Functional Characterisation of the 19q12 Amplicon in Grade 3 Breast Cancers.
Rachael Natrajan, Daniel Wetterskog, Maryou B Lambros, Felipe C Geyer, Alan Mackay, Radost Vatcheva, Jorge S Reis-Filho. Institute of Cancer Research, London, United Kingdom
Background: Grade III (GIII) invasive ductal carcinomas of no special type (IDC-NST) comprise up to 60% of all invasive breast cancers and have an aggressive clinical behavior. Amplification of 19q12 is found in a subgroup of estrogen receptor (ER)-negative HER2 amplified (4%) and basal-like (16%) subtypes of breast cancer. This amplicon comprises 9 genes, including cyclin E1 (CCNE1), which has been proposed as its driver due to a strong association with mRNA over-expression. The aims of this study were to identify functionally the genes within the 19q12 amplicon whose expression is required for the survival of cancer cells harboring this amplification.
Design: We analysed a series of 297 frozen breast cancers with high-resolution microarray CGH to assess the frequency of 19q12 amplification. A subset of 48 GIII micro-dissected IDC-NST were subjected to mRNA expression arrays, and data integrated with aCGH to find those genes that were significantly over-expressed when amplified. Breast cancer cell lines with (MDAMB157 and HCC1569) and without (Hs578T, MCF7, MDAMB231 and ZR75.1) 19q12 amplification were screened with an RNAi library targeting the genes mapping to the smallest region of amplification on 19q12. Individual short interfering RNA (siRNA) SMARTpools targeting these genes were plated in 96-well plates and cell viability was assessed after 9 days using CellTiter-Glo Luminescent Cell viability assay.
Results: We identified 19q12 amplification in 5% of breast cancers, which was significantly associated with high grade and ER negative tumors. Of the 9 genes mapping to the smallest region of amplification, UQCRFS1, POP4, C19ORF12, CCNE1 and C19ORF2 were significantly over-expressed when amplified (P<0.05). Silencing of POP4, PLEKHF1, CCNE1 and ZNF537 selectively inhibited cell viability in 19q12-amplified cells compared to control cells. Over-expression of CCNE1 in amplified cells was found to confer sensitivity to doxorubicin, cisplatin and paclitaxel.
Conclusions: Expression of POP4, PLEKHF1, CCNE1 and ZNF537 are required for the survival of cancer cells displaying their amplification and suggest that the 19q12 amplicon may harbor more than one 'driver'. Furthermore identification of tumors with CCNE1 amplification may identify a subgroup of patients that are responsive to common chemotherapy agents.
Tuesday, March 1, 2011 1:00 PM
Poster Session IV # 20, Tuesday Afternoon