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Human epidermal growth factor receptor 2 (HER2) aberrations are observed in various cancers. In non-small cell lung cancer, genetic alterations activating HER2, mostly exon 20 insertion mutations, occur in approximately 2-4% of cases. Trastuzumab deruxtecan (T-DXd), a HER2-targeted antibody-drug conjugate has been approved as the first HER2-targeted drug for HER2-mutant lung cancer. However, some cases are not responsive to T-DXd and the primary resistant mechanism remains unclear. In this study, we assessed sensitivity to T-DXd in JFCR-007, a patient-derived HER2-mutant lung cancer cell line. Although JFCR-007 was sensitive to HER2 tyrosine kinase inhibitors, it showed resistance to T-DXd in attachment or spheroid conditions. Accordingly, we established a three-dimensional (3D) layered co-culture model of JFCR-007, where it exhibited a lumen-like structure and became sensitive to T-DXd. In addition, an in-house inhibitor library screening revealed that G007-LK, a tankyrase inhibitor, was effective when combined with T-DXd. G007-LK increased the cytotoxicity of topoisomerase-I inhibitor, DXd, a payload of T-DXd and SN-38. This combined effect was also observed in H2170, an HER2-amplified lung cancer cell line. These results suggest that the proposed 3D co-culture system may help in evaluating the efficacy of T-DXd and may recapitulate the tumor microenvironment., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Ryohei Katayama reports financial support was provided by Japan Society for the Promotion of Science. Ryohei Katayama reports financial support was provided by Japan Agency for Medical Research and Development. Ryohei Katayama reports financial support was provided by the Nakatomi Foundation. Ryohei Katayama reports financial support was provided by Nippon Foundation. Ryohei Katayama reports financial support was provided by Takeda Science Foundation. Ryohei Katayama reports financial support was provided by Kobayashi Foundation for Cancer Research. Ken Uchibori reports financial support was provided by Japan Society for the Promotion of Science. Akari Shiraishi reports financial support was provided by Japan Science and Technology Agency. Ryohei Katayama reports a relationship with Chugai Pharmaceutical Co Ltd that includes: funding grants. Ryohei Katayama reports a relationship with Toppan Inc that includes: funding grants. Yuki Takahashi reports a relationship with Toppan Inc that includes: employment. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Recently approved RET tyrosine kinase inhibitors (TKIs) have shown promising therapeutic effects against RET-rearranged non-small cell lung cancer (NSCLC) or RET-mutated thyroid cancer. However, resistance develops, limiting long-term efficacy. Although many RET-TKI resistance mechanisms, such as secondary mutations in RET or activation of bypass pathways, are known, some primary or acquired resistance mechanisms are unclear. Here, human genome-wide CRISPR/Cas9 screening was performed to identify genes related to drug-tolerant persister cells. Patient-derived cells with RET-fusion were introduced genome-wide sgRNA library and treated with RET-TKI for 9 days, resulting in the discovery of several candidate genes. Knockout of MED12 or MIG6 significantly increased residual drug-tolerant persister cells under RET-TKI treatment. MIG6 loss induced significant EGFR activation even with low concentrations of EGFR ligands and led to resistance to RET-TKIs. EGFR inhibition with afatinib or cetuximab in combination with RET TKIs was effective in addressing drug persistence. By contrast, a KIF5B-RET positive cells established from a RET-rearranged NSCLC patient, showed significant resistance to RET-TKIs and high dependence on EGFR bypass signaling. Consistently, knocking out EGFR or RET led to high sensitivity to RET or EGFR inhibitor respectively. Here, we have provided a comprehensive analysis of adaptive and acquired resistance against RET-rearranged NSCLC., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests.R. Katayama received research grants from Chugai, and TOPPAN. The other authors have no conflicts of interest to declare., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: It is difficult to predict gene mutations individually based on clinical background alone. Tumor markers may help to predict each gene mutation. Identifying tumor markers that can predict gene mutation will facilitate timely genetic testing and intervention., Methods: We selected 134 cases of advanced or recurrent ALK-positive and 172 cases of advanced or recurrent EGFR-positive lung cancer from our clinical database. The cutoff values for the tumor markers were defined as 5.0 ng/mL or higher for carcinoembryonic antigen (CEA) and 3.5 ng/mL or higher for soluble fragment of cytokeratin 19 (CYFRA21-1) in accordance with the institutional standards. A positive CYFRA21-1:CEA ratio was defined as 0.7 or higher., Results: The CEA-positivity rate was 49% for ALK-positive lung cancers and 73% for EGFR-positive lung cancers, which was significantly different (p < 0.001). The CYFRA21-1 positivity rate was significantly higher in ALK-positive lung cancer (36%) compared with EGFR-positive lung cancer (23%) (p = 0.034). The median CYFRA21-1:CEA ratio was 0.395 for the ALK group, which was significantly higher compared with 0.098 for the EGFR group (p < 0.001). These trends were similar when excluding histology other than adenocarcinoma. The median time-to-treatment failure (TTF) for initial tyrosine kinase inhibitor (TKI) therapy was 308 days for the high CYFRA21-1:CEA ratio group and 617 days for the low CYFRA21-1:CEA ratio group for ALK-positive lung cancer (p = 0.100)., Conclusions: A higher proportion of patients with ALK-positive lung cancer were CYFRA21-1 positive and had higher CYFRA21-1:CEA ratios compared with EGFR-positive lung cancer patients., (© 2024 The Authors. Thoracic Cancer published by John Wiley & Sons Australia, Ltd.)

