SIVA mutation, a candidate metastasis gene identified from clonally related bilateral breast cancers, promotes breast cancer cell aggressiveness in vitro and in vivo

Our purpose is to identify candidate genes involved in the early steps of breast cancer metastasis and examine their pro-invasive functions both in vitro and in vivo. A percentage of bilateral breast cancers were clonally related based on copy number variation profiling. Whole exome sequencing and comparative sequence analysis revealed that a limited number of somatic mutations were acquired in this “breast to breast” metastasis. These mutations might promote breast cancer distant spread. The pro-invasive functions of a candidate metastasis gene were assessed in vitro by its abilities to promote proliferation, migration and invasion and in vivo as tumor xenografts in immunocompromised mice or a syngeneic orthotopic mouse breast cancer model. RNAseq analysis was performed to probe the transcription programs modulated by this candidate metastasis gene. SIVA1-D160N was one somatic mutation acquired in the breast to breast metastasis. Over-expression of SIVA1-D160N promoted migration and invasion of human MB-MDA-231 breast cancer cells in vitro, consistent with a dominant negative interfering function. When introduced via tail vein injection, 231 cells over-expressing SIVA1-D160N displayed enhanced distant spread on IVIS imaging. Over-expression of SIVA1-D160N promoted anchorage independent growth of mouse 4T1 breast cancer cells in vitro. When introduced orthotopically via mammary fat pad injection in syngeneic Balb/c mice, over-expression of SIVA1-D160N in 4T1 cells increased mammary gland tumor growth as well as liver metastasis. We conclude clonally related bilateral breast cancers represent a novel system to investigate metastasis and revealed a role of SIVA1-D160N in breast cancer metastasis. Overall design: Differential gene expression analysis and pathway enrichment analysis of OVCAR8, SKOV3, HCC1954, and MDA-MB-231 parental cell lines as compared to their SIVA1-WT or SIVA1-D160N transformed counterparts.