Masahiro Morise

Therapeutic Priority of the PI3K/AKT/mTOR Pathway in Small Cell Lung Cancers as Revealed by a Comprehensive Genomic Analysis

Introduction: The information regarding therapeutically relevant genomic alterations in small cell lung cancer (SCLC) is not well developed. We analyzed the SCLC genome using an integrative approach to stratify the targetable alterations. Methods: We performed whole exon sequencing (n = 51) and copy number analysis (n =47) on surgically resected tumors and matched normal tissue samples from treatment-naive Japanese SCLC patients. Results: The demographics of the 51 patients included in this study were as follows: median age, 67 years (range, 42–86 years); female, 9 (18%); history of smoking, 50 (98%); and pathological stage I/II/III/IV, 28/13/9/1, respectively. The average number of nonsynonymous mutations was 209 (range, 41–639; standard deviation, 130). We repeatedly confirmed the high prevalence of inactivating mutations in TP53 and RB1, and the amplification of MYC family members. In addition, genetic alterations in the PI3K/AKT/mTOR pathway were detected in 36% of the tumors: PIK3CA, 6%; PTEN, 4%; AKT2, 9%; AKT3, 4%; RICTOR, 9%; and mTOR, 4%. Furthermore, the individual changes in this pathway were mutually exclusive. Importantly, the SCLC cells harboring active PIK3CA mutations were potentially targetable with currently available PI3K inhibitors. Conclusions: The PI3K/AKT/mTOR pathway is distinguishable in SCLC genomic alterations. Therefore, a sequencing-based comprehensive analysis could stratify SCLC patients by potential therapeutic targets.

Heterologous regulation of anion transporters by menthol in human airway epithelial cells.

The present study concerns previously unreported effects of menthol, a cyclic terpene alcohol produced by the peppermint herb, on anion transporters in polarized human airway Calu-3 epithelia. Application of menthol (0.01-1mM) attenuated transepithelial anion transport, estimated as short-circuit currents (I(SC)), after stimulation by forskolin (10microM) but not before. In contrast, menthol potentiated forskolin-stimulated and -unstimulated apical Cl(-) conductance, which reflected the cystic fibrosis transmembrane conductance regulator (CFTR: the cAMP-regulated Cl(-) channel)-mediated conductance, without correlation to changes in cytosolic cAMP levels. These results indicate that menthol-induced attenuation of forskolin-induced I(SC) despite CFTR up-regulation was due to cAMP-independent inhibition of basolateral anion uptake, which is the rate-limiting step for transepithelial anion transport. Analyses of the responsible basolateral anion transporters revealed that forskolin increased both bumetanide (an inhibitor of the basolateral Na(+)-K(+)-2Cl(-) cotransporter [NKCC1])- and DNDS (an inhibitor of basolateral HCO(3)(-)-dependent anion transporters [NBC1/AE2])-sensitive I(SC) in the control whereas only the former was prevented by the application of menthol. Neither the bumetanide- nor DNDS-sensitive component was, however, reduced by menthol without forskolin. These heterologous effects of menthol were reproduced by latrunculin B, an inhibitor of actin polymerization. F-actin staining showed that menthol prevented forskolin-stimulated rearrangements of actin microfilaments without affecting the distribution of forskolin-unstimulated microfilaments. Collectively, menthol functions as an activator of CFTR and prevents activation of NKCC1 without affecting NBC1/AE although all of these transporters are commonly cAMP-dependent. The heterologous effects may be mediated by the actin cytoskeleton, which interacts with CFTR and NKCC1.

Factors Affecting the Diagnostic Yield of Transbronchial Biopsy Using Endobronchial Ultrasonography with a Guide Sheath in Peripheral Lung Cancer.

