Abstract 3483: Inhibition of Monocarboxylate Transporter 4 (MCT4) targets stem-like cells in glioblastoma

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Glioblastomas (GBM) are the most common adult malignant brain tumors and contain a hypoxic/necrotic core surrounded by proliferative cells. To unmask genes important in hypoxia, we exposed 2 GBM neurosphere lines, HSR-GBM1 and JHH-GBM10, to 1% and 21% oxygen levels for 24 hours and compared gene expression using Agilent oligonucleotide microarrays. We identified SLC16A3 (Monocarboxylate transporter-4, MCT4) as one of the most upregulated genes in response to hypoxia. To investigate the clinical importance of MCT4 in GBM, we examined the Kaplan-Meier survival curves of glioma patients using public databases. We found that patients with upregulation of MCT4 (≥2.0X) have a significantly shorter survival (p<0.0001) than patients with intermediate expression. Consistent with this, MCT4 upregulation correlated with the aggressive mesenchymal subset of GBM (p<0.0001). We next examined MCT4 protein levels using immunohistochemical analysis of tissue microarrays, confirming that MCT4 protein levels were increased in high-grade as compared to lower grade astrocytomas (p<0.0001). These data clearly demonstrated that MCT4 is a clinically relevant target. We next tested the requirement of MCT4 in vitro. We found that when neurospheres were transduced with viruses encoding short hairpin RNA (shRNA) against MCT4, cell growth was inhibited by 30-69% in hypoxia. Interestingly, similar results were observed in normoxia, suggesting that MCT4 may be critical to tumor growth and survival independent of oxygen levels. We next explored the role that MCT4 may play in the maintenance of the stem-like population in our neurosphere lines. We found that MCT4 is overexpressed in sorted CD133-positive cells compared to CD133-negative cells in both HSR-GBM1 and JHH-GBM10. Furthermore, flow cytometric analysis of shMCT4-expressing neurospheres showed a decrease in the percentage of CD133-positive cells as compared to controls. This suggested to us that MCT4 silencing might inhibit stem-like cell proliferation/survival, offering a potential explanation for growth inhibition following MCT4 knockdown in normoxia. To test this hypothesis, we examined the effect of MCT4 inhibition functionally using a clonogenic assay. Cells were exposed to normoxia or hypoxia for 48 hours followed by recovery in normoxia. We found that MCT4 silencing resulted in significant reduction in sphere number and size in both normoxic and hypoxic cells, consistent with a loss of clonogenicity. In agreement with what we found in vitro, MCT4 silencing also slowed GBM intracranial xenograft growth in vivo (p=0.009). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3483. doi:1538-7445.AM2012-3483