Evaluation of the Safety and Toxicological Profile of RheoSprayTM on the Vaginal Mucosa

The vaginal mucosa is a common site for local and systemic delivery of medication. This study aims to compare the safety and toxicological profile of RheoSpray, a proprietary sprayable topical gel compounding base, to the profile of the positive control Gynoll II, using a 3D model of the human vaginal mucosa. Results have demonstrated that RheoSpray presented an ET50 superior to 24 hr, as opposed to an ET50 of less than 4 hr for the Gynol II. Therefore, it may be concluded that RheoSpray does not cause toxicity to the vaginal tissues. Compounded medicines including this sprayable topical gel base may then be considered safe to be applied to the vaginal mucosa for over 24 hr. Introduction: Vaginal delivery of medication is advantageous in allowing for the medication to avoid first-pass metabolism and gastrointestinal degradation [1]. Lined with noncornified, stratified squamous epithelium, the vaginal mucosa offers a large surface area and rich blood supply, making it a promising site for delivery of medication in the treatment of several conditions and also in hormone replacement therapy [2]. The aim of this study was to evaluate the safety and toxicological profile of RheoSpray, a proprietary sprayable topical gel compounding base, in comparison to the positive control Gynol II [nonoxynol-9 (3%)], an irritant of the vaginal mucosa, using a 3-dimensional (3D) in vitro model of the human vaginal mucosa. Vaginal Tissue Model The EpiVaginalTM tissue model by MatTek Corporation (Ashland, MA) is a highly differentiated tissue cultured from normal, human-derived vaginal epithelial and dendritic cells. Its tissue structure and cellular physiology closely parallels in vivo vaginal epithelial tissue. It is therefore an ideal in vitro research tool for safety and toxicological testing of feminine products. The EpiVaginalTM tissue containing epithelial VEC-100 cells was the model used in this study (Figure 1) [3]. Figure 1. Illustration of the EpiVaginalTM tissue model. (Adapted from MatTek Corporation, 2016) Methodology: Upon receipt of the EpiVaginalTM tissue model, the VEC100 cells (Lot 25009) were maintained in the supplied culture media and stored in accordance to the manufacturer’s protocol until the initiation of the study. Following preparation of the cells, the EpiVaginalTM tissues were treated in triplicate with 100 μL of the test product (RheoSpray, Lot #7542733) and another set of tissues were treated with Gynol II for 1, 4 and 24 hr. A triplicate set of EpiVaginalTM tissues was also left untreated to serve as negative control. Following the exposure period, the dosing solutions were removed and the cells were analyzed for cell viability by the MTT Effective Time 50 (ET50) assay. The MTT ET50 assay consists of measuring the reduction of MTT (3-[4,5-dimethylthiazol-2yl]-2,5diphenyltetrazolium bromide) by the cells. Succinate dehydrogenase enzymes within the mitochondria of viable cells have the ability to reduce soluble yellow tetrazonium salt of MTT to an insoluble purple formazan derivative. MTT is therefore an indicator of cell viability as the tissues are evaluated for their ability to reduce soluble-MTT (yellow) to formazan-MTT (purple) [4]. The MTT solution was prepared in the provided medium and added to the basal side of each tissue, followed by an incubation period of the tissues for 3 hr at 37°C. The purple formazan product was then extracted using the provided extractant, which was previously applied to both the apical and basal side of the tissues. Sample absorbance was read at 570 nm and reference absorbance at 650 nm with CLARIOstar – BMG Labtech Plate reader. Results and Discussion: Viability of the vaginal cells following exposure to the test products is represented by the absorbance of the respective extracts and expressed in percentage relative to the negative control (tissues left untreated), as follows: % Cell Viability=100 x [OD(test product) / OD(negative control)] ©2017 PCCA Science | 99381 | Page 1 of 2 T E C H N I C AL R E P O R T