Objective Endobronchial ultrasonography with a guide sheath (EBUS-GS) and virtual bronchoscopic navigation (VBN) improves the diagnostic yield in patients with peripheral pulmonary lesions (PPLs). Most previous reports on EBUS-GS-guided transbronchial biopsy (TBB) have included patients with benign and malignant diseases. We aimed to determine the factors that predicted a successful diagnosis by EBUS-GS-guided TBB diagnostic in patients with small peripheral lung cancer, with a focus on the high-resolution computed tomography (HRCT) findings before bronchoscopy. Methods We retrospectively reviewed the medical records of 173 consecutive patients with 175 small (≤30 mm) PPLs who were diagnosed with primary lung cancer between June 2010 and October 2013 at Nagoya University Hospital. All patients underwent EBUS-GS-guided TBB with VBN using a ZioStation computer workstation (Ziosoft, Osaka, Japan). We analyzed the patient characteristics, HRCT findings, diagnostic yield, and the diagnostic factors in small peripheral lung carcinoma. Results The EBUS probe position was within the PPL in 83 of the 175 lesions (47%) and 112 (64.0%) cases were successfully diagnosed by EBUS-GS-guided TBB. A univariate analysis revealed that the following factors were associated with a significantly higher diagnostic yield: CT bronchus sign positivity, a lesion of >20 mm in diameter, a solid nodule, and a probe position that was within the lesion. The following factors were not significant: the lesion location, the number of biopsies, and the lung cancer histology. A multivariate analysis revealed that the following factors significantly affected the diagnostic yield: CT bronchus sign positivity [odds ratio (OR) =2.479]; a probe position that was within the lesion (OR=2.542); and a solid nodule (OR=2.304). Conclusion The significant factors that were significantly associated with a successful diagnosis using EBUS-GS-guided TBB in small peripheral lung carcinoma were as follows: CT bronchus sign positivity, a solid nodule, and a probe position that was within the lesion.

Dual Pathway Activated by tert-Butyl Hydroperoxide in Human Airway Anion Secretion

We analyzed the mechanisms underlying the ion transport induced by tert-butyl hydroperoxide (t-BOOH), a membrane-permeant oxidant that has been widely used as a model of oxidative stress, in human airway epithelial cells (Calu-3). We found that t-BOOH induced a short-circuit current that was composed of two distinct components, a peaked component (PC) and a sustained component (SC). Both components were reduced by the presence of H-89 (N-[2-(4-bromocinnamylamino)ethyl]-5-isoquinoline) [10 μM, a protein kinase A (PKA) inhibitor] and clofilium (100 μM, a cAMP-dependent K+ channel inhibitor) but not by charybdotoxin (50 nM, a human intermediate conductance Ca2+-activated K+ channel inhibitor), suggesting that both PC and SC were generated through a common PKA-dependent/Ca2+-independent pathway. Notwithstanding, analyses of the physiological properties revealed that PC and SC were attributable to different pathways. PC, but not SC, was correlated with apical membrane Cl- conductance and was inhibited by the cyclooxygenase (COX)-2 inhibitor NS-398 (N-[2-(cyclohexyloxyl)-4-nitrophenyl]-methane sulfonamide; 10 μM). In contrast, SC, but not PC, was composed of a component sensitive to bumetanide (50 μM), an inhibitor of the basolateral Na+-K+-2Cl- cotransporter (NKCC1), and was abolished by the cytoskeleton dysfunction induced by cytochalasin D (10 μM) and (R)-(+)-trans-N-(4-pyridyl)-4-(1-aminoethyl)-cyclohexane carboxamide (Y-27632; 20 μM). Collectively, t-BOOH induces PKA-related anion secretion through two independent pathways: rapid activation of apical anion efflux through a COX-2-dependent/cytoskeleton-independent pathway and relatively delayed activation of NKCC1 for basolateral anion uptake through a COX-2-independent/cytoskeleton-dependent pathway.

Patients With Antithyroid Antibodies Are Prone To Develop Destructive Thyroiditis by Nivolumab: A Prospective Study

