Abstract
Expression of estrogen receptor alpha and beta (ER-α and ER-β) in the skin contributes to wound healing, hair follicle morphogenesis, and non-melanoma skin cancer development. A recently identified ER-α variant known as ER-α36 increases the growth of gastric, endometrial, and ER-negative breast cancers, although its function in cutaneous squamous cell carcinoma (SCC) has not been reported. An inhibitor of ER-α36 named icaritin, targets breast cancer stem cells and is in clinical trials for breast cancer and hepatocellular carcinoma. We hypothesized that ER-α36 signaling expands the cancer stem cell (CSC) population in cutaneous SCC to promote tumor growth, such that targeting of ER-α36 with icaritin will inhibit skin cancer growth. Immunohistochemistry for ER-α36 in a set of human and mouse skin and skin cancers revealed increased levels of ER-α36 expression in SCCs. Silencing of ER-α36 in tumor sphere assay used as a measure of the CSC/progenitor population, decreased the size and number of spheroids by 61% and 46%, respectively, and decreased expression of the CSC markers aldehyde dehydrogenase (ALDH1) and cytokeratin 19 (KRT19) in human SCC cell line SCC13. Conversely, over-expression of ER-α36 resulted in a 3.5-fold increase in the number of spheroids and reduced differentiation. Treatment of SCC13 cells with icaritin at 1 µM, a concentration that did not impact the growth of SCC13 cells in monolayer, reduced both the size and number of spheres, as well as CSC marker expression. Xenografts of SCC13 cells in immunocompromised NCG mice were treated i.p. twice per week with icaritin or vehicle. By 6 weeks of treatment, icaritin-treated tumors were 55% smaller than the vehicle-treated tumors. Histological and immunofluorescence analyses of these tumors revealed that the icaritin-treated tumors had reduced cell proliferation, increased differentiation, and decreased phospho-EGFR activity. Taken together, our results demonstrate a role for ER-α36 in skin cancer stem/progenitor cells and suggest that targeting of ER-α36 with icaritin may be a useful strategy for SCC treatment.