Herein we report the formulation and vaginal delivery of CN54gp140 within solid dosage forms; lyophilized equivalents of the Carbopol®, RSV and modified RSV semi-solid formulations. The innovative, robust, lyophilized solid dosage formulations (LSDFs) in this study were more conducive to CN54gp140 stability with the potential to offer improved patient acceptability for vaginal administration than the equivalent semi-solid formulations. In addition, the viability of the LSDFs as delivery modalities for vaginal immunization was demonstrated by the ability of the vaginally administered lyophilized formulations containing CN54gp140 to boost subcutaneously primed mice.

Polyvinylpyrollidone (PVP) (Plasdone® K-90, Mv 1.3 M) and Polycarbophil (PC) (Noveon® AA1, divinyl crosslinked polyacrylic I-BET151 ic50 acid) were kindly donated by International Speciality Products (Ohio, USA) and Noveon Pharma GmbH & Co KG (Raubling, Germany), respectively. HEC (Natrosol 250 HHX and 250 G) and sodium NaCMC (Blanose® 7LF, 7MF, and 7HF) were also kindly donated by Aqualon (Warrington, UK). GMP manufactured Carbopol® 974P gel, formulation #2449 was kindly donated by Particle Sciences (Bethlehem, PA, USA). Galanthus nivalis (GNA) was obtained from Vector Laboratories (Peterborough, England).

3,3′,5,5′-Tetramethylbenzidine Z-VAD-FMK concentration peroxidase substrate (TMB/E) was obtained from Cygnus Technologies Inc. (North Carolina, USA). CN54gp140 (gp120 plus the ectodomain of gp41) was encoded by the CN54gp140REKE HIV-1 envelope gene cassette derived from the clade-C/B′ HIV-1 molecular clone p97CN54 of Chinese origin developed by Wolf and Wagner, University of Regensburg, Germany [15] and [16]. CN54gp140 was produced as a recombinant product in CHO cells by S. Jeffs, Imperial Org 27569 College, London, and manufactured to GMP specification by Polymun Scientific (Vienna, Austria) who also donated the HIV-1 gp41 specific monoclonal antibody 5F3 (HuMab 5F3). Sodium hydroxide, phosphate buffered saline containing Tween 20 (PBS-T), sterile-filtered porcine serum and goat anti-human horseradish

peroxidase (HRP)-conjugated IgG were purchased from Sigma–Aldrich (Poole, Dorset, UK). Goat anti-mouse HRP-conjugated IgA and biotinylated goat anti-mouse IgA were obtained from AbD Serotec (UK). HRP-conjugated streptavidin was purchased from R&D Systems (MN, USA). 25X protease inhibitor cocktail was obtained from Roche (Hertfordshire, UK). Reactibind 96 well microplates were obtained from Perbio Science (Northumberland, England). Nunc Maxisorp 96 well microplates were obtained from Nalge Nunc International (Rochester, NY). Nalgene tubing (PVC, 3 mm internal diameter, 5 mm outer diameter, 1 mm Wall) was purchased from VWR International Ltd. (Dublin, Ireland) and blister packs were kindly supplied by Almac (Craigavon, UK) and Warner Chilcott (Larne, UK). Ultra-pure water was obtained using an Elga Purelab Maxima system.