Hiroyuki Yasuda

Lung cancer is the leading cause of cancer-related death. The identification of epidermal growth factor receptor (EGFR) somatic mutations defined a new, molecularly classified subgroup of non-small-cell lung cancer (NSCLC). Classic EGFR activating mutations, such as inframe deletions in exon 19 or the Leu858Arg (L858R) point mutation in exon 21 are associated with sensitivity to first generation quinazoline reversible EGFR tyrosine kinase inhibitors (TKIs). EGFR exon 20 insertion mutations, which are typically located after the C-helix of the tyrosine kinase domain of EGFR, may account for up to 4% of all EGFR mutations. Preclinical models have shown that the most prevalent EGFR exon 20 insertion mutated proteins are resistant to clinically achievable doses of reversible (gefitinib, erlotinib) and irreversible (neratinib, afatinib, PF00299804) EGFR TKIs. Growing clinical experience with patients whose tumours harbour EGFR exon 20 insertions corresponds with the preclinical data; very few patients have had responses to EGFR TKIs. Despite the prevalence and biological importance of EEGFR exon 20 insertions, few reports have summarised all preclinical and clinical data on these mutations. Here, we review the literature and provide an update with an emphasis on the structural, molecular, and clinical implications of EGFR exon 20 insertions.

Crystal structure and detailed analysis of different EGFR mutants explain why some mutations in exon 20 make lung cancers resistant to EGFR inhibitors and others make them more sensitive.

Patients with non-small cell lung cancer (NSCLC) that harbors epidermal growth factor receptor (EGFR) mutations initially respond to EGFR-tyrosine kinase inhibitors (TKI) but eventually experience relapse. Acquired resistance to EGFR-TKIs is strongly associated with patient mortality. Thus, elucidation of the mechanism of acquired resistance to EGFR-TKIs is of great importance. In this study, gefitinib-resistant cell line models were established by long-term exposure to gefitinib using the gefitinib-sensitive lung cancer cell lines, PC9 and HCC827. Expression analyses indicated that both FGFR1 and FGF2 were increased in PC9 gefitinib-resistant (PC9 GR) cells as compared with PC9 naïve (PC9 na) cells. Importantly, proliferation of gefitinib-resistant cells was dependent on the FGF2 -FGFR1 pathway. Mechanistically, inhibition of either FGF2 or FGFR1 by siRNA or FGFR inhibitor (PD173074) restored gefitinib sensitivity in PC9 GR cells. These data suggest that FGF2 -FGFR1 activation through an autocrine loop is a novel mechanism of acquired resistance to EGFR-TKIs.

<h2>Summary</h2> Gastroenteropancreatic (GEP) neuroendocrine neoplasm (NEN) that consists of neuroendocrine tumor and neuroendocrine carcinoma (NEC) is a lethal but under-investigated disease owing to its rarity. To fill the scarcity of clinically relevant models of GEP-NEN, we here established 25 lines of NEN organoids and performed their comprehensive molecular characterization. GEP-NEN organoids recapitulated pathohistological and functional phenotypes of the original tumors. Whole-genome sequencing revealed frequent genetic alterations in <i>TP53</i> and <i>RB1</i> in GEP-NECs, and characteristic chromosome-wide loss of heterozygosity in GEP-NENs. Transcriptome analysis identified molecular subtypes that are distinguished by the expression of distinct transcription factors. GEP-NEN organoids gained independence from the stem cell niche irrespective of genetic mutations. Compound knockout of TP53 and RB1, together with overexpression of key transcription factors, conferred on the normal colonic epithelium phenotypes that are compatible with GEP-NEN biology. Altogether, our study not only provides genetic understanding of GEP-NEN, but also connects its genetics and biological phenotypes.

Most clinically available small-molecule kinase inhibitors are multi-targeted and can inhibit multiple kinases. Our driving hypothesis was that one of these multi-targeted tyrosine kinase inhibitors (TKIs) would have antiproliferative activity against ROS1 translocated non-small-cell lung cancer (NSCLC).We selected NSCLC cell lines--A549 (KRAS G12S), NCI-H3255 (EGFR L858R), NCI-H3122 (EML4-ALK E13;A20), and HCC78 (SLC34A2-ROS1)-to evaluate the antiproliferative effects of submicromolar concentrations of the multitargeted TKIs imatinib, sorafenib, erlotinib, and crizotinib.Imatinib and sorafenib were unable to significantly inhibit proliferation of the aforementioned cell lines. Erlotinib only inhibited EGFR mutated NCI-H3255, as expected. Crizotinib displayed dose-dependent inhibition of anaplastic lymphoma kinase translocated NCI-H3122 and also ROS1--translocated HCC78. The SLC34A2-ROS1 translocated HCC78 cell line had phosphorylated levels of ROS1, AKT, and ERK inhibited by submicromolar doses of crizotinib, and subsequently underwent apoptosis.The ROS1-translocated HCC78 cell line was sensitive to inhibition by the multitargeted ALK/MET/RON/ROS1 inhibitor crizotinib. Preclinical data supports the clinical development of crizotinib for ROS1-translocated NSCLC.

Alterations in the processing of the genetic information in carcinogenesis result from stable genetic mutations or epigenetic modifications. It is becoming clear that nucleosomal histones are central to proper gene expression and that aberrant DNA methylation of genes and histone methylation plays important roles in tumor progression. To date, several histone lysine methyltransferases (HKMTs) have been identified and histone lysine methylation is now considered to be a critical regulator of transcription. However, still relatively little is known about the role of HKMTs in tumorigenesis.We observed differential HKMT expression in a lung cancer model in which normal human bronchial epithelial (NHBE) cells expressing telomerase, SV40 large T antigen, and Ras were immortal, formed colonies in soft agar, and expressed specific HKMTs for H3 lysine 9 and 27 residues but not for H3 lysine 4 residue. Modifications in the H3 tails affect the binding of proteins to the histone tails and regulate protein function and the position of lysine methylation marks a gene to be either activated or repressed. In the present study, suppression by siRNA of HKMTs (EZH2, G9A, SETDB1 and SUV39H1) that are over-expressed in immortalized and transformed cells lead to reduced cell proliferation and much less anchorage-independent colony growth. We also found that the suppression of H3-K9, G9A and SUV39H1 induced apoptosis and the suppression of H3-K27, EZH2 caused G1 arrest.Our results indicate the potential of these HKMTs in addition to the other targets for epigenetics such as DNMTs and HDACs to be interesting therapeutic targets.

EGFR mutated lung cancer accounts for a significant subgroup of non-small-cell lung cancer (NSCLC).Over the last decade, multiple EGFR tyrosine kinase inhibitors (EGFR-TKIs) have been developed to target mutated EGFR.However, there is little information regarding mutation specific potency of EGFR-TKIs against various types of EGFR mutations.The purpose of this study is to establish an in vitro model to determine the "therapeutic window" of EGFR-TKIs against various types of EGFR mutations, including EGFR exon 20 insertion mutations.The potency of 1 st (erlotinib), 2 nd (afatinib) and 3 rd (osimertinib and rociletinib) generation EGFR-TKIs was compared in vitro for human lung cancer cell lines and Ba/F3 cells, which exogenously express mutated or wild type EGFR.An in vitro model of mutation specificity was created by calculating the ratio of IC50 values between mutated and wild type EGFR.The in vitro model identified a wide therapeutic window of afatinib for exon 19 deletions and L858R and of osimertinib and rociletinib for T790M positive mutations.The results obtained with our models matched well with previously reported preclinical and clinical data.Interestingly, for EGFR exon 20 insertion mutations, most of which are known to be resistant to 1 st and 2 nd generation EGFR-TKIS, osimertinib was potent and presented a wide therapeutic window.To our knowledge, this is the first report that has identified the therapeutic window of osimertinib for EGFR exon 20 insertion mutations.In conclusion, this model will provide a preclinical rationale for proper selection of EGFR-TKIs against clinically-relevant EGFR mutations.

EGFR-mutated lung cancers account for a significant subgroup of non-small cell lung cancers overall. Third-generation EGFR tyrosine kinase inhibitors (TKI) are mutation-selective inhibitors with minimal effects on wild-type EGFR. Acquired resistance develops to these agents, however, the mechanisms are as yet uncharacterized. In this study, we report that the Src-AKT pathway contributes to acquired resistance to these TKI. In addition, amplification of EGFR wild-type alleles but not mutant alleles was sufficient to confer acquired resistance. These findings underscore the importance of signals from wild-type EGFR alleles in acquiring resistance to mutant-selective EGFR-TKI. Our data provide evidence of wild-type allele-mediated resistance, a novel concept of acquired resistance in response to mutation-selective inhibitor therapy in cancer treatment. Cancer Res; 77(8); 2078-89. ©2017 AACR.

IntroductionThe multitargeted tyrosine kinase inhibitor (TKI) crizotinib is active against ALK translocated non-small-cell lung cancer (NSCLC); however acquired resistance invariably develops over time. ALK mutations have previously been implicated in only a third of resistant tumors. We sought to evaluate alternative mechanisms of resistance and preclinical strategies to overcome these in a cell line driven by EML4-ALK.MethodsWe selected the NSCLC cell line NCI-H3122 (H3122: EML4-ALK E13;A20) and derived resistant variants that were able to grow in the presence of 1 μM crizotinib. These were analyzed for ALK mutations, sensitivity to crizotinib in combination with other TKIs, and for activation of alternative tyrosine kinases.ResultsAll H3122 crizotinib resistant (CR) clones lacked amplification or mutations in the kinase domain of ALK. To evaluate if possible alternative kinases functioned as “bypass” tracks for downstream signaling activation in these resistance cells, we performed of phosho-receptor tyrosine kinase array that demonstrated that CR clones had higher phospho-EGFR signals than H3122 cells before and after exposure to crizotinib. A functional approach of dual ALK TKI (with crizotinib) with combinatory TKI inhibition was used as a secondary screen for possible targets. Crizotinib + erlotinib (reversible EGFR TKI) and crizotinib + afatinib (irreversible EGFR/ERBB2 TKI) were able to inhibit the growth of H3122 CR clones, confirming EGFR activation as a mechanism of resistance. The removal of crizotinib from the culture media re-sensitized CR cells to crizotinib.ConclusionsWe identified activation of EGFR as a mechanism of resistance to crizotinib in preclinical models of ALK translocated NSCLC. If EGFR activation is confirmed as a predominant mechanism of ALK TKI-induced resistance in patient-derived tumors, the use of ALK plus EGFR TKIs could be explored for this important cohort of NSCLCs.

Although the main cellular target of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is thought to be alveolar cells, the absence of their tractable culture system precludes the development of a clinically relevant SARS-CoV-2 infection model. Here, we establish an efficient human alveolosphere culture method and sphere-based drug testing platform for SARS-CoV-2. Alveolospheres exhibit indolent growth in a Wnt- and R-spondin-dependent manner. Gene expression, immunofluorescence, and electron microscopy analyses reveal the presence of alveolar cells in alveolospheres. Alveolospheres express ACE2 and allow SARS-CoV-2 to propagate nearly 100,000-fold in 3 days of infection. Whereas lopinavir and nelfinavir, protease inhibitors used for the treatment of human immunodeficiency virus (HIV) infection, have a modest anti-viral effect on SARS-CoV-2, remdesivir, a nucleotide prodrug, shows an anti-viral effect at the concentration comparable with the circulating drug level. These results demonstrate the validity of the alveolosphere culture system for the development of therapeutic agents to combat SARS-CoV-2.

Human lung cancer is a constellation of tumors with various histological and molecular properties. To build a preclinical platform that covers this broad disease spectrum, we obtained lung cancer specimens from multiple sources, including sputum and circulating tumor cells, and generated a living biobank consisting of 43 lines of patient-derived lung cancer organoids. The organoids recapitulated the histological and molecular hallmarks of the original tumors. Phenotypic screening of niche factor dependency revealed that EGFR mutations in lung adenocarcinoma are associated with the independence from Wnt ligands. Gene engineering of alveolar organoids reveals that constitutive activation of EGFR-RAS signaling provides Wnt independence. Loss of the alveolar identity gene NKX2-1 confers Wnt dependency, regardless of EGFR signal mutation. Sensitivity to Wnt-targeting therapy can be stratified by the expression status of NKX2-1. Our results highlight the potential of phenotype-driven organoid screening and engineering for the fabrication of therapeutic strategies to combat cancer.

