Uncovering the Molecular Determinants of Metastatic Recurrence and Impaired NK Cell Function

PROJECT SUMMARY Our proposed research employs innovative, unbiased approaches and novel preclinical models to elucidate how disseminated HER2+ breast cancer cells cells survive in the brain microenvironment and initiate metachronous metastasis. Through functional characterization of phenotypically stable preclinical models of HER2+ breast cancer brain metastasis, we discovered latent/dormant (Lat) HER2+ cells display stem cell-like characteristics, downregulate immune activating sensors and survive in equilibrium with innate immune surveillance, while brain metastatic cells escape and metastasize. Moreover, metabolically distinct HER2+ brain-tropic Lat cells and metachronous brain metastatic (M-BM) cells are resistant to radiation and systemic HER2 targeted therapies. AXL, a member of the TAM (TYRO3, AXL, MERTK) receptor tyrosine kinase family is enriched in Lat and M-BMs. AXL expression is enriched in metachronous brain metastatic lesions compared to matched primary tumors from HER2+ breast cancer patients. Of note, AXL is predominantly nuclear in these brain metastatic lesions and in our preclinical brain metastatic model systems. CRISPR affinity purification of in situ regulatory elements revealed enrichment of TEAD transcription factor at the AXL promoter region in Lat and M-BMs. AXL immunoprecipitation-mass spectrometry analysis identified WRN Helicase Interacting Protein 1 (WRNIP1) among other nuclear proteins that interact with AXL. WRNIP1 aids maintenance of genomic stability under replicative stress and promotes survival of Lat and M-BM cells. Depletion of AXL in Lat and M-BM cells or administration of small molecule AXL inhibitor (BGB324) results in attenuated metastasis initiating capacity. Noticeably, increased AXL expression and reduced cytotoxicity was also observed in tumor trained/exposed NK cells, while administration of BGB324 to augmented cytotoxicity. Thus, our central hypothesis is therapy resistant brain-tropic HER2+ breast cancer cells are dependent on nuclear AXL signaling response for survival and membranous AXL expression in NK cell results in dysfunction. The proposed aims will delineate how TEAD signaling response promotes brain metastasis and assess the impact of AXL inhibitors in limiting tumor cell survival and reactivating NK mediated innate immune surveillance.