Brigatinib-based therapy was effective against osimertinib-resistant EGFR C797S mutants and is undergoing clinical studies. However, tumor relapse suggests additional resistance mutations might emerge. Here, we first demonstrated the binding mode of brigatinib to the EGFR-T790M/C797S mutant by crystal structure analysis and predicted brigatinib-resistant mutations through a cell-based assay including N-ethyl-N-nitrosourea (ENU) mutagenesis. We found that clinically reported L718 and G796 compound mutations appeared, consistent with their proximity to the binding site of brigatinib, and brigatinib-resistant quadruple mutants such as EGFR-activating mutation/T790M/C797S/L718M were resistant to all the clinically available EGFR-TKIs. BI-4020, a fourth-generation EGFR inhibitor with a macrocyclic structure, overcomes the quadruple and major EGFR-activating mutants but not the minor mutants, such as L747P or S768I. Molecular dynamics simulation revealed the binding mode and affinity between BI-4020 and EGFR mutants. This study identified potential therapeutic strategies using the new-generation macrocyclic EGFR inhibitor to overcome the emerging ultimate resistance mutants., (© 2024. The Author(s).)

Background: As an epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI), osimertinib has emerged as a standard EGFR-mutation positive treatment for non-small cell lung cancer (NSCLC). However, the efficacy of osimertinib for malignant pleural effusion (MPE) remains understudied. This study aimed to evaluate the impact of osimertinib on time to treatment failure (TTF) and overall survival (OS) in patients with EGFR-mutation positive NSCLC, comparing those with and without MPE., Methods: This retrospective analysis included patients with advanced or recurrent NSCLC treated with osimertinib at our hospital between April 2016 and June 2021. TTF was defined as the duration from osimertinib initiation to discontinuation, and OS as the duration until death, irrespective of the reason., Results: Among 229 patients receiving osimertinib, 84 had MPE before administration, 39 acquired EGFR exon20 T790M mutation following previous EGFR-TKI therapy, and 45 were EGFR-TKI-naive. Among EGFR-TKI-naive patients, median TTF was 14.8 and 19.8 months for those with and without MPE, respectively (hazard ratio [HR] 1.40; 95% confidence interval [CI]: 0.90-2.18; p = 0.12). Median OS was 32.0 and 42.0 months for patients with and without MPE, respectively (HR 1.43; 95% CI: 0.86-2.38; p = 0.16). Among patients with T790M mutation, median TTF was 12.3 and 13.1 months for patients with and without MPE, respectively (HR 1.03; 95% CI: 0.69-1.55; p = 0.88). Median OS for patients with and without MPE was 23.2 and 24.7 months, respectively (HR 1.09; 95% CI: 0.72-1.67; p = 0.68)., Conclusion: Among patients with EGFR-mutation positive NSCLC, the evidence of MPE has little effect on survival with osimertinib., (© 2024 The Authors. Thoracic Cancer published by John Wiley & Sons Australia, Ltd.)

Although tyrosine kinase inhibitor (TKI) therapy shows marked clinical efficacy in patients with anaplastic lymphoma kinase-positive (ALK+) and ROS proto-oncogene 1-positive (ROS1+) non-small cell lung cancer (NSCLC), most of these patients eventually relapse with acquired resistance. Therefore, genome-wide CRISPR/Cas9 knockout screening was performed using an ALK+ NSCLC cell line established from pleural effusion without ALK-TKI treatment. After 9 days of ALK-TKI therapy, sequencing analysis was performed, which identified several tumor suppressor genes, such as NF2 or MED12, and multiple candidate genes. Among them, this study focused on ERRFI1, which is known as MIG6 and negatively regulates EGFR signaling. Interestingly, MIG6 loss induced resistance to ALK-TKIs by treatment with quite a low dose of EGF, which is equivalent to plasma concentration, through the upregulation of MAPK and PI3K/AKT/mTOR pathways. Combination therapy with ALK-TKIs and anti-EGFR antibodies could overcome the acquired resistance in both in vivo and in vitro models. In addition, this verified that MIG6 loss induces resistance to ROS1-TKIs in ROS1+ cell lines. This study found a potentially novel factor that plays a role in ALK and ROS1-TKI resistance by activating the EGFR pathway with low-dose ligands.