Abstract Context Immune checkpoint inhibitors, including anti–programmed cell death-1 (PD-1) antibodies, have become promising treatments for a variety of advanced malignancies. However, these medicines can cause immune-related adverse events (irAEs), including endocrinopathies. Objective This study examined the incidence of endocrine irAEs induced by nivolumab. Patients and Main Outcome Measured Sixty-six patients treated with nivolumab at Nagoya University Hospital were prospectively evaluated for pituitary hormones, thyroid function, antithyroid antibodies (Abs), and glucose levels every 6 weeks after the initiation of nivolumab for 24 weeks. Results Four out of 66 patients developed destructive thyroiditis, and three patients developed hypothyroidism requiring levothyroxine replacement. The prevalence of positive anti-thyroglobulin Abs (TgAbs) and/or anti–thyroid peroxidase Abs (TPOAbs) at baseline was significantly higher in the group that developed destructive thyroiditis (3/4) compared with the group that did not develop thyroiditis (3/62; P = 0.002). There were no significant differences in other clinical variables between the groups. There were no endocrine irAEs other than destructive thyroiditis during the 24 weeks. The prevalence of TgAbs and/or TPOAbs at baseline was not associated with the development of other irAEs, including pneumonitis, colitis, or skin reactions. Conclusions Our real-world data showed that destructive thyroiditis was an endocrine irAE that was frequently induced by nivolumab and was significantly associated with positive TgAbs and/or TPOAbs before treatment. Our findings indicate that evaluating these Abs before treatment may help identify patients with a high risk of thyroidal irAEs and may have important clinical benefit.

Low-dose Irinotecan as a Second-line Chemotherapy for Recurrent Small Cell Lung Cancer

OBJECTIVE Irinotecan is a potent inhibitor of deoxyribonucleic acid topoisomerase 1 and the weekly schedule of 100-125 or 350 mg/m(2) administration on Day 1 every 3 weeks is recommended for recurrent small cell lung cancer. However, severe gastrointestinal toxic effects and myelosuppression are often observed in this dose setting. We conducted a retrospective study to evaluate the efficacy and safety of low-dose irinotecan monotherapy (60 mg/m(2) on Days 1, 8 and 15 every 4 weeks) as second-line chemotherapy for small cell lung cancer. METHODS The medical charts of small cell lung cancer patients who had received second-line chemotherapy at the National Cancer Center Hospital East between April 2003 and June 2012 were reviewed. Consecutive 57 patients who were treated with low dose of irinotecan (60 mg/m(2) on Days 1, 8 and 15 every 4 weeks) were analyzed in this study. RESULTS Median age was 70 years (range, 51-83). Fifty-two (91%) were male, 36 (63%) had an Eastern Cooperative Oncology Group performance status 0-1 and 26 (46%) were sensitive relapse. The median number of chemotherapy cycles was 2. The objective response rate was 32% (95% confidence interval: 20-45%).The median progression-free survival and the median overall survival were 2.9 months (95% confidence interval: 1.9-3.4 months) and 5.3 months (95% confidence interval: 3.6-7.6 months), respectively. The incidence of Grade 3/4 neutropenia, diarrhea and nausea/vomiting was 21, 4 and 5%, respectively. CONCLUSIONS Low-dose irinotecan monotherapy for recurrent small cell lung cancer might be effective with favorable toxicity. Randomized trial of 60 mg/m(2) versus standard dose of irinotecan is warranted.

Impact of mild to moderate COPD on feasibility and prognosis in non-small cell lung cancer patients who received chemotherapy

Background Non-small cell lung cancer (NSCLC) is the predominant cause of death in patients with COPD, and the severity of COPD in NSCLC patients is classified mainly as mild to moderate. Most advanced NSCLC patients with mild to moderate COPD are treated with chemotherapy; however, the feasibility for and prognosis after chemotherapy of these patients are not well understood. The aim of this study was to elucidate the impact of mild to moderate COPD on the feasibility for and prognosis after chemotherapy in NSCLC patients. Patients and methods A retrospective review was performed on 268 NSCLC patients who received first-line chemotherapy from 2009 to 2014 in our institution. Finally, 85 evaluable patients were included in this study. The clinical characteristics, toxicity profile, objective response rate, and prognosis were analyzed and compared between patients with mild to moderate COPD and those without COPD (non-COPD). Results Forty-three patients were classified as COPD (27 cases mild and 16 cases moderate) and 42 patients as non-COPD. The COPD group were older and had fewer never-smokers than the non-COPD group. The objective response rate did not differ between groups (p=0.14). There was no significant difference in overall survival between COPD and non-COPD groups (15.0 and 17.0 months, log-rank test p=0.57). In the multivariate Cox’s proportional hazard model, the adjusted hazard ratio (HRadj) was statistically significant for male sex (HRadj =5.382, 95% CI: 1.496–19.359; p=0.010), pathological diagnosis of adenocarcinoma (HRadj =0.460, 95% CI: 0.223–0.948; p=0.035), and epithelial growth factor receptor negative mutation (HRadj =6.040, 95% CI: 1.158–31.497; p=0.033), but not for the presence of COPD (HRadj =0.661, 95% CI: 0.330–1.325; p=0.24). Toxicity profile in COPD group was favorable, as in the non-COPD group. Conclusion Mild to moderate COPD did not have a significant deleterious impact on toxicity and prognosis in NSCLC patients.

