Zhijie Wang

Potential Clinical Significance of a Plasma-Based KRAS Mutation Analysis in Patients with Advanced Non–Small Cell Lung Cancer

Purpose: Non–small cell lung cancer (NSCLC) with KRAS mutation may be resistant to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKI). This study aims to evaluate a plasma-based KRAS mutation analysis and the clinical significance of plasma KRAS mutation as a predictive marker for tumor resistance to EGFR-TKIs in patients with NSCLC. Experimental Design: DNA extracted from plasma and matched tumor tissues were obtained from 273 patients with advanced stage NSCLC. Patients were followed up prospectively for treatment outcomes. KRAS mutations in codon 12 and 13 were detected using PCR-restriction fragment length polymorphism. Mutations in plasma and matched tumors were compared. Associations between KRAS mutation status and patients clinical outcomes were analyzed. Results: KRAS mutation was found in 35 (12.8%) plasma samples and 30 (11.0%) matched tumor tissues. The consistency of KRAS mutations between plasma and tumors is 76.7% (23 of 30; κ = 0.668; P < 0.001). Among 120 patients who received EGFR-TKI treatment, the response rate was only 5.3% (1 of 19) for patients with plasma KRAS mutation compared with 29.7% for patients with no KRAS mutation in plasma DNA (P = 0.024). The median progression-free survival time of patients with plasma KRAS mutation was 2.5 months compared with 8.8 months for patients with wild-type KRAS (P < 0.001). Conclusions: KRAS mutation in plasma DNA correlates with the mutation status in the matched tumor tissues of patients with NSCLC. Plasma KRAS mutation status is associated with a poor tumor response to EGFR-TKIs in NSCLC patients and may be used as a predictive marker in selecting patients for such treatment. Clin Cancer Res; 16(4); 1324–30

Quantification of mutant alleles in circulating tumor DNA can predict survival in lung cancer

Purpose We aimed to investigate the feasibility of droplet digital PCR (ddPCR) for the quantitative and dynamic detection of EGFR mutations and next generation sequencing (NGS) for screening EGFR-tyrosine kinase inhibitors (EGFR-TKIs) resistance-relevant mutations in circulating tumor DNA (ctDNA) from advanced lung adenocarcinoma (ADC) patients. Results Detection limit of EGFR mutation in ctDNA by ddPCR was 0.04%. Taking the EGFR mutation in tumor tissue as the golden standard, the concordance of EGFR mutations detected in ctDNA was 74% (54/73). Patients with EGFR mutation in ctDNA (n = 54) superior progression-free survival (PFS, median, 12.6 vs. 6.7 months, P < 0.001) and overall survival (OS, median, 35.6 vs. 23.8 months, P = 0.028) compared to those with EGFR wild type in ctDNA (n = 19). Patients with high EGFR-mutated abundance in ctDNA (> 5.15%) showed better PFS compared to those with low EGFR mutated abundance (≤ 5.15%) (PFS, median, 15.4 vs. 11.1 months, P = 0.021). NGS results showed that 66.6% (8/12) total mutational copy number were elevated and 76.5% (26/34) mutual mutation frequency increased after disease progression. Methods Seventy-three advanced ADC patients with tumor tissues carrying EGFR mutations and their matched pre- and post-EGFR-TKIs plasma samples were enrolled in this study. Absolute quantities of plasma EGFR mutant and wild-type alleles were measured by ddPCR. Multi-genes testing was performed using NGS in 12 patients. Conclusions Dynamic and quantitative analysis of EGFR mutation in ctDNA could guide personalized therapy for advanced ADC. NGS shows good performance in multiple genes testing especially novel and uncommon genes.

Comprehensive Analysis of the Discordance of EGFR Mutation Status between Tumor Tissues and Matched Circulating Tumor DNA in Advanced Non-Small Cell Lung Cancer.