In Brief Study Design. A prospective observational study. Objective. To evaluate the paravertebral muscle (PVM) degeneration in patients with degenerative lumbar scoliosis (DLS), using magnetic resonance imaging. Summary of Background Data. Several studies have described the histological and morphological changes to the PVM in patients with chronic low back pain and lumbar radiculopathy. However, there is little knowledge about the PVM changes in patients with DLS. Methods. Fifty-seven patients with lumbar spinal stenosis (LSS) with DLS (DLS group) and 50 control patients with LSS without DLS (LSS group) were examined. The cross-sectional area (CSA) and percentage of fat infiltration area (%FIA) of the bilateral multifidus and longissimus muscles at the L1–S1 levels were measured using T2-weighted axial magnetic resonance imaging and computer software. A multifidus muscle biopsy and histological evaluation were performed in some patients. Results. In the DLS group, the CSA of the multifidus muscle was significantly smaller and the %FIA of both muscles was significantly higher on the concave side than on the convex side at all levels (P < 0.0001 for each). These differences were also found in the longissimus muscles at the L4–L5 and L5–S1 levels (P < 0.0001 for each). Histologically, the multifidus muscle exhibited reductions in the muscle fiber size and number of nuclei on the concave side. In the LSS group, the total CSA and %FIA did not differ significantly between the left and right sides. However, in patients with unilateral radiculopathy, the CSA of the multifidus muscle was significantly smaller (P < 0.05) and the %FIA of both muscles was significantly higher (P < 0.05) on the symptomatic side, especially at 1 level below. Conclusion. This observational study with magnetic resonance imaging and histology showed that muscle degeneration was more common on the concave side in patients with DLS. Radiculopathy and spinal deformity may contribute to the PVM degeneration. In this study, paravertebral muscles were analyzed using magnetic resonance image in patients with lumbar spinal stenosis. Paravertebral muscle degeneration was more severe in concave side of group with degenerative lumbar scoliosis and in symptomatic side of group with lumbar spinal stenosis with unilateral symptom. Radiculopathy and spinal deformity may contribute to the paravertebral muscle degeneration.

Retrospective review.Evaluation of the impact of diabetes on lumbar spine surgery.Characteristics of diabetes that increase the risk of postoperative complications and poor surgical outcomes after lumbar spine surgery remain unclear.The demographic and clinical data of diabetic and nondiabetic patients, 50 years or older, undergoing lumbar spine surgery were reviewed. Japanese Orthopaedic Association and visual analogue scale scores for low back pain, leg pain, and numbness were assessed as perioperative outcomes. Analysis of covariance was used for comparison of perioperative outcomes to adjust for differences between the groups, and a proportional odds model was used to compute the odds ratio of poor improvement in each outcome.Forty-one patients with diabetes were compared with 124 patients without diabetes. Visual analogue scale scores of final low back pain was higher for patients with than without diabetes (29.3 vs. 17.9, P = 0.013). Complications were similar in patients with and without diabetes except for nonunion after fusion surgery (20% vs. 3%, P = 0.095). When stratified by surgical procedure, final low back pain was significantly higher for patients with diabetes who underwent fusion surgery (39.1 vs. 17.4, P = 0.001). Patients with glycosylated hemoglobin of 6.5% or more displayed a 2-fold increase only in the odds ratio (OR) of poor improvement of low back pain (OR = 2.37; 95% confidence interval [CI], 0.99-5.70). Patients having diabetes for 20 years or more were more likely to experience poor improvement of low back pain and leg numbness (OR = 4.95; 95% CI, 1.69-14.5 and OR = 2.80; 95% CI, 0.98-7.94, respectively). Insulin use was associated with an increased OR for poor improvement of leg numbness (OR = 4.49; 95% CI, 1.24-16.3).Longstanding diabetes, poor glycemic control, and insulin use might be associated with poor postoperative improvement.

The discovery of somatic mutations in EGFR and development of EGFR tyrosine kinase inhibitors (TKI) have revolutionized treatment for lung cancer. However, resistance to TKIs emerges in almost all patients and currently no effective treatment is available. Here, we show that β-catenin is essential for development of EGFR-mutated lung cancers. β-Catenin was upregulated and activated in EGFR-mutated cells. Mutant EGFR preferentially bound to and tyrosine phosphorylated β-catenin, leading to an increase in β-catenin-mediated transactivation, particularly in cells harboring the gefitinib/erlotinib-resistant gatekeeper EGFR-T790M mutation. Pharmacologic inhibition of β-catenin suppressed EGFR-L858R-T790M mutated lung tumor growth, and genetic deletion of the β-catenin gene dramatically reduced lung tumor formation in EGFR-L858R-T790M transgenic mice. These data suggest that β-catenin plays an essential role in lung tumorigenesis and that targeting the β-catenin pathway may provide novel strategies to prevent lung cancer development or overcome resistance to EGFR TKIs.

The niche surrounding stem cells regulate their fate during homeostasis and after injury or infection. The 3D organoid assay has been widely used to study stem cells behavior based on its capacity to evaluate self-renewal, differentiation and the effect of various medium supplements, drugs and co-culture with supportive cells. We established an assay to study both lung and trachea stem cells in vitro. We characterized their proliferation and differentiation spectrum at baseline then evaluated the effect of co-culturing with fibroblasts and endothelial cells and/or treating with several biologically relevant substances as possible contributors to their niche. We found that lung epithelial (but not tracheal basal) stem cells require co-culture with stromal cells to undergo clonal proliferation and differentiation. Fibroblasts were more efficient than endothelial cells in offering this support and the pattern of support varied based on the tissue origin of the stromal cells. Treating distal lung epithelial or basal stem cells with FGF2, FGF9, FGF10, LIF as well as ALK5 and ROCK inhibitors increased their colony formation efficiency and resulted in variable effects on colonies number, size and differentiation spectrum. This model and findings pave the way for better understanding of lung stem cell niche components and factors that can manipulate lung stem cell behavior.

Activation of the EGFR pathway is one of the mechanisms inducing acquired resistance to anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKI) such as crizotinib and alectinib. Ceritinib is a highly selective ALK inhibitor and shows promising efficacy in non-small cell lung cancers (NSCLC) harboring the ALK gene rearrangement. However, the precise mechanism underlying acquired resistance to ceritinib is not well-defined. This study set out to clarify the mechanism in ALK-translocated lung cancer and to find the preclinical rationale overcoming EGFR pathway-induced acquired resistance to ALK-TKIs. To this end, ceritinib-resistant cells (H3122-CER) were established from the H3122 NSCLC cell line harboring the ALK gene rearrangement via long-term exposure to ceritinib. H3122-CER cells acquired resistance to ceritinib through EGFR bypass pathway activation. Furthermore, H3122 cells that became resistant to ceritinib or alectinib through EGFR pathway activation showed cross-resistance to other ALK-TKIs. Ceritinib and afatinib combination treatment partially restored the sensitivity to ceritinib.This study proposes a preclinical rationale to use ALK-TKIs and afatinib combination therapy for ALK-translocated lung cancers that have acquired resistance to ALK-TKIs through EGFR pathway activation. Mol Cancer Res; 15(1); 106-14. ©2016 AACR.

Alectinib is a highly selective ALK inhibitor and shows promising efficacy in non-small cell lung cancers (NSCLC) harboring the EML4-ALK gene rearrangement. The precise mechanism of acquired resistance to alectinib is not well defined. The purpose of this study was to clarify the mechanism of acquired resistance to alectinib in ALK-translocated lung cancer cells. We established alectinib-resistant cells (H3122-AR) from the H3122 NSCLC cell line, harboring the EML4-ALK gene rearrangement, by long-term exposure to alectinib. The mechanism of acquired resistance to alectinib in H3122-AR cells was evaluated by phospho-receptor tyrosine kinase (phospho-RTK) array screening and Western blotting. No mutation of the ALK-TK domain was found. Phospho-RTK array analysis revealed that the phosphorylation level of EGFR was increased in H3122-AR cells compared with H3122. Expression of TGFα, one of the EGFR ligands, was significantly increased and knockdown of TGFα restored the sensitivity to alectinib in H3122-AR cells. We found combination therapy targeting ALK and EGFR with alectinib and afatinib showed efficacy both in vitro and in a mouse xenograft model. We propose a preclinical rationale to use the combination therapy with alectinib and afatinib in NSCLC that acquired resistance to alectinib by the activation of EGFR bypass signaling.

Despite the worldwide approval of three generations of EGFR tyrosine kinase inhibitors (TKI) for advanced non-small cell lung cancers with EGFR mutations, no TKI with a broad spectrum of activity against all clinically relevant mutations is currently available. In this study, we sought to evaluate a covalent mutation-specific EGFR TKI, TAS6417 (also named CLN-081), with the broadest level of activity against EGFR mutations with a prevalence of ≥1%. Lung cancer and genetically engineered cell lines, as well as murine xenograft models were used to evaluate the efficacy of TAS6417 and other approved/in-development EGFR TKIs (erlotinib, afatinib, osimertinib, and poziotinib). We demonstrate that TAS6417 is a robust inhibitor against the most common EGFR mutations (exon 19 deletions and L858R) and the most potent against cells harboring EGFR-T790M (first/second-generation TKI resistance mutation). In addition, TAS6417 has activity in cells driven by less common EGFR-G719X, L861Q, and S768I mutations. For recalcitrant EGFR exon 20 insertion mutations, selectivity indexes (wild-type EGFR/mutant EGFR ratio of inhibition) favored TAS6417 in comparison with poziotinib and osimertinib, indicating a wider therapeutic window. Taken together, we demonstrate that TAS6417 is a potent EGFR TKI with a broad spectrum of activity and a wider therapeutic window than most approved/in-development generations of EGFR inhibitors. IMPLICATIONS: TAS6417/CLN-081 is a potent EGFR TKI with a wide therapeutic window and may be effective in lung cancer patients with clinically relevant EGFR mutations.

Abstract Purpose: EGFR exon 20 insertions account for up to 10% of all EGFR mutations in lung adenocarcinomas, representing the third most common cluster of mutations. The management of advanced cancers with these mutations remains elusive, without an approved inhibitor. Experimental Design: Preclinical models of a representative set of EGFR exon 20 insertion mutations to evaluate the efficacy of different inhibitors and description of the clinical outcome of an advanced lung cancer. Results: We show that select first-, second-, and third-generation EGFR inhibitors are unable to deter common EGFR exon 20 insertion mutants in concentrations that spare the wild-type kinase. Nonetheless, EGFR exon 20 insertion mutants associate with the Hsp90 chaperone system. We exploit this vulnerability to show that the nongeldanamycin Hsp90 inhibitor luminespib (formerly AUY922) degrades EGFR exon 20 mutations, downstream targets, and induces apoptosis. In addition, a patient whose EGFR inhibitor–insensitive lung adenocarcinoma harbored an EGFR exon 20 insertion mutation had a confirmed radiographic response to luminespib. Conclusions: The report confirms that EGFR exon 20 mutations are dependent on Hsp90 and are readily inhibited by the Hsp90 inhibitor luminespib; a treatment strategy that has been pursued in a confirmatory clinical trial (NCT01854034) for this group of lung adenocarcinomas that currently represent an unmet clinical need in precision oncology.

