The authors of the article entitled, “Distinct mutational features across preinvasive and invasive subtypes identified through comprehensive profiling of surgically resected lung adenocarcinoma” (Modern Pathology 2022;35(9):1181-1192; DOI: https://doi.org/10.1038/s41379-022-01076-w) have discovered an error in the published Acknowledgements statement related to the funding number of the National Natural Science Foundation of China by CX. The correct number should be as follows: 82003154. A corrected statement in full is included below. Corrected acknowledgements in full: The authors thank all the patients who participated in this study and their families. We also thank the investigators, study coordinators, operation staff, and the whole project team who worked on this study. This work was supported by grants from the Shanghai Municipal Health Commission Intelligent Medical Research Project (Grant no. 2018ZHYL0213 to YH); Clinical Research Project (Grant No. 20194Y0053 to CX); National Natural Science Foundation of China (Grant No. 82003154 to CX); and Shanghai Jiao Tong University Medical Engineering Cross Fund (Grant no. YG2019QNA50 to CX). The funders had no role in the study design, participant recruitment, data collection, data analysis, data interpretation, manuscript preparation, and the decision to submit the paper for publication. Distinct mutational features across preinvasive and invasive subtypes identified through comprehensive profiling of surgically resected lung adenocarcinomaModern PathologyVol. 35Issue 9PreviewLung adenocarcinoma (LUAD) is a heterogeneous disease. Our study aimed to understand the unique molecular features of preinvasive to invasive LUAD subtypes. We retrospectively analyzed the clinical, histopathological, and molecular data of 3,254 Chinese patients with preinvasive lesions (n = 252), minimally invasive adenocarcinomas (n = 479), and invasive LUAD (n = 2,523). Molecular data were elucidated using a targeted 68-gene next-generation sequencing panel. Our findings revealed four preinvasive lesion-predominant gene mutations, including MAP2K1 insertion-deletions (indels), BRAF non-V600E kinase mutations, and exon 20 insertions (20ins) in both EGFR and ERBB2, which we referred to as mutations enriched in AIS (MEA). Full-Text PDF Open Access

Background The feasibility of DNA methylation-based assays in detecting minimal residual disease (MRD) and postoperative monitoring remains unestablished. We aim to investigate the dynamic characteristics of cancer-related methylation signals and the feasibility of methylation-based MRD detection in surgical lung cancer patients. Methods Matched tumor, tumor-adjacent tissues, and longitudinal blood samples from a cohort (MEDAL) were analyzed by ultra-deep targeted sequencing and bisulfite sequencing. A tumor-informed methylation-based MRD (timMRD) was employed to evaluate the methylation status of each blood sample. Survival analysis was performed in the MEDAL cohort ( n = 195) and validated in an independent cohort (DYNAMIC, n = 36). Results Tumor-informed methylation status enabled an accurate recurrence risk assessment better than the tumor-naïve methylation approach. Baseline timMRD-scores were positively correlated with tumor burden, invasiveness, and the existence and abundance of somatic mutations. Patients with higher timMRD-scores at postoperative time-points demonstrated significantly shorter disease-free survival in the MEDAL cohort (HR: 3.08, 95% CI: 1.48–6.42; P = 0.002) and the independent DYNAMIC cohort (HR: 2.80, 95% CI: 0.96–8.20; P = 0.041). Multivariable regression analysis identified postoperative timMRD-score as an independent prognostic factor for lung cancer. Compared to tumor-informed somatic mutation status, timMRD-scores yielded better performance in identifying the relapsed patients during postoperative follow-up, including subgroups with lower tumor burden like stage I, and was more accurate among relapsed patients with baseline ctDNA-negative status. Comparing to the average lead time of ctDNA mutation, timMRD-score yielded a negative predictive value of 97.2% at 120 days prior to relapse. Conclusions The dynamic methylation-based analysis of peripheral blood provides a promising strategy for postoperative cancer surveillance. Trial registration This study (MEDAL, ME thylation based D ynamic A nalysis for L ung cancer) was registered on ClinicalTrials.gov on 08/05/2018 (NCT03634826). https://clinicaltrials.gov/ct2/show/NCT03634826 .

Background: Patient-specific flexible gene panels designed based on whole-exome sequencing (WES) of resected tumor tissues is a promising strategy for ctDNA-based detection of molecular residual disease (MRD) in early-stage NSCLC. Flexible gene panels could potentially overcome the limitations of fixed panels by incorporating more unique genomic regions that might be absent in fixed panels; however, no study has reported a head-to-head comparison of these two approaches in postoperative disease monitoring. In this study, we investigated the clinical utility of a novel Patient-specific pROgnostic and Potential tHErapeutic marker Tracking (PROPHET) tumor-informed ctDNA assay. Using the same set of longitudinal blood samples, we further compared the performance of PROPHET assay with tumor-informed (TI) and tumor-naïve (TN) fixed panels for predicting MRD and prognosis in surgical NSCLC patients. Methods: Fifty-three patients with stage I-III resected NSCLC from the MEDAL study (NCT03634826) with adequate samples and median follow-up of 647 days were analyzed. Matched surgical tumor tissue and blood samples collected at various time points, including before surgery (baseline), 3-days (B) and 1-month (C) postoperative time points before any adjuvant therapy and subsequent follow-up time points (F) were analyzed. PROPHET assay involved four major steps: identify somatic mutations using WES, customized design of a patient-specific panel consisting of 50 single nucleotide variants, ultra-deep unique molecular-identifier-based next-generation sequencing (UMI-NGS) of serial blood samples using the patient-specific panel, and MRD risk prediction. Fixed panel assay of serial blood samples was performed using UMI-NGS with 168 gene panel spanning 273 kb of human genome. Results: At 1-month post-surgery, PROPHET assay accurately predicted MRD-positive cases among relapsed patients (50%, 13/26), whereas all disease-free patients were MRD-negative (100%, 21/21). Three-year prognostication with PROPHET assay at B+C yielded higher sensitivity (59% vs 26% vs 22%), negative predictive value (66% vs 51% vs 50%), and hazard ratio (7.15, 95%CI [3.2-15.9] vs 4.48 [1.9-10.9] vs 5.58 [2.1-14.7]) as compared with TI and TN fixed panel assays. Disease monitoring using PROPHET assay at B/C/F accurately predicted the MRD risk in 70% (21/30) of relapsed patients at a median lead time of 318 days (range: 20-751), whereas TI assay predicted 43% (13/30) at 282 days (range 20-716) and TN assay predicted 37% (11/30) at 282 days (range: 20-634). Conclusion: Patient-specific tumor-informed ctDNA-based postoperative monitoring enables risk stratification at early postoperative settings better than fixed panel, which paves an alternative strategy in the individualized management of surgical NSCLC patients. Citation Format: Kezhong Chen, Haifeng Shen, Shuailai Wu, Pengfei Zhu, Chenyang Wang, Analyn Lizaso, Guannan Kang, Yang Wang, Juan Lv, Shuai Fang, Wenjun Wu, Fujun Qiu, Yuan Sun, Qiang Lu, Heng Zhao, Shannon Chuai, Fan Yang, Zhihong Zhang. Tumor-informed patient-specific panel outperforms tumor-naïve and tumor-informed fixed panel for circulating tumor DNA (ctDNA)-based postoperative monitoring of non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5916.

