Co-targeting BET bromodomain BRD4 and RAC1 suppresses growth, stemness and tumorigenesis by disrupting the c-MYC-G9a-FTH1axis and downregulating HDAC1 in molecular subtypes of breast cancer.

BET bromodomain BRD4 and RAC1 oncogenes are considered important therapeutic targets for cancer and play key roles in tumorigenesis, survival and metastasis. However, combined inhibition of BRD4-RAC1 signaling pathways in different molecular subtypes of breast cancer including luminal-A, HER-2 positive and triple-negative breast (TNBC) largely remains unknown. Here, we demonstrated a new co-targeting strategy by combined inhibition of BRD4-RAC1 oncogenic signaling in different molecular subtypes of breast cancer in a context-dependent manner. We show that combined treatment of JQ1 (inhibitor of BRD4) and NSC23766 (inhibitor of RAC1) suppresses cell growth, clonogenic potential, cell migration and mammary stem cells expansion and induces autophagy and cellular senescence in molecular subtypes of breast cancer cells. Mechanistically, JQ1/NSC23766 combined treatment disrupts MYC/G9a axis and subsequently enhances FTH1 to exert antitumor effects. Furthermore, combined treatment targets HDAC1/Ac-H3K9 axis, thus suggesting a role of this combination in histone modification and chromatin modeling. C-MYC depletion and co-treatment with vitamin-C sensitizes different molecular subtypes of breast cancer cells to JQ1/NSC23766 combination and further reduces cell growth, cell migration and mammosphere formation. Importantly, co-targeting RAC1-BRD4 suppresses breast tumor growth in vivo using xenograft mouse model. Clinically, RAC1 and BRD4 expression positively correlates in breast cancer patient's samples and show high expression patterns across different molecular subtypes of breast cancer. Both RAC1 and BRD4 proteins predict poor survival in breast cancer patients. Taken together, our results suggest that combined inhibition of BRD4-RAC1 pathways represents a novel and potential therapeutic approach in different molecular subtypes of breast cancer and highlights the importance of co-targeting RAC1-BRD4 signaling in breast tumorigenesis via disruption of C-MYC/G9a/FTH1 axis and down regulation of HDAC1.

Estrogen signaling differentially alters iron metabolism in monocytes in an Interleukin 6-dependent manner.

The ability of monocytes to release or sequester iron affects their role in cancer and inflammation. Previous work has shown that while IL-6 upregulates hepcidin synthesis and enhances iron sequestration, E2 reduces hepcidin synthesis and increases iron release. Given that E2 upregulates IL-6 production in monocytes, it is likely that the exact effect of E2 on iron metabolism in monocytes is shaped by its effect on IL-6 expression. To address this issue, the expression of key iron regulatory proteins was assessed in E2-treated U937, HuT-78, THP-1 and Hep-G2 cells. Iron status was also evaluated in U937 cells treated with the ERα agonist PPT, the ER antagonist ICI-182780, dexamethasone + E2, IL-6 + E2 and in IL-6-silenced U937 cells. E2 treatment reduced hepcidin synthesis in HuT-78, THP-1 and Hep-G2 cells but increased hepcidin synthesis and reduced FPN expression in U937 cells. E2-treated U937 cells also showed reduced HIF-1α and FTH expression and increased TFR1 expression, which associated with increased labile iron content as compared with similarly treated Hep-G2 cells. While treatment of U937 cells with interleukin 6 (IL-6) resulted in increased expression of hepcidin, dexamethasone treatment resulted in reduced hepcidin synthesis relative to E2- or dexamethasone + E2-treated cells; IL-6 silencing also resulted in reduced hepcidin synthesis in U937 cells. Lastly, while iron depletion resulted in increased cell death in U937 cells, E2 treatment resulted in enhanced cell survival and reduced apoptosis. These findings suggest that E2 differentially alters iron metabolism in monocytes in an IL-6 dependent manner.