Background: KRAS mutation positive lung cancer is known to be clinically characterized by older age, males, and smokers. It is reported to be more common in mucinous adenocarcinoma, but all reports are based on analysis of tissue samples. Recently, blood samples have become available for analysis, suggesting a low detection rate of circulating tumor DNA in histological types, especially mucinous adenocarcinoma. In this study, we investigated the clinical characteristics of KRAS mutation-positive cases in the analysis of blood specimens, as these remain unclear., Methods: The clinical background of patients with KRAS mutation among those who underwent next-generation sequencing (NGS) analysis using blood samples was evaluated., Results: NGS analysis was performed on 214 blood samples. KRAS mutations were detected in blood samples in 33 cases (15.4%), of which 31 cases (14.5%) had a histological pathology diagnosis. Mucinous adenocarcinoma accounted for 28.6% of cases with positive blood and tissue specimens, 10.0% of cases with positive blood specimens only, and 57.1% of cases with positive tissue specimens only. Mucinous adenocarcinoma tended to be less common in cases with positive blood specimens. In KRAS-positive patients with lung metastasis only, only one nonmucinous adenocarcinoma had a positive blood sample, and the others all had mucinous adenocarcinomas with positive tissue samples only., Conclusion: The results showed that the detection rate of KRAS-positive lung cancers detected by blood and tissue samples differs, and that the detection rate of blood samples may be poor, especially in the case of mucinous adenocarcinoma with lung metastases only., (© 2023 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Soluble interleukin-2 receptor (sIL-2R) suppresses effector T-cells. Few studies have assessed serum sIL-2R in patients receiving immunotherapy. We evaluated the association between serum sIL-2R levels and the efficacy of anti-programmed cell death 1/ programmed death-ligand 1 (anti-PD-1/PD-L1) antibody combined with chemotherapy in non-small cell lung cancer (NSCLC) patients. We prospectively enrolled NSCLC patients who received anti-PD-1/PD-L1 antibody combined with platinum-based chemotherapy between 8/2019 and 8/2020 and measured their serum sIL-2R. The patients were divided into high and low sIL-2R groups based on the median of sIL-2R levels at pretreatment. Progression-free survival (PFS) and overall survival (OS) of patients in the high and low sIL-2R groups were compared. The Kaplan-Meier curves of PFS and OS were evaluated using the log-rank test. The multivariate analysis of PFS and OS was performed using the Cox proportional hazard models. Among 54 patients (median age 65, range 34-84), 39 were male and 43 had non-squamous cell carcinoma. The sIL-2R cut-off value was 533 U/mL. Median PFS was 5.1 months (95% CI, 1.8-7.5 months) and 10.1 months (95% CI, 8.3-not reached [NR] months) in the high and low sIL-2R groups (P = 0.007), respectively. Median OS was 10.3 months (95% CI, 4.0-NR months) and NR (95% CI, 10.3-NR months) in the high and low sIL-2R groups (P = 0.005), respectively. Multivariate Cox regression analysis showed that high sIL-2R was significantly associated with shorter PFS and OS. SIL-2R may be a biomarker for the poor efficacy of anti-PD-1/PD-L1 antibody combined with chemotherapy., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Introduction: Only a few reports have determined whether recently advanced anticancer drugs, particularly next-generation tyrosine kinase inhibitors (TKIs) and immune checkpoint inhibitors (ICIs), prolong the survival of patients with NSCLC in the real world., Methods: To evaluate the association between recently advanced drugs and patient survival, survival data of 2078 patients with stage IV NSCLC from 1995 to 2022 were analyzed in the present study. The patients were classified into the following six groups in terms of the date of diagnosis: period A, 1995 to 1999; period B, 2000 to 2004; period C, 2005 to 2009; period D, 2010 to 2014; period E, 2015 to 2019; and period F, 2000 to 2022. They were further grouped in terms of EGFR mutation and ALK fusion., Results: The median overall survival (mOS) times were 8.9, 11.0, 13.6, 17.9, and 25.2 months in periods A to E, respectively, and the mOS time was not reached in period F. This time was significantly longer in period E than in period D (25.2 versus 17.9 mo, p < 0.005). Moreover, the mOS times of patients with EGFR mutation, those with ALK fusion, and those without both alterations were significantly longer in period E than in period D (46.0 versus 32.0 mo, p < 0.005; not reached versus 36.2 mo, p  = 0.018; 14.6 versus 11.7 mo, p < 0.005). The history of treatment with next-generation TKIs and ICIs was found to be associated with overall survival., Conclusions: The survival of patients with NSCLC was improved from period D to period E, regardless of the presence of driver gene alteration. We found that next-generation TKIs and ICIs may be associated with improvements in overall survival., (© 2023 The Authors.)

