Quantitative Analysis of HER2 Amplification by Droplet Digital PCR in the Follow-Up of Gastric Cancer Patients Being Treated with Trastuzumab after Surgery

Abstract

Background. Circulating tumor DNA (ctDNA) derived from tumors is a promising biomarker for monitoring tumor status and evaluating therapeutic effects and prognosis. We studied the plasma human epidermal growth factor receptor 2 (HER2) amplification in gastric cancer (GC) patients by droplet digital PCR (ddPCR) during therapy with trastuzumab. Methods. A total of 12 patients were recruited after surgery. All patients received FOLFOX chemotherapy combined with trastuzumab as a treatment regimen. During the 12 months of the follow-up period, using elongation factor Tu GTP binding domain containing 2 (EFTUD2) as a reference gene, plasma HER2 to EFTUD2 ratios (the HER2 ratio) were determined for each patient every 2 months by ddPCR. Results. The concordance rate of HER2 amplification examined in plasma and formalin-fixed paraffin-embedded (FFPE) samples with ddPCR was 81.4%, with a sensitivity of 76.5% and a specificity of 83.8%. Plasma HER2 ratios were correlated with the primary tumor size (p<0.01). A significant decrease in the plasma HER2 ratio was found after two months of treatment (p<0.0001). Nine patients experienced partial response, and three patients had stable disease. Seven patients had progressive disease (PD) during follow-up, and four of them had died. The median progression-free survival (PFS) was 9.8 months. For each patient who developed PD, the plasma HER2 ratio was approximately 2.3-4.1 times higher than the cut-off value at the time of PD, which was the highest during the whole follow-up period. Conclusion. Longitudinal monitoring for the plasma HER2 ratio by ddPCR in the clinical courses of GC patients holds great promise for use as an indicator of tumor progression and treatment efficacy.

HER2 Gene Assessment in Liquid Biopsy of Gastric and Esophagogastric Junction Cancer Patients Qualified for Surgery

“…This level of concordance between the results of the HER2 gene tests in cancer tissue and in liquid biopsy was also found in our study and in study by Kim et al [9]. In addition Kinuasa et al found that patients who had high number of HER2 gene copies in ct-DNA showed signi cantly shorter survival compared with HER2-negative patients [21]. The median HER2 gene copy number in ct-DNA in Kinugasa et al study was 1.15.…”

“…Liu et al studied the plasma HER2 gene ampli cation in gastric cancer patients by droplet digital PCR (ddPCR) during 12 months of chemotherapy with uorouracil and oxaliplatin combined with trastuzumab [21]. They observed that the concordance rate of results of HER2 gene examination in plasma and FFPE tissue samples by ddPCR was 81.4%, with a sensitivity of 76% and a speci city of 84%.…”

Comparative analysis of HER2 copy number between plasma and tissue samples in gastric cancer using droplet digital PCR

“…Two research teams covered HER2 ddPCR with ctDNA, and one team measured HER2 ratio using ribonuclease P RNA component H1 (RPPH1) as a reference gene and the sensitivity and specificity of plasma HER2 ddPCR were 73.3% and 93.3%, respectively 22 . The other used elongation factor Tu GTP binding domain containing 2 (EFTUD2) as a reference gene and demonstrated the sensitivity and specificity were 76.5% and 83.8% 23 . Compared with the previous studies that measured HER2 ratios of gastric cancer patients using ddPCR [19][20][21][22][23][24] , our study covered the largest cohort to the best of our knowledge; moreover, we worked with paired samples of formalin-fixed and paraffin-embedded (FFPE) tissues and preoperative plasma cfDNA.…”

“…The other used elongation factor Tu GTP binding domain containing 2 (EFTUD2) as a reference gene and demonstrated the sensitivity and specificity were 76.5% and 83.8% 23 . Compared with the previous studies that measured HER2 ratios of gastric cancer patients using ddPCR [19][20][21][22][23][24] , our study covered the largest cohort to the best of our knowledge; moreover, we worked with paired samples of formalin-fixed and paraffin-embedded (FFPE) tissues and preoperative plasma cfDNA. We used EIF2C1 as the reference gene, and ddPCR using the HER2/EIF2C1 ratio demonstrated that it can assess HER2 status accurately 25 .…”

“…Further prospective studies with larger cohorts will be needed. In addition, HER2 measurement in patients after www.nature.com/scientificreports www.nature.com/scientificreports/ gastrectomy and during the follow-up period will give more information about relapse or treatment effects of trastuzumab 23,31 . The other limitation is that there has been no consensus about setting cutoffs for HER2 amplification.…”