Identification of proteasomal catalytic subunit PSMA6 as a therapeutic target for lung cancer

To identify potential therapeutic targets for lung cancer, we performed semi‐genome‐wide shRNA screening combined with the utilization of genome‐wide expression and copy number data. shRNA screening targeting 5043 genes in NCI‐H460 identified 51 genes as candidates. Pathway analysis revealed that the 51 genes were enriched for the five pathways, including ribosome, proteasome, RNA polymerase, pyrimidine metabolism and spliceosome pathways. We focused on the proteasome pathway that involved six candidate genes because its activation has been demonstrated in diverse human malignancies, including lung cancer. Microarray expression and array CGH data showed that PSMA6, a proteasomal subunit of a 20S catalytic core complex, was highly expressed in lung cancer cell lines, with recurrent gene amplifications in some cases. Therefore, we further examined the roles of PSMA6 in lung cancer. Silencing of PSMA6 induced apoptosis or G2/M cell cycle arrest in cancer cell lines but not in an immortalized normal lung cell line. These results suggested that PSMA6 serves as an attractive target with a high therapeutic index for lung cancer.

An EGFR -mutated Lung Adenocarcinoma Undergoing Squamous Cell Carcinoma Transformation Exhibited a Durable Response to Afatinib

Squamous cell carcinoma (SCC) transformation has been identified as a mechanism of resistance to first-generation epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs), gefitinib or erlotinib, in EGFR-mutated lung cancer. However, whether second- or third-generation TKIs can overcome resistance due to SCC transformation remains unclear. We herein report an EGFR-mutated lung adenocarcinoma undergoing transformation into SCC that exhibited a durable response to afatinib, which is a second-generation irreversible EGFR-TKI. We suggest that afatinib can be considered as a treatment option for EGFR-mutated tumor undergoing SCC transformation, particularly in the absence of a T790M mutation.

eIF2β, a subunit of translation-initiation factor EIF2, is a potential therapeutic target for non-small cell lung cancer

To identify novel therapeutic targets for non‐small cell lung cancer (NSCLC), we conducted an integrative study in the following 3 stages: (i) identification of potential target gene(s) through shRNA functional screens in 2 independent NSCLC cell lines; (ii) validation of the clinical relevance of identified gene(s) using public databases; and (iii) investigation of therapeutic potential of targeting the identified gene(s) in vitro. A semi‐genome‐wide shRNA screen was performed in NCI‐H358 cells, and was integrated with data from our previous screen in NCI‐H460 cells. Among genes identified in shRNA screens, 24 were present in both NCI‐H358 and NCI‐H460 cells and were considered potential targets. Among the genes, we focused on eIF2β, which is a subunit of heterotrimeric G protein EIF2 and functions as a transcription initiation factor. The eIF2β protein is highly expressed in lung cancer cell lines compared with normal bronchial epithelial cells, and gene copy number analyses revealed that eIF2β is amplified in a subset of NSCLC cell lines. Gene expression analysis using The Cancer Genome Atlas (TCGA) dataset revealed that eIF2β expression is significantly upregulated in lung cancer tissues compared with corresponding normal lung tissues. Furthermore, high eIF2β expression was correlated with poor survival in patients with lung adenocarcinoma, as shown in other cohorts using publicly available online tools. RNAi‐mediated depletion of eIF2β suppresses growth of lung cancer cells independently of p53 mutation status, in part through G1 cell cycle arrest. Our data suggest that eIF2β is a therapeutic target for lung cancer.