Objectives: We investigated the effect of tyrosine kinase inhibitors (TKIs) and immunotherapy on survival after postoperative recurrence of non-small cell lung cancer (NSCLC)., Methods: This single-center retrospective study included patients with NSCLC who underwent lobectomy or more with complete pathological resection between 2008 and 2018 (N = 2254). Median follow-up was 5.1 years. Survival trends and the effect of TKIs/immunotherapy were analyzed using Joinpoint (National Cancer Institute) and Cox regression., Results: In 443 (19.7%) postoperative recurrences, median time to recurrence was 1.1 years; epidermal growth factor receptor mutation (EGFR+), 191 (43.1%); anaplastic lymphoma kinase rearrangement (ALK+), 13 (2.9%); not detected or unknown (ND), 239 (54.0%). In multivariable analysis, age, time to recurrence, adenocarcinoma, symptomatic recurrence, any treatment for recurrence, use of the epidermal growth factor receptor TKI, use of the anaplastic lymphoma kinase TKI, and use of immunotherapy were significant prognostic factors. Survival was significantly better in the EGFR+/ALK+ group than in the ND group (median, 4.7 vs 2.1 years; P < .01). Between 2010 and 2018, 2-year postrecurrence survival improved significantly (annual percentage change [APC], 4.2; 95% CI, 1.5-7.0). In subset analyses, neither change in 2-year survival nor TKI use was significant over time in the EGFR+/ALK+ group, but the ND group experienced significant improvement in 2-year survival (APC, 13.5; 95% CI, 5.4-22.2) and increasing trend in immunotherapy use (APC, 23.0; 95% CI, -5.9 to 60) after 2013., Conclusions: Survival after postoperative recurrence of NSCLC has improved significantly since 2010. Use of immunotherapy in patients without driver mutations may have contributed to that improvement. Prognosis in patients with driver mutations remains favorable with the TKIs introduced before the study period., (Copyright © 2022 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.)

Data on the re-administration of epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) after osimertinib failure in patients with T790M-positive non-small cell lung cancer (NSCLC) is limited. EGFR-TKI re-administration efficacy may vary between patients with T790M loss and those with T790M persistent with re-biopsy after osimertinib treatment. Patients who received EGFR-TKI re-administration (gefitinib, erlotinib, afatinib, dacomitinib, and osimertinib) after osimertinib failure were identified from our database. T790M mutation status before EGFR-TKI re-administration was analyzed via repeat biopsy. We retrospectively evaluated the efficacy of EGFR-TKI re-administration, especially differences according to the T790M mutation status, via repeat biopsy. Until June 2020, 28 patients received EGFR-TKI re-administration and 17 underwent repeat biopsy after osimertinib failure. Patients were divided into three groups, including the T790M loss group, where active mutation persisted and T790M was lost (13/17); T790M remaining group, where both the active mutation and T790M persisted (3/17); and active mutation loss group where both the active mutation and T790M were lost (1/17). The overall response rate (ORR) of EGFR-TKI re-administration in the T790M loss group was 31% and the disease control rate (DCR) was 54%, which were higher than the ORR of 21% and DCR of 43% in the entire patient population. ORR and DCR of the not re-biopsy group were low (9% and 27%, respectively). The therapeutic effect of EGFR-TKI re-administration in patients with T790M-positive NSCLC after osimertinib failure is limited. EGFR-TKI re-administration may be considered in cases of T790M loss after repeat biopsy., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Genomic profiling of tumors from cancer patients facilitates molecular-guided therapy. The turnaround time is one of important issues to deliver results timely for clinical decisions. The Ion Torrent™ Genexus™ Integrated Sequencer automates all next generation sequencing (NGS) workflows and delivers results within a day., Methods: In this study, we conducted a feasibility study to evaluate the detection rate of genomic alterations from cell-free total nucleic acid (cfTNA, containing cfDNA and cfRNA) of 119 non-small cell lung cancer using Oncomine Precision Assay on Genexus™ Integrated Sequencer. Oncomine Precision Assay (OPA) covers actionable mutations, copy number variations and fusion genes and that are applicable for the selection of targeted therapy. cfTNA isolated from plasma (derived from 14 ml of blood) were subjected to the Genexus system for library construction, templating, sequencing, and data analyses., Results: The sequencing resulted in median overall depth of 35,773× and median molecular coverage of 2,192× with cfTNA input ranged from 11 to 36 ng. Among the 119 samples evaluated, we detected at least one genomic alteration in plasma cfTNA of 79 cases (66%). When comparing to standard-of-care testing, the sensitivity and specificity of mutation detection in non-small cell lung cancer related genes using liquid biopsy with Genexus-OPA ranged between 49-67% and 93-100%, respectively. 