Despite recent advances in diagnosis and treatment, breast cancer remains the most common cause of cancer-related deaths among women globally. Most deaths from breast cancer are due to metastatic and recurrent disease that occurs years after primary tumor regression from treatment. Consequently, there is a pressing need to better understand the mechanisms that underly metastasis and therapy resistance. Recently, our laboratory performed whole-exome sequencing of both matched metastatic and primary tumors and unmatched metastatic tumors from breast cancer patients. We identified copy number (CN) gain of a 77.5kbp region encompassing the full exon sequence of PAQR8 that was enriched in the metastases compared to primary tumors. PAQR8 encodes a membrane progesterone receptor (mPR) that has never before been implicated in breast cancer metastasis or recurrence. Moreover, CN gain of this region was mutually exclusive of mutations in estrogen receptor (ESR1) and progesterone receptor (PGR) among patients who received anti- estrogen therapy. But intriguingly, it was found in equal frequency among patients who were treated with non- hormonal therapies as those with hormonal therapy. Plus, both CN gain and increased RNA levels of Paqr8 were identified in recurrent compared to primary tumors of several genetically engineered mouse (GEM) models of oncogene-driven breast cancers, none of which express hormone receptors. Given these findings, I hypothesize that PAQR8 gain promotes breast cancer metastasis and therapy resistance.
Aim 1 addresses whether PAQR8 gain promotes metastasis. I will orthotopically transplant an equal mixture of PAQR8- overexpressing and control cells into immunocompromised mice and track the formation of metastatic lesions by in vivo luciferase imaging. Comparing the ratio of the two different cell types in metastatic and primary lesions, I expect to find an enrichment for PAQR8-overexpressing cells in the metastases compared to primary tumors. Next, I will use immunofluorescence to detect active mediators predicted to be downstream of PAQR8 signaling in both primary and metastatic lesions.
Aim 2 addresses whether PAQR8 gain promotes therapy resistance. I will perform a recurrence free survival assay to determine whether mice injected with PAQR8- overexpressing cells recur earlier following treatment than those injected with control cells. To investigate the composition of residual and recurrent tumors, I will orthotopically inject a mixture of PAQR8-overexpressing and control cells as in aim 1. Primary tumors will be treated with targeted therapy or chemotherapy, and both residual lesions and recurrent tumors will be assayed for the ratio of PAQR8-overexpressing cells to controls. I expect PAQR8-overexpressing cells to be enriched in both the residual and recurrent tumors. Finally, an in vitro cell competition assay will determine whether PAQR8 overexpression mediates therapy resistance in a progesterone dose-dependent manner. Results from the proposed experiments will yield novel insight into the role of PAQR8 membrane progesterone receptor in breast cancer metastasis and therapy resistance.
Despite significant advances in the treatment of primary breast cancer, metastatic and recurrent breast cancer continues to be a major cause of death among women worldwide. Membrane progesterone receptors like PAQR8 have never before been implicated in breast cancer metastasis and therapy resistance, though hormones have long been known to be a major determining factor of prognosis and treatment. Completion of this project further advances our understanding of the mechanisms behind metastatic and recurrent breast cancer, and may potentially shed light on a novel therapeutic target.
