Abstract
Many if not all tumors are highly heterogeneous, consisting of several populations of cells including cancer stem cells (CSCs). CSCs, functionally defined by aldehyde dehydrogenase (ALDH) activity and the expression of various cell surface markers, are a small population of tumor initiating cells that display several characteristics common of normal stem cells. CSCs, for example, can undergo self-renewal, as well as form tumors by differentiating into other cancer cell types. CSCs have also been shown to possess various genetic and cellular adaptations that make them resistant to conventional cytoreductive therapies. Radiation and chemotherapy, therefore, effectively debulk tumors by killing proliferating cells (non-CSCs), but leave behind the quiescent CSCs; thereby promoting tumor metastasis and recurrence. CSCs haven't yet been identified in all tumor types, but their presence in several cancers indicates an urgent need to develop a therapy regimen that specifically targets CSCs so that the survival of patients with metastatic or recurrent disease can be improved. As such, the overall purpose of this project was to investigate whether combining radiation therapy or proteasome inhibition with direct NK cell immunotherapy is an efficient strategy to target the CSC population within Ewing's sarcoma (A673), breast (MDA-231 ), and pancreatic cancers (P ANC-1 ). Cell surface marker expression and ALDH activity, assessed via flow cytometry and the aldefluor assay, respectively, were the sole criteria used to identify CSCs within the A673, MDA-231, and PANC-1 cell lines. In agreement with other published findings, the results of this project showed that NK cells, cytotoxic lymphocytes that function as part of the innate immune system, can preferentially kill the CSC population within each cell line. Our preliminary data on brain tumor CSCs showed that this preferential killing is likely mediated through NKG2D, DR5, and FasL. Indeed, we found that irradiating the tumor cells not only enriches for the CSC population, but also causes a selective upregulation ofMICA/B (NKG2D ligands), DR5, and FasL on CSCs relative to non-CSCs. We observed a similar effect after culturing the tumor cells in various concentrations of the proteasome inhibitor bortezomib. As such, both irradiation and proteasome inhibition sensitize the CSCs to NK cell attack in vitro and can therefore be used in combination with NK cell immunotherapy to specifically target the CSCs within residual tumors; thereby significantly, if not completely, reducing the risk of tumor metastasis and recurrence.