59% of actionable mutations, which were present in tumor tissues, were detected by the Genexus- Oncomine Precision Assay using plasma cfTNA. Among the 5 mutations detected from liquid biopsy only, three mutations are of level 1 evidence according to OncoKB database, highlighting the clinical utilities of liquid biopsy in addressing tumor heterogeneity. Extrathoracic metastasis and levels of lactate dehydrogenase (LDH), C-reactive protein (CRP) and Carcinoembryonic Antigen (CEA) are found to be associated with increased circulating tumor DNA detection., Conclusions: The Genexus™ Integrated Sequencer system is an automated, accurate NGS system with short turnaround time (TAT) that could assist clinicians to make more timely decision., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-21-981/coif). SKL serves as a scientific advisor and received honorarium from Cancer Precision Medicine Inc. RA receives personal fees for lectures from AstraZeneca, Chugai Pharmaceutical. KU receives personal fees for lectures from AstraZeneca, Brystol Myers Squibb, Chugai Pharmaceutical, Daiichi Sankyo, Eli Lilly, Novartis, Ono Pharmaceutical, and Takeda Pharmaceutical Company Limited, and for manuscript writing from Chugai-Igakusha, Nankodo Co., Ltd., Nanzando Co., Ltd., Yodosha Co., Ltd. KU’s spouse is an employee of Daiichi Sankyo. RH and YMC are Employees of Cancer Precision Medicine, Inc. NY is a medical advisor of Chugai Pharmaceutical and receives lecture’s fee from Ono Pharmaceutical, Bristol Myers Squibb, Pfizer, Chugai Pharmaceutical, Eli Lilly, Taiho Pharmaceutical, AstraZeneca, MSD, and Takeda Pharmaceutical Company Limited. YN serves as a scientific advisor of Oncotherapy Science Inc., and possesses stock from Oncotherapy Science Inc. MN receives grants and personal fees from Ono Pharmaceutical, Bristol Myers Squibb, Pfizer, Chugai Pharmaceutical, Eli Lilly, Taiho Pharmaceutical, AstraZeneca, MSD, Novartis, Daiichi Sankyo, and Takeda Pharmaceutical Company Limited, personal fees from Boehringer-Ingelheim, Merck Biopharma, Teijin Pharma Limited, and AbbVie. The other authors have no conflicts of interest to declare., (2022 Translational Lung Cancer Research. All rights reserved.)

Background: A standard treatment regimen for advanced non-small cell lung cancer (NSCLC) patients with interstitial lung disease (ILD) has not been established since most clinical trials exclude such patients because of the high risk of acute exacerbation of ILD. This study aimed to prospectively investigate the efficacy and safety of carboplatin and nab-paclitaxel as a first-line regimen for NSCLC patients with ILD., Methods: The enrolled patients had treatment-naïve advanced NSCLC with ILD. The patients received 4-6 cycles of carboplatin (area under the curve = 5) on day 1 and nab-paclitaxel 100 mg/m 2 on days 1, 8, and 15 every 4 weeks. The primary endpoint was the completion rate of four or more cycles. Secondary endpoints included toxicity, overall response rate (ORR), disease control rate (DCR), progression-free survival (PFS), and overall survival (OS)., Results: Twenty-five patients were enrolled in this study. Nine patients had adenocarcinoma, 11 had squamous cell carcinoma, one had large cell carcinoma, and four had NSCLC, not otherwise specified. The completion rate of ≥4 cycles was 76% (95% confidence interval: 56.2%-88.8%), which met the primary endpoint. The ORR and DCR were 44% and 88%, respectively. The median PFS and OS were 5.8 months and 15.8 months, respectively. Three patients experienced grade ≥2 pneumonitis, and one patient met the acute exacerbation criteria., Conclusion: The 4-week modified regimen of carboplatin and nab-paclitaxel showed tolerable toxicity with favorable efficacy in NSCLC patients with ILD. This regimen may be an effective treatment option for patients in real clinical settings., (© 2022 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Anaplastic lymphoma kinase (ALK) fusion is found in ~3%-5% of patients with non-small-cell lung cancers (NSCLCs). Although the third-generation ALK tyrosine kinase inhibitor (TKI) lorlatinib shows high clinical efficacy in ALK-positive NSCLC, most of the patients eventually relapse with acquired resistance. Recently, drug-tolerant persister (DTP) cells have been considered an important seed of acquired resistance cells. In this study, we established lorlatinib intermediate resistant cells from a patient-derived cell model. Glycogen synthase kinase 3 (GSK3) inhibitions significantly suppressed lorlatinib intermediate resistant cell growth. GSK3 inhibition also sensitized acquired resistance cells derived from alectinib-treated patients with or without secondary mutations to lorlatinib. Therefore, GSK3 plays a crucial role in developing acquired resistance against lorlatinib in ALK-positive NSCLC mediated by lorlatinib intermediate resistant cells and could be a potential molecular target to prevent acquired lorlatinib resistance and overcome ALK-TKI resistance., (© 2022. The Author(s).)