OBJECTIVES:Ovarian cancer is a fatal gynecological cancer due to the lack of effective screening strategies at early stage. This study explored the utility of DNA methylation profiling of blood samples for the detection of ovarian cancer.METHODS:Targeted bisulfite sequencing was performed on tissue (n = 152) and blood samples (n = 373) obtained from healthy women, women with benign ovarian tumors, or malignant epithelial ovarian tumors. Based on the tissue-derived differentially-methylated regions, a supervised machine learning algorithm was implemented and cross-validated using the blood-derived DNA methylation profiles of the training cohort (n = 178) to predict and classify each blood sample as malignant or non-malignant. The model was further evaluated using an independent test cohort (n = 184).RESULTS:Comparison of the DNA methylation profiles of normal/benign and malignant tumor samples identified 1272 differentially-methylated regions, with 49.4% hypermethylated regions and 50.6% hypomethylated regions. Five-fold cross-validation of the model using the training dataset yielded an area under the curve of 0.94. Using the test dataset, the model accurately predicted non-malignancy in 96.2% of healthy women (n = 53) and 93.5% of women with benign tumors (n = 46). For patients with malignant tumors, the model accurately predicted malignancy in 44.4% of stage I-II (n = 9), 86.4% of stage III (n = 59), 100.0% of stage IV tumors (n = 6), and 81.8% of tumors with unknown stage (n = 11). Overall, the model yielded a predictive accuracy of 89.5%.CONCLUSIONS:Our study demonstrates the potential clinical application of blood-based DNA methylation profiling for the detection of ovarian cancer.

Lung adenocarcinoma (LUAD) is a heterogeneous disease. Our study aimed to understand the unique molecular features of preinvasive to invasive LUAD subtypes. We retrospectively analyzed the clinical, histopathological, and molecular data of 3,254 Chinese patients with preinvasive lesions (n = 252), minimally invasive adenocarcinomas (n = 479), and invasive LUAD (n = 2,523). Molecular data were elucidated using a targeted 68-gene next-generation sequencing panel. Our findings revealed four preinvasive lesion-predominant gene mutations, including MAP2K1 insertion-deletions (indels), BRAF non-V600E kinase mutations, and exon 20 insertions (20ins) in both EGFR and ERBB2, which we referred to as mutations enriched in AIS (MEA). The detection rate of MEA in invasive tumors was relatively lower. MAP2K1 missense mutations, which were likely passenger mutations, co-occurred with oncogenic driver mutations, while small indels were mutually exclusive from other genes regardless of the invasion level. BRAF non-V600E kinase-mutant invasive adenocarcinomas (IAC) had significantly higher mutation rates in tumor suppressor genes but lower frequency of co-occurring oncogenic driver mutations than non-kinase-mutant IAC, suggesting the potential oncogenic activity of BRAF non-V600E kinase mutations albeit weaker than BRAF V600E. Moreover, similar to the extremely low frequency of MAP2K1 indels in IAC, BRAF non-V600E kinase domain mutations co-occurring with TSC1 mutations were exclusively found in preinvasive lesions. Compared with EGFR L858R and exon 19 deletion, patients with preinvasive lesions harboring 20ins in either EGFR or ERBB2 were significantly younger, while those with IAC had similar age. Furthermore, our study demonstrated distinct mutational features for subtypes of oncogene mutations favored by different invasion patterns in adenocarcinomas. In conclusion, our data demonstrate distinct mutational features between preinvasive lesions and invasive tumors with MEA, suggesting the involvement of MEA in the early stages of tumorigenesis. Further pre-clinical studies are required to establish the role of these genes in the malignant transformation of LUAD.

