Intra-Tumor Heterogeneity of Colorectal Cancer Necessitates the Multi-Regional Sequencing for Comprehensive Mutational Profiling

Shaohua Guo; Yumeng Ye; Xinyi Liu; Yuan Gong; Mingyan Xu; Lele Song; Hongyi Liu

doi:10.2147/CMAR.S327596

Intra-Tumor Heterogeneity of Colorectal Cancer Necessitates the Multi-Regional Sequencing for Comprehensive Mutational Profiling

Cancer Manag Res. 2021 Dec 16:13:9209-9223. doi: 10.2147/CMAR.S327596. eCollection 2021.

Authors

Shaohua Guo^#^{1

2}, Yumeng Ye^#³, Xinyi Liu⁴, Yuan Gong⁵, Mingyan Xu⁴, Lele Song^{4

6}, Hongyi Liu¹

Affiliations

¹ Department of General Surgery, The First Medical Center of the Chinese PLA General Hospital, Beijing, People's Republic of China.
² Department of General Surgery, The Eighth Medical Center of the Chinese PLA General Hospital, Beijing, People's Republic of China.
³ Department of Experimental Pathology, Beijing Institute of Radiation Medicine, Beijing, People's Republic of China.
⁴ Department of Medical Division, HaploX Biotechnology, Shenzhen, People's Republic of China.
⁵ Department of Gastroenterology, The Second Medical Center of the Chinese PLA General Hospital, Beijing, People's Republic of China.
⁶ Department of Radiotherapy, The Eighth Medical Center of the Chinese PLA General Hospital, Beijing, People's Republic of China.

^# Contributed equally.

Abstract

Background: The panorama and details of quantitative intratumor heterogeneity have not been fully investigated in colorectal cancer (CRC) patients with solitary lesion without distal metastasis, and its influences on sequencing interpretation and therapeutic strategies have not been explored.

Methods: Cancer tissues and matched blood from 70 sporadic CRC patients were collected and were divided into two cohorts. Four individual tissue biopsies were obtained from each of the 47 patients (multi-sample cohort). One random cancer tissue biopsy was obtained from each of the rest 23 patients (single-sample cohort). A 10 mL of blood was collected from all patients and the circulating cell-free DNA (cfDNA) was extracted. A 605-gene panel was used for targeted sequencing with tissue and paired blood.

Results: Mutational landscape revealed significantly higher mutational frequency in APC, CARD11 and CSMD3 in multi-sample cohort than single-sample cohort (P<0.05). The number of mutations and the ratio of trunk, shared and branch mutations showed extensive heterogeneity in multi-sample cohort, and the percentage of trunk mutations in major driver genes, including APC, TP53 and KRAS, was higher than 70%. A total of 929 mutations were detected in tissue/blood in multi-sample group, with 921(99.1%) from tissue and 472(50.8%) from blood (464 common mutations,49.9%). In contrast, 394 mutations were detected in tissue/blood in single-sample group, with 231 (58.6%) from tissue and 219 (55.6%) from blood (56 common mutations, 11.9%). The number of mutations of major driver genes detected in tissue was higher than that in blood in the multi-sample cohort, while it was similar in the single-sample group. Quantification analysis revealed differential correlation between tissue and blood VAF in trunk, shared and branch mutations. Meanwhile, VAF of trunk mutations was significantly higher than shared mutations and branch mutations. VAF exhibited significant differences among distinct stages, locations, differentiation and sex status.

Conclusion: Characteristic extensive heterogeneity was revealed for solitary CRC without distal metastasis. Multi-regional biopsy was necessary for comprehensive mutation detection in CRC.

Keywords: APC; KRAS; TP53; circulating tumor DNA; colorectal cancer; ctDNA; intra-tumor heterogeneity; multi-region sequencing.

Grants and funding

This study was supported by the Special Funds for Strategic Emerging Industry Development of Shenzhen (Grant Number 20170922151538732), and the Science and Technology Project of Shenzhen (Grant Number JSGG20180703164202084). This study was also supported by the Special health research projects of 2019 funded by the Chinese PLA general hospital, under the project “Screening of early-stage CRC and adenoma by combined methylated SEPT9 and FIT assays in military aging population” (NLBJ-2019003). All funders did not participate in the study design, study implementation, data collection, data analysis, data interpretation and manuscript writing of the study.