Supplementary methods and Figures S1-S7 from Nintedanib Is a Highly Effective Therapeutic for Neuroendocrine Carcinoma of the Pancreas (PNET) in the Rip1Tag2 Transgenic Mouse Model

Supplementary methods and Figures S1-S7. Supplementary Methods. Detailed description of reagents and protocols used for immunohistochemical and metastasis analysis, and immunofluorescence microscopy analysis. Supplementary Figure S1. Histopathological staging of insulinoma development in Rip1Tag2 transgenic mice (A); quantification of tumor cell apoptosis (B) and tumor cell proliferation (C) in nintedanib and vehicle-treated Rip1Tag2 mice; determination of the IC50 of nintedanib in repressing cultured insulinoma cells (D). Supplementary Figure S2. Quantification of the number of NG2-positive pericytes (A) and the percentage of perivascular cells not associated with blood vessels (B) in tumors of nintedanib and vehicle-treated Rip1Tag2 mice. Supplementary Figure S3. Quantification and localization of hypoxic areas in tumors of nintedanib and vehicle-treated Rip1Tag2 mice (A-D). Supplementary Figure S4. Quantification of microvessel density (A), tumor volumes (B) and tumor grading (C) in tumors of nintedanib, PTK/ZK, sunitinib and vehicle-treated Rip1Tag2 mice. Supplementary Figure S5. Quantification of liver metastasis in mice treated open-end with nintedanib. Supplementary Figure S6. Quantification of microvessel density (A, C) and tumor volumes (B, D) of tumors from Rip1Tag2 mice treated for 5 days with nintedanib (A, B) or for 3 weeks with sunitinib (C, D). Supplementary Figure S7. Quantification of microvessel density (A, C) and volumes (B, D) of tumors from Rip1Tag2 mice treated with PTK/ZK following the onset of tumorigenesis for 3, 4, 5, and 6 weeks (A, B) or for 5 days at a later stage of tumor development (C, D).