Pharmacogenetics, molecular correlates and multiparameter high throughput evaluation of the response of inflamatory breast cancer cells to anticancer agents

4637 Inflammatory breast cancer (IBC) is an aggressive, fast-growing and highly invasive tumor, and therapeutically has the worst outcome among breast cancers. Understanding the molecular and genetic determinants of IBC therapeutic response and the identification of novel agents and strategies for treating IBC are thus urgently needed. Using two IBC cell lines, SUM 149 and SUM 159, we optimized a multi-parameter high throughput system for both cell- and target-based screening of anticancer agents against IBC cells. The target-based screen utilizes either IBC cell extracts or the purified target, while the cell-based screening simultaneously evaluates cell viability and apoptotic response in a single assay. We used the system to determine the growth kinetics and to characterize the differential sensitivity of mechanistically different anticancer agents (cisplatin, carboplatin, adriamycin, paclitaxel and vinorelbine) against the two IBC cell lines. The growth rate of SUM 159 (8.5 x 10-2.Hr-1) was approximately 2-fold that of SUM 149. At clinically relevant concentrations, both cell lines showed the highest sensitivity to cisplatin and the highest resistance to vinorelbine, when tested as single agents. Overall, the IC50 of all five agents were higher for SUM 159 than SUM 149. Because previous studies have shown that expression of GSTP1, a phase II metabolizing enzyme and regulator of cellular signaling in response to stress, is a strong determinant of therapeutic outcome and survival in breast and other cancers, we examined the expression of the GSTP1 gene by real time quantitative PCR and western blotting. Single nucleotide polymorphisms in GSTP1 and three other GST genes (GSTM1, GSTM3 and GSTT1) were determined using an allele-specific TaqMan RT-PCR assay. The results showed SUM149 to have a lower level of GSTP1 expression and to lack the Ala‡Val transition in exon 6 of the GSTP1 gene. In addition, unlike SUM159, which is homozygous for GSTM1*B, the GSTM1 gene was deleted in SUM149. These genotypes and phenotype are consistent with the higher drug sensitivity of SUM149 compared to SUM 159. The pharmacogenetic differences between the cell lines correlated with differential effects of siRNA-mediated GSTP1 suppression on their drug sensitivity. Screening of the cell lines against GSTP1 inhibitors is ongoing. Our data and the high throughput target- and cell-based screening system we describe provide a useful tool for the discovery of novel targeted agents and for rapidly evaluating novel combination therapy strategies for IBC. The system is suitable for screening engineered cells and cells with specific molecular defects and should complement in vivo systems in the rational drug discovery of novel IBC therapeutics. Supported by RO1 CA90462, P50 CA69438 and the Cancer Center Core Grant P30 CA124322.