoxide was added for the final hour. Alternatively, astrocytes were incubated with tert-butyl hydroperoxide for 30 min and then with propofol for another 30 min. Control cells received drug vehicle rather than propofol. The rate of uptake of glutamate, the efflux of the nonmetabolizable analog D-aspartate, and the intracellular concentration of the endogenous antioxidant glutathione were measured. RESULTS: Tert-butyl hydroperoxide decreased the glutathione concentration and inhibited glutamate uptake but had a negligible effect on D-aspartate efflux. At clinically relevant concentrations, propofol did not affect the glutathione concentration but did prevent the effect of tert-butyl hydroperoxide on glutamate transport. Furthermore, the addition of propofol after tert-butyl hydroperoxide reversed the inhibition of glutamate uptake. CONCLUSIONS: Propofol prevents and reverses the inhibition of excitatory amino acid uptake in astrocytes exposed to tert-butyl hydroperoxide. The ability of propofol to defend against peroxide-induced inhibition of glutamate clearance may prevent the pathologic increase in extracellular glutamate at synapses, and thus delay or prevent the onset of excitotoxic neuronal death.