Abstract Cancer‐associated fibroblasts (CAFs) are known to promote tumourigenesis through various mechanisms. Fibroblast growth factor (FGF)/FGF receptor (FGFR)‐dependent lung cancers have been described. We have developed a mouse model of lung adenocarcinoma that was constructed through the induction of Fgf9 overexpression in type 2 alveolar cells. The expression of Fgf9 in adult lungs resulted in the rapid development of multiple adenocarcinoma‐like tumour nodules. Here, we have characterised the contribution of CAFs and the Fgf/Fgfr signalling pathway in maintaining the lung tumours initiated by Fgf9 overexpression. We found that CAF‐secreted Fgf2 contributes to tumour cell growth. CAFs overexpressed Tgfb , Mmp7 , Fgf9 , and Fgf2 ; synthesised more collagen, and secreted inflammatory cell‐recruiting cytokines. CAFs also enhanced the conversion of tumour‐associated macrophages (TAMs) to the tumour‐supportive M2 phenotype but did not influence angiogenesis. In vivo inhibition of Fgfrs during early lung tumour development resulted in significantly smaller and fewer tumour nodules, whereas inhibition in established lung tumours caused a significant reduction in tumour size and number. Fgfr inhibition also influenced tumour stromal cells, as it significantly abolished TAM recruitment and reduced tumour vascularity. However, the withdrawal of the inhibitor caused a significant recurrence/regrowth of Fgf/Fgfr‐independent lung tumours. These recurrent tumours did not possess a higher proliferative or propagative potential. Our results provide evidence that fibroblasts associated with the Fgf9‐induced lung adenocarcinoma provide multiple means of support to the tumour. Although the Fgfr blocker significantly suppressed the tumour and its stromal cells, it was not sufficient to completely eliminate the tumour, probably due to the emergence of alternative (resistance/maintenance) mechanism(s). This model represents an excellent tool to further study the complex interactions between CAFs, their related chemokines, and the progression of lung adenocarcinoma; it also provides further evidence to support the need for a combinatorial strategy to treat lung cancer. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley &amp; Sons, Ltd.

Significance A variety of rare mutations account for 10–20% of EGFR mutations in nonsmall cell lung cancer. However, due to high diversity, proper medication for patients with such mutations is impossible in daily clinic. To appropriately treat lung cancer patients harboring such rare EGFR mutations, a robust prediction model to predict sensitivities of rare EGFR mutants to existing drugs is strongly needed. Using molecular dynamics simulation-based model, we successfully predicted diverse sensitivities of EGFR exon 20 insertion mutants to existing inhibitors. The findings suggest the usefulness of in silico simulation to overcome mutation diversity at a clinically relevant level. The present in silico model will help in selecting effective drugs for these patients.

Epidermal growth factor receptor (EGFR) mutation-positive lung cancer accounts for a significant subgroup of non-small cell lung cancers (NSCLC). Approximately 4-10% of EGFR mutations in NSCLC are EGFR exon 20 insertion mutations, which are reportedly associated with resistance to EGFR tyrosine kinase inhibitor (EGFR-TKI) treatment. NSCLC patients carrying these mutations are rarely treated with EGFR-TKIs. The purpose of this study was to evaluate the efficacy of afatinib or osimertinib plus cetuximab combination therapy in experimental NSCLC models with EGFR exon 20 insertion mutations.The EGFR mutations examined in this study were A763_Y764insFQEA, Y764_V765insHH, A767_V769dupASV, and D770_N771insNPG. Ba/F3 cells constitutively expressing wild type or mutated EGFR were used to determine the efficacy of afatinib or osimertinib plus cetuximab combination therapy in vitro. To determine the efficacy of the combination therapy in vivo, female BALB/c-nu mice were injected subcutaneously with 1 million Ba/F3 cells carrying EGFR A767_V769dupASV or Y764_V765insHH.We observed a mild but significant (P < 0.05) additive effect of the combination therapy against several EGFR exon 20 insertion mutations in vitro. Regarding EGFR A767_V769dupASV and EGFR Y764_V765insHH, cetuximab and afatinib single treatment did not induce significant inhibition of tumor formation; however, afatinib plus cetuximab combination treatment induced significant (P < 0.05) tumor growth inhibition without significant body weight loss or skin rash.The combination therapy induced a more potent inhibitory effect against several EGFR exon 20 insertion mutations than either therapy alone. Cetuximab can potentially increase the efficacy of afatinib or osimertinib in NSCLC with EGFR exon 20 insertion mutations.

Several preclinical data proposed a potential efficacy of osimertinib, a third-generation EGFR tyrosine kinase inhibitor, for EGFR exon 20 insertion (EGFR ex20ins)-positive non-small cell lung cancer (NSCLC). However, reported case series and a retrospective study proposed controversial efficacy. The efficacy of osimertinib in EGFR ex20ins-positive NSCLC have not been well evaluated in prospective clinical trials. In this study, we performed a prospective, single-arm, multi-center, open-label, non-randomized phase I/II study to evaluate efficacy of osimertinib for EGFR ex20ins-positive NSCLC.From August 2018 to January 2020, 14 NSCLC patients with EGFR ex20ins were enrolled, of whom 2 were excluded because they did not meet the inclusion criteria. Efficacy and safety of 80 mg osimertinib were evaluated. In addition, we performed a translational exploratory study to clarify the association of mutation type-specific drug sensitivity, osimertinib pharmacokinetic data, and clinical efficacy.Of the evaluated patients, none experienced objective response, 7 experienced stable disease (58.3%), and 5 experienced disease progression (41.7%). The median progression free survival (PFS) was 3.8 months, and the median overall survival was 15.8 months. Interestingly, the exploratory study demonstrated statistically significant positive correlation between plasma osimertinib concentration/in vitro IC50 ratio and PFS (R = 0.9912, P = 0.0001), highlighting the mutation type-specific concentration-dependent efficacy of osimertinib for EGFR ex20ins-positive NSCLC.Regular dose, 80 mg/day, of osimertinib has limited clinical activity in NSCLC patients with EGFR ex20ins. The translational study proposed the potential efficacy of higher dose osimertinib in a subgroup of EGFR ex20ins-positive NSCLC.

Crohn's disease, a chronic and recurrent gastrointestinal disease, frequently causes intestinal fibrosis. Transient receptor potential melastatin 2 (TRPM2), a non-selective cation channel, is activated by reactive oxygen species. This study investigated the role of TRPM2 in acute colitis and chronic colitis-associated fibrosis progression. Acute colitis and chronic colitis-associated fibrosis were induced in TRPM2-deficient (TRPM2KO) and wild-type (WT) mice through single and repeated intrarectal injections of 2,4,6-trinitrobenzene sulfonic acid (TNBS). Bone marrow-derived macrophages (BMDMs) from WT and TRPM2KO mice were stimulated using H2O2. In WT mice, a single TNBS injection induced acute colitis with upregulated inflammatory cytokines/chemokines and Th1/Th17-related cytokines, while repeated TNBS injections induced chronic colitis-associated fibrosis with upregulation of fibrogenic factors and Th2-related cytokines. Acute colitis and chronic colitis-associated fibrosis with cytokines/chemokine upregulation and fibrogenic factors were considerably suppressed in TRPM2KO mice. Treating BMDMs with H2O2 increased cytokine/chemokine expression and JNK, ERK, and p38 phosphorylation; however, these responses were significantly less in TRPM2KO than in WT mice. These findings suggest that TRPM2 contributes to acute colitis progression via Th1/Th17-mediated immune responses. Furthermore, TRPM2 may be directly involved in colitis-associated fibrosis induction, likely due to the regulation of Th2/TGF-β1-mediated fibrogenesis in addition to a consequence of acute colitis progression.

Articular cartilage defects that do not repair spontaneously induce osteoarthritic changes in joints over a long period of observation. In this study, we examined the usefulness of transplanting culture-expanded bone marrow mesenchymal cells into osteochondral defects of joints with cartilage defects. First, we performed experiments on rabbits and up on obtaining good results proceeded to perform the experiments on humans. Macroscopic and histological repair with this method was good, and good clinical results were obtained although there was no significant difference with the control group. Recent reports have indicated that this procedure is comparable to autologous chondrocyte implantation, and concluded that it was a good procedure because it required one step less than that required by surgery, reduced costs for patients, and minimized donor site morbidity. Although some reports have previously shown that progenitor cells formed a tumor when implanted into immune-deficient mice after long term in vitro culture, the safety of the cell transplantation was confirmed by our clinical experience. Thus, this procedure is useful, effective, and safe, but the repaired tissues were not always hyaline cartilage. To obtain better repair with this procedure, treatment approaches using some growth factors during in vitro culture or gene transfection are being explored.

Recent advances in the treatment of non-small cell lung cancer (NSCLC) with new agents require accurate histological subtyping at diagnosis to avoid the higher risk of an adverse response and to obtain the maximum therapeutic response. However, interobserver variability, tumor hetero­geneity and the degree of differentiation may affect the decision concerning a pathological diagnosis of NSCLC. Therefore, the aim of this study was to identify specific microRNAs (miRNAs) as standardized biomarkers with high sensitivity and specificity in order to distinguish between squamous cell carcinoma (SCC) and adenocarcinoma (AC). Quantitative polymerase chain reaction (qPCR)‑based miRNA array analysis was performed to identify microRNAs differentially expressed between SCC and AC using 86 resected NSCLC samples in addition to adjacent normal tissues. The results were confirmed by independent qRT-PCR assays with the same test samples and 88 additional validation samples, and from this we evaluated the usefulness of the identified miRNAs as biomarkers to distinguish between SCC and AC. Three miRNAs (hsa-miR-196b, hsa-miR-205 and hsa-miR-375) were identified. Discriminant analysis combining the three miRNAs appeared to distinguish SCC from AC accurately in the test and validation samples, demonstrating a sensitivity and specificity of 76 and 80%, and 85 and 83%, respectively. hsa-miR-196b, hsa-miR-205 and hsa-miR-375 were identified as biomarkers capable of distinguishing between lung SCC and lung AC. These newly identified miRNAs may prove to be highly valuable molecular markers for the classification of NSCLC histological subtypes and may contribute to the pathogenesis of each subtype of NSCLC.

In many countries, patients are generally allowed to have clear fluids until 2–3 h before surgery. In Japan, long preoperative fasting is still common practice. To shorten the preoperative fasting period in Japan, we tested the safety and efficacy of oral rehydration therapy until 2 h before surgery. Three hundred low-risk patients scheduled for morning surgery in six university-affiliated hospitals were randomly assigned to an oral rehydration solution (ORS) group or to a fasting group. Patients in the ORS group consumed up to 1,000 ml of ORS containing balanced glucose and electrolytes: 500 ml between 2100 the night before surgery and the time they woke up the next morning and 500 ml during the morning of surgery until 2 h before surgery. Patients in the fasting group started fasting at 2100 the night before surgery. Primary endpoints were gastric fluid volume and pH immediately after anesthesia induction. Several physiological measures of hydration and electrolytes including the fractional excretion of sodium (FENa) and the fractional excretion of urea nitrogen (FEUN) were also evaluated. Mean (SD) gastric fluid volume immediately after anesthesia induction was 15.1 (14.0) ml in the ORS group and 17.5 (23.2) ml in the fasting group (P = 0.30). The mean difference between the ORS group and fasting group was −2.5 ml. The 95% confidence interval ranged from −7.1 to +2.2 ml and did not include the noninferior limit of +8 ml. Mean (SD) gastric fluid pH was 2.1 (1.9) in the ORS group and 2.2 (2.0) in the fasting group (P = 0.59). In the ORS group, mean FENa and FEUN immediately after anesthesia induction were both significantly greater than those in the fasting group (P < 0.001 for both variables). The ORS group reported they had been less thirsty and hungry before surgery (P < 0.001, 0.01). Oral rehydration therapy until 2 h before surgery is safe and feasible in the low-risk Japanese surgical population. Physicians are encouraged to use this practice to maintain the amount of water in the body and electrolytes and to improve the patient's comfort.

