Non-invasive Analysis of Genomic Copy Number Variation in Patients with Hepatocellular Carcinoma by Next Generation DNA Sequencing

Abstract

To explore new molecular diagnosis approaches for early detection and differential diagnosis of hepatocellular carcinoma (HCC), we analyzed genomic copy number variations (CNV) using plasma cell-free DNA from patients with HCC by next generation DNA sequencing. Plasma samples from 31 patients with HCC and 8 patients with chronic hepatitis or cirrhosis were analyzed. In HCC group, most samples with large tumor size (tumor dimension greater than 50 mm) showed CNVs that are visually recognizable at chromosome CNV plots, few samples with small tumor and none samples with chronic liver diseases showed CNVs recognizable at CNV plots. CNV Z score analysis showed significant CNVs in samples with HCC and chronic liver diseases although more significant changes were found in HCC group, some are differentially valuable (such as gain in 1q, 7q, and 19q in HCC), while others are less differentially valuable (such as loss in 4q, 13q, gain in 17q, 22q). We proposed a CNV scoring method that generated positive result in 26 of the 31 HCC patients (83.9%) or 11 of the 16 HCC with tumor dimension 50 mm or less (68.8%) or 4 of the 7 HCC with tumor dimension 30 mm or less (57.1%), while all the 8 samples with chronic hepatitis or cirrhosis scored negative. Ten HCC patients had normal or low serum AFP levels, among them, 7 were scored positive by CNV analysis, including 4 with tumor dimension 50 mm or less. Our study suggested that non-invasive genomic CNV analysis using plasma samples could be a valuable tool for early detection and differential diagnosis of HCC. Although CNV analysis itself cannot establish the diagnosis, it can help identify patients at high risk for HCC among patients with chronic liver diseases, which would prompt closer and more frequent surveillance for early tumor detection and intervention.

NTRK gene amplification in patients with metastatic cancer

“…Gene amplification, similar to gene mutation, plays a significant role in tumorigenesis in many types of cancer, such as gastric cancer, ovarian cancer, hepatocellular carcinoma, colon cancer, and others [22,23]. Thus, targeting the "driver genes" that are amplified may provide novel opportunities for precision medicine [20].…”

Gastric cancer and gene copy number variation: emerging cancer drivers for targeted therapy

“…12 DNA CNVs include gene amplification, gain, loss and deletion. In addition to gene mutation, CNV has a significant role in tumorigenesis in many cancers, such as GC, 13 ovarian cancer, 14 hepatocellular carcinoma, 15 testicular germ cell tumors, 16 colorectal carcinoma, 17 bladder cancer 18 and so on. The accumulation of CNVs during gastric oncogenesis may be a result of preferential selection by which transforming cells gain evolutionary advantage.…”

Chromosomal Instability Estimation Based on Next Generation Sequencing and Single Cell Genome Wide Copy Number Variation Analysis

“…Z scores were calculated for each gene by comparing the test sample and WBC controls, , where X is the normalized copy number of the test sample, is the average copy number from the universal WBC reference, and S is the standard deviation from the universal reference. The single cell level cut-off for gene amplification is Z score >3; for deletion it is Z score <-3 [36]. At the patient level, amplification or deletion of a single gene needed to be observed on at least two CTCs for classification as alteration.…”