Immune checkpoint therapy targeting the PD-1/PD-L1 axis is a potentially novel development in anticancer therapy and has been applied to clinical medicine. However, there are still some problems, including a relatively low response rate, innate mechanisms of resistance against immune checkpoint blockades, and the absence of reliable biomarkers to predict responsiveness. In this study of in vitro and in vivo models, we demonstrate that PD-L1-vInt4, a splicing variant of PD-L1, plays a role as a decoy in anti-PD-L1 antibody treatment. First, we showed that PD-L1-vInt4 was detectable in clinical samples and that it was possible to visualize the secreting variants with IHC. By overexpressing the PD-L1-secreted splicing variant on MC38 cells, we observed that an immune-suppressing effect was not induced by their secretion alone. We then demonstrated that PD-L1-vInt4 secretion resisted anti-PD-L1 antibody treatment, compared with WT PD-L1, which was explicable by the PD-L1-vInt4's decoying of the anti-PD-L1 antibody. The decoying function of PD-L1 splicing variants may be one of the reasons for cancers being resistant to anti-PD-L1 therapy. Measuring serum PD-L1 levels might be helpful in deciding the therapeutic strategy.

Background: Most patients treated with anaplastic lymphoma kinase (ALK)-tyrosine kinase inhibitors for ALK-positive non-small cell lung cancer (NSCLC) develop resistance, leading to metastasis, with progression to the central nervous system (CNS) being a primary concern. Although alectinib has better CNS penetration than crizotinib, patients treated with alectinib also develop CNS progression. CNS metastases more likely occurs during crizotinib treatment due to less blood-brain barrier (BBB) penetration capability than alectinib. CNS progression pattern may be different during crizotinib and alecitinib treatment. Understanding the characteristics of CNS progression is important for developing treatment strategies., Aims: We compared the clinical-radiographic characteristics of CNS metastases among patients undergoing crizotinib and alectinib treatment for ALK-positive NSCLCs., Methods and Results: We retrospectively analyzed the radiographic and clinical characteristics of CNS progression in ALK-positive NSCLC patients treated with crizotinib or alectinib at our hospital between July 2011 and May 2020. CNS and systemic tumor progression were evaluated using computed tomography or magnetic resonance imaging. Fifty-three and 65 patients were treated with crizotinib and alectinib, respectively. Baseline CNS metastasis was observed in 18 and 27 patients in the crizotinib and alectinib groups, respectively. Among the patients in the crizotinib and alectinib groups who developed disease progression, 15/49 (30.6%) and 9/44 (20.5%) had CNS progression, respectively (P = .344). Intra-CNS progression-free survival was significantly longer in the alectinib group than in the crizotinib group (median: 14.0 vs 5.6 months, P = .042). The number of CNS metastases sized ≥3 cm, rate of peritumoral brain edema, and the second progression pattern after treatment continuation was not significantly different between the groups., Conclusion: We observed no significant difference in the clinical-radiographic characteristics of CNS progression between patients undergoing crizotinib and alectinib treatments. Local therapy, including stereotactic radiosurgery, for CNS progression may be suitable and important following alectinib and crizotinib treatment., (© 2021 The Authors. Cancer Reports published by Wiley Periodicals LLC.)