Nivolumab, an immune checkpoint inhibitor, is now a standard treatment for previously treated advanced non-small-cell lung cancer based on the results from phase III clinical trials. We evaluated the real-world efficacy and safety of nivolumab in a nonselected population and identified the clinical characteristics that influence efficacy.A total of 142 patients with advanced non-small-cell lung cancer who were administered nivolumab at Keio University and affiliated hospitals in Japan from January to July 2016 were enrolled. The treatment efficacy and adverse events were retrospectively reviewed, and the clinical characteristics associated with the nivolumab response were evaluated using univariate and stratified analyses and the Cochran-Mantel-Haenszel test.The objective response rate was 17.0% (95% confidence interval [CI], 12.0%-24.0%), the median progression-free survival (PFS) was 58 days (95% CI, 50-67 days), and the proportion of patients with adverse events of any grade was 45.0%. EGFR/ALK mutation status was inversely associated with the treatment response (P < .05), and the difference in PFS for the mutation-positive versus mutation-negative patients was statistically significant (49 vs. 63 days; hazard ratio, 1.9; 95% CI, 1.1-5.2; P = .029). Previous radiotherapy also had a positive association with the treatment response (P = .012).The objective response rate, PFS, and adverse event profiles were comparable to those observed in previous clinical trials. EGFR/ALK mutation-negative status and previous radiotherapy might be key clinical characteristics associated with a positive treatment response. Our findings could aid in the efficient immunotherapeutic management of lung cancer.

ObjectivesFibroblast growth factor (FGF) 9 is a member of the FGF family, which modulates cell proliferation, differentiation, and motility. Recent studies show that the activation of FGF signals including FGF9 is associated with the pathogenesis of several cancers; however, its clinicopathological and biological significance in non-small cell lung cancer (NSCLC) is unclear. The purpose of this study was to clarify the characteristics of NSCLC with FGF9 expression.Materials and MethodsWe evaluated the expression of FGF9 in resected NSCLC specimens and corresponding non-tumorous lung tissue samples using cDNA microarray and evaluated its clinicopathological characteristics.ResultsNine out of 90 NSCLC specimens (10%) had “high” FGF9 expression compared with corresponding non-cancerous lung tissues. Histologically, of the 9 NSCLC specimens with high FGF9 expression, 5 were adenocarcinoma, whereas none were squamous cell carcinoma. FGF9 expression was not associated with sex, smoking history, or clinical stage. However, in patients with high and low FGF9 expression, the postoperative recurrence rates were 78% and 24% (p = 0.033), respectively. Overall survival was significantly shorter in patients with high FGF9 expression than in those with low FGF9 expression (p < 0.001).ConclusionOur data indicate that FGF9 may be a novel unfavorable prognostic indicator and a candidate therapeutic target of NSCLC.

<h2>Abstract</h2><h3>Objectives</h3> Over the past decade, the biological and clinical characteristics of lung cancer with epidermal growth factor receptor (<i>EGFR</i>) mutation have been well studied. However, most studies have focused on advanced inoperable cancer, and not on resected early-stage lung adenocarcinoma. We aimed to elucidate the differences in the clinicopathological characteristics and postoperative prognosis according to the <i>EGFR</i> mutation status in early-stage lung adenocarcinoma. <h3>Materials and methods</h3> We retrospectively collected clinical and pathological data from 369 patients with pathological stage I or II lung adenocarcinoma who underwent complete resection. Clinicopathological characteristics and postoperative prognosis were compared depending on the <i>EGFR</i> mutation status, using the Chi-squared test and the log-rank test, respectively. <h3>Results and conclusion</h3> Of the 369 patients, 160 (43.3%) had <i>EGFR</i> mutation, of which 64 (40.0%) were exon 19 deletion (Del-19) and 90 (56.3%) were exon 21 point mutation L858R. Although there was no difference in overall survival (OS) between patients with and without <i>EGFR</i> mutation (<i>p</i>=0.086), tumors with <i>EGFR</i> mutation were associated with a lower consolidation to tumor ratio (CTR) (<i>p <</i>0.001) and a higher incidence of a lepidic growth pattern by pathological evaluation (<i>p <</i>0.001) compared to those without <i>EGFR</i> mutation. Among tumors with <i>EGFR</i> mutation, there was no difference in OS (<i>p</i>=0.140) between Del-19 and L858R. Tumors with L858R were associated with a lower CTR (<i>p</i>=0.046), and tended to have a higher incidence of a lepidic growth pattern by pathological evaluation (<i>p</i>=0.073) compared to those with Del-19. In conclusion, although <i>EGFR</i> mutation status was not a prognostic indicator after surgery in early-stage lung adenocarcinoma, L858R and Del-19 had different radiological and pathological features.

EGFR-mutated lung cancer accounts for a significant proportion of lung cancer cases worldwide. For these cases, osimertinib, a third-generation EGFR tyrosine kinase inhibitor, is extensively used as a first-line or second-line treatment. However, lung cancer cells acquire resistance to osimertinib in 1 to 2 years. Thus, a thorough clarification of resistance mechanisms to osimertinib is highly anticipated. Recent next-generation sequencing (NGS) of lung cancer samples identified several genetically defined resistance mechanisms to osimertinib, such as EGFR C797S or MET amplification. However, nongenetically defined mechanisms are not well evaluated. For a thorough clarification of osimertinib resistance, both genetic and nongenetic mechanisms are essential. By using our comprehensive protein phosphorylation array, we detected IGF1R bypass pathway activation after EGFR abolishment. Both of our established lung cancer cells and patient-derived lung cancer cells demonstrated IGF2 autocrine-mediated IGF1R pathway activation as a mechanism of osimertinib resistance. Notably, this resistance mechanism was not detected by a previously performed NGS, highlighting the essential roles of living cancer cells for a thorough clarification of resistance mechanisms. Interestingly, the immunohistochemical analysis confirmed the increased IGF2 expression in lung cancer patients who were treated with osimertinib and met the established clinical definition of acquired resistance. The findings highlight the crucial roles of cell-autonomous ligand expression in osimertinib resistance. Here, we report for the first time the IGF2 autocrine-mediated IGF1R activation as a nongenetic mechanism of osimertinib resistance in lung cancer at a clinically relevant level. IMPLICATIONS: Using comprehensive protein phosphorylation array and patient-derived lung cancer cells, we found that IGF2 autocrine-mediated IGF1R pathway activation is a clinically relevant and common mechanism of acquired resistance to osimertinib.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes the disease COVID-19 can lead to serious symptoms, such as severe pneumonia, in the elderly and those with underlying medical conditions. While vaccines are now available, they do not work for everyone and therapeutic drugs are still needed, particularly for treating life-threatening conditions. Here, we showed nasal delivery of a new, unmodified camelid single-domain antibody (VHH), termed K-874A, effectively inhibited SARS-CoV-2 titers in infected lungs of Syrian hamsters without causing weight loss and cytokine induction. In vitro studies demonstrated that K-874A neutralized SARS-CoV-2 in both VeroE6/TMPRSS2 and human lung-derived alveolar organoid cells. Unlike other drug candidates, K-874A blocks viral membrane fusion rather than viral attachment. Cryo-electron microscopy revealed K-874A bound between the receptor binding domain and N-terminal domain of the virus S protein. Further, infected cells treated with K-874A produced fewer virus progeny that were less infective. We propose that direct administration of K-874A to the lung could be a new treatment for preventing the reinfection of amplified virus in COVID-19 patients.

Potential novel strategies for adverse event (AE) management of osimertinib therapy, including therapeutic drug monitoring and the use of biomarkers, have not yet been fully investigated. This study aimed to evaluate (1) the relationship between exposure to osimertinib, especially its active metabolites (AZ5104 and AZ7550), and AEs, and (2) the relationship between germline polymorphisms and AEs.We conducted a prospective, longitudinal observational study of 53 patients with advanced non-small cell lung cancer receiving osimertinib therapy from February 2019 to April 2022. A population pharmacokinetic model was developed to estimate the area under the serum concentration-time curve from 0 to 24 h (AUC0-24) of osimertinib and its metabolites. Germline polymorphisms were analyzed using TaqMan® SNP genotyping and CycleavePCR® assays.There was a significant association between the AUC0-24 of AZ7550 and grade ≥ 2 paronychia (p = 0.043) or anorexia (p = 0.011) and between that of osimertinib or AZ5104 and grade ≥ 2 diarrhea (p = 0.026 and p = 0.049, respectively). Furthermore, the AUC0-24 of AZ5104 was significantly associated with any grade ≥ 2 AEs (p = 0.046). EGFR rs2293348 and rs4947492 were associated with severe AEs (p = 0.019 and p = 0.050, respectively), and ABCG2 rs2231137 and ABCB1 rs1128503 were associated with grade ≥ 2 AEs (p = 0.008 and p = 0.038, respectively).Higher exposures to osimertinib, AZ5104, and AZ7550 and polymorphisms in EGFR, ABCG2, and ABCB1 were related to higher severity of AEs; therefore, monitoring these may be beneficial for osimertinib AE management.

Companion diagnostic tests play a crucial role in guiding treatment decisions for patients with non-small cell lung cancer (NSCLC). The Oncomine Dx Target Test (ODxTT) Multi-CDx System has emerged as a prominent companion diagnostic method. However, its efficacy in detecting driver gene mutations, particularly rare mutations, warrants investigation.This study aimed to assess the performance of the ODxTT in detecting driver gene mutations in NSCLC patients. Specifically, we aimed to evaluate its sensitivity in detecting epidermal growth factor receptor (EGFR) mutations, a key determinant of treatment selection in NSCLC.We conducted a retrospective analysis of NSCLC patients who underwent testing with the ODxTT at Keio University Hospital between May 2020 and March 2022. Patient samples were subjected to both DNA and RNA tests. Driver gene mutation status was assessed, and instances of missed mutations were meticulously examined.Of the 90 patients, five had nucleic acid quality problems, while 85 underwent both DNA and RNA tests. Driver gene mutations were detected in 56/90 (62.2%) patients. Of the 34 patient specimens, driver mutations were not detected using the ODxTT; however, epidermal growth factor receptor (EGFR) mutations were detected using polymerase chain reaction-based testing in two patients, and a KRAS mutation was detected by careful examination of the sequence data obtained using the ODxTT in one patient. For the above three cases, carefully examining the gene sequence information obtained using the ODxTT could identify driver mutations that were not mentioned in the returned test results. Additionally, we confirmed comparable instances of overlook results for EGFR mutations in the dataset from South Korea, implying that this type of oversight could occur in other countries using the ODxTT. EGFR mutation was missed in ODxTT in Japan (6.3%, 2/32), South Korea (2.0%, 1/49), and overall (3.7%, 3/81).Even if sufficient tumor samples are obtained, rare EGFR mutations (which are excluded from the ODxTT's genetic mutation list) might not be detected using the current ODxTT system due to the program used for sequence analysis. However, such rare EGFR mutations can still be accurately detected on ODxTT's sequence data using next-generation sequencing.

A case of an epidermoid cyst in the frontal base which showed homogeneous high density in noncontrast computed tomography, simulating a meningioma with calcification, is reported. Operative findings and histological examination suggested that this high density was caused by spontaneous hemorrhage into the cyst.

Third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (EGFR-TKIs) were developed to overcome EGFR T790M-mediated resistance to first-and second-generation EGFR-TKIs.Third-generation EGFR-TKIs, such as osimertinib and nazartinib, are effective for patients with the EGFR T790M mutation.However, there are no direct comparison data to guide the selection of a thirdgeneration EGFR-TKI for patients with different EGFR mutations.We previously established an in vitro model to estimate the therapeutic windows of EGFR-TKIs by comparing their relative efficacies against cells expressing mutant or wild type EGFRs.The present study used this approach to characterize the efficacy of third-generation EGFR-TKIs and compare them with that of other EGFR-TKIs.Treatment efficacy was examined using human lung cancer-derived cell lines and Ba/F3 cells, which were transduced with clinically relevant mutant EGFRs.Interestingly, mutation-related differences in EGFR-TKI sensitivity were observed.For classic EGFR mutations (exon 19 deletion and L858R, with or without T790M), osimertinib showed lower IC50 values and wider therapeutic windows than nazartinib.For less common EGFR mutations (G719S or L861Q), afatinib showed the lowest IC50 values.For G719S+T790M or L861Q+T790M, the IC50 values of osimertinib and nazartinib were around 100 nM, which was 10-to 100-fold higher than those for classic+T790M mutations.On the contrary, osimertinib and nazartinib showed similar efficacies in cells expressing EGFR exon 20 insertions.The findings highlight the diverse mutation-related sensitivity pattern of EGFR-TKIs.These data may help in the selection of EGFR-TKIs for non-small cell lung cancer patients harboring EGFR mutations.www.impactjournals.