Background: Approximately 2%-8% of non-small-cell lung cancer (NSCLC) harbors concurrent epidermal growth factor receptor (EGFR) sensitizing mutation and mesenchymal-epithelial transition factor (MET) amplification prior to EGFR-tyrosine kinase inhibitor (EGFR-TKI) therapy. This study aimed to investigate the optimal first-line therapeutic options for patients with concurrent EGFR-mutant, MET-overexpressed/amplified advanced NSCLC. Methods: A total of 104 treatment-naive patients with EGFR-mutant de novo MET-overexpressed advanced NSCLC were identified using immunohistochemistry and stratified to four groups according to treatment regimen: EGFR-TKI monotherapy (n = 48), EGFR-TKI combined with either crizotinib (n = 9) or chemotherapy (n = 12), and chemotherapy (n = 35). A subpopulation of 28 patients was also tested with next-generation sequencing (NGS). Objective response rate (ORR) and progression-free survival (PFS) outcomes were analyzed according to treatment strategies and molecular features. Results: All the patients (n = 104) achieved ORR of 36.5% and median PFS (mPFS) of 7.0 months. Baseline clinicopathologic characteristics were similar among the four treatment groups. Compared with chemotherapy, EGFR-TKI monotherapy or EGFR-TKI combination therapy achieved significantly higher ORR (P < 0.001) and longer mPFS (P = 0.003). No ORR or PFS difference was observed between EGFR-TKI monotherapy and combination therapy. In the NGS-identified population (n = 28), patients who received EGFR-TKI plus crizotinib (n = 9) achieved similar ORR (88.9% versus 57.9%, P = 0.195) and mPFS (9.0 versus 8.5 months, hazard ratio 1.10, 95% confidence interval 0.43-2.55, P = 0.45) than those who received EGFR-TKI monotherapy (n = 19), regardless of MET copy number status. Grade 3/4 rashes were significantly more among patients who received EGFR-TKI plus crizotinib (P = 0.026). Conclusions: Our findings provided clinical evidence that patients with concurrent EGFR sensitizing mutation and de novo MET amplification/overexpression could benefit from first-line EGFR-TKI monotherapy.

Background Comprehensive genomic profiling has become standard clinical practice in the management of advanced lung cancer. In addition to tissue and plasma, other body fluids are also being actively explored as alternative sources of tumor DNA. This study investigated the utility of induced sputum obtained from patients with non-small-cell lung cancer (NSCLC) for somatic variation profiling. Methods Our study included 41 treatment-naïve patients diagnosed with locally advanced to advanced NSCLC between October 2018 and June 2019. Capture-based targeted sequencing was performed on matched tumor, plasma, and induced sputum samples of 41 patients using a 168-gene panel. We analyzed the somatic variations detected from each sample type and the concordance of variations detected between matched samples. The concordance rate was defined as the proportion of the total number of variations detected from one sample type relative to the reference sample type. Results Comparative analysis on the somatic variation detection using matched tumor samples as a reference revealed detection rates of 76.9% for plasma, 72.4% for sputum-supernatant, and 65.7% for sputum-sediment samples. Plasma, sputum-supernatant, and sputum-sediment achieved positive predictive values of 73.3%, 80.4%, and 55.6% and sensitivities of 50.0%, 36.9%, 31.3%, respectively, relative to tumor samples for 168 genes. Sputum-supernatants had significantly higher concordance rates relative to matched tumor samples (69.2% vs. 37.8%; P=0.031) and maximum allelic fraction (P<0.001) than their matched sputum-sediments. Sputum-supernatants had comparable detection rates (71.4% vs. 67.9%; P=1.00) but with significantly higher maximum allelic fraction than their matched plasma samples (P=0.003). Furthermore, sputum-supernatant from smokers had a significantly higher maximum allelic fraction than sputum-supernatant from non-smokers (P=0.021). Conclusions Our study demonstrated that supernatant fraction from induced sputum is a better sampling source than its sediment and performs comparably to plasma samples. Induced sputum from NSCLC patients could serve as an alternative media for next-generation sequencing (NGS)-based somatic variation profiling.

This retrospective study investigated the association between the pattern of disease progression and molecular mechanism of acquired resistance in a large cohort of 49 patients with ROS1-rearranged advanced non-small-cell lung cancer treated with first-line crizotinib. We found that treatment-emergent ROS1 point mutations were the major molecular mechanism of crizotinib resistance, particularly for patients who developed extracranial-only disease progression. Our findings highlight the importance of rebiopsy and gene testing for subsequent-line therapeutic management.

BACKGROUND Due to the rarity of mesenchymal-epithelial transition factor (MET) fusions, the clinical efficacy of crizotinib has only been described in a few patients with MET fusions involving various fusion partners. Herein, we report the clinical response to crizotinib of a patient with advanced poorly differentiated non-small cell carcinoma (NSCLC) having concurrent MET fusions. CASE SUMMARY A 46-year-old woman was diagnosed with poorly differentiated NSCLC (T4N3M1). With no classic driver mutations, she was treated with two cycles of gemcitabine and cisplatin without clinical benefit. Targeted sequencing revealed the detection of two concurrent MET fusions, KIF5B-MET and novel MET-CDR2. Crizotinib was initiated at a dose of 250 mg twice daily. Within 4 wk of crizotinib therapy, repeat computed chromatography revealed a dramatic reduction in primary and metastatic lesions, assessed as partial response. She continued to benefit from crizotinib for 3 mo until disease progression and died within 1 mo despite receiving nivolumab therapy. CONCLUSION Crizotinib sensitivity was observed in an advanced poorly differentiated NSCLC patient with concurrent MET fusions KIF5B-MET and MET-CDR2. Crizotinib can serve as a therapeutic option for patients with MET fusions. In addition, our case also highlights the importance of comprehensive genomic profiling particularly in patients with no classic driver mutation for guiding alternative therapeutic decisions.

