Quantifying in Vivo Collagen Reorganization During Immunotherapy in Murine Melanoma with Second Harmonic Generation Imaging

Significance:Increased collagen linearization and deposition during tumorigenesis can impede immune cell infiltration and lead to tumor metastasis. Although melanoma is well studied in immunotherapy research, studies that quantify collagen changes during melanoma progression and treatment are lacking. Aim:Image in vivo collagen in preclinical melanoma models during immunotherapy and quantify the collagen phenotype in treated and control mice. Approach:Second harmonic generation imaging of collagen was performed in mouse melanoma tumors in vivo over a treatment time-course. Animals were treated with a curative radiation and immunotherapy combination. Collagen morphology was quantified over time at an image and single fiber level using CurveAlign and CT-FIRE software. Results:In immunotherapy-treated mice, collagen reorganized toward a healthy phenotype, including shorter, wider, curlier collagen fibers, with modestly higher collagen density. Temporally, collagen fiber straightness and length changed late in treatment (Day 9 and 12) while width and density changed early (Day 6) compared to control mice. Single fiber level collagen analysis was most sensitive to the changes between treatment groups compared to image level analysis. Conclusions:Quantitative second harmonic generation imaging can provide insight into collagen dynamics in vivo during immunotherapy, with key implications in improving immunotherapy response in melanoma and other cancers.