Osimertinib is a third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) that is effective in treating both naïve and T790M-mutated EGFR-TKI-resistant non-small cell lung cancer patients. The EGFR C797S mutation is the major osimertinib resistance mechanism. The present study monitored the EGFR C797S mutation during osimertinib treatment in Japanese patients using droplet digital PCR (ddPCR). In our first cohort, C797S detection was validated with tumor specimens and/or plasma samples from 26 patients using ddPCR with custom-designed probes detecting and discriminating T790M and C797S in cis and trans positions. In our second cohort, 18 patients with EGFR-T790M who were going to start osimertinib were analyzed using ddPCR by collecting the plasma samples every month from the beginning of the course of osimertinib. In the first cohort, C797S was detected in 15.4% of patients. C797S and T790M in cis and trans positions were distinguished using ddPCR. In the second cohort, serial cfDNA evaluation revealed that the rate of EGFR mutation changes with disease state. Increases of EGFR mutation were detected, including C797S several months before the diagnosis of disease progression. As with the first cohort, C797S and T790M in cis and trans position were distinguished by ddPCR at disease progression. Coincidentally, in the first cohort, next generation sequencing detected NRAS Q61K mutation and the resistance with NRAS Q61K mutation was overcome by trametinib. In the second cohort, serial cfDNA analysis was useful for evaluating bone oligo-progression and local radiation therapy., (© 2021 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Approximately 15-30% of patients with lung cancer harbor mutations in the EGFR gene. Major EGFR mutations (>90% of EGFR-mutated lung cancer) are highly sensitive to EGFR tyrosine kinase inhibitors (TKIs). Many uncommon EGFR mutations have been identified, but little is known regarding their characteristics, activation, and sensitivity to various EGFR-TKIs, including allosteric inhibitors. We encountered a case harboring an EGFR-L747P mutation, originally misdiagnosed with EGFR-del19 mutation using a routine diagnostic EGFR mutation test, which was resistant to EGFR-TKI gefitinib. Using this minor mutation and common EGFR-activating mutations, we performed the binding free energy calculations and microsecond-timescale molecular dynamic (MD) simulations, revealing that the L747P mutation considerably stabilizes the active conformation through a salt-bridge formation between K745 and E762. We further revealed why several EGFR inhibitors, including the allosteric inhibitor, were ineffective. Our computational structural analysis strategy would be beneficial for future drug development targeting the EGFR minor mutations.

Background/aim: The Renin-Angiotensin system (RAS) induces immunosuppression in the tumor microenvironment, and RAS inhibitors (RASi) improve the tumor immune microenvironment. We evaluated the impact of RASi on the efficacy anti-programmed cell death-1/Ligand-1 (anti-PD-1/PD-L1) antibodies., Patients and Methods: This retrospective study analyzed non-small cell lung cancer (NSCLC) patients who received anti-PD-1/PD-L1 antibodies monotherapy as second- or later-line treatment. We classified patients into those with or without use of RASi., Results: A total of 256 NSCLC patients were included and 37 patients used RASi. The median PFS of patients treated with RASi was significantly longer than that of patients treated without (HR=0.59, 95%CI=0.40-0.88). The median OS of patients treated with RASi tended to be longer than that of patients treated without (HR=0.71, 95%CI=0.45-1.11)., Conclusion: The use of RASi was associated with a significantly longer PFS in NSCLC patients treated with anti-PD-1/PD-L1 antibodies. RASi use may enhance the efficacy of anti-PD-1/PD-L1 antibodies., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background: We assessed the efficacy and safety of bevacizumab and S-1 chemotherapy for patients with previously treated advanced non-squamous non-small-cell lung cancer (NSCLC)., Methods: This was a prospective single-arm study, including patients with non-squamous NSCLC who had received at least one chemotherapy regimen along with a platinum-based regimen. Bevacizumab 15 mg/kg was intravenously administered every 3 weeks, and S-1 40 mg/m 2 was orally administered twice daily from day 1 (evening) through day 15 (morning). The treatment continued for 3 weeks/cycle until disease progression or until unacceptable toxicities occurred. During the lead-in part, six patients were evaluated for dose-limiting toxicity (DLT) rate. In phase II, the primary endpoint was objective response rate (ORR). Secondary endpoints were progression-free survival (PFS), overall survival (OS), and safety., Results: In the lead-in part, we evaluated the safety in the first six patients and observed no DLT. In phase II, a total of 46 patients were enrolled from September 2012 to December 2018. The median follow-up duration was 13.7 months [95% confidence interval (CI) 1.4-72.0]. The ORR was 28.3%. The median PFS and OS were 4.3 (95% CI 2.9-5.9) and 15.0 months (95% CI 9.8-30.3), respectively. The most common adverse events were hypertension (65.2%), diarrhea (47.8%), mucositis oral (45.7%), and proteinuria (43.5%), and the most common grade 3 adverse events were hypertension (23.9%) and proteinuria (6.5%). Grade 4/5 adverse events were not observed., Conclusion: Bevacizumab and S-1 combination chemotherapy showed high activity and were well tolerated in patients with previously treated advanced non-squamous NSCLC.

