Development and pharmacokinetics of a combination vaginal ring for sustained release of dapivirine and the protein microbicide 5P12-RANTES.

The past fifteen years have witnessed a resurgence of interest in vaginal ring technologies for drug delivery applications, mostly driven by the impetus for development of vaginally-administered antiretroviral microbicides to help reduce the high acquisition rates for human immunodeficiency virus (HIV) among Sub-Saharan African women. Currently, the lead candidate microbicide is a 28-day silicone elastomer vaginal ring releasing dapivirine (Ring-004), an experimental non-nucleoside reverse transcriptase inhibitor. The ring was tested in two pivotal Phase III clinical studies in 2016 and is currently undergoing review by the European Medicines Agency. Recently, we described a new type of silicone elastomer vaginal ring offering sustained release of the protein molecule 5P12-RANTES, a potent experimental chemokine analogue that potently blocks the HIV CCR5 coreceptor. Building on our previous work, here we report the preclinical development of a new combination vaginal ring that offers sustained release of both 5P12-RANTES and dapivirine, in which the 5P12-RANTES is incorporated into an exposed core within the ring body and the dapivirine in the sheath. In this way, in vitro release of dapivirine matches closely that for Ring-004. Also, we report the pharmacokinetic testing of this combination ring formulation in sheep, where vaginal concentrations of both drugs are maintained over 28 days at levels potentially useful for preventing HIV infection in women.

Vaginal rings with exposed cores for sustained delivery of the HIV CCR5 inhibitor 5P12-RANTES.

Antiretroviral-releasing vaginal rings are at the forefront of ongoing efforts to develop microbicide-based strategies for prevention of heterosexual transmission of the human immunodeficiency virus (HIV). However, traditional ring designs are generally only useful for vaginal administration of relatively potent, lipophilic, and small molecular weight drug molecules that have sufficient permeability in the non-biodegradable silicone elastomer or thermoplastic polymers. Here, we report a novel, easy-to-manufacture 'exposed-core' vaginal ring that provides sustained release of the protein microbicide candidate 5P12-RANTES, an experimental chemokine analogue that potently blocks the HIV CCR5 coreceptor. In vitro release, mechanical, and stability testing demonstrated the utility and practicality of this novel ring design. In a sheep pharmacokinetic model, a ring containing two »-length excipient-modified silicone elastomer cores - each containing lyophilised 5P12-RANTES and exposed to the external environment by two large windows - provided sustained concentrations of 5P12-RANTES in vaginal fluid and vaginal tissue between 10 and 10,000 ng/g over 28days, at least 50 and up to 50,000 times the reported in vitro IC50 value.

Delayed release of chemokine CCL25 with bioresorbable microparticles for mobilization of human mesenchymal stem cells.

Chemokines are guiding cues for directional trafficking of mesenchymal stem cells (MSC) upon injury and local chemokine delivery at injury sites is an up-to-date strategy to potentiate and prolong recruitment of MSC. In this study we present the chemokine CCL25, also referred to as thymus-expressed chemokine, to mobilize human MSC along positive but not along negative gradients. We hence proceeded to design a biodegradable and injectable release device for CCL25 on the basis of poly(lactic-co-glycolic acid) (PLGA). The conducted studies had the objective to optimize PLGA microparticle fabrication by varying selected formulation parameters, such as polymer type, microparticle size and interior phase composition. We found that microparticles of DV,50∼75 µm and fabricated using end-capped polymers, BSA as carrier protein and vortex mixing to produce the primary emulsion yielded high chemokine loading and delayed CCL25 release. To determine bioactivity, we investigated CCL25 released during the microparticle erosion phase and showed that deacidification of the release medium was required to induce significant MSC mobilization. The designed PLGA microparticles represent an effective and convenient off-the-shelf delivery tool for the delayed release of CCL25. However, continuative in vivo proof-of-concept studies are required to demonstrate enhanced recruitment of MSC and/or therapeutical effects in response to CCL25 release microparticles. STATEMENT OF SIGNIFICANCE: With the discovery of chemokines, particularly CXCL12, as stimulators of stem cell migration, the development of devices that release CXCL12 has proceeded quickly in the last few years. In this manuscript we introduce CCL25 as chemokine to induce mobilization of human MSC. This study proceeds to demonstrate how selection of key formulation parameters of CCL25 loading into PLGA microparticles exerts considerable influence on CCL25 release. This is important for a broad range of efforts in in situ tissue engineering where the candidate chemokine and the delivery device need to be selected carefully. The use of such a cell-free CCL25 release device may provide a new therapeutic option in regenerative medicine.