We previously reported low expression of miR-375 in squamous-cell carcinoma (SCC) and high expression in adenocarcinoma (AC) of the lung. miR-375's target genes and its function in non-small-cell lung cancer (NSCLC) have not been elucidated. Therefore, the present study was designed to identify the targets of miR-375 and to characterize its function in NSCLC.Candidate targets of miR-375 were determined using a prediction database and previous data on differential gene expression between SCC and AC. We evaluated miR-375 and target-gene expression levels in 12 NSCLC cell lines. The effect of miR-375 overexpression and knockdown was evaluated in NSCLC cell lines by transfecting them with an miR-375 precursor or inhibitor. A luciferase-reporter assay was performed to confirm a direct interaction between miR-375 and its target gene. Further, a wound-healing assay was performed to evaluate the effect of miR-375 overexpression on the migration of SK-MES-1 cells. Finally, to assess the clinical relevance, 63 clinical NSCLC samples were analyzed.Claudin-1 (CLDN1) has 4 putative miR-375 target sites in its 3'-untranslated region, and this gene was determined to be a target of miR-375. CLDN1 messenger RNA and protein expression were attenuated by overexpression of miR-375 and increased by knockdown of miR-375 in NSCLC cell lines. In a luciferase-reporter assay, miR-375 overexpression resulted in a 3-fold repression of luciferase activity (P<0.001). Cell migration was promoted by miR-375 overexpression, suggesting a high potential for invasion and metastasis in NSCLC expressing high levels of miR-375. In clinical NSCLC samples, there was a negative correlation between miR-375 and CLDN1 expression (r=-0.35, P=0.005). In addition, high miR-375 expression was correlated with a shorter survival time among the clinical samples (P=0.043).CLDN1 is a novel target of miR-375, and high miR-375 expression shortens survival in NSCLC.

Background. Recent clinical trials proven the clinically significant efficacy and tolerability of nivolumab, a programmed death 1 (PD-1) inhibitor, in previously treated patients with non-small cell lung cancer (NSCLC). Case Presentation. Here, we describe the case of a patient who experienced possible "disease flare" immediately after initiation of nivolumab treatment. A 54-year-old man was diagnosed with Stage IIB (T2N1M0) lung adenocarcinoma. After 7 years from recurrence, 10th line chemotherapy, nivolumab, was initiated. Six weeks later, after 3 cycles of nivolumab treatment, rapid lung cancer progression was observed with an increase in the size of the primary lesion, multiple novel nodules on both lungs, and multiple novel brain metastases. Conclusion. We believe that physicians should be made aware that, in a subset of NSCLC patients, disease flare might occur on nivolumab treatment.

BACKGROUND CONTEXT The incidence of adjacent vertebral fracture (AVFs) is reported to be 10%–38% after balloon kyphoplasty. However, no reports have established a system for prediction of AVF occurrence. PURPOSE To establish a scoring system for predicting AVF occurrence after balloon kyphoplasty for osteoporotic vertebral fractures (OVFs). DESIGN A prospective cohort study. PATIENT SAMPLE Consecutive elderly patients aged 65 years and older who underwent balloon kyphoplasty for OVFs within 2 months after the onset. OUTCOME MEASURES AVF was confirmed by X-ray. METHODS From 2015 to 2017, 116 consecutive patients from 10 participating hospitals who underwent balloon kyphoplasty were enrolled in this study. Prior to study enrollment, each patient underwent plain X-ray, computed tomography (CT), and magnetic resonance imaging (MRI) of the thoracic and lumbar spine. Severity of pain was subjectively assessed using a visual analog scale (VAS) based on the average level of back pain that the patient had experienced in the preceding week. After enrollment, subjects underwent balloon kyphoplasty. Quality of life was evaluated using SF-36. Patients were followed up for at least 6 months. RESULTS Of the 116 patients enrolled, 109 patients with all the required data at the time of enrolment and the 6-month follow-up were included in the study. A total of 32 patients (29%) showed AVFs within the 6-month follow-up. No significant differences were observed in each clinical outcome at 6-month follow-up, although higher VAS score for back pain at 1-month follow-up was observed in the AVF group (37.5) than in the non-AVF group (20.8, p<.001). Wedge angle of vertebrae before surgery was greater in the AVF group (21.6°) than in the non-AVF group (15.7°, p<.001). The change in wedge angle between pre- and postsurgery was greater in the AVF group than in the non-AVF group, whereas the change in local kyphosis was not significantly different. The multiple logistic regression model showed increased odds ratio (OR) of thoracic or thoracolumbar spine, old OVF presence, >25° kyphosis before surgery, and >10°correction for AVF. Based on this result, a simple scoring system for predicting AVF occurrence was developed. The total AVF score was calculated as the sum of the individual scores, which varied from 0 to 6. All patients with 5–6 points sustained AVF. CONCLUSIONS More severe wedge angle before surgery, correction degree, old OVF presence, and thoracolumbar level were predictive factors for AVF. All patients with AVF risk score of 5 or more showed AVF. This information may aid preoperative risk assessment, informed shared decision-making, and consideration of potential alternative management strategies.

Lu and colleagues recently reviewed mortality-related risk factors of COVID-19.1 COVID-19 was first reported in Wuhan, China, in December 2019 and subsequently spread globally, leading to a pandemic;2 as of August 25, 2020, more than 23 million people worldwide had been confirmed to have COVID-19 infections, and more than 810,000 patients had died.3 Although approximately 80% of COVID-19 cases are classified as mild or asymptomatic, 15% of adults infected with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) developed severe illness and required oxygen supplementation, and an additional 5% progressed to a critical state.

Not all non-small cell lung cancer (NSCLC) patients possess drug-targetable driver mutations, and response rates to immune checkpoint blockade therapies also remain unsatisfactory. Therefore, more effective treatments are still needed. Here, we report the results of a phase 2 clinical trial of adoptive cell therapy using zoledronate-expanded autologous Vγ9Vδ2 T-cells for treatment-refractory NSCLC.NSCLC patients who had undergone at least two regimens of standard chemotherapy for unresectable disease or had had at least one treatment including chemotherapy or radiation for recurrent disease after surgery were enrolled in this open-label, single-arm, multicenter, phase 2 study. After preliminary testing of Vγ9Vδ2 T-cell proliferation, autologous peripheral blood mononuclear cells were cultured with zoledronate and IL-2 to expand the Vγ9Vδ2 T-cells. Cultured cells (>1×109) were intravenously administered every 2 weeks for six injections. The primary endpoint of this study was progression-free survival (PFS), and secondary endpoints included overall survival (OS), best objective response rate (ORR), disease control rate (DCR), safety and immunomonitoring. Clinical efficacy was defined as median PFS significantly >4 months.Twenty-five patients (20 adenocarcinoma, 4 squamous cell carcinoma and 1 large cell carcinoma) were enrolled. Autologous Vγ9Vδ2 T-cell therapy was administered to all 25 patients, of which 16 completed the foreseen course of 6 injections of cultured cells. Median PFS was 95.0 days (95% CI 73.0 to 132.0 days); median OS was 418.0 days (179.0-479.0 days), and best overall responses were 1 partial response, 16 stable disease (SD) and 8 progressive disease. ORR and DCR were 4.0% (0.1%-20.4%) and 68.0% (46.5%-85.1%), respectively. Severe adverse events developed in nine patients, mostly associated with disease progression. In one patient, pneumonitis and inflammatory responses resulted from Vγ9Vδ2 T-cell infusions, together with the disappearance of a massive tumor.Although autologous Vγ9Vδ2 T-cell therapy was well tolerated and may have an acceptable DCR, this trial did not meet its primary efficacy endpoint.UMIN000006128.

Epidermal growth factor receptor (EGFR) mutations are strong determinants of tumor response to EGFR tyrosine kinase inhibitors in non-small cell lung cancers (NSCLCs). Currently available methods of EGFR mutation detection rely on direct sequencing. Here, we describe the use of an alternative way to screen EGFR mutations.A total of 109 frozen tumor specimens from NSCLC patients were obtained. For mutational analysis of EGFR exons 18, 19, and 21, reverse transcription-polymerase chain reaction was performed on the cDNA using original primers designed for restriction fragment length polymorphism (RFLP).EGFR mutations were detected in 37 patients (34%) by both RFLP and direct sequencing except one case in which it was detected only by RFLP. EGFR mutations were more frequently observed to be significant by multivariate analysis in patients with adenocarcinoma (OR = 5.56), no-smoking history (OR = 4.34), and 65-year-old or younger (OR = 2.64), but not in women (OR = 1.14). Among 37 patients, 18 were treated with gefitinib and 9 responded to the treatment. One patient without any mutation responded.RFLP is a useful method for screening EGFR mutations and can also be applied to predicting the sensitivity of NSCLC patients to EGFR-tyrosine kinase inhibitors.

Purpose We sought to determine the mechanisms of downregulation of the airway transcription factor Foxa2 in lung cancer and the expression status of Foxa2 in non-small-cell lung cancer (NSCLC). Methods A series of 25 lung cancer cell lines were evaluated for Foxa2 protein expression, FOXA2 mRNA levels, FOXA2 mutations, FOXA2 copy number changes and for evidence of FOXA2 promoter hypermethylation. In addition, 32 NSCLCs were sequenced for FOXA2 mutations and 173 primary NSCLC tumors evaluated for Foxa2 expression using an immunohistochemical assay. Results Out of the 25 cell lines, 13 (52%) had undetectable FOXA2 mRNA. The expression of FOXA2 mRNA and Foxa2 protein were congruent in 19/22 cells (p = 0.001). FOXA2 mutations were not identified in primary NSCLCs and were infrequent in cell lines. Focal or broad chromosomal deletions involving FOXA2 were not present. The promoter region of FOXA2 had evidence of hypermethylation, with an inverse correlation between FOXA2 mRNA expression and presence of CpG dinucleotide methylation (p < 0.0001). In primary NSCLC tumor specimens, there was a high frequency of either absence (42/173, 24.2%) or no/low expression (96/173, 55.4%) of Foxa2. In 130 patients with stage I NSCLC there was a trend towards decreased survival in tumors with no/low expression of Foxa2 (HR of 1.6, 95%CI 0.9–3.1; p = 0.122). Conclusions Loss of expression of Foxa2 is frequent in lung cancer cell lines and NSCLCs. The main mechanism of downregulation of Foxa2 is epigenetic silencing through promoter hypermethylation. Further elucidation of the involvement of Foxa2 and other airway transcription factors in the pathogenesis of lung cancer may identify novel therapeutic targets.

Clinical microarray datasets were analyzed to search for new therapeutic targets and prognostic markers of non-small cell lung cancer (NSCLC).Microarray datasets from 90 lung cancer specimens, were analyzed with focus on the FOXD1 gene. Levels of FOXD1 mRNA were assessed in lung cancer cell lines and these levels were correlated with survival.FOXD1-knockdown led to suppression of cell proliferation. Moreover, patients with high FOXD1 expression survived for a significantly shorter time than those with low FOXD1 expression.The expression status of FOXD1 is a novel prognostic factor and may lead to new treatment strategies for NSCLC.

