Early detection of squamous cell carcinoma in carcinogen induced oral cancer rodent model by ratiometric activatable cell penetrating peptides

•MMP sensitive RACPP-PLGC(Me)AG detects SCCA in an immune-competent carcinogen induced oral cancer mouse model.•RACPP-PLGC(Me)AG identifies cancer with excellent sensitivity and specificity.•RACPP-PLGC(Me)AG detects small cancerous lesions that are not visible by white light examination.•RACPP may be useful for intraoperative as well as non-invasive identification of cancer. Ratiometric cell-penetrating-peptides (RACPP) are hairpin-shaped molecules that undergo cleavage by tumor-associated proteases resulting in measurable Cy5:Cy7 fluorescence ratiometric change to label cancer in vivo. We evaluated an MMP cleavable RACPP for use in the early detection of malignant lesions in a carcinogen-induced rodent tumor model. Wild-type immune-competent mice were given 4-nitroquinoline-oxide (4NQO) for 16weeks. Oral cavities from live mice that had been intravenously administered MMP cleavable PLGC(Me)AG-RACPP were serially imaged from week 11 through week 21 using white-light reflectance and Cy5:Cy7 ratiometric fluorescence. In an initial study we found that at week 21 nearly all mice (13/14) had oral cavity lesions, of which 90% were high-grade dysplasia or invasive carcinoma. These high-grade lesions were identifiable with white light reflectance and RACPP Cy5:Cy7 ratiometric fluorescence with similar detectability, Area Under Curve (AUC) for RACPP detection was 0.97 (95% Confidence interval (CI)=0.92–1.02, p