Abstract P3-06-35: Topoisomerase 1 gene copy aberrations in 52 human breast cancer cell lines and association to gene expression
Authors: J Stenvang, M Smid, S Nielsen, M Timmermans, M Rømer, D Nielsen, J Foekens, N Brünner, J Martens
Journal: Cancer Research (2012)

Abstract Background: Topoisomerase 1 (TOP1) targeting drugs, e.g. irinotecan, are included in treatment regimens for different cancer types. Two clinical phase II studies1,2 suggested a role for irinotecan in the treatment of metastatic breast cancer patients being refractory to anthracyclines and taxanes. Access to predictive biomarker for irinotecan would increase response rates and secure that only patients with a high likelihood of obtaining benefit will receive the drug. However, no predictive biomarkers for TOP1 targeting drugs are available. Experimental design: A TOP1/CEN-20 fluorescence in situ hybridization (FISH) probe mix3 (Dako A/S, Denmark) was applied to analyze formalin fixed paraffin embedded (FFPE) tissue sections from 52 human breast cancer cell lines4. The FISH data were correlated to the gene expression levels (Affymetrix u133a GeneChip) of ER, HER2, TOP2A and molecular subtypes. Furthermore, gene expression data from 8 TOP1 high copy and 8 TOP1 low copy cell lines were applied for Kegg pathway analysis. Results: To categorize the cell lines, we initially analyzed the TOP1 copy number and CEN-20 counts in 100 normal breast epithelium sections and defined the normal diploid range as the mean plus/minus 0.5 times the haploid gene copy number. Thereby the normal diploid range for TOP1 was defined as 1.7 (range 1.45–1.90) and for CEN-20 it was 1.6 (range 1.38–1.82). By applying these cut-off values we found that among the 52 breast cancer cell lines 61.5% had TOP1 gene gains and 59.6% had increased CEN-20 copy numbers. However, the Pearson correlation between TOP1 and CEN-20 counts was 0.54 resulting in only 11.5% of the cell lines having TOP1/CEN-20 ratios &amp;gt; 1.5. A significant association exist between TOP1 gene copy number (FISH) and TOP1 copy numbers obtained from SNP array analysis (p &amp;lt; 0.0001). Furthermore, a significant correlation between TOP1 gene copy number as determined by FISH and TOP1 mRNA was observed (Spearman correlation coefficient 0.5, p = 0.0012). This suggests that the TOP1 gene is actively transcribed. No significant correlations were observed between TOP1 copy number and ESR1, TOP2a and HER2 gene expression or between TOP1 copy numbers and breast cancer subtypes. Using Kegg pathway analysis, TOP1 copy numbers were found to be significantly associated with mismatch repair and DNA replication (multiple testing corrected p-value 0.016 and 0.018, respectively). Conclusion: This study shows that TOP1 DNA aberrations are a frequent finding in human breast cancer cell lines and that these aberrations are not associated with ERS1, HER2 and TOP2a gene expression or breast cancer subtypes. Moreover, TOP1 gene copy number was strongly associated with mismatch repair pathways. This study suggests that TOP1 copy numbers may be applicable as predictive biomarker for irinotecan-response. The potential predictive value of TOP1 FISH is currently being tested in the cell lines and in a clinical phase II study with metastatic breast cancer patients refractory to anthracyclines and taxanes. References 1. Perez EA et al. J Clin Oncol. 2004, 22(14):2849–55. 2. Lee KS et al. Invest New Drugs. 2012 May 5. 3. Rømer MU et al. Scand J Gastroenterol. 2012, 47(1):68–79. 4. Hollestelle A et al. Breast Cancer Res Treat. 2010, 121(1):53–64. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P3-06-35.