The third-generation tyrosine kinase inhibitor osimertinib is approved to treat patients with T790M-positive non-small cell lung cancer (NSCLC) who have developed resistance to earlier-generation drugs. Acquired C797S mutation has been reported to mediate osimertinib resistance in some patients. However, the remaining resistance mechanisms are largely unknown. We performed mutation profiling using targeted next-generation sequencing (NGS) for 416 cancer-relevant genes on 93 osimertinib-resistant lung cancer patients' samples, mainly cell-free DNAs (cfDNAs), and matched pretreatment samples of 12 patients. experiments were conducted to functionally study the secondary mutations identified. G796/C797, L792, and L718/G719 mutations were identified in 24.7%, 10.8%, and 9.7% of the cases, respectively, with certain mutations coexisting in one patient with different prevalence. L792 and L718 mutants markedly increased the half inhibitory concentration (IC) of osimertinib , among which the L718Q mutation conferred the greatest resistance to osimertinib, as well as gefitinib resistance when not coexisting with T790M. Further analysis of the 12 matched pretreatment samples confirmed that these mutations were acquired during osimertinib treatment. Alterations in parallel or downstream oncogenes such as , and were also discovered, potentially contributing to the osimertinib-resistance in patients without secondary mutations. We present comprehensive mutation profiles of a large cohort of osimertinib-resistance lung cancer patients using mainly cfDNA. Besides C797 mutations, novel secondary mutations of L718 and L792 residues confer osimertinib resistance, both and , and are of great clinical and pharmaceutical relevance..

Accurately evaluating minimal residual disease (MRD) could facilitate early intervention and personalized adjuvant therapies. Here, using ultradeep targeted next-generation sequencing (NGS), we evaluate the clinical utility of circulating tumor DNA (ctDNA) for dynamic recurrence risk and adjuvant chemotherapy (ACT) benefit prediction in resected non-small cell lung cancer (NSCLC). Both postsurgical and post-ACT ctDNA positivity are significantly associated with worse recurrence-free survival. In stage II-III patients, the postsurgical ctDNA positive group benefit from ACT, while ctDNA negative patients have a low risk of relapse regardless of whether or not ACT is administered. During disease surveillance, ctDNA positivity precedes radiological recurrence by a median of 88 days. Using joint modeling of longitudinal ctDNA analysis and time-to-recurrence, we accurately predict patients’ postsurgical 12-month and 15-month recurrence status. Our findings reveal longitudinal ctDNA analysis as a promising tool to detect MRD in NSCLC, and we show pioneering work of using postsurgical ctDNA status to guide ACT and applying joint modeling to dynamically predict recurrence risk, although the results need to be further confirmed in future studies.

Purpose: Immune checkpoint inhibitors (ICI) have revolutionized cancer management. However, molecular determinants of response to ICIs remain incompletely understood. Experimental Design: We performed genomic profiling of 78 patients with non-small cell lung cancer (NSCLC) who underwent anti-PD-(L)1 therapies by both whole-exome and targeted next-generation sequencing (a 422-cancer-gene panel) to explore the predictive biomarkers of ICI response. Tumor mutation burden (TMB), and specific somatic mutations and copy-number alterations (CNA) were evaluated for their associations with immunotherapy response. Results: We confirmed that high TMB was associated with improved clinical outcomes, and TMB quantified by gene panel strongly correlated with WES results (Spearman's r ¼ 0.81). Compared with wild-type, patients with FAT1 mutations had higher durable clinical benefit (DCB, 71.4% vs. 22.7%, P ¼ 0.01) and objective response rates (ORR, 57.1% vs. 15.2%, P ¼ 0.02). On the other hand, patients with activating mutations in EGFR/ERBB2 had reduced median progression-free survival (mPFS) compared with others [51.0 vs. 70.5 days, P ¼ 0.0037, HR, 2.47; 95% confidence interval (CI), 1.32-4.62]. In addition, copy-number loss in specific chromosome 3p segments containing the tumorsuppressor ITGA9 and several chemokine receptor pathway genes, were highly predictive of poor clinical outcome (survival rates at 6 months, 0% vs. 31%, P ¼ 0.012, HR, 2.08; 95% CI, 1.09-4.00). Our findings were further validated in two independently published datasets comprising multiple cancer types. Conclusions: We identified novel genomic biomarkers that were predictive of response to anti-PD-(L)1 therapies. Our findings suggest that comprehensive profiling of TMB and the aforementioned molecular markers could result in greater predictive power of response to ICI therapies in NSCLC.

