Rectal cancer profiling identifies distinct subtypes in India based on age at onset, genetic, epigenetic and clinicopathological characteristics

Abstract

Rectal cancer is a heterogeneous disease that develops through multiple pathways characterized by genetic and epigenetic alterations. India has a comparatively higher proportion of rectal cancers and early-onset cases. We analyzed genetic (KRAS, TP53 and BRAF mutations, and MSI), epigenetic alterations (CpG island methylation detection of 10 tumor-related genes/loci), the associated clinicopathological features and survival trend in 80 primary rectal cancer patients from India. MSI was detected using BAT 25 and BAT 26 mononucleotide markers and mutation of KRAS, TP53, and BRAF V600E was detected by direct sequencing. Methyl specific polymerase chain reaction was used to determine promoter methylation status of the classic CIMP panel markers (P16, hMLH1, MINT1, MINT2, and MINT31) as well as other tumor specific genes (DAPK, RASSF1, BRCA1, and GSTP1). MSI and BRAF mutations were uncommon but high frequencies of overall KRAS mutations (67.5%); low KRAS codon 12 and a novel KRAS G15S mutation with concomitant RASSF1 methylation in early onset cases were remarkable. Hierarchical clustering as well as principal component analysis identified three distinct subgroups of patients having discrete age at onset, clinicopathological, molecular and survival characteristics: (i) a KRAS associated CIMP-high subgroup; (ii) a significantly younger MSS, CIMP low, TP53 mutant group having differential KRAS mutation patterns, and (iii) a CIMP-negative, TP53 mutated group. The early onset subgroup exhibited the most unfavorable disease characteristics with advanced stage, poorly differentiated tumors and had the poorest survival compared to the other subgroups. Genetic and epigenetic profiling of rectal cancer patients identified distinct subtypes in Indian population.

Targeted sequencing-based analyses of candidate gene variants in ulcerative colitis-associated colorectal neoplasia

“…Moreover, little is known about the molecular alterations aiding the malignant transformation in UC patients in Indian population. Recently, few reports have highlighted close association of molecular signature patterns identified in CRC in India and that of the western population (Gupta et al , 2010; Laskar et al , 2015). In colorectal carcinoma, the malignant transformation occurs by acquiring series of driver mutations in sequential manner.…”

“…At present, little is known about the molecular alterations that govern the colitis-derived neoplasia in several population. UC-associated CRC studies have shown sequential mutations in the KRAS, BRAF and TP53 genes in an Indian population (Shivakumar et al , 2012; Laskar et al , 2015). Previously, our group has reported few molecular signatures using conventional Sanger sequencing, PCR-RFLP and multiplex PCR (Shivakumar et al , 2012).…”

Exome sequencing identifies ARID2 as a novel tumor suppressor in early-onset sporadic rectal cancer

“…With several decades of CRC molecular biology research and more than a decade of studies involving targeted and whole exome/genome sequencing, one would expect to have identified all important CRC genes. Nevertheless, by exclusively targeting tumors that a) originated in the rectum, b) were from patients aged 60 years or lower (EOSRC, hugely under-represented in most CRC genomic studies) and c) were from a population shown previously to exhibit deviations from the canonical CRC dogma 28, 29, 42 , we increased the likelihood of identifying a hitherto poorly studied albeit important CRC gene.…”

Promoter Hypermethylation Profiling Identifies Subtypes of Head and Neck Cancer with Distinct Viral, Environmental, Genetic and Survival Characteristics

“…Promoter hypermethylation profile of tumor-related genes was anticipated to be crucial and frequent in different cancers. Many epigenetic events in carcinogenic pathways have been studied recently and revealed the methods for detecting CpG island promoter methylation pattern to stratify high risk groups among different cancers [ 15 , 17 , 18 , 34 ]. This helps in detection of early onset of cancer, and predicts clinical outcomes.…”

“…Promoter methylation status of tumor-related genes ( RASSF1 , DAPK , ECAD , BRCA1 , MLH1 , p16 and GSTP1 ) and three methylated loci ( MINT1 , MINT2 , and MINT31 ) were analyzed using Methylation Specific PCR (MSP) primers ( S2 Table ). For MSP assay, DNA samples were subjected to modification using Imprint1 DNA Modification kit (Sigma–Aldrich, St. Louis, MO), following instructions as described by manufacturer’s [ 18 ]. In this procedure, DNA denaturation and bisulfite modification are carried out simultaneously.…”