Comparative genomic hybridisation and transcriptome microarray analysis in triple negative breast cancer: An INDIAN study.

Abstract

Triple-negative breast cancer (TNBC), which accounts for approximately 15–20% of all breast cancers, defined as lack of expression of estrogen receptor, progesterone receptor and Her-2 neu receptors. TNBC has two subtypes basal like and non-basal like, the former characterised by aggressive biology with limited therapeutic options. This study explored molecular markers involved in pathogenesis of TNBC and investigated novel potential diagnostic and therapeutic targets by CGH array and transcriptome array. aCGH analysis in TNBC demonstrated genes amplified were 3888, number of pathway hits was 1554 and major pathways amplified was found to be WNT signalling pathway and Cadherin signalling pathway. Among all metastatic sites and remission, activation of WNT signalling pathway is commonly observed. TNBC exhibited 1486 copy number variations (CNVR) which is approximately 250 times higher than controls. More than 20 CNVR was observed in all chromosomes and more than 80 CNVR was observed in Chr 1 to Chr 4, Chr 7, Chr 11 and Chr X. Common CNVR associated with amplified regions in Chr 22, Chr 14, Chr 8 and Chr 2 was observed in TNBC and CNVR associated with Chr 22q11.22–23, Chr 6p21.32–33, Chr11q12.2, Chr14q32.22–23, Chr 8p11.22–23, was observed in metastatic disease. In transcriptome array analysis a total of 11,359 differentially expressed genes with fold change 2.0 were in observed in TNBC comprise of 7639 upregulated genes and 3720 downregulated genes. Further, with fold change 10, 1526 upregulated genes and 839 down regulated genes were identified. Panther pathway analysis identified the main pathways of upregulated genes were Wnt signalling pathway, Integrin signalling pathway and Cadherin signalling pathway. The main pathways of down regulated genes were Inflammation mediated by chemokine and cytokine signalling pathways. PPI network shows that COL12A1, COL6A3, FN1, MMP3, WNT5A were key upregulated genes and ITGB7, PTPRC, ITGA4, LCK and CD247 were key down regulated genes. Cytoscape analysis followed by multiple list comparator tool identified top 5 significant hub genes were FN1, MMP3, COLL11A1, COL12A1 and COL3A1. The significant pathway genes obtained by CGH array and transcriptome array when compared, exhibited 5 common genes COL4A1, FN1, COL6A3, COL5A2 and PCDH7. These genes were not overexpressed in Controls and therefore involved in pathogenesis of TNBC. Expressions of these genes were validated by studying protein expression by immunohistochemistry. FN1 and COL6A3 protein over expression predicted worse DFS in TNBC and can be considered as therapeutic targets at diagnosis to reduce the disease metastases. These findings provide new insights into the pathogenesis of TNBC and guide for selection of targets related to diagnosis, prognosis and prediction of treatment in TNBC.