Improved Dual-Modality Bioequivalence Evaluation of Topical Formulations Within Human Skin Using Stimulated Raman Scattering Microscopy.

Abstract

Background: The use of optical microscopic techniques has gained increasing attention in recent years for studying the bioavailability (BA) and bioequivalence (BE) of topical drugs. Stimulated Raman scattering (SRS), one type of optical imaging technique, probes chemical-specific information and has excellent spatiotemporal resolution. It uses intrinsic molecular vibrational signatures, and therefore, labeling samples or other treatments is unnecessary to track a molecule. Because of its unique advantages, studies have used SRS for BA evaluations and, more recently, for BE evaluations. In BE evaluation, low data variance within a treatment group is important to ensure sensitivity and specificity in comparing treatment groups. Methods: When measuring forward-direction SRS signals transmitted through skin, the signal intensity is susceptible to variance due to several factors, such as the microscope system's performance, the different optical features of topical drug products, and the heterogeneity of skin in transmitting light. This work closely investigated the effects of these factors on an SRS signal and developed solutions to reduce their effects on the data variance. Specifically, we constructed a method using a dual-modality detector built in-house, which simultaneously measured both the SRS signal and total light transmission synchronized in time and co-registered in space. Results: We developed equations to normalize SRS signals using the transmission intensity, and the results demonstrated a clear improvement in the SRS signal via a reduction in the signal variance (up to a 9.46% CV value decrease) that is otherwise caused by various factors associated with the use of topical drugs and the composition of the skin. We carried out an exploratory BE study using tretinoin-containing topical products and observed improvements in BE assessment with the developed method (could achieve a reduction of 0.11 in the CI value). Conclusions: This work has led to a better understanding of the factors that affect SRS imaging and has provided an effective method to compensate for these factors in BE assessments. This is a critical initial effort for better practical implementation of SRS in cutaneous pharmacokinetics (cPKs) studies of topical drugs.