Background Precise methods for postoperative risk stratification to guide the administration of adjuvant chemotherapy (ACT) in localized colorectal cancer (CRC) are still lacking. Here, we conducted a prospective, observational, and multicenter study to investigate the utility of circulating tumor DNA (ctDNA) in predicting the recurrence risk. Methods From September 2017 to March 2020, 276 patients with stage II/III CRC were prospectively recruited in this study and 240 evaluable patients were retained for analysis, of which 1290 serial plasma samples were collected. Somatic variants in both the primary tumor and plasma were detected via a targeted sequencing panel of 425 cancer-related genes. Patients were treated and followed up per standard of care. Results Preoperatively, ctDNA was detectable in 154 of 240 patients (64.2%). At day 3–7 postoperation, ctDNA positivity was associated with remarkably high recurrence risk (hazard ratio [HR], 10.98; 95%CI, 5.31–22.72; P < 0.001). ctDNA clearance and recurrence-free status was achieved in 5 out of 17 ctDNA-positive patients who were subjected to ACT. Likewise, at the first sampling point after ACT, ctDNA-positive patients were 12 times more likely to experience recurrence (HR, 12.76; 95%CI, 5.39–30.19; P < 0.001). During surveillance after definitive therapy, ctDNA positivity was also associated with extremely high recurrence risk (HR, 32.02; 95%CI, 10.79–95.08; P < 0.001). In all multivariate analyses, ctDNA positivity remained the most significant and independent predictor of recurrence-free survival after adjusting for known clinicopathological risk factors. Serial ctDNA analyses identified recurrence with an overall accuracy of 92.0% and could detect disease recurrence ahead of radiological imaging with a mean lead time of 5.01 months. Conclusions Postoperative serial ctDNA detection predicted high relapse risk and identified disease recurrence ahead of radiological imaging in patients with stage II/III CRC. ctDNA may be used to guide the decision-making in postsurgical management.

ObjectiveTo monitor trastuzumab resistance and determine the underlying mechanisms for the limited response rate and rapid emergence of resistance of HER2+ metastatic gastric cancer (mGC).DesignTargeted sequencing of 416 clinically relevant genes was performed in 78 paired plasma and tissue biopsy samples to determine plasma-tissue concordance. Then, we performed longitudinal analyses of 97 serial plasma samples collected from 24 patients who were HER2+  to track the resistance during trastuzumab treatment and validated the identified candidate resistance genes.ResultsThe results from targeted sequencing-based detection of somatic copy number alterations (SCNA) of HER2 gene were highly consistent with fluorescence in situ hybridisation data, and the detected HER2 SCNA was better than plasma carcinoembryonic antigen levels at predicting tumour shrinkage and progression. Furthermore, most patients with innate trastuzumab resistance presented high HER2 SCNA during progression compared with baseline, while HER2 SCNA decreased in patients with acquired resistance. PIK3CA mutations were significantly enriched in patients with innate resistance, and ERBB2/4 genes were the most mutated genes, accounting for trastuzumab resistance in six (35.3%) and five (29.4%) patients in baseline and progression plasma, respectively. Patients with PIK3CA/R1/C3 or ERBB2/4 mutations in the baseline plasma had significantly worse progression-free survival. Additionally, mutations in NF1 contributed to trastuzumab resistance, which was further confirmed through in vitro and in vivo studies, while combined HER2 and MEK/ERK blockade overcame trastuzumab resistance.ConclusionLongitudinal circulating tumour DNA sequencing provides novel insights into gene alterations underlying trastuzumab resistance in HER2+mGC.