Background: Preliminary studies have identified cancer stem cells (CSCs) in various cancers and there are several ongoing clinical studies targeting these cells. CD44 (standard or variant isoforms) and/or aldehyde dehydrogenase (ALDH) expression is the most commonly used markers for the identification of CSCs. The goal of the current study was to examine the ability of CD44v, either alone or in combination with ALDH, to identify CSCs within human lung cancer cells lines. Methods: We examined several lung adenocarcinoma cell lines for the ability of CD44v and/or ALDH expression to enrich for cells with CSC characteristics such as in vitro differential proliferation rate, chemotherapeutic-resistance, tumorsphere formation, and in vivo tumorigenicity. We also compared their in vivo secondary tumor formation, and histological characteristics of their xenograft tumors, and examined their expression of PD-L1, EGFR, xCT, and reactive oxygen species (ROS). Results: Both CD44vhigh/ALDHhigh and CD44vhigh/ALDHlow cells were enriched in cells with CSC characteristics, with the CD44vhigh/ALDHlow cells being more proliferative and more resistant to chemotherapeutics, whereas CD44vhigh/ALDHhigh cells were more efficient in forming tumorspheres in vitro, in making primary xenograft tumors, and in propagating secondary tumors in vivo. Applying stricter sorting gates to select for cells with the highest CD44v/ALDH expression caused the CD44vhigh/ALDHlow cells to lose their high proliferation rates and chemotherapeutic resistance ability, but enriched for the tumorsphere-forming cells among the CD44vhigh/ALDHhigh and CD44vhigh/ALDHlow cells. CD44vhigh expression was associated with PD-L1 and xCT expression in both H1650 and HCC827 cells. This association was not modified by ALDH expression in the H1650 cell line. However, in the HCC827 cell line, ALDH expression was negatively associated with PD-L1 and positively associated with xCT expression. Conclusion: Lung adenocarcinoma cells with high CD44v expression are enriched for CSCs. Addition of ALDH as an enrichment marker bestowed some CSCs characteristics to CD44vhigh/ALDHlow cells and others to CD44vhigh/ALDHhigh cells. We propose that lung adenocarcinoma contains different CSCs, each of them endowed with different CSC characteristics.

Abstract Multiple epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (EGFR-TKI) have been developed to effectively inhibit EGFR-derived signals in non–small cell lung cancer (NSCLC). In this study, we assessed the efficacy of EGFR-TKIs, including a novel third-generation inhibitor naquotinib (ASP8273), in clinically relevant EGFR mutations, including L858R, exon 19 deletion, L858R+T790M, exon 19 deletion+T790M with or without a C797S mutation, and several exon 20 insertion mutations. Using structural analyses, we also elucidated the mechanism of activation and sensitivity/resistance to EGFR-TKIs in EGFR exon 20 insertion mutations. The efficacy of naquotinib in cells with L858R, exon 19 deletion and exon 19 deletion+T790M was comparable with that of osimertinib. Interestingly, naquotinib was more potent than osimertinib for L858R+T790M. Additionally, naquotinib and osimertinib had comparable efficacy and a wide therapeutic window for cells with EGFR exon 20 insertions. Structural modeling partly elucidated the mechanism of activation and sensitivity/resistance to EGFR-TKIs in two EGFR exon 20 insertion mutants, A767_V769dupASV and Y764_V765insHH. In summary, we have characterized the efficacy of EGFR-TKIs for NSCLC using in vitro and structural analyses and suggested the mechanism of activation and resistance to EGFR-TKIs of EGFR exon 20 insertion mutations. Our findings should guide the selection of appropriate EGFR-TKIs for the treatment of NSCLC with EGFR mutations and help clarify the biology of EGFR exon 20 insertion mutations. Mol Cancer Ther; 17(4); 740–50. ©2018 AACR.

Prognostic understanding in advanced cancer patients and their caregivers may have an impact on the delivery of effective care. The aims of this study were to explore prognostic understanding at diagnosis in both patients with advanced lung cancer and their caregivers and to investigate correlates of their understanding.A total of 193 patients with newly diagnosed advanced lung cancer and their 167 caregivers were enrolled at 16 hospitals in Japan. We assessed their perceptions of prognosis and goals of therapy and examined their associations with their sociodemographic characteristics, clinical status, quality of life, mood symptoms, and the status of disclosure of information by their treating physicians.One fifth of patients and caregivers (21.7% and 17.6%, respectively) mistakenly believed that the patients' cancer was "completely curable." Substantial proportions of them (16.9% and 10.3%, respectively) mistakenly believed that the primary goal of therapy was to remove all the cancer. Levels of anxiety and depression in both patients and caregivers were significantly higher among those who had accurate understanding of prognosis. In multivariate analyses, inaccurate perceptions of prognosis in patients were associated with sex, better emotional well-being, and lower lung cancer-specific symptom burden. Caregivers' inaccurate perceptions of patients' prognoses were associated with better performance status and better emotional well-being of patients.Substantial proportions of advanced lung cancer patients and their caregivers misunderstood their prognosis. Interventions to improve their accurate prognostic understanding should be developed with careful attention paid to its associated factors.This study demonstrated that substantial proportions of patients with newly diagnosed advanced lung cancer and their caregivers had misunderstandings about their prognosis. Accurate perceptions of prognosis, which are indispensable in the delivery of effective care, were associated with elevated levels of anxiety and depression in both patients and caregivers, warranting psychosocial care and support for them immediately after diagnosis. Inaccurate perceptions of prognosis in patients were associated with better emotional well-being and lower lung cancer-specific symptom burden. Illness understanding in caregivers was associated with patients' physical and mental status. Those findings provide insight into how they obtain accurate illness understanding.

Study Design. A multicenter, prospective, single-arm, intervention study. Objective. The aim of this study was to investigate efficacy of balloon kyphoplasty (BKP) for acute osteoporotic vertebral fractures (OVFs) in patients with poor prognostic factors. Summary of Background Data. The indications for BKP remain unclear. Characteristic magnetic resonance imaging (MRI) findings (high-intensity or diffuse low-intensity area in fractured vertebrae on T2-weighted images) were reportedly predictive of delayed union. Methods. This study enrolled 106 patients with poor prognostic MRI findings who underwent BKP within 2 months after injury, and 116 controls with acute OVFs and the same poor prognostic factors who underwent conservative treatment. Patients were propensity score matched in a logistic regression model adjusted for age, sex, number of baseline old fractures, and fracture level. The primary outcome was reduction in activities of daily living (ADLs) at 6 months after fracture, and the secondary outcomes were improvement in short-form (SF)-36 subscales, back pain visual analog scale (VAS) score, and vertebral body deformity. Results. A decrease in ADLs occurred in 5.6% of patients in the BKP group and 25.6% of patients in the conservative treatment group ( P &lt; 0.001). The SF-36 vitality subscale score improved by 26.9 ± 25.9 points in the BKP group and 14.5 ± 29.4 points in the control group ( P = 0.03). The VAS pain score improved by 43.4 ± 34.4 in the BKP group and 52.2 ± 29.8 in the control group ( P = 0.44). The vertebral body wedge angle improved by 5.5 ± 6.2° in the BKP group and −6.3 ± 5.0° in the control group ( P &lt; 0.0001). The percent vertebral body height improved by 15.2 ± 19.2% in the BKP group and −20.6 ± 14.2% in the control group ( P &lt; 0.0001). Conclusion. ADLs, quality of life, and vertebral deformity showed greater improvement with BKP intervention for acute OVF with poor prognostic factors than with conservative treatment at 6 months after injury. Our treatment strategy uses BKP intervention according to the presence or absence of poor prognostic MRI findings. Level of Evidence: 4

The authors compared the clinical outcomes of microscopic bilateral decompression via a unilateral approach (MBDU) for the treatment of degenerative lumbar scoliosis (DLS) and for lumbar canal stenosis (LCS) without instability. The authors also compared postoperative spinal instability in terms of different approach sides (concave or convex) following the procedure.The authors retrospectively reviewed data obtained in 50 consecutive patients (25 in the DLS group and 25 in the LCS group) who underwent MBDU; the minimum follow-up period was 2 years. Patients with DLS were divided into 2 subgroups according to the surgical approach side: a concave group (23 segment) and a convex group (17 segments). The Japanese Orthopaedic Association Scale scores for the assessment of low-back pain were evaluated before surgery and at final follow-up. The Japanese Orthopaedic Association Scale scores and recovery rates were compared between the DLS and LCS groups, and between the convex and concave groups. Cobb angle and scoliotic wedging angle (SWA) were evaluated on standing radiographs before surgery and at final follow-up. Facet joint preservation (the percentage of preservation) was assessed on pre- and postoperative CT scans, compared between the LCS and DLS groups, and compared between the concave and convex groups. The influence of approach side on postoperative progression of segmental instability was also examined in the DLS group.The mean recovery rate was 58.7% in the DLS and 62.0% in the LCS group. The mean recovery rate was 58.6% in the convex group and 60.6% in the concave group. There were no significant differences in recovery rates between the LCS and DLS groups, or between the DLS subgroups. The mean Cobb angles in the DLS group were significantly increased from 12.7° preoperatively to 14.1° postoperatively (p < 0.05), and mean preoperative SWAs increased significantly from 6.2° at L3-4 and 4.1° at L4-5 preoperatively to 7.4° and 4.9°, respectively, at final follow-up (p < 0.05). There was no significant difference in percentage of preservation between the DLS and LCS groups. The mean percentages of preservation on the approach side in the DLS group at L3-4 and L4-5 were 89.0% and 83.1% in the convex group, and those in the concave group were 67.3% and 77.6%, respectively. The percentage of preservation at L3-4 was significantly higher in the convex than the concave group. The mean SWA had increased in the concave group (p = 0.01) but not the convex group (p = 0.15) at final follow-up.The MBDU can reduce postoperative segmental spinal instability and achieve good postoperative clinical outcomes in patients with DLS. The convex approach provides surgeons with good visibility and improves preservation of facet joints.

PTPRH is a receptor-type protein tyrosine phosphatase thought to be a potential regulator of tumorigenesis. The aim of the present study was to clarify the significance of PTPRH expression and its regulation by DNA methylation in non-small cell lung cancer (NSCLC), especially in lung adenocarcinoma (LADC). PTPRH mRNA expression was examined in 89 NSCLC and corresponding non-cancerous tissues. The correlation between DNA methylation and PTPRH gene expression was investigated in another cohort that consisted of 145 patients with LADC, a major NSCLC subtype. Gene regulation by DNA methylation was assessed using a DNA methylation inhibitor. PTPRH mRNA expression was significantly upregulated in NSCLC. PTPRH DNA methylation was reduced in LADC samples and inversely correlated with mRNA expression. 5-Aza-2'-deoxycytidine treatment of lung cancer cell lines with low PTPRH expression, restored mRNA PTPRH expression levels. Furthermore, low PTPRH methylation was associated with shorter recurrence-free survival (P=1.64x10-4) and overall survival (P=5.54x10-5). Multivariate analysis revealed that PTPRH DNA methylation was an independent prognostic factor (P=6.88x10-3). It was confirmed that PTPRH is overexpressed in NSCLC. Furthermore, we determined that PTPRH is epigenetically regulated by DNA hypomethylation, with prognostic implications for LADC.

Significant advances have been made since the development of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) targeting EGFR mutations in non-small-cell lung cancer (NSCLC), however, lung cancer cells eventually acquire resistance to those agents. Osimertinib (AZD9291) has been developed as 3rd generation EGFR-TKI with activities against sensitizing mutations and T790 M resistance mutation, which account for about 50% of the mechanisms of acquired resistance to 1st or 2nd generation EGFR-TKIs. A recent phase I/II clinical trial with osimertinib for advanced NSCLC patients with known sensitizing EGFR mutations and documented disease progression on prior EGFR-TKIs revealed promising effect with acceptable toxicities. Areas covered: This article summarizes current understanding and available preclinical and clinical data on osimertinib and also discusses future directions. The literature search included PubMed and the latest articles from international conferences. Expert commentary: The development of osimertinib has provided new therapeutic options for NSCLC patients harboring T790 M. Compared with other EGFR-TKIs including rociletinib, osimertinib seems to possess an advantage with respect to the effect and safety profile among existing EGFR-TKIs. However, tumor progression still occurs even when treating with osimertinib. A further understanding of the mechanisms of resistance is eagerly anticipated in order to develop next generation EGFR-TKIs.

