A Functional Role for CERT in Cancer Drug Induced Autophagy and Prognosis in Her2 Positive Breast Cancer.

Abstract

Abstract CERT was recently identified in a functional genomic siRNA screen for regulators of multi-drug sensitivity. CERT encodes a ceramide transporter which shuttles the pro-apoptotic lipid ceramide from the endoplasmic reticulum to the golgi. Silencing CERT, promoted sensitivity to paclitaxel, doxorubicin, cisplatin and 5-FU in breast, colon and non-small cell lung cancer cell lines. Further analysis of CERT expression in 18 NCI-60 cancer cell lines together with drug sensitivity data for over 5000 drugs within the NCI database, revealed that elevated CERT expression is associated with multi-drug resistance to diverse cancer cytotoxics. CERT mRNA was significantly elevated in HER2 positive breast cancer cell lines. Consistent with these data, in a primary breast cancer tissue microarray cohort of 356 primary breast cancers with outcome data, CERT protein expression was significantly greater in Her2 positive disease (p<0.0001). Survival analysis indicates that increased CERT expression correlates with significantly poorer long-term survival across the whole cohort and in Her2 positive patients.In order to explore the downstream molecular targets of CERT activity, we used the Affymetrix GeneChip Exon Array to detect gene expression changes following siRNA mediated CERT silencing. Genes that were differentially expressed following CERT silencing were identified and validated by real-time PCR. Significant representation of genes involved in the autophagy pathway were identified. One of these genes, LAMP2, a protein involved in autophagosome-lysosome fusion, is up-regulated following CERT silencing. LAMP2 silencing promotes paclitaxel resistance when depleted from cancer cells. Consistent with a role for CERT silencing in mediating an autophagy response following drug exposure, CERT depletion together with Paclitaxel treatment induces alterations in the autophagy response marker-LC3 similar to those seen in starvation-induced autophagy and an increase in lysosomal activity as visualised by microscopy. These data indicate a role for autophagy in regulating cytotoxic drug response in cancer.The pharmacological targeting of CERT, a poor prognostic marker in breast cancer, may be a rational approach to enhance drug response and patient outcome. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 1137.