Chemopreventive Effects of Black Raspberry Gel on Oral Epithelial Dysplasia

It is known that reduced expression of pro-apoptotic and terminal differentiation genes in conjunction with increased levels of proinflammatory and angiogenesis-inducing enzymes cyclooxygenase 2 (COX2) and incucible nitric oxide synthase (iNOS) correlate with malignant transformation of oral intraepithelial neoplasia (IEN) Objective: This study assessed the effects of a 10% (w/w) freeze dried black raspberry (FBR) gel on oral IEN histopathology, gene expression profiles and intraepithelial COX2 and iNOS proteins. Our laboratories have previously demonstrated that FBR possess antioxidant properties and also induce keratinocyte apoptosis and terminal differentiation. Methods: Oral IEN lesions were hemisected to obtain pretreatment diagnoses and establish baseline biochemical & molecular parameters. Dosing to the remaining lesional tissue (0.5 grams qid for 6 weeks) was initiated 1 week after the initial biopsy. RNA was isolated from snap frozen IEN lesions for microarray analyses, followed by QRT-PCR validation. Additional epithelial gene specific QRT-PCR analyses facilitated assessment of target tissue treatment effects. Surface epithelial COX2 and iNOS protein levels and microvascular densities were determined by image analysis quantified immunohistochemistry. Results: The topical berry gel application uniformly suppressed genes associated with RNA processing, growth factor recycling and inhibition of apoptosis. Histopathologic results showed: 35% decreased in grade, 20% increased in grade, and 45% retained stable disease. Treated lesional epithelium showed a significant reduction in the proinflammatory protein COX-2 (p<0.005) and a reduction (p=0.053) in iNOS levels (Wilcoxon sign rank test). Microarray and QRT-PCR analyses showed gene expression profile modifications towards a less biochemically active, more differentiated tissue. Conclusions: These data show that berry gel topical application modulated oral IEN gene expression profiles, and reduced epithelial COX2 and iNOS proteins. In a patient subset, berry gel application also reduced vascular density in the superficial connective tissues and induced genes associated with keratinocyte terminal differentiation. NIH/NCI R21 CA111210