Introduction: This study aimed to address the underlying reasons for and clinical significance of the discordant EGFR mutation (EGFRm) status between tumor tissue (TT) and circulating tumor DNA (ctDNA). Methods: Three groups of EGFR tyrosine kinase inhibitor (EGFR TKI)‐treated patients whose EGFRm status was determined by the amplification refractory mutation system (ARMS) were included (group A, TT‐positive/ctDNA‐positive EGFRm status; group B, TT‐negative/ctDNA‐positive EGFRm status; and group C, TT‐positive/ctDNA‐negative EGFRm status). Patients with discordant EGFRm status were reevaluated by droplet digital polymerase chain reaction (ddPCR) and next‐generation sequencing. Meanwhile, surgical tumor specimens were microdissected for EGFRm detection by ddPCR. Results: Of the 2463 patients with matched TT and ctDNA specimens, 1017 patients carried EGFRm in TT and/or ctDNA by the ARMS. Of these 1017 patients, 472 received EGFR TKIs, including 264, 28, and 180 in groups A, B, and C, respectively. The median progression‐free survivals of those receiving EGFR TKIs across the three groups were similar (p = 0.062). Through ddPCR and next‐generation sequencing of biopsy specimens (n = 22) and microdissected surgical specimens (n = 5), 27 patients in group B were identified as harboring EGFRm. After reevaluation by ddPCR, 64 patients in group C tested positive for EGFRm in their ctDNA. ctDNA as a screen for EGFRm then tissues as supplement (ctDNA→TT pattern) had similar detection efficiency and saved about 30% of TT compared with TT for initial EGFRm detection followed by ctDNA (TT→ctDNA pattern). Conclusions: Intratumor heterogeneity and the relatively low sensitivity of the ARMS contributed to discordant EGFRm status between TT specimens and ctDNA. The ctDNA→TT pattern might be a rational clinical procedure for EGFRm determination.

Real world study of regimen containing bevacizumab as first-line therapy in Chinese patients with advanced non-small cell lung cancer: Bevacizumab as first-line for NSCLC

Large scale randomized controlled trials have demonstrated that the use of bevacizumab in addition to chemotherapy in patients with advanced non‐small cell lung cancer (NSCLC) conveys significant survival benefits. We explored the clinical impact of a first‐line regimen containing bevacizumab (B+) versus a non‐bevacizumab regimen (non‐B) in advanced non‐squamous NSCLC (NS‐NSCLC) patients in a real world setting.

Detection of EGFR mutations in plasma circulating tumour DNA as a selection criterion for first-line gefitinib treatment in patients with advanced lung adenocarcinoma (BENEFIT): a phase 2, single-arm, multicentre clinical trial

BACKGROUNDnDetection of EGFR mutations in tumour tissue is the gold-standard approach to ascertain if a patient will benefit from treatment with an EGFR tyrosine kinase inhibitor. However, if tissue is scant, another strategy is to use circulating tumour DNA (ctDNA), but this method needs validation in clinical trials. We did a prospective clinical trial to assess ctDNA-based EGFR mutation detection as a selection criterion for patients with lung adenocarcinoma receiving gefitinib as first-line treatment.nnnMETHODSnBENEFIT is a multicentre, single-arm, phase 2 clinical trial at 15 centres in China. Patients aged 18-75 years with stage IV metastatic lung adenocarcinoma and EGFR mutations detected in ctDNA were given oral gefitinib 250 mg once daily as first-line treatment. The primary endpoint was the proportion achieving an objective response. Secondary endpoints included median progression-free survival and safety. Next-generation sequencing (NGS) of a 168-gene panel was used for genetic analysis of baseline blood samples. The primary efficacy analysis was done by intention to treat in patients who had at least one post-baseline tumour assessment. The safety analysis was done in all patients who received at least one dose of study treatment. This trial is registered with ClinicalTrials.gov, number NCT02282267.nnnFINDINGSnBetween Dec 25, 2014, and Jan 16, 2016, 426 patients were screened for the trial, of whom 188 with EGFR mutations in ctDNA were enrolled and received gefitinib. 183 patients had one or more post-baseline tumour assessment and were included in the primary efficacy analysis. Median follow-up was 14·5 months (IQR 12·2-16·5). At the time of data cutoff (Jan 31, 2017), 152 patients had progressive disease or had died. The proportion achieving an objective response was 72·1% (95% CI 65·0-78·5). Median progression-free survival was 9·5 months (95% CI 9·07-11·04). Of 167 patients with available blood samples, 147 (88%) showed clearance of EGFR mutations in ctDNA at week 8, and median progression-free survival was longer for these patients than for the 20 patients whose EGFR mutations persisted at week 8 (11·0 months [95% CI 9·43-12·85] vs 2·1 months [1·81-3·65]; hazard ratio [HR] 0·14, 95% CI 0·08-0·23; p<0·0001). From baseline NGS data in 179 patients, we identified three subgroups of patients: those with EGFR mutations only (n=58), those with mutations in EGFR and tumour-suppressor genes (n=97), and those with mutations in EGFR and oncogenes (n=24). Corresponding median progression-free survival in these subgroups was 13·2 months (95% CI 11·5-15·0), 9·3 months (7·6-11·0), and 4·7 months (1·9-9·3), respectively (EGFR mutations only vs mutations in EGFR and tumour-suppressor genes, HR 1·78, 95% CI 1·23-2·58; p=0·002; EGFR mutations only vs mutations in EGFR and oncogenes, 2·66, 1·58-4·49; p=0·0003). The most common grade 3 or 4 adverse events were hepatic function abnormalities (n=24). Serious adverse events were reported in 17 (9%) patients. No unexpected safety events for gefitinib were recorded.nnnINTERPRETATIONnDetection of EGFR mutations in ctDNA is an effective method to identify patients who might benefit from first-line gefitinib treatment. Further analyses of dynamic alterations of EGFR mutations and accompanying gene aberrances could predict resistance to gefitinib.nnnFUNDINGnGuangdong Association of Clinical Trials, AstraZeneca, National Natural Sciences Foundation Key Programme, and National Key Research and Development Programme of China.

Correlation among genetic variations of c-MET in Chinese patients with non-small cell lung cancer

Background The purpose of our research was to determine the correlation of amplification, protein expression and somatic mutation of c-MET in IIIb-IV stage NSCLC (Non-small cell lung cancer). We also explored correlation of c-MET variation with clinical outcome. Results c-MET expression was observed in 28.6% (56/196) cases, and among those 13.8% (27/196) were shown to be FISH positive. Only 2.67% patients in this study carried the c-MET mutation. Cases with c-MET FISH positive were all IHC positive ,but in IHC positive cases, only half were FISH positive. Among patients with IHC2+ staining, 35.5% was FISH positive, while cases with IHC3+ staining,64% was FISH positive. Both protein expression and copy number of c-MET did not significantly correlate with clinical prognosis in these patients treated with EGFR-TKIs. Conclusions IHC could be used as a preliminary screening method for c-MET copy number amplification and should be confirmed by FISH only in IHC positive case which facilitate selection of ALK or MET inhibitor therapy. Methods c-MET gene copy number, protein expression and somatic mutation for exon 14 were detected by fluorescent- In-Situ-Hybridization (FISH), Immunohistochemistry (IHC), and Denaturing-High-Performance-Liquid-Chromatography (DHPLC), respectively, in 196 NSCLC patients. The relationship between c-MET abnormalities and clinical outcome of targeted therapy was analyzed by McNemars test.

Prediction of chemotherapeutic efficacy in non-small cell lung cancer by serum metabolomic profiling

Purpose: No validated biomarkers that could identify the subset of patients with lung adenocarcinoma who might benefit from chemotherapy have yet been well established. This study aimed to explore potential biomarker model predictive of efficacy and survival outcomes after first-line pemetrexed plus platinum doublet based on metabolomics profiling. Experimental Design: In total, 354 consecutive eligible patients were assigned to receive first-line chemotherapy of pemetrexed in combination with either cisplatin or carboplatin. Prospectively collected serum samples before initial treatment were utilized to perform metabolomics profiling analyses under the application of LC/MS-MS. Binary logistic regression analysis was carried out to establish discrimination models. Results: There were 251 cases randomly sorted into discovery set, the rest of 103 cases into validation set. Seven metabolites including hypotaurine, uridine, dodecanoylcarnitine, choline, dimethylglycine, niacinamide, and l-palmitoylcarnitine were identified associated with chemo response. On the basis of the seven-metabolite panel, a discriminant model according to logistic regression values g(z) was established with the receiver operating characteristic curve (AUC) of 0.912 (Discovery set) and 0.909 (Validation set) in differentiating progressive disease (PD) groups from disease control (DC) groups. The median progression-free survival (PFS) after chemotherapy in patients with g(z) ≤0.155 was significantly longer than that in those with g(z) > 0.155 (10.3 vs.4.5 months, P < 0.001). Conclusions: This study developed an effective and convenient discriminant model that can accurately predict the efficacy and survival outcomes of pemetrexed plus platinum doublet chemotherapy prior to treatment delivery. Clin Cancer Res; 24(9); 2100–9. ©2018 AACR.

Analysis of topoisomerase I expression and identification of predictive markers for efficacy of topotecan chemotherapy in small cell lung cancer: Topoisomerase I in SCLC

We evaluated topoisomerase I (TOPO1) expression in patients with small cell lung cancer (SCLC) and identified predictive factors for the efficacy of second‐line topotecan chemotherapy.

Real world study of the continuation of bevacizumab beyond disease progression after first-line treatment containing bevacizumab in Chinese patients with advanced non-small cell lung cancer: Continuation of bevacizumab in NSCLC

Bevacizumab (Bev) plus platinum‐based chemotherapy is a standard first‐line treatment option for advanced non‐squamous non‐small cell lung cancer (NS‐NSCLC). We evaluated the efficacy and safety of continuing Bev in Chinese patients with advanced NS‐NSCLC progression after first‐line treatment containing Bev in a real‐world setting.

The Tumor Mutational Burden of Chinese Advanced Cancer Patients Estimated by a 381-cancer-gene Panel

Purpose: Tumor mutational burden (TMB) calculated by whole-exome sequencing (WES) is proved to be effective to predict the clinical benefit of immune checkpoint blockades. However, WES is not commonly used in China. We aimed to determine if a 381-caner-gene panel (CGP) could be used to estimate TMB, delineate the landscape of TMB of Chinese patients and identify mutated genes and pathways related to higher TMB. Methods: We first evaluated the correlation between the TMB estimated by a 381-cancer-gene panel MasterView and WES using the data from the melanoma sample cohort. 3023 formalin fixed, paraffin-embedded tumor specimens from 2932 Chinese patients with advanced solid tumor were profiled for 381 gene sequencing, the baits of which covered 4,557 exons of 365 cancer-related genes and 47 introns of 25 genes frequently rearranged in cancer (All performed in a lab who achieved full marks five times in the external quality assessment by College of American Pathologists [CAP]). Using the sequencing data, we estimated the TMB of Chinese advanced solid tumor and identified mutated genes and pathways related to higher TMB level. Results: 381-CGP-mutational burden was strongly associated with those calculated by WES (R2 = 0.978). The median TMB for each tumor type was 5.65 (colorectal cancer), 4.84 (lung cancer), 4.03 (hepatobiliary cancer), 4.03 (gastric carcinoma), 4.03 (breast cancer) mutations/mb respectively. No correlation was observed between TMB level and age (P = 0.577) or gender (P = 0.307). The TMB of patients with mismatch repair (MMR) or DNA repair response (DDR) pathway deficiency was significantly higher than that without MMR or DDR pathway deficiency (P < 0.001). Conclusion: The 381-cancer gene panel is a clinical practicable method to assess tumor mutational burden compared with whole exome sequencing. MMR and DDR deficiency are correlated with higher tumor mutational burden of Chinese patients with advanced solid tumors.