e Oral or topical daily administration of antiretroviral (ARV) drugs to HIV-1-negative individuals in vulnerable populations is a promising strategy for HIV-1 prevention. Adherence to the dosing regimen has emerged as a critical factor determining efficacy outcomes of clinical trials. Because adherence to therapy is inversely related to the dosing period, sustained release or long-acting ARV formulations hold significant promise for increasing the effectiveness of HIV-1 preexposure prophylaxis (PrEP) by reducing dosing frequency. A novel, subdermal implant delivering the potent prodrug tenofovir alafenamide (TAF) with controlled, sustained, zero-order (linear) release characteristics is described. A candidate device delivering TAF at 0.92 mg day ؊1 in vitro was evaluated in beagle dogs over 40 days for pharmacokinetics and preliminary safety. No adverse events related to treatment with the test article were noted during the course of the study, and no significant, unusual abnormalities were observed. The implant maintained a low systemic exposure to TAF (median, 0.85 ng ml ؊1 ; interquartile range [IQR], 0.60 to 1.50 ng ml ؊1 ) and tenofovir (TFV; median, 15.0 ng ml ؊1 ; IQR, 8.8 to 23.3 ng ml ؊1 ), the product of in vivo TAF hydrolysis. High concentrations (median, 512 fmol/10 6 cells over the first 35 days) of the pharmacologically active metabolite, TFV diphosphate, were observed in peripheral blood mononuclear cells at levels over 30 times higher than those associated with HIV-1 PrEP efficacy in humans. Our report on the first sustained-release nucleoside reverse transcriptase inhibitor (NRTI) for systemic delivery demonstrates a successful proof of principle and holds significant promise as a candidate for HIV-1 prophylaxis in vulnerable populations. O ral or topical daily administration of antiretroviral (ARV) drugs to HIV-1-negative individuals in vulnerable populations is a promising strategy for HIV-1 prevention, but clinical outcomes have varied widely (1-3). Adherence to frequent dosing is burdensome to the user and has emerged as a key factor in explaining the heterogeneous efficacy outcomes of HIV-1 preexposure prophylaxis (PrEP) clinical trials (4-7). It is well established across different delivery methods that adherence to therapy is inversely related with the dosing period (8-11). Sustained-release or long-acting ARV formulations hold significant promise as a means of reducing dosing frequency, thereby increasing the effectiveness of HIV-1 PrEP.Long-acting preexposure prophylaxis (LA-PrEP) is an alternative regimen to daily dosing designed to mitigate the abovedescribed adherence challenges (12, 13). LA-PrEP has been based primarily on ARV nanoparticles for parenteral administration as injections (12,14). Dosing intervals of 1 month or longer for injectable, long-acting, nanomilled formulations of the integrase strand-transfer inhibitor cabotegravir (GSK1265744) and the nonnucleoside reverse transcriptase inhibitor (NNRTI) rilpivirine are undergoing clinical evaluation as possible regimens for H...

Using both rational and random mutagenesis, we have created the first known broad substrate range, nicotinamide cofactor dependent, and highly stereoselective D-amino acid dehydrogenase. This new enzyme is capable of producing D-amino acids via the reductive amination of the corresponding 2-keto acid with ammonia. This biocatalyst was the result of three rounds of mutagenesis and screening performed on the enzyme meso-diaminopimelate D-dehydrogenase. The first round targeted the active site of the wild-type enzyme and produced mutants that were no longer strictly dependent on the native substrate. The second and third rounds produced mutants that had an increased substrate range including straight-and branched-aliphatic amino acids and aromatic amino acids. The very high selectivity towards the D-enantiomer (95 to > 99% e.e) was shown to be preserved even after the addition of the five mutations found in the three rounds of mutagenesis and screening. This new enzyme could complement and improve upon current methods for D-amino acid synthesis.

With more than 7,000 new HIV infections daily worldwide, there is an urgent need for non-vaccine biomedical prevention (nBP) strategies that are safe, effective, and acceptable. Clinical trials have demonstrated that pre-exposure prophylaxis (PrEP) with antiretrovirals (ARVs) can be effective at preventing HIV infection. In contrast, other trials using the same ARVs failed to show consistent efficacy. Topical (vaginal and rectal) dosing is a promising regimen for HIV PrEP as it leads to low systematic drug exposure. A series of titration studies were carried out in bone marrow/liver/thymus (BLT) mice aimed at determining the adequate drug concentrations applied vaginally or rectally that offer protection against rectal or vaginal HIV challenge. The dose-response relationship of these agents was measured and showed that topical tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC) can offer 100% protection against rectal or vaginal HIV challenges. From the challenge data, EC50 values of 4.6 μM for TDF and 0.6 μM for FTC for HIV vaginal administration and 6.1 μM TDF and 0.18 μM for FTC for rectal administration were obtained. These findings suggest that the BLT mouse model is highly suitable for studying the dose-response relationship in single and combination ARV studies of vaginal or rectal HIV exposure. Application of this sensitive HIV infection model to more complex binary and ternary ARV combinations, particularly where agents have different mechanisms of action, should allow selection of optimal ARV combinations to be advanced into pre-clinical and clinical development as nBP products.

Millions of intravaginal rings (IVRs) are used by women worldwide for contraception and for the treatment of vaginal atrophy. These devices also are suitable for local and systemic sustained release drug delivery, notably for antiviral agents in human immunodeficiency virus pre-exposure prophylaxis. Despite the widespread use of IVRs, no studies have examined whether surfaceattached bacterial biofilms develop in vivo, an important consideration when determining the safety of these devices. The present study used scanning electron microscopy, fluorescence in situ hybridization and confocal laser scanning microscopy to study biofilms that formed on the surface of IVRs worn for 28 days by six female pig-tailed macaques, an excellent model organism for the human vaginal microbiome. Four of the IVRs released the nucleotide analogue reverse transcriptase inhibitor tenofovir at a controlled rate and the remaining two were unmedicated. Large areas of the ring surfaces were covered with monolayers of epithelial cells. Two bacterial biofilm phenotypes were found to develop on these monolayers and both had a broad diversity of bacterial cells closely associated with the extracellular material. Phenotype I, the more common of the two, consisted of tightly packed bacterial mats approximately 5 mm in thickness. Phenotype II was much thicker, typically 40 mm, and had an open architecture containing interwoven networks of uniform fibres. There was no significant difference in biofilm thickness and appearance between medicated and unmedicated IVRs. These preliminary results suggest that bacterial biofilms could be common on intravaginal devices worn for extended periods of time.