Pharmacokinetics of the Protein Microbicide 5P12-RANTES in Sheep following Single-Dose Vaginal Gel Administration.

5P12-RANTES, a chemokine analogue that potently blocks the HIV CCR5 coreceptor, is being developed as both a vaginal and rectal microbicide for prevention of sexual transmission of HIV. Here, we report the first pharmacokinetic data for 5P12-RANTES following single-dose vaginal gel administration in sheep. Aqueous gel formulations containing low (1.24-mg/ml), intermediate (6.18-mg/ml), and high (32.0-mg/ml; suspension-type gel) concentrations of 5P12-RANTES were assessed via rheology, syringeability, and in vitro release testing. Following vaginal gel administration to sheep, 5P12-RANTES concentrations were measured in vaginal fluid, vaginal tissue, and serum over a 96-h period. All gels showed non-Newtonian pseudoplastic behavior, with the high-concentration gels exhibiting a greater viscosity and cohesive structure than the intermediate- and low-concentration gels. In in vitro release testing, >90% 5P12-RANTES was released from the low- and intermediate-concentration gels after 72 h. For the high-concentration gel, ∼50% 5P12-RANTES was detected, attributed to protein denaturation during lyophilization and/or subsequent solvation of the protein within the gel matrix. In sheep, 5P12-RANTES concentrations in vaginal fluid, vaginal tissue, and serum increased in a dose-dependent manner. The highest concentrations were measured in vaginal fluid (105 to 107 ng/ml), followed by vaginal tissue (104 to 106 ng/ml). Both of these concentration ranges are several orders of magnitude above the reported half-maximal inhibitory concentrations. The lowest concentration was measured in serum (<102 ng/ml). The 5P12-RANTES pharmacokinetic data are similar to those reported previously for other candidate microbicides. These data, coupled with 5P12-RANTES's potency at picomolar concentrations, its strong barrier to resistance, and the full protection that it was observed to provide in a rhesus macaque vaginal challenge model, support the continued development of 5P12-RANTES as a microbicide.

Eotaxin but not MCP-3 induces eosinophil influx into nasal fluid in allergic patients.

BACKGROUND: Eotaxin and MCP-3 (CC chemokines), owing to their preferential action on eosinophils, seem to be the very importance in the patophysiology of allergic rhinitis and asthma. The purpose of this study was to examine the effect of intranasally administered eotaxin and MCP-3 after specific allergen priming on the influx of inflammatory cells and their soluble mediators into the nasal mucosa. METHODS: Eotaxin and MCP-3 have been applied intranasally at the increasing doses of 1, 5 and 10 microg to allergic patients after allergen priming. The 'nasal pool' technique was used. The cell count and biochemical parameters in nasal lavage were evaluated before 30 min, and 4 and 24 h after the challenge with chemokines. RESULTS: Both eotaxin and MCP-3 induced the increase in clinical 'score' lasting till 24 h. Eosinophil influx into nasal mucosa after provocation with eotaxin was also observed. The challenge with MCP-3 did not induce any significant changes in nasal lavage fluid. CONCLUSIONS: Eotaxin is likely to play an important role in the pathogenesis of allergic conditions in humans. MCP-3 did not induce inflammatory cell influx into nasal mucosa. The role of this chemokine in the pathogenesis of allergic inflammation is difficult to assess and requires further studies.