Background: Several companion diagnostic (CDx) tests for epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) have been approved. In our institute, the CDx test for EGFR-TKIs was changed from the Therascreen test (Therascreen) to the Cobas EGFR v2 test (Cobas) because only Cobas was approved for the use of osimertinib in patients with EGFR-mutated non-small cell lung cancer (NSCLC) with T790M mutations. The clinical influence of switching the CDx test has not yet been examined comprehensively., Methods: All serial patients with lung cancer tested for EGFR mutations with CDx tests between February 2014 and February 2016 at the Cancer Institute Hospital of the Japanese Foundation for Cancer Research (JFCR) were enrolled in this analysis., Results: Therascreen was used as a CDx test for EGFR-TKI therapy in 607 patients between February 2014 and January 2015, and Cobas was used in 621 patients between February 2015 and February 2016. EGFR mutations were detected in 218 patients (35.9%) and 244 patients (39.3%) tested with Therascreen and Cobas, respectively. At the initial diagnosis, 400 and 459 patients were tested with Therascreen and Cobas, respectively. EGFR mutation subtypes, including del19, L858R, and others, were detected in 13.0%, 17.0%, and 2.5% of patients using Therascreen and 17.4%, 14.4%, and 1.5% of patients using Cobas, respectively., Conclusions: No significant impact of switching from Therascreen to Cobas as the CDx test for EGFR mutations in clinical practice was observed. However, the detection pattern of the EGFR mutation subtypes between the two CDx tests was slightly different., Key Points: SIGNIFICANT FINDINGS OF THE STUDY: We examined the influence of changing the EGFR test in 1228 patients in total. The detection rate of EGFR mutations was similar. However, the detection pattern for EGFR subtype mutations was slightly different between the two tests., What This Study Adds: Switching CDx tests from target polymerase chain reaction (PCR)- to next-generation sequencing (NGS)-based methods may lead to obvious changes in clinical practice. When the CDx test is required to change, the investigation of this influence is warranted in future studies., (© 2020 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Background: The usefulness of the Oncomine Dx Target test (Oncomine Dx), a next-generation sequencing (NGS) test, has already been proven in clinical trials. However, NGS requires high-quality tumor samples and takes a long time to generate results. The feasibility of NGS for use in advanced non-small cell lung cancer (NSCLC) patients in clinical practice has not yet been determined., Methods: Patients serially diagnosed with advanced NSCLC were evaluated in our hospital. The Oncomine Dx, Cobas EGFR mutation test (Cobas EGFR), and ALK-IHC were performed. The patients were divided into four sets: the full analysis set (FAS) that referred to patients diagnosed with NSCLC, the intent to perform companion diagnostics (CDx) set (IPS) that referred to patients in which CDx had been ordered regardless of sample quality, the per-performed CDx set (PPS) that referred to patients who could undergo CDx regardless of the results, and the per-completed CDx set (CCS) that referred to patients in which informative results were received from the CDx., Results: The total number of patients analyzed in the study was 167. The IPS/FAS of Oncomine Dx (80.2%) was lower than that of the ALK-IHC (85.0%) and Cobas EGFR (92.8%). The CCS/FAS of Oncomine Dx (65.9%) was lower than that of the ALK-IHC (82.0%) and Cobas EGFR (92.2%). PPS/IPS and CCS/PPS of the Oncomine Dx with nonsurgical biopsy ranged between 78.6% and 90.9%, which was lower than those patients who underwent surgical resection (95.0% and 100%)., Conclusions: The feasibility of Oncomine Dx in clinical practice was lower than the other CDx. The feasibility of Oncomine Dx will increase by improving the biopsy procedure., Key Points: SIGNIFICANT STUDY FINDINGS: The usefulness of a next-generation sequencing (NGS) test has been proven in clinical trials. The feasibility of NGS is lower than other diagnostics in clinical practice especially with regard to nonsurgical biopsy., What This Study Adds: It is necessary to improve the feasibility of NGS in clinical practice. To improve NGS feasibility, turnaround time must be shortened, and larger samples must be obtained during surgical procedures., (© 2020 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.)

Purpose: Patients with epidermal growth factor receptor (EGFR)-mutated non-small cell lung cancer with BIM deletion polymorphism may have a limited response to EGFR tyrosine kinase inhibitors (EGFR-TKIs). However, some results of previous reports are discordant. It is necessary to evaluate the relationship between BIM polymorphism and the efficacy of EGFR-TKIs., Methods: We retrospectively analyzed patients treated with EGFR-TKIs. We collected serum samples from patients before EGFR-TKI administration. We analyzed BIM deletion polymorphism and BIM single nucleotide polymorphism in exon 5 c465C > T by the Invader ® assay., Results: BIM deletion polymorphism was identified in 27 of 194 patients (13.9%). BIM single nucleotide polymorphism was identified in 29 of 194 patients (14.9%). The overall response ratio was 81.5% in patients with BIM deletion polymorphism, 89.7% with BIM single nucleotide polymorphism, and 83.6% with BIM wild type. Median progression-free survival was 10.3 months with BIM deletion polymorphism, 8.5 months with BIM single nucleotide polymorphism, and 10.4 months with BIM wild type. Overall survival was 38.4 months with BIM deletion polymorphism, 29.1 months with BIM single nucleotide polymorphism, and 31.6 months with BIM wild type. There were no significant differences between the groups in overall response ratio, progression-free survival, and overall survival., Conclusions: BIM polymorphism does not affect EGFR-TKI efficacy.