Introduction: Meta-analysis shows women with diabetes have a 20% increased risk of breast cancer. Diabetes increases the risk of breast cancer mortality. Additionally, patients with breast cancer and preexisting diabetes have an increased risk for distant metastasis compared with non-diabetic counterparts. The molecular mechanisms for distant metastasis in breast cancer patients with diabetes are not very well understood. Our preliminary data suggested that hyperglycemia enhances stem cell activity in triple negative breast cancer (TNBC) cells. Here, we report miR-424 to be a key regulator of breast cancer stem cell pool dynamics under hyperglycemic environment.

Materials and Methods: MDA-MB-231, TNBC cell line was maintained under euglycemic (5mM) and hyperglycemic (10mM) culture conditions. RT-PCR based microRNA array was performed followed by validation of significantly altered microRNA to confirm the effect of hyperglycemia on the global microRNA profile. Flow cytometric analysis for CD44+/CD24- was performed to assess the alterations in breast cancer stem cell population. In addition, sphere-forming assay were also performed. Mir-424 over-expressing or knocked down cell lines were established from parental MDA-MB-231 cells and maintained in hyperglycemic or euglycemic condition respectively. Promoter analysis for PR/SET domain 14 (prdm14) was done. Western blotting and immunofluorescence assays were performed for cdc42 (cell division control 42), pStat5 (phospho signal transducer and activator of transcription factor 5) and prdm14.

Results: Loss of miR-424 in TNBC cells under euglycemic conditions led to enhanced stem cell activity; whereas over expression of miR- 424 under hyperglycemic conditions resulted in suppressed stem cell activity. Introduction of cdc42, (a miR-424 target gene) in miR-424 over expressing TNBC cells under hyperglycemic conditions leads to increased stem cell activity demonstrating the significance of miR-424-cdc42 signaling in hyperglycemia. Over-expression of miR-424 in TNBC dramatically reduced its metastatic abilities in vivo. Mechanistically, we found that miR-424-cdc42 signaling in hyperglycemia promotes Prdm14 activation, a stem cell regulator through increased phosphorylation of Stat5.

Conclusions: Our findings establish a key molecular signaling cascade (miR-424→cdc42→prdm14) that promotes TNBC stem cell activity under hyperglycemic conditions.

Citation Format: Sushmita B. Nandy, Alexis Orozco, Gautham Prabhakar, Viktoria Stewart, Stephanie Jones, Paloma Munoz, Ramadevi Subramani, Diego Pedroza, Rajkumar Lakshmanaswamy. miR-424-cdc42, key signaling axis in hyperglycemic regulation of stemness in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1461. doi:10.1158/1538-7445.AM2017-1461