A 67-year-old man with a 20 pack-year smoking history presented to our hospital with a month-long persistent cough. Blood tests showed elevated serum tumour biomarkers including neuron-specific enolase and carcinoembryonic antigen up to 10-fold the normal range. CT examination of the chest and whole abdomen revealed a 5 cm mass located on the left upper lobe with diffuse metastasis in both lobes of the lung. In addition to the mass in the left upper lung lobe, we also observed extra lung tissue with notable interstitial markings and the presence …

ObjectivesVarious genomic alterations and genomic signatures, including ERBB2 amplification, mutations in PIK3CA, AKT1, and ESR1, and tumor mutational burden (TMB), have become important biomarkers for treatment selection in breast cancer (BC). This study aimed to investigate the mutational features of Chinese early-stage BC patients.MethodsTumors and matched blood samples collected from 589 Chinese patients with early-stage BC were sequenced using a commercial gene panel consisting of 520 cancer-related genes to analyze all types of genomic alterations and estimate the TMB status.ResultsA total of 18 genes were found to be more frequently mutated (P<0.05) or amplified (P<0.05) in stage T3–4 tumors as compared with T1–2 tumors. A total of 18 genes were found to be differentially mutated (P<0.05) or amplified (P<0.05) in patients with lymph node metastasis than those without lymph node metastasis. Younger patients (≤35 years) were more frequently identified with mutations or gene amplifications in eleven genes (P<0.05). TMB >10mutations/Mb were found in 5.7% of our cohort. Although the TMB was similar for various molecular subtypes between our cohort and the BC cohort of The Cancer Genome Atlas (TCGA) study, the TMB were statistically different for HR+/HER-, HR+/HER2+, and triple-negative subtypes between our cohort and African Americans in the TCGA study. As compared to the TCGA BC cohort, our cohort had a much earlier median age of diagnosis (48 vs. 58 years, P<0.001), and had significantly lower frequency of triple-negative subtype (11.5% vs. 18.4%, P<0.001) and invasive lobular BC (2.4% vs. 19.0%, P<0.001). Further subgroup analyses revealed that mutation rates in various genes including TP53, ERBB2, and PIK3CA were distinct for patients who were younger (≤35 years), had triple-negative or invasive lobular BC in our cohort than in the TCGA cohort.ConclusionsThis study revealed distinct mutational features of various molecular subtypes of early-stage BC among Chinese patients. Moreover, we provide new insights into the differences in early-stage BC between the East and West.

This study evaluated the efficacy and safety of anlotinib combined with programmed cell death protein 1 (PD-1) blockade for the treatment of small-cell lung cancer (SCLC) and non-small-cell lung cancer (NSCLC). SCLC (n = 28) and NSCLC (n = 177) patients who received treatment at Hunan Cancer Hospital between June 1, 2019, and July 1, 2020, were retrospectively analyzed. Progression-free survival (PFS) and treatment responses were compared among patients who received combination therapy of anlotinib plus PD-1 inhibitor, or monotherapy of either chemotherapy or PD-1 inhibitor. Independent prognostic factors were identified by Cox regression analysis. Patients with relapsed SCLC who received anlotinib plus PD-1 inhibitor as a ≥ second-line therapy (n = 14) had a significantly longer PFS than those who received PD-1 inhibitor alone (n = 14, 5.0 vs. 3.0 months; P = 0.005). For patients with previously untreated wild-type NSCLC, the combination therapy in the first-line setting (n = 6) provided a marginally longer PFS than mono-chemotherapy (n = 6, 8.0 vs. 3.0 months; P = 0.075). For patients with relapsed NSCLC, the combination therapy in the  ≥ second-line setting (n = 62) resulted in significantly higher objective response rate (19.3 vs. 5.0 vs. 2.4%; P = 0.013) and longer PFS (8.0 vs. 2.0 vs. 2.0 months; P <0.001) as compared to monotherapy of either chemotherapy (n = 41) or PD-1 inhibitor (n = 62). Anlotinib and PD-1 blockade combination therapy was an independent predictive factor of longer PFS (P <0.001). The combination of anlotinib and PD-1 inhibitor has promising efficacy and manageable toxicity as a second- or later-line treatment of relapsed NSCLC and possibly for relapsed SCLC.

BACKGROUND:Serial profiling of circulating tumor DNA (ctDNA) could reflect dynamic molecular changes in response to treatment and potentially predict impending disease progression (PD). Herein, we investigated the molecular factors and dynamic changes in ctDNA that can serve as predictors of survival outcomes of patients with epidermal growth factor receptor (EGFR)-mutated advanced non-small cell lung cancer (NSCLC) who received osimertinib therapy after progression from prior EGFR inhibitor regimen.METHODS:Capture-based targeted sequencing was performed on the baseline and longitudinal plasma samples collected from 72 and 57 patients, respectively, using a 168-gene panel.RESULTS:Analysis revealed that inferior overall survival (OS) was correlated with various baseline molecular features including high allelic fraction (AF) of EGFR sensitizing mutations (P=0.045), high maximum AF (maxAF, P=0.060), or harboring concurrent genomic alterations such as copy number amplification (CNA) in EGFR (P=0.026) or in other genes (P=0.026), and genes involved in the cell cycle (P=0.004) or TP53 signaling pathway (P=0.032). Moreover, ctDNA clearance at first follow-up after 6 weeks of osimertinib therapy was correlated with significantly longer progression-free survival (PFS) (P=0.022) and OS (P=0.009). Molecular PD, reflected by the emergence of new mutation or increased AF of existing mutations, was detected at an average lead time of 2.5 months prior to radiological PD. Patients with molecular PD were more likely to harbor CNA (P=0.035) and TP53 mutations (P=0.023).CONCLUSIONS:Molecular factors derived from serial ctDNA profiling can serve as predictive and prognostic markers, which could allow early detection of PD, preceding imaging modalities by 2.5 months.

EGFR exon 18 G724S mutation has been described to be associated with cancer progression and mediating resistance to first- and third-generation EGFR tyrosine kinase inhibitors(TKIs). In vitro experiments have provided compelling evidence that EGFR G724S retains its sensitivity for second-generation inhibitors, including afatinib. However, limited data has reported the clinical efficacy of afatinib in Non-small cell lung cancer (NSCLC) patients harboring EGFR G724S mutation. In our study, we aimed to investigate the treatment response and survival outcome of advanced lung adenocarcinoma patients with EGFR G724S mutation who were treated with afatinib.