Objectives Tumor-associated macrophages (TAMs) are known to promote tumorigenesis but the mechanism(s) remain elusive. We have developed a mouse model of lung cancer that is initiated through an inducible overexpression of fibroblast growth factor 9 (FGF9) in type-2 pneumocytes. Expression of FGF9 in adult lungs resulted in a rapid development of multiple adenocarcinoma-like tumor nodules, and is associated with an intense immunological reaction. The purpose of this study is to characterize the immune response to the FGF9-induced lung adenocarcinoma and to determine the contribution of TAMs to growth and survival of these tumors. Materials and methods We used flow cytometry, immunostaining, RT-PCR and in vitro culture system on various cell populations isolated from the FGF9-induced adenocarcinoma mouse lungs. Results Immunostaining demonstrated that the majority of the inflammatory cells recruited to FGF9-induced lung tumors were macrophages. These TAMs were enriched for the alternatively activated (M2) macrophage subtype. TAMs performed a significantly high immune suppressive function on T-cells and displayed high levels of arginase-1 expression and activity. The growth and colony forming potential of tumor cells was induced by co-culture with TAMs. Additionally, TAMs were shown to promote fibroblast proliferation and angiogenesis. TAMs had high expression of Tgf-β, Vegf, Fgf2, Fgf10, Fgfr2 and several matrix metalloproteinases; factors that play multiple roles in supporting tumor growth, immune protection, fibroblast activation and angiogenesis. Conclusion Our results provide evidence that the Fgf9-induced lung adenocarcinoma is associated with recruitment and activation of M2-biased TAMs, which provided multiple means of support to the tumor. This model represents an excellent means to further study the complex interactions between TAMs, their related chemokines, and progression of lung adenocarcinoma, and adds further evidence to support the importance of TAMs in tumorigenesis.

We investigated the effect of paravertebral muscle (PVM) on poor prognosis in osteoporotic vertebral fracture (OVF) and remaining lower back pain (LBP) in the thoracolumbar and lower lumbar regions. Additional OVF occurrence in the thoracolumbar and remaining LBP in the lumbar region was significantly related to PVM fat infiltration percentage.Paravertebral muscle (PVM) is an important component of the spinal column. However, its role in the healing process after osteoporotic vertebral fracture (OVF) is unclear. This study aimed to clarify the effect of PVM in thoracolumbar and lower lumbar regions on OVF clinical and radiological outcomes.This was a multicenter prospective cohort study from 2012 to 2015. Patients ≥ 65 years old who presented within 2 weeks after fracture onset were followed up for 6 months. PVM was measured at the upper edge of the L1 and L5 vertebral body in the magnetic resonance imaging (MRI) T2-axial position at registration. The cross-sectional area (CSA), relative CSA (rCSA), and fat infiltration percentage (FI%) were measured. Severe vertebral compression, delayed union, new OVF, and remaining low back pain (LBP) were analyzed.Among 153 patients who were followed up for 6 months, 117 with measurable PVM were analyzed. Their average age was 79.1 ± 7.2 years, and 94 were women (80.3%). There were 48 cases of severe vertebral compression, 21 delayed unions, 11 new OVF, and 27 remaining LBP. Among all poor prognoses, only the FI% of the PVM was significantly associated with new OVF (p = 0.047) in the thoracolumbar region and remaining LBP (p = 0.042) in the lumbar region.The occurrence of additional OVF in the thoracolumbar region and remaining LBP in the lumbar region was significantly related to the FI% of the PVM. Physicians should be aware that patients with such fatty degeneration shown in acute MRI may require stronger treatment.

Transdifferentiation of lung adenocarcinoma to small cell lung cancer (SCLC) has been reported in a subset of lung cancer cases that bear EGFR mutations. Several studies have reported the prerequisite role of TP53 and RB1 alterations in transdifferentiation. However, the mechanism underlying transdifferentiation remains understudied, and definitive additional events, the third hit, for transdifferentiation have not yet been identified. In addition, no prospective experiments provide direct evidence for transdifferentiation. In this study, we show that FGF9 upregulation plays an essential role in transdifferentiation. An integrative omics analysis of paired tumor samples from a patient with transdifferentiated SCLC exhibited robust upregulation of FGF9. Furthermore, FGF9 upregulation was confirmed at the protein level in four of six (66.7%) paired samples. FGF9 induction transformed mouse lung adenocarcinoma-derived cells to SCLC-like tumors in vivo through cell autonomous activation of the FGFR pathway. In vivo treatment of transdifferentiated SCLC-like tumors with the pan-FGFR inhibitor AZD4547 inhibited growth. In addition, FGF9 induced neuroendocrine differentiation, a pathologic characteristic of SCLC, in established human lung adenocarcinoma cells. Thus, the findings provide direct evidence for FGF9-mediated SCLC transdifferentiation and propose the FGF9-FGFR axis as a therapeutic target for transdifferentiated SCLC. SIGNIFICANCE: This study demonstrates that FGF9 plays a role in the transdifferentiation of lung adenocarcinoma to small cell lung cancer.

Study Design: A cross-sectional study. Objective: This study was aimed to quantify the extent of lumbar endplate and facet joint erosion in patients with rheumatoid arthritis (RA) using magnetic resonance imaging, to calculate the prevalence of erosion, and sought correlated factors. Summary of Background Data: Few studies have examined the lumbar spine in RA, especially the relationship between endplate and facet joint erosions and lumbar lesions induced by RA. Methods: A total of 201 patients with RA were enrolled. Lumbar endplate and facet joint erosion were defined as irregularities and low-intensity change on magnetic resonance imaging, and graded utilizing the Rheumatoid Arthritis Magnetic Resonance Imaging Score. Lumbar lesions were defined as scoliosis, spondylolisthesis, and vertebral fracture on plain x-ray. Multivariable logistic regression analysis was used to seek correlations between the erosion and spinal level, RA-related factors and x-ray findings. Results: Lumbar endplate and facet erosion were detected in 70.6% and 76.6% of individuals, respectively, and at 33.8% and 38.7% of lumbar intervertebral levels, respectively. The severity of erosion in individual patients correlated with lumbar lesions. Endplate and facet erosion at each level correlated with high disease activity, and were most common at mid-lumbar and lower-lumbar levels. Strong correlations were observed between endplate erosion and adjacent vertebral body fracture or disk degeneration, and between facet erosion and spondylolisthesis. Conclusions: Lumbar endplate and facet erosion are common in RA, and are observed more frequently at mid-low levels and when RA is poorly controlled. This pattern of erosion may play a crucial role in the generation of lumbar lesions in RA.

There has been no report regarding the results of two-level keyhole foraminotomy. The purpose of this study was to detail clinical outcomes following consecutive two-level cervical foraminotomy (tandem keyhole foraminotomy (TKF)) in patients with radiculopathy. The authors conducted a retrospective review of 35 cases involving patients treated by a single surgeon using TKF. Clinical symptoms, data of physical examinations, pathology and clinical outcomes were detailed and discussed about this surgical method. Patients consisted of cervical disc herniation (CDH) (19/35), cervical spondylotic radiculopathy (CSR) (13/35), and cervical spondylotic amyotrophy (CSA) (3/35). TKF was performed from C3 to C5 in 2 patients (6%), from C4 to C6 in 7 patients (20%), from C5 to C7 in 23 patients (66%), and from C6 to T1 in 3 patients (8%). The mean operative duration was 99.2 min (range, 72 to 168 min). The mean estimated blood loss was 55.8 g (range, 0 to 200 g). Radicular pain was relieved within 3 months in 88% (29/32) and in 97% (31/32) at final follow-up. Resolution of muscle weakness was recognized within 6 months after operation in all CSA cases. Sixty-six percent of patients showed a greater than 20% deficit in grip weakness on the affected side compared with the normal side. After pain was relieved, grip strength improved by more than 15%. TKF is a safe and highly effective procedure for patients with cervical radiculopathy and does not require invasive preoperative examinations. Further investigation is required to determine the effects of consecutive facetectomy.

Fibroblast growth factor 9 (FGF9) is essential for lung development and is highly expressed in a subset of human lung adenocarcinomas. We recently described a mouse model in which FGF9 expression in the lung epithelium caused proliferation of the airway epithelium at the terminal bronchioles and led to rapid development of adenocarcinoma. Here, we used this model to characterize the effects of prolonged FGF9 induction on the proximal and distal lung epithelia, and examined the propagation potential of FGF9-induced lung tumours. We showed that prolonged FGF9 over-expression in the lung resulted in the development of adenocarcinomas arising from both alveolar type II and airway secretory cells in the lung parenchyma and airways, respectively. We found that tumour cells harboured tumour-propagating cells that were able to form secondary tumours in recipient mice, regardless of FGF9 expression. However, the highest degree of tumour propagation was observed when unfractionated tumour cells were co-administered with autologous, tumour-associated mesenchymal cells. Although the initiation of lung adenocarcinomas was dependent on activation of the FGF9-FGF receptor 3 (FGFR3) signalling axis, maintenance and propagation of the tumour was independent of this signalling. Activation of an alternative FGF-FGFR axis and the interaction with tumour stromal cells is likely to be responsible for the development of this independence. This study demonstrates the complex role of FGF-FGFR signalling in the initiation, growth and propagation of lung cancer. Our findings suggest that analysing the expressions of FGF-FGFRs in human lung cancer will be a useful tool for guiding customized therapy.

Objectives. Cervical spine instability (CSI) is commonly involved in patients with rheumatoid arthritis (RA). Although the treatment for RA has dramatically changed due to methotrexate and biologics, it is unclear whether this change contributes to the prevalence of CSI or not. Our objectives were to update the current prevalence of CSI and to investigate the factors associated with CSI. Methods. A cross-sectional study of patients with RA was conducted in our outpatient clinic. Clinical information and symptoms related to CSI were obtained. Plain radiography and magnetic resonance imaging were performed. CSI included atlantoaxial subluxation (AAS), vertical subluxation (VS), and subaxial subluxation (SAS). Results. Two hundred and twenty patients were analyzed, 93 (42%) of whom had CSI. A ≥ 10-year disease duration, Steinbrocker stage III, and three or more narrowed disc spaces from C2/3 to C6/7 were significantly associated with CSI. A neck pain VAS was associated with VS, but not with AAS and SAS. In contrast, methotrexate and biologics had no effect on CSI. Conclusion. The prevalence of CSI in this study was lower compared to previous reports before the approval of biologic, although we failed to detect the effect of biologics. Further prospective studies are needed to elucidate the efficacy.

Background: Osimertinib, a third-generation epidermal growth factor receptor-tyrosine kinase inhibitor, exerts remarkable effects against EGFR T790M resistance mutation-positive nonsmall cell lung cancer.Identifying T790M mutation by re-biopsy is essential before prescribing osimertinib.Tissue biopsy is the golden standard for this purpose, but several factors limit its success rate.The liquid biopsy with blood, using circulating tumor DNA, has been an alternative method.However, the true biological meaning and equivalence of liquid biopsy and tumor biopsy are still under investigation.Especially, the usefulness of serum samples to detect T790M mutation is not yet been known.Patients and methods: We prospectively evaluated the sensitivity, specificity, and parallelism of the detection of EGFR mutations in tissue re-biopsy and liquid biopsy (plasma and serum), simultaneously, from June 2016 to May 2017.EGFR mutations in tumor re-biopsy were evaluated by COBAS ver2 and PNA-LNA PCR clamp method, and those in liquid biopsy were evaluated with COBAS ver2.Results: Fifteen patients were enrolled.In 10 patients whose EGFR mutation was detected in liquid biopsy, the original EGFR mutation (exon 19 del or L858R) was detected in all patients.Detection of EGFR mutation by COBAS ver2 and by PNA-LNA method was almost the same in tissue re-biopsy.The detection rate of T790M was lower than that of the original EGFR mutation in liquid biopsy compared to that in tissue re-biopsy.The detection of T790M in serum exhibited a higher specificity (67%) and positive predictive value (50%) than that in plasma (50% and 40%, respectively).The detection sensitivity was similar in plasma and serum.Conclusion: Plasma, serum, and tissue genotyping can have complementary roles for detecting EGFR-T790M using COBAS ver2.Repeated tests with different samples and different methods may improve accuracy of T790M detection and will lead to the maximum benefit for the patient.

