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Identification of Novel Mutations in Colorectal Cancer Patients Using AmpliSeq Comprehensive Cancer Panel.

ABSTRACT: Biomarker discovery would be an important tool in advancing and utilizing the concept of precision and personalized medicine in the clinic. Discovery of novel variants in local population provides confident targets for developing biomarkers for personalized medicine. We identified the need to generate high-quality sequencing data from local colorectal cancer patients and understand the pattern of occurrence of variants. In this report, we used archived samples from Saudi Arabia and used the AmpliSeq comprehensive cancer panel to identify novel somatic variants. We report a comprehensive analysis of next-generation sequencing results with a coverage of >300X. We identified 466 novel variants which were previously unreported in COSMIC and ICGC databases. We analyzed the genes associated with these variants in terms of their frequency of occurrence, probable pathogenicity, and clinicopathological features. Among pathogenic somatic variants, 174 were identified for the first time in the large intestine. APC, RET, and EGFR genes were most frequently mutated. A higher number of variants were identified in the left colon. Occurrence of variants in ERBB2 was significantly correlated with those of EGFR and ATR genes. Network analyses of the identified genes provide functional perspective of the identified genes and suggest affected pathways and probable biomarker candidates. This report lays the ground work for biomarker discovery and identification of driver gene mutations in local population.

SUBMITTER: Almuzzaini B

PROVIDER: S-EPMC8230213 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Ataxia-telangiectasia (A-T), or Louis-Bar syndrome, is a rare neurodegenerative disorder associated with immunodeficiency. For families with at least one affected child, timely A-T genotyping during any subsequent pregnancy allows the parents to make an informed decision about whether to continue to term when the fetus is affected. Mutations in the ATM gene, which is 150?kb long, give rise to A-T; more than 600 pathogenic variants in ATM have been characterized since 1990 and new mutations continue to be discovered annually. Therefore, limiting genetic screening to previously known SNPs by PCR or hybridization with microarrays may not identify the specific pathogenic genotype in ATM for a given A-T family. However, recent developments in next-generation sequencing technology offer prompt high-throughput full-length sequencing of genomic fragments of interest. This allows the identification of the whole spectrum of mutations in a gene, including any novel ones. We report two A-T families with affected children and current pregnancies. Both families are consanguineous and originate from Caucasian regions of Russia and Azerbaijan. Before our study, no ATM mutations had been identified in the older children of these families. We used ion semiconductor sequencing and an Ion AmpliSeq™ Inherited Disease Panel to perform complete ATM gene sequencing in a single member of each family. Then we compared the experimentally determined genotype with the affected/normal phenotype distribution in the whole family to provide unambiguous evidence of pathogenic mutations responsible for A-T. A single novel SNP was allocated to each family. In the first case, we found a mononucleotide deletion, and in the second, a mononucleotide insertion. Both mutations lead to truncation of the ATM protein product. Identification of the pathogenic mutation in each family was performed in a timely fashion, allowing the fetuses to be tested and diagnosed. The parents chose to continue with both pregnancies as both fetuses had a healthy genotype and thus were not at risk of A-T.

Project description:Background The identification of driver mutations has greatly promoted the precise diagnosis and treatment of non-small cell lung cancer (NSCLC), but there is lack of targeted sequencing panels specifically designed and applied to Chinese NSCLC patients. This study aimed to design and validate of a novel sequencing panel for comprehensive characterization of driver mutations in Chinese NSCLC patients, facilitating further exploration of downstream pathway alterations and therapeutic utility. Methods A novel target sequencing panel including 21 driver genes was designed and examined in a cohort of 260 Chinese NSCLC patients who underwent surgery in Peking Union Medical College Hospital (PUMCH). Genetic alterations were identified and further analyzed for driver mutations, downstream pathways and therapeutic utilities. Results The most frequently identified driver mutations in PUMCH NSCLC cohort were on genes TP53 (28%), EGFR (27%) and PIK3CA (19%) for lung adenocarcinoma (LUAD), and TP53 (41%), PIK3CA (14%) and CDKN2A (13%) for lung squamous cell carcinoma (LUSC), respectively. Downstream pathway analysis revealed common pathways like G1_AND_S1_PHASES pathway were shared not only between LUAD and LUSC patients, but also among three different NSCLC cohorts, while other pathways were subtype-specific, like the unique enrichment of SHC1_EVENT_IN_EGFR_SIGNALING pathway in LUAD patients, and P38_ALPHA_BETA_DOWNSTREAM pathway in LUSC patients, respectively. About 60% of both LUAD and LUSC patients harbored driver mutations as sensitive biomarkers for different targeted therapies, covering not only frequent mutations like EGFR L858R mutation, but also rare mutations like BRAF D594N mutation. Conclusions Our study provides a novel target sequencing panel suitable for Chinese NSCLC patients, which can effectively identify driver mutations, analyze downstream pathway alterations and predict therapeutic utility. Overall it is promising to further optimize and apply this panel in clinic with convenience and effectiveness.

Project description:This study aims at identifying common pathogenic somatic mutations at different stages of colorectal carcinogenesis in Egyptian patients. Our cohort included colonoscopic biopsies collected from 120 patients: 20 biopsies from patients with inflammatory bowel disease, 38 from colonic polyp patients, and 62 from patients with colorectal cancer. On top of this, the cohort included 20 biopsies from patients with non-specific mild to moderated colitis. Targeted DNA sequencing using a customized gene panel of 96 colorectal related genes running on the Ion Torrent NGS technology was used to process the samples. Our results revealed that 69% of all cases harbored at least one somatic mutation. Fifty-seven genes were found to carry 232 somatic non-synonymous variants. The most frequently pathogenic somatic mutations were localized in TP53, APC, KRAS, and PIK3CA. In total, 16 somatic mutations were detected in the CRC group and in either the IBD or CP group. In addition, our data showed that 51% of total somatic variants were CRC-specific variants. The average number of CRC-specific variants per sample is 2.4. The top genes carrying CRC-specific mutations are APC, TP53, PIK3CA, FBXW7, ATM, and SMAD4. It seems obvious that TP53 and APC genes were the most affected genes with somatic mutations in all groups. Of interest, 85% and 28% of the APC and TP53 deleterious somatic mutations were located in Exon 14 and Exon 3, respectively. Besides, 37% and 28% of the total somatic mutations identified in APC and TP53 were CRC-specific variants, respectively. Moreover, we identified that, in 29 somatic mutations in 21 genes, their association with CRC patients was unprecedented. Ten detected variants were likely to be novel: six in PIK3CA and four variants in FBXW7. The detected P53, Wnt/βcatenin, Angiogenesis, EGFR, TGF-β and Interleukin signaling pathways were the most altered pathways in 22%, 16%, 12%, 10%, 9% and 9% of the CRC patients, respectively. These results would contribute to a better understanding of the colorectal cancer and in introducing personalized therapies for Egyptian CRC patients.

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Identification of Novel Mutations in Colorectal Cancer Patients Using AmpliSeq Comprehensive Cancer Panel.

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