Abstract Growing efforts are being invested in investigating various molecular approaches to detect minimal residual disease (MRD) and predict disease recurrence. In our study, we investigated the utility of parallel longitudinal analysis of mutation and DNA methylation profiles for predicting MRD in postoperative non‐small‐cell lung cancer (NSCLC) patients. Tumor tissues and longitudinal blood samples were obtained from 65 patients with resected stage IA‐IIIB NSCLC. Somatic mutation and DNA methylation profiling were performed using ultra‐deep targeted sequencing and targeted bisulfite sequencing, respectively. Dynamic changes in plasma‐based mutation and tumor‐informed methylation profiles, reflected as MRD score, were observed from before surgery (baseline) to postoperative follow‐up, reflecting the decrease in tumor burden of the patients with resected NSCLC. Mutations were detected from plasma samples in 63% of the patients at baseline, which significantly reduced to 23‐25% during post‐operative follow‐ups. MRD score positive rate was 95.7% at baseline, which reduced to 74% at the first and 70% at the second follow‐up. Among the 5 relapsed patients with parallel longitudinal analysis of mutation and methylation profile, elevated MRD score was observed at follow‐up between 0.5‐7 months prior to radiologic recurrence for all 5 patients. Of them, 4 patients also had concomitant increase in allelic fraction of mutations in at least 1 follow‐up time point, but one patient had no mutation detected throughout all follow‐ups. Our results demonstrate that longitudinal profiling of mutation and DNA methylation may have potential for detecting MRD and predicting recurrence in postoperative NSCLC patients.

BackgroundMET proto-oncogene amplification (amp) is an important mechanism underlying acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs). However, the optimal treatment strategy after acquiring MET-amp-mediated EGFR-TKI resistance remains controversial. Our study compared three treatment strategies for patients with EGFR-mutant non-small-cell lung cancer (NSCLC) who were detected with MET-amp at EGFR-TKI progression using next-generation sequencing. MethodsOf the 70 patients included in the study, 38 received EGFR-TKI + crizotinib, 10 received crizotinib monotherapy, and 22 received chemotherapy. Clinical outcomes and molecular profiles were analyzed. ResultsThe objective response rate was 48.6% for EGFR-TKI + crizotinib group, 40.0% for crizotinib monotherapy group, and 18.2% for chemotherapy group. Patients who received EGFR-TKI + crizotinib had significantly longer progression-free survival than those who received crizotinib or chemotherapy (5.0 vs. 2.3 vs. 2.9 months, p = 0.010), but overall survival was comparable (10.0 vs. 4.1 vs. 8.5 months, p = 0.088). TP53 mutation (58.5%) and EGFR-amp (42.9%) were frequent concurrent mutations of the cohort. Progression-free survival was significantly longer for patients with either concurrent TP53 mutation (n = 17) (6.0 vs. 2.3 vs. 2.9 months, p = 0.009) or EGFR-amp (n = 13) (5.0 vs. 1.2 vs. 2.4 months, p = 0.016) in the EGFR-TKI + crizotinib group than the other two regimen. Potential acquired resistance mechanisms to EGFR-TKI + crizotinib included EGFR-T790M (n = 2), EGFR-L718Q (n = 1), EGFR-S645C (n = 1), MET-D1228H (n = 1), BRAF-V600E (n = 1), NRAS-Q61H (n = 1), KRAS-amp (n = 1), ERBB2-amp (n = 1), CDK4-amp (n = 1), and MYC-amp (n = 1). ConclusionOur study provides real-world clinical evidence from a large cohort that simultaneous inhibition of EGFR and MET could be a more effective therapeutic strategy for patients with MET-amp acquired from EGFR-TKI therapy.

Background The combination of bevacizumab and epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) could prolong progression-free survival (PFS) in patients with EGFR -mutant advanced non-small-cell lung cancer (NSCLC). Our study investigated the clinical and molecular factors that affect the efficacy of first-generation EGFR-TKI with or without bevacizumab and identify the subset of patients who can benefit from combination therapy. Methods Our study included 318 patients with EGFR -mutant locally advanced/advanced NSCLC treated with either first-generation EGFR-TKI combined with bevacizumab (A+T; n = 159) or EGFR-TKI monotherapy (T; n = 159). Two nomogram models to predict PFS and overall survival (OS), respectively, were constructed using two factors that impact EGFR-TKI efficacy: metastatic site and presence of concurrent mutations. The study cohort was stratified into 2 cohorts for training ( n = 176) and validation ( n = 142) of the nomogram model. Using the median score from the nomogram, the patients were stratified into two groups to analyze their survival outcome. Results The A+T group had significantly longer PFS (14.0 vs. 10.5 months; p < 0.001) and OS (37.0 vs. 26.0 months; p = 0.042) than the T group. Among the patients with concurrent mutations in tumor suppressor genes, those in the A+T group had significantly longer PFS and OS than the T group (PFS 14.5 vs. 8.0 months, p < 0.001; OS 39.0 vs. 20.0 months, p = 0.003). The higher scores from the nomograms were associated with the presence of brain/liver/pleural metastasis or concomitant gene mutations, which indicated a higher likelihood of shorter PFS and OS. The validation of the nomogram revealed that patients with lower scores had significantly longer PFS for the T group than those with higher scores (15.0 vs. 9.0 months, p = 0.002), but not for the A+T group (15.9 vs. 13.9 months, p = 0.256). Conclusions Using a nomogram, our study demonstrated that the addition of bevacizumab may enhance the therapeutic effectiveness of EGFR-TKI by overcoming the negative impact of certain clinical and molecular factors on the efficacy of EGFR-TKI.

