Natural Compounds Can Target SHH Signaling To Block MED12 Mutant Breast Cancer Oncogenesis

Abstract

Current statistics show that breast cancer is the second leading cause of cancer deaths among American women. Therefore, there is a strong need to find improved treatment strategies for women suffering from this lethal disease. Though radiation and chemotherapy have been the frontline choice of treatment for cancer over the past few decades, due to the detrimental side effects, personalized treatment is rapidly rising as a superior treatment method. This is largely due to recent advances in genome‐wide DNA sequencing that have allowed for the identification of cancer‐related mutational landscapes. Through these studies, it has been found that up to 67% of breast cancer tumors carry a mutation in the Mediator subunit MED12 thus indicating that MED12 likely has a critical tumor suppressive role in breast cancer. Previous results from our lab, and others, have indicated that MED12 plays a critical role in restricting GLI3‐dependent SHH signaling. This finding is of particular interest to this study since hyper‐activated SHH signaling is known to play a major role in promoting breast cancer oncogenesis. Due to these findings, we hypothesize that mutations in MED12 cause hyper‐activated SHH signaling in breast cancer to promote oncogenesis and, furthermore, that natural compounds could provide useful as a personalized treatment for MED12 mutant breast cancer. To study the effect of mutant MED12, a lentivirus carrying an shRNA against MED12 was generated and infected into MCF‐7 cells. Since the vast majority of breast cancer‐associated MED12 mutations lead to loss of protein function, the knockdown strategy through lentiviral shRNA is assumed to mimic the MED12 mutant setting. Proliferation assays were utilized to confirm that downregulated MED12 plays a role in the increased proliferation of breast cancer cells. Next, quantitative PCR was performed to determine the effect of downregulated MED12 on the expression of genes that are known to be regulated by GLI3. Finally, a screening strategy was employed by using a natural compound library to find potential novel treatment strategies for MED12 mutant breast cancer. Our findings confirmed that downregulation of MED12 increases proliferation of breast cancer cells in vitro thereby providing a strong argument that mutant MED12 would have the same effect. Through quantitative PCR it was confirmed that GLI3 target genes are upregulated in cancer cells when MED12 expression is low. Importantly, our natural compound screen identified several novel therapeutic compounds that specifically target MED12 downregulated breast cancer cells through a mechanism that involves SHH signaling. We therefore uncovered that MED12 mutations promote GLI3‐dependent SHH signaling in breast cancer and, importantly, we identified potential novel therapeutic strategies for MED12 mutant breast cancer patients.