ROBO1 deletion as a novel germline alteration in breast and colorectal cancer patients

Abstract

Despite one third of breast (BC) and colorectal cancer (CRC) cases having a hereditary component, only a small proportion can be explained by germline mutations. The aim of this study was to identify potential genomic alterations related to cancer predisposition. Copy number variations (CNVs) were interrogated in 113 unrelated cases fulfilling the criteria for hereditary BC/CRC and presenting non-pathogenic mutations in BRCA1, BRCA2, MLH1, MSH2, TP53, and CHEK2 genes. An identical germline deep intronic deletion of ROBO1 was identified in three index patients using two microarray platforms (Agilent 4x180K and Affymetrix CytoScan HD). The ROBO1 deletion was confirmed by quantitative PCR (qPCR). Six relatives were also evaluated by CytoScan HD Array. Genomic analysis confirmed a co-segregation of the ROBO1 deletion with the occurrence of cancer in two families. Direct sequencing revealed no pathogenic ROBO1 point mutations. Transcriptomic analysis (HTA 2.0, Affymetrix) in two breast carcinomas from a single patient revealed ROBO1 down-expression with no splicing events near the intronic deletion. Deeper in silico analysis showed several enhancer regions and a histone methylation mark in the deleted region. The ROBO1 deletion in a putative transcriptional regulatory region, its down-expression in tumor samples, and the results of the co-segregation analysis revealing the presence of the alteration in affected individuals suggest a pathogenic effect of the ROBO1 in cancer predisposition.

Contribution of rare germline copy number variations and common susceptibility loci in Lynch syndrome patients negative for mutations in the mismatch repair genes

“…Despite the CNVs harboring intronic regions being found in three of these CRC related‐genes ( ROBO1 , CADM1 and CNTNAP2 ), their potential pathogenic effect could not be excluded since intronic alterations may affect the splicing process leading to a loss of gene function, as previously reported for MLH1 and MSH2 genes . In addition, our group has previously detected identical ROBO1 intronic deletions (as presented in this study) in two cases of family breast cancer, providing further evidence for their potential pathogenicity …”

Germline copy number variations are associated with breast cancer risk and prognosis

“…Germline CNVs are a class of structural variations, and are defined as loss or gain of genomic DNA in size range of 50 bp to 1 Mb 14 . Germline CNVs are studied as genetic determinants for susceptibility for familial breast cancer 15 – 20 and also cancers of prostate 21 – 23 , ovary 18 , 24 – 26 , pancreas 27 – 29 , colon, rectum 16 , 30 – 34 , endometrium 35 , lung 36 – 38 and melanoma 39 , 40 .…”

Genetic susceptibility markers for a breast-colorectal cancer phenotype: Exploratory results from genome-wide association studies

“…From our data and analyses, the association signal for ROBO1 , comprised a cluster of 53 SNPs in intron 1, spanning 87.866 kb (79,703,548–79,791,414 hg 18) and the SNP closest to the deletion was located 675.51 kb away from the deletion. Furthermore, unlike the rare deletion identified by Villacis and colleagues [ 34 ], these were common SNPs with a minor allele frequency ranging between 0.39–0.46.…”

“…Evidence in support of ROBO1 as a susceptibility gene for a hereditary breast-colorectal cancer (HBCC) phenotype comes from a study by Villacis and colleagues [ 34 ]. The aim of their study was to identify genomic alterations (copy number variations, CNVs) related to cancer predisposition in patients with a suggestive HBCC phenotype who did not carry high risk mutations in the major genes known to be implicated in hereditary breast or colorectal cancer (i.e., patients who met HBCC criteria as defined by Naseem et al [ 4 ]).…”