Assessment of HER2 (ERBB2) amplification (HER2amp) using blood-based circulating tumor DNA (ctDNA) next generation sequencing (NGS) and correlation with tissue-based testing in metastatic colorectal cancer (mCRC).
Authors: Raghav Kanwal Pratap Singh , Nakamura Yoshiaki , Marsoni Silvia , Strickler John H. , Yaeger Rona , Shah Aakash Tushar , Okamoto Wataru , Crisafulli Giovanni , Nagy Rebecca , Raymond Victoria M. , Routbort Mark , Siena Salvatore , Corcoran Ryan Bruce , Bardelli Alberto , Kopetz Scott , Yoshino Takayuki 
Published: June 2021

3589 Background: HER2 amplified mCRC has emerged as a unique clinical subset, characterized by resistance to anti-EGFR therapy and response to anti-HER2 strategies. Accurate identification and quantification of HER2amp has predictive value for efficacy of anti-HER2 therapies and appropriate patient selection. Despite availability and use of various tumor tissue-based and blood-based assays for detecting HER2amp, data on cross-performance of these platforms are lacking. Methods: Leveraging a multicenter international consortium (Italy, Japan and USA), we generated a large cohort (N = 353) of mCRC patients (pts), tested for HER2amp using both tissue and blood. Tissue testing was done using immunohistochemistry (IHC), in-situ hybridization (ISH) and (NGS). ctDNA NGS was performed using CLIA-certified Guardant360 ctDNA assay, capable of detecting HER2 copy number (CN) variations. The primary endpoint was to correlate HER2 gene CNs in tissue (tCN) and plasma (pCN). Descriptive statistics, spearman correlation (r) and Fisher’s exact test were used. Results: Baseline tumors characteristics included right-sided primary in 234 (23%), proficient mismatch repair in 264 (98%) and RAS/BRAF wild type (WT) genotype in 194 (67%) pts. Tissue testing was done by IHC, ISH and NGS in 76%, 64% and 74% pts, respectively. A total of 177 pts had HER2amp detected by at least one test: 116 (66%), 157 (89%) and 96 (54%) of which had tissue +, ctDNA +, and both tissue and ctDNA + disease, respectively. Discordant cases consisted of 20 (6%) with positivity in tumor only and 61 (17%) in ctDNA only. Sensitivity, specificity, positive and negative predictive values of ctDNA assay (vis-à-vis tissue) were 83%, 74%, 61% and 90% respectively. Among HER2amp pts, median (range) HER2/CEP17 (ISH) ratio, tCN and pCN were 5.2 (2–12), 11.6 (2–700) and 3.5 (2–122), respectively. The pCN showed strong correlation with ISH ratio (r = 0.69) and tCN (r = 0.68) (P < 0.001). Median pCN differed significantly between pts with HER2 IHC 3+ (12.0), 2+ (2.2) and 0/1+ (2.0) tumors (P < 0.001). High HER2amp (pCN > 4.0) appeared to be enriched with tissue + cases (69% vs 8% [OR 24.6, P < 0.001]), tumor tissue HER2 + status (IHC3+ [75%] vs IHC2+ISH+ [50%] vs IHC2+/ISH- or IHC0/1+ [12%], P < 0.001), HER2 tCN > 6 (79% vs 31% [OR 8.7, P < 0.001]) and RAS/BRAF WT tumors (41% vs 17% [OR 3.5, P = 0.064) but not left sidedness (41% vs 38%; OR 1.1; P = 0.82). Conclusions: In this large diverse cohort of mCRC, we demonstrated correlation of HER2 tCN and pCN obtained by tissue-based and blood-based ctDNA assay. Further prospective efforts are needed to standardize this cross-platform quantification of HER2amp to facilitate robust clinical application of HER2 therapies. This effort shows the value of strategic international partnership in furthering research for rare cancer subsets.