MLN8237 Alisertib were collected at sacrifice 24 h after a single dose and were divided

sue viability, MLN8237 Alisertib transepithelial electrical resistance, and histology using an organotypic human VEC as previously reported.25,30,31 Gynol II, which contains the nonionic surfactant nonoxynol 9, was used as a positive control and PBS as a negative control. Additional controls were the ISG placebo gel and the universal HEC placebo gel.32,33 Candidate SG, ISG, and BG compositions were evaluated in these studies. In Vivo Pharmacokinetics Following vaginal administration of the SG, ISG, and BG, the PK of TFV and UC781 were evaluated in female New Zealand White HraSPF rabbits weighing approximately 2.5 kg each. The study was approved by the MPI Research Institutional Animal Care andUse Committee and followed the protocol reported previously for assessment of the UC781 only gels,25 with the exception that 1.
0mL of gel was administered intravaginally to the rabbits in this study. Each gel containing 1.0% TFV and 0.1%UC781 was administered 8 cm into the abdominal vagina via a polyurethane catheter. Vaginal tissues were collected at sacrifice 24 h after a single dose and were divided into distal and proximal sections. Another group of rabbits was administered the appropriate gel once daily for 7 cox2 inhibitor days, 24 h following the last dose, the rabbits were sacrificed and tissues divided as described above. The tissues were swabbed three times to remove any remaining gel, without damaging the epithelium. Plasma and vaginal tissue samples were processed and analyzed for UC781 using an LC MS MS method described previously.25 Likewise, plasma and tissues were analyzed for free TFV and total TFV using LC MS MS methods described below.
Tissues were snap frozen in liquid nitrogen and stored at 0. For analysis, tissue was ground in a coffee grinder type small blade homogenizer with dry ice to make a finely ground LY315920 tissue sample. The sample was then placed in a freezer at 0 overnight to allow the dry ice to sublime. The samples were then transferred to a cryogenic tube for storage at 0. The ground tissue was diluted 10 fold with the extraction solvent in a 2mL amber cryovial. All samples were then sonicated in an ice bath for 15 min. Bioanalytical Methods Plasma UC781 Measurement of UC781 in rabbit plasma and vaginal tissues of rabbits was based on a validated LC MS MS method as reported previously.25 Plasma TFV The plasma samples were added to a 96 well plate and 50 :L working internal standard TFVd6 was added.
Then, 0.5% formic acid was added to each plate well and the plate was gentlymixed by hand for approximately 20 s. The samples were then transferred to preconditioned SPE plates and drawn through using minimal vacuum. Next, formic acid was added to each well and drawn through with vacuum until the bed surface appeared dry, the same process was repeated using 800 :L methanol. This stepwas followed by application of full vacuum for 2 min. Elution was performed with 600 :L methanol/NH4OH and the samples evaporated to dryness under a stream of N2 at 50. The samples were reconstituted with 200 :L water. The plate was then capped and gently vortexed for 20 s. Tenofovir in samples prepared as described above was measured using an Agilent 1100 Series HPLC system coupled to an Applied Biosystems/MDS Sciex API 5000. MS MS data were obtained in the positive po

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