Balloon kyphoplasty or vertebroplasty is widely performed as a surgical intervention for osteoporotic vertebral fracture (OVF) and the effects have been investigated in many previous studies. However, the influence of the timing of the procedure on patient outcomes has not been studied formally. The purpose of this study was to investigate differences in the surgical outcomes of OVFs according to the timing of balloon kyphoplasty.This was a multicenter cohort study. Participants comprised 72 consecutive patients who underwent balloon kyphoplasty between January 2012 and January 2016. Patients were analyzed in two groups according to the timing of kyphoplasty after onset (Early group: ≤2 months; Late group: >2 months). Follow-up continued for more than 6 months.A total of 72 patients were effectively analyzed. Of these, 27 (38%) patients underwent kyphoplasty within 2 months after symptom onset. The Late group showed greater angular motion of fractured vertebrae (p = 0.005) and compression of anterior vertebral height (p = 0.001) before surgery. Final outcomes adjusted for age and preoperative outcome showed lower visual analog scale (VAS) scores for low back pain in the Early group than in the Late group (19.9 vs. 30.4, p = 0.049). Final relative anterior vertebral height and kyphotic angle were more preserved in the Early group than in the Late group (p = 0.002 and p = 0.020, respectively), although absolute differences were not significant.Vertebral height and kyphotic angle before and after balloon kyphoplasty were greater in patients who underwent kyphoplasty within 2 months after onset, and the VAS score for low back pain at final follow-up was better. Our results support kyphoplasty within 2 months.

Abstract Background Patients with non‐small cell lung cancer (NSCLC) harboring activating EGFR mutations are sensitive to epidermal growth factor receptor‐tyrosine kinase inhibitors (EGFR‐TKIs) but inevitably develop resistance to the inhibitors mostly through acquisition of the secondary T790M mutation. Although third‐generation EGFR‐TKIs overcome this resistance by selectively inhibiting EGFR with EGFR‐TKI‐sensitizing and T790M mutations, acquired resistance to third‐generation EGFR‐TKIs invariably develops. Methods Next‐generation sequencing (NGS) and fluorescence in situ hybridization (FISH) analysis were performed in an EGFR T790M ‐mutated NSCLC patient who had progressed after a third‐generation EGFR‐TKI, TAS‐121. EGFR ‐mutated cell lines were subjected to a cell proliferation assay and western blotting analysis with EGFR‐TKIs and a heat shock protein 90 (HSP90) inhibitor. Results NGS and FISH analysis revealed EGFR amplification in the resistant cancer cells. While EGFR L858R/T90M ‐mutated cell line was sensitive to osimertinib or TAS‐121 in vitro, EGFR ‐overexpressing cell lines displayed resistance to these EGFR‐TKIs. Western blot analysis showed that EGFR phosphorylation and overexpression of EGFR in cell lines was not suppressed by third‐generation EGFR‐TKIs. In contrast, an HSP90 inhibitor reduced total and phosphorylated EGFR and inhibited the proliferation of resistant cell lines. Conclusions EGFR amplification confers resistance to third‐generation EGFR‐TKIs which can be overcome by HSP90 inhibition. The results provide a preclinical rationale for the use of HSP90 inhibitors to overcome EGFR amplification‐mediated resistance.

Abstract Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKI) show antitumor activity in a subset of non–small cell lung cancer (NSCLC) patients. However, the initial tumor response is followed by recurrence. Several studies have suggested the importance of other receptor tyrosine kinases (RTK) and downstream kinases as potential targets in the treatment of NSCLC. We used the multiple-RTK inhibitor AEE788, which inhibits EGFR, vascular endothelial growth factor receptor, and human epidermal growth factor receptor 2, with and without the downstream kinase inhibitor RAD001 (an inhibitor of mammalian target of rapamycin). AEE788 inhibited cell growth more effectively than did erlotinib in three NSCLC cell lines examined (A549, H1650, and H1975). However, in the EGFR-TKI–resistant cell line H1975 harboring T790M resistance mutation, cell growth inhibition by AEE788 was only mild, and the phosphorylation of its leading targets such as EGFR and vascular endothelial growth factor receptor 2 was not inhibited. In H1975, AEE788 induced significantly greater cell growth inhibition when combined with RAD001 than when used alone. This cooperative effect was not seen with the combination of erlotinib and RAD001. We found that c-Met was highly phosphorylated in this cell line, and the phosphorylated c-Met was inhibited effectively by AEE788. Using a phospho-RTK array, the phosphorylation of c-Met and insulin-like growth factor-I receptor was inhibited by AEE788. These results suggest that upstream RTK inhibitor overcomes the acquired resistance to EGFR-TKI when combined with downstream kinase inhibitor. Thus, the combined inhibition of upstream and downstream RTKs is a promising strategy for the treatment of NSCLC. Mol Cancer Res; 8(8); 1142–51. ©2010 AACR.

Study Design: A cross-sectional imaging study. Objective: To investigate the relationship between cervical and lumbar spine lesions in patients with rheumatoid arthritis (RA), and to analyze associated factors in those with concurrent cervical and lumbar endplate erosion (EE). Summary of Background Data: Both the lumbar and cervical spines are often involved in RA, but little is known about the relationship between cervical and lumbar lesions. EE is often found in the spine of RA patients, but its prevalence and associated factors have not been well studied. Methods: We enrolled 201 RA patients in this study. Cervical lesions (horizontal and vertical atlantoaxial dislocation, subaxial subluxation) and lumbar lesions (vertebral fracture, scoliosis, spondylolisthesis) were evaluated on plain radiographs. EE was evaluated on sagittal T1-weighted magnetic resonance images, and graded into 4 categories. The prevalence of each lesion was calculated, and correlations between general cervical and lumbar lesions and between cervical and lumbar EE were analyzed. To assess the clinical condition of RA, we evaluated disease duration, Steinbrocker stage, modified Stanford Health Assessment Questionnaire results, and Disease Activity Score for 28 joints with the erythrocyte sedimentation rate. Factors associated with concurrent lumbar and cervical EE were analyzed using multiple logistic regression analysis. Results: Cervical lesions were found in 42.3% of patients and lumbar lesions in 56.2%. There was no significant correlation between the presence of cervical and the presence of lumbar lesions, but patients with cervical subaxial subluxation were significantly more likely to have lumbar spondylolisthesis. Cervical EE (≥1 points) was found in 61.2% of patients and lumbar EE in 39.3%. Total cervical EE score was significantly correlated with lumbar EE score. Moderate/high disease activity, Steinbrocker stage IV, and severe cervical or lumbar disk degeneration were associated with concurrent cervical and lumbar EE. Conclusions: Some cervical lesions are significantly associated with lumbar spinal lesions. Concurrent cervical and lumbar EE are related to RA disease activity and peripheral joint deterioration, suggesting that RA activity may play an important role in total spinal involvement.

OBJECTIVE Increasing soft-tissue mass posterior to the odontoid process causes spinal cord compression. Retro-odontoid pseudotumors are considered to be associated with atlantoaxial instability in patients with rheumatoid arthritis (RA), but the exact mechanism by which these lesions develop has not been elucidated. The purpose of this study was to identify the relationships between retro-odontoid soft-tissue (ROST) thickness and radiological findings or clinical data in patients with RA. METHODS A total of 201 patients with RA who had been followed up at the outpatient clinic of the authors' institution were enrolled in this study. ROST thickness was evaluated on midsagittal T1-weighted MRI. The correlations between ROST thickness and radiographic findings or clinical data on RA were examined. The independent factors related to ROST thickness were analyzed using stepwise multiple regression analysis. RESULTS The average thickness of ROST was 3.0 ± 1.4 mm. ROST thickness showed an inverse correlation with disease duration (r = −0.329, p &lt; 0.01), Steinbrocker stage (r = −0.284, p &lt; 0.01), the atlantodental interval (ADI) in the neutral position (r = −0.326, p &lt; 0.01), the ADI in the flexion position (r = −0.383, p &lt; 0.01), and the ADI in the extension position (r = −0.240, p &lt; 0.01). On stepwise multiple regression analysis, ADI in the flexion position and Steinbrocker stage were independent factors associated with ROST thickness. CONCLUSIONS Although the correlations were not strong, ROST thickness in patients with RA was inversely correlated with ADI and Steinbrocker stage. In other words, ROST thickness tends to be smaller as atlantoaxial instability and peripheral joint destruction worsen. Clinical trial registration no.: UMIN000000980 (UMIN Clinical Trials Registry)

Abstract EGFR mutation-positive patients with non–small cell lung cancer (NSCLC) respond well to treatment with EGFR–tyrosine kinase inhibitors (EGFR–TKI); however, treatment with EGFR–TKIs is not curative, owing to the presence of residual cancer cells with intrinsic or acquired resistance to this class of drugs. Additional treatment targets that may enhance the efficacy of EGFR–TKIs remain elusive. Using a CRISPR/Cas9-based screen, we identified the leucine-rich repeat scaffold protein SHOC2 as a key modulator of sensitivity to EGFR–TKI treatment. On the basis of in vitro assays, we demonstrated that SHOC2 expression levels strongly correlate with the sensitivity to EGFR–TKIs and that SHOC2 affects the sensitivity to EGFR–TKIs in NSCLC cells via SHOC2/MRAS/PP1c and SHOC2/SCRIB signaling. The potential SHOC2 inhibitor celastrol phenocopied SHOC2 depletion. In addition, we confirmed that SHOC2 expression levels were important for the sensitivity to EGFR–TKIs in vivo. Furthermore, IHC showed the accumulation of cancer cells that express high levels of SHOC2 in lung cancer tissues obtained from patients with NSCLC who experienced acquired resistance to EGFR–TKIs. These data indicate that SHOC2 may be a therapeutic target for patients with NSCLC or a biomarker to predict sensitivity to EGFR–TKI therapy in EGFR mutation-positive patients with NSCLC. Our findings may help improve treatment strategies for patients with NSCLC harboring EGFR mutations. Implications: This study showed that SHOC2 works as a modulator of sensitivity to EGFR–TKIs and the expression levels of SHOC2 can be used as a biomarker for sensitivity to EGFR–TKIs.

Introduction The rapid spread of COVID-19 posed a global burden. Substantial number of people died of the disease in the acute phase of infection. In addition, a significant proportion of patients have been reported to suffer from post-acute phase symptoms, sequelae of COVID-19, which may negatively influence the quality of daily living and/or socioeconomic circumstances of the patients. However, no previous study has comprehensively and objectively assessed the quality of life of patients by using existing international scales. Further, evidence of socioeconomic consequences among patients with COVID-19 is scarce. To address the multidimensional issues from sequelae of COVID-19, evidence from comprehensive surveys beyond clinical perspectives is critical that investigates health, and social determinants of disease progression as well as socioeconomic consequences at a large scale. Methods and analysis In this study, we plan to conduct a nationwide and comprehensive survey for the sequelae of COVID-19 in a total of 1000 patients diagnosed at 27 hospitals throughout Japan. This study will evaluate not only the health-related status of patients from clinical perspectives but also the Health-related Quality of Life (HRQoL) scores, socioeconomic status and consequences to discuss the sequelae of the disease and the related risk factors. The primary endpoint is the frequency of long-term complications of COVID-19 infection. The secondary endpoints are risk factors for progression to sequelae of COVID-19 infection. The study will provide robust and important evidence as a resource to tackle the issues from the sequelae of COVID-19 from the multi-dimensional perspectives. Ethics and dissemination This trial was approved by the Keio University School of Medicine Ethics Committee (20200243, UMIN000042299). The results of this study will be reported at a society meeting or published in a peer-reviewed journal.

Neutropenia, a rare immune-related adverse event, affects patients receiving treatment with immune checkpoint inhibitors (ICIs). We herein report a case of pembrolizumab-induced agranulocytosis. An 83-year-old man was diagnosed with advanced-stage lung carcinoma concomitant with splenomegaly complicated by hypersplenism, causing pancytopenia. To avoid the risk of bone marrow suppression due to cytotoxic chemotherapy, pembrolizumab monotherapy was chosen. However, the patient developed agranulocytosis despite the resolution of pancytopenia through splenectomy performed after the fourth pembrolizumab cycle. Neutrophil counts improved after steroid treatment but not after granulocyte colony-stimulating factor treatment. This case demonstrated that neutropenia can sometimes develop abruptly after several ICI therapy cycles.

UV-B radiation