Identification of pro-metastatic pathways and molecules in mouse models with reduced expression or signaling of the insulin-like growth factor type 1 receptor

We performed single-cell RNA sequencing of multiple IGF-1R loss-of function mouse mammary tumor models to uncover how IGF-1R signaling regulates intrinsic epithelial cell signaling to suppress metastasis. We identify key pathways necessary for promoting metastasis, determined IGF-1R is required to maintain a metastatic suppressive tumor microenvironment and demonstrate that attenuated epithelial IGF-1R signaling in the MMTV-Wnt1 mouse tumor model is sufficient for metastatic invasion. We further show that adherence between luminal and basal tumor cells is necessary for tumor growth at the secondary site and that reduced IGF-1R signaling in tumor epithelial cells inhibits secondary tumor epithelial cell growth due to dysregulated E-cadherin and P-cadherin and loss of cell-cell adhesion. Overall design: To test how loss of IGF-1R alters the primary tumor phenotype, we made use of two distinct mouse models. In one model IGF-1R function is reduced through expression of an MMTV-driven dominant-negative IGF-1R transgene in the MMTV-Wnt1 (Wnt1) basal-like breast cancer tumor model (DN-Wnt1). The second model has a mammary luminal epithelial-specific (Keratin 8-Cre) conditional Igf1r knockout (K8iKOR) in the MMTV-Wnt1 tumor mouse line Deletion of the Igf1r in the K8iKOR line was initiated by tamoxifen injections into 8-week old female mice to avoid alterations in pubertal mammary gland development. These two lines were compared to MMTV-Wnt1 tumors with intact IGF-1R. Tumors were identified in mice between 12 weeks- 1 year of age (latency dependent on genotype and stochastic tumor development) by palpation and were harvested and dissociated for single cell RNA-Seq analysis when they reached 1.5 cm2. Three tumors (one per genotype) were subjected to initial scRNA-Seq. Additional tumors from each genotype were similarly harvested for validation studies.