Background Germline DNA mismatch repair (MMR) gene aberrations are associated with colorectal cancer (CRC) predisposition and high tumor mutation burden (TMB-H), with increased likelihood of favorable response to immune checkpoint inhibitors (ICIs). Case presentation We present a 32-year old male patient diagnosed with constitutional MMR deficiency (CMMRD) CRC whose MMR immunohistochemistry (IHC) revealed inconsistent results from two tumor blocks. Targeted sequencing of two tumor specimens used in MMR-IHC and plasma-derived circulating tumor DNA consistently revealed the detection of bi-allelic germline MSH6 c.3226C > T (p.R1076C) mutation, TMB-H as well as the genetic heterogeneity of the tumor samples. Unexpectedly, both blocks were microsatellite stable (MSS) after PCR confirmation. Interestingly, the patient failed to show response to ICI monotherapy or dual therapy, but clinically benefitted from combined therapy of ICI pembrolizumab plus multi-kinase inhibitor regorafenib. Conclusion Our case reported a CMMRD patient with heterogeneous MMR results who showed complicated response to ICIs, highlighting the importance of accurate diagnosis using targeted sequencing with multiple specimens to reveal the possible mechanism of response to ICI in patients with CMMRD.

NSCLC harboring gene rearrangements involving the ALK and retaining the ALK tyrosine kinase domain benefits from the antitumor activity generated by various ALK inhibitors, including crizotinib, alectinib, and brigatinib.1Kwak E.L. Bang Y.J. Camidge D.R. et al.Anaplastic lymphoma kinase inhibition in non-small-cell lung cancer [published correction appears in N Engl J Med. 2011;364:588].N Engl J Med. 2010; 363: 1693-1703Crossref PubMed Scopus (3743) Google Scholar, 2Hida T. Nokihara H. Kondo M. et al.Alectinib versus crizotinib in patients with ALK-positive non-small-cell lung cancer (J-ALEX): an open-label, randomised phase 3 trial.Lancet. 2017; 390: 29-39Abstract Full Text Full Text PDF PubMed Scopus (595) Google Scholar, 3Mok T. Camidge D.R. Gadgeel S.M. et al.Updated overall survival and final progression-free survival data for patients with treatment-naive advanced ALK-positive non-small-cell lung cancer in the ALEX study.Ann Oncol. 2020; 31: 1056-1064Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar Despite durable responses, as with all targeted therapeutics, most patients treated with crizotinib or alectinib would eventually experience disease progression. In this issue of the Journal of Thoracic Oncology, Nishio et al.4Nishio M. Yoshida T. Kumagai T. et al.Brigatinib in Japanese patients with ALK-positive NSCLC previously treated with alectinib and other tyrosine kinase inhibitors: outcomes of the phase 2 J-ALTA trial.J Thorac Oncol. 2021; 16: 452-463Abstract Full Text Full Text PDF PubMed Scopus (23) Google Scholar reported the promising results from the J-ALTA study, a phase 2, single-arm study, and the first prospective clinical study that evaluated the efficacy and safety of brigatinib in patients with advanced ALK-rearranged NSCLC who progressed from previous alectinib and other ALK inhibitor therapies. Among the 47 Japanese patients who received brigatinib at the standard dose after progression from alectinib with or without history of crizotinib use, objective response rate (ORR) and disease control rate (DCR) were 34% (16 of 47) and 79% (37 of 47), respectively, with a median progression-free survival (PFS) of 7.3 months. Brigatinib is a newer second-generation selective inhibitor that has been found to have clinical efficacy and safety in patients with treatment-naive and crizotinib-refractory ALK-positive NSCLC.5Kim D.W. Tiseo M. Ahn M.J. et al.Brigatinib in patients with crizotinib-refractory anaplastic lymphoma kinase-positive non-small-cell lung cancer: a randomized, multicenter phase II trial.J Clin Oncol. 2017; 35: 2490-2498Crossref PubMed Scopus (416) Google Scholar, 6Camidge D.R. Kim H.R. Ahn M.J. et al.Brigatinib versus crizotinib in ALK-positive non-small-cell lung cancer.N Engl J Med. 2018; 379: 2027-2039Crossref PubMed Scopus (476) Google Scholar, 7Huber R.M. Hansen K.H. Paz-Ares Rodríguez et al.Brigatinib in crizotinib-refractory ALK+ NSCLC: 2-year follow-up on systemic and intracranial outcomes in the phase 2 ALTA trial.J Thorac Oncol. 2020; 15: 404-415Abstract Full Text Full Text PDF PubMed Scopus (77) Google Scholar In crizotinib-refractory NSCLCs, brigatinib at the standard dose of 180 mg once daily with a 7-day lead-in at 90 mg had an ORR of 56%, robust PFS of 16.7 months, and overall survival (OS) of 34.1 months (n = 110).7Huber R.M. Hansen K.H. Paz-Ares Rodríguez et al.Brigatinib in crizotinib-refractory ALK+ NSCLC: 2-year follow-up on systemic and intracranial outcomes in the phase 2 ALTA trial.J Thorac Oncol. 2020; 15: 404-415Abstract Full Text Full Text PDF PubMed Scopus (77) Google Scholar Moreover, the ALTA trial reported an intracranial ORR of 67% (12 of 18) and duration of intracranial response of 16.6 months among 18 patients with measurable brain lesions who received brigatinib after crizotinib progression.7Huber R.M. Hansen K.H. Paz-Ares Rodríguez et al.Brigatinib in crizotinib-refractory ALK+ NSCLC: 2-year follow-up on systemic and intracranial outcomes in the phase 2 ALTA trial.J Thorac Oncol. 2020; 15: 404-415Abstract Full Text Full Text PDF PubMed Scopus (77) Google Scholar In contrast, among the 14 patients with measurable brain metastasis at baseline who received brigatinib after alectinib progression from the J-ALTA study, the intracranial ORR and DCR were much lower at only 21% (three of 14) and 93% (13 of 14), respectively.4Nishio M. Yoshida T. Kumagai T. et al.Brigatinib in Japanese patients with ALK-positive NSCLC previously treated with alectinib and other tyrosine kinase inhibitors: outcomes of the phase 2 J-ALTA trial.J Thorac Oncol. 2021; 16: 452-463Abstract Full Text Full Text PDF PubMed Scopus (23) Google Scholar The efficacy of brigatinib as reflected by the overall response and intracranial response of alectinib-refractory NSCLCs from the J-ALTA study seems low as compared with the reported efficacy in crizotinib-refractory NSCLCs in the ALTA trial. However, the observations from the J-ALTA study are more promising than the observed clinical outcomes from the retrospective study conducted by Lin et al.8Lin J.J. Zhu V.W. Schoenfeld A.J. et al.Brigatinib in patients with alectinib-refractory ALK-positive NSCLC.J Thorac Oncol. 2018; 13: 1530-1538Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar They suggested that brigatinib had limited clinical activity in patients with alectinib-refractory, ALK-positive NSCLC on the basis of the low ORR of 17% (three of 22), DCR of 55% (12 of 22), and short median PFS of 4.4 months.8Lin J.J. Zhu V.W. Schoenfeld A.J. et al.Brigatinib in patients with alectinib-refractory ALK-positive NSCLC.J Thorac Oncol. 2018; 13: 1530-1538Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar As discussed by Nishio et al.,4Nishio M. Yoshida T. Kumagai T. et al.Brigatinib in Japanese patients with ALK-positive NSCLC previously treated with alectinib and other tyrosine kinase inhibitors: outcomes of the phase 2 J-ALTA trial.J Thorac Oncol. 2021; 16: 452-463Abstract Full Text Full Text PDF PubMed Scopus (23) Google Scholar the discrepancy in their observations was brought on by the following two main factors: the difference in the treatment history (18% of patients received at least 3 ALK inhibitor previously in a retrospective study versus none in the J-ALTA study) and the difference in baseline clinical characteristics of the patients, particularly the higher rate of baseline brain metastasis (82% in retrospective study versus 40% in the J-ALTA study).8Lin J.J. Zhu V.W. Schoenfeld A.J. et al.Brigatinib in patients with alectinib-refractory ALK-positive NSCLC.J Thorac Oncol. 2018; 13: 1530-1538Abstract Full Text Full Text PDF PubMed Scopus (46) Google Scholar Approximately 25% of untreated patients diagnosed with having advanced NSCLCs present with brain metastasis at initial diagnosis9Cagney D.N. Martin A.M. Catalano P.J. et al.Incidence and prognosis of patients with brain metastases at diagnosis of systemic malignancy: a population-based study.Neuro Oncol. 2017; 19: 1511-1521Crossref PubMed Scopus (262) Google Scholar and between 20% and 40% of patients with NSCLC will develop brain metastasis during the disease course.10Mujoomdar A. Austin J.H. Malhotra R. et al.Clinical predictors of metastatic disease to the brain from non-small cell lung carcinoma: primary tumor size, cell type, and lymph node metastases.Radiology. 2007; 242: 882-888Crossref PubMed Scopus (161) Google Scholar Therefore, it is important to monitor and evaluate the therapeutic efficacy not only in the primary lung lesions but also in the brain lesions. Nevertheless, brigatinib has been found to have substantial intracranial responses in patients with baseline brain metastasis who previously received crizotinib or alectinib.4Nishio M. Yoshida T. Kumagai T. et al.Brigatinib in Japanese patients with ALK-positive NSCLC previously treated with alectinib and other tyrosine kinase inhibitors: outcomes of the phase 2 J-ALTA trial.J Thorac Oncol. 2021; 16: 452-463Abstract Full Text Full Text PDF PubMed Scopus (23) Google Scholar,7Huber R.M. Hansen K.H. Paz-Ares Rodríguez et al.Brigatinib in crizotinib-refractory ALK+ NSCLC: 2-year follow-up on systemic and intracranial outcomes in the phase 2 ALTA trial.J Thorac Oncol. 2020; 15: 404-415Abstract Full Text Full Text PDF PubMed Scopus (77) Google Scholar,11Camidge D.R. Kim D.W. Tiseo M. et al.Exploratory analysis of brigatinib activity in patients with anaplastic lymphoma kinase-positive non-small-cell lung cancer and brain metastases in two clinical trials.J Clin Oncol. 2018; 36: 2693-2701Crossref PubMed Scopus (93) Google Scholar We also anticipate the results from the ongoing global phase 2 ALTA-2 trial (NCT03535740) evaluating the efficacy of brigatinib after progression from alectinib or ceritinib.12Kim E.S. Ou S.H.I. Barlesi F. et al.Phase 2 study of brigatinib in patients (pts) with anaplastic lymphoma kinase (ALK)–positive, advanced non–small cell lung cancer (NSCLC) that progressed on alectinib or ceritinib.J Clin Oncol. 2019; 37 (TPS9115–TPS9115)Google Scholar In preclinical studies, brigatinib potently inhibited a broad array of ALK mutants including secondary ALK mutations associated with resistance to crizotinib, ceritinib, or alectinib.13Zhang S. Anjum R. Squillace R. et al.The potent ALK inhibitor brigatinib (AP26113) overcomes mechanisms of resistance to first- and second-generation ALK inhibitors in preclinical models.Clin Cancer Res. 2016; 22: 5527-5538Crossref PubMed Scopus (212) Google Scholar Among the 16 ALK mutations tested including mutations in positions I1171, L1196, and G1202 typically acquired during crizotinib, ceritinib, or alectinib therapy, brigatinib had robust inhibitory activity against all mutants, with G1202R being the most resistant.13Zhang S. Anjum R. Squillace R. et al.The potent ALK inhibitor brigatinib (AP26113) overcomes mechanisms of resistance to first- and second-generation ALK inhibitors in preclinical models.Clin Cancer Res. 2016; 22: 5527-5538Crossref PubMed Scopus (212) Google Scholar In the J-ALTA study, brigatinib has efficacy in patients harboring L1196M, I1171N, V1180L, and even G1202R. It would be interesting to perform a subanalysis of the efficacy for each secondary mutation or ALK fusion partner. Furthermore, it would be clinically meaningful to identify other molecular factors that could affect the treatment efficacy of brigatinib, such as the presence of nonreciprocal/reciprocal ALK translocations, which affect crizotinib efficacy.14Zhang Y. Zeng L. Zhou C. et al.Detection of nonreciprocal/reciprocal ALK translocation as poor predictive marker in patients with first-line crizotinib-treated ALK-rearranged NSCLC.J Thorac Oncol. 2020; 15: 1027-1036Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar Other unanswered questions are the resistance mechanisms acquired after progression with brigatinib post alectinib or crizotinib and whether the currently available ALK inhibitors are sufficient in overcoming the resistance. Currently, no large-scale analysis of resistance mechanisms with brigatinib therapy has been reported. In People's Republic of China, owing to the health insurance coverage, crizotinib, and alectinib remain as the standard of care for Chinese patients with ALK-rearranged NSCLC, whereas alectinib is the subsequent treatment after crizotinib progression. As per the guidelines of the National Comprehensive Cancer Network, alectinib is now the standard of care for ALK-rearranged NSCLCs in Japan and the rest of the world. The promising results from the J-ALTA trial are clinically significant and contribute a step in favor of brigatinib as the treatment option after alectinib progression with or without previous crizotinib therapy. However, the question still remains, what then would be the optimal treatment option after developing progression from brigatinib? With the approval of brigatinib as first-line therapy, the question on the optimal treatment sequencing of ALK inhibitors still remains. The question on the treatment sequencing of ALK inhibitors lies on which specific sequence would yield the best clinical outcome. From the J-ALTA and ALTA studies, brigatinib therapy confers different clinical outcomes in alectinib-refractory and crizotinib-refractory NSCLCs, suggesting the choice of ALK inhibitors in various treatment settings is critical in long-term survival outcome. More clinical studies with larger sample size are required to validate their findings. However, because most ALK inhibitors have just become commercially available, the OS data are still largely immature. Hence, in the future, it would be interesting to compare the cumulative PFS or OS with sequential use or systematic use of various ALK inhibitors to help identify the most optimal treatment algorithm that could offer the best clinical outcomes. Dr. Zhang contributed to the conceptualization, organization, data collection, auditing, project administration, and writing—reviewing and editing. Drs. Song and Xu contributed to data curation and writing—reviewing and editing. Dr. Lizaso contributed to formal analysis and writing—reviewing and editing. All authors approved the final version of the manuscript. Brigatinib in Japanese Patients With ALK-Positive NSCLC Previously Treated With Alectinib and Other Tyrosine Kinase Inhibitors: Outcomes of the Phase 2 J-ALTA TrialJournal of Thoracic OncologyVol. 16Issue 3PreviewThis phase 2 trial evaluated the efficacy and safety of brigatinib in patients with advanced ALK-positive NSCLC refractory to alectinib or other ALK tyrosine kinase inhibitors (TKIs). Full-Text PDF Open Access

Abstract Introduction: Two phase 3 clinical studies, NEJ026 and CTONG1509, have consistently demonstrated better anti-tumor activity of adding bevacizumab to erlotinib regimen in the treatment of patients with epidermal growth factor receptor (EGFR)-mutant advanced non-squamous non-small-cell lung cancer (NSCLC); however, the subset of patients that benefits more with the addition of bevacizumab to their EGFR-TKI regimen still remains to be identified. In this study, we aimed to investigate the clinical and molecular factors that affect the efficacy of first-generation EGFR-TKI regimen with or without bevacizumab as first-line treatment of patients with EGFR-mutant NSCLC. Methods: A total of 176 patients with EGFR-mutant stage IIIB-IV relapsed or metastatic NSCLC and received first-generation EGFR-TKI gefitinib or erlotinib monotherapy (T; n=88) or combined with bevacizumab (A+T; n=88) were included in our study. The two groups had comparable baseline clinical characteristics. Retrospective analysis was performed on clinical data, survival outcomes, and mutation profiles at baseline and progression. A nomogram model to predict PFS was constructed with the following variables identified from Cox multivariate analysis: treatment group (A+T) and mutations in EGFR L858R, exon 19 deletion, TP53, CDK4, FGF3, and NFkB signaling pathway. Another nomogram was constructed to predict overall survival with treatment group (A+T) and mutations in TP53 signaling pathway as the variables. Results: As compared to the T group, progression-free survival (PFS) was significantly longer among patients who received A+T and harbored concurrent EGFR amplification (16.1 vs 9.0 months; HR: 0.267, 95%CI: 0.110-0.651; p=0.004), and hotspot and loss-of-function mutations in TP53 (16.1 vs 8.0 months; HR: 0.215, 95%CI: 0.108-0.426; p<0.001). At progression, EGFR T790M, detected from 35% in the A+T group and 42% in the T group, was the major resistance mechanism in both groups. No statistical difference was found between the acquired resistance mechanisms of the two groups (p=0.787). Cox multivariate analysis demonstrated that receiving A+T is an independent predictor of longer PFS (HR: 0.51, 95%CI: 0.35-0.75; p=0.001) and overall survival (HR: 0.62, 95%CI: 0.38-0.99; p=0.045). Cross-validation of both nomograms using data from 144 evaluable patients in the cohort stratified into low-risk and high-risk groups based on the variables predicted the risk for disease progression (c-index: 0.676; p<0.001) and the risk for poor prognosis (c-index: 0.593; p<0.001) of the high-risk groups. Conclusions: Our study demonstrated the clinical value of the nomogram model in predicting the subset of patients who benefits from first-generation EGFR-TKI regimen with or without bevacizumab. Citation Format: Yongchang Zhang, Analyn Lizaso. Molecular features-based model for predicting benefit from Bevacizumab combined with first-generation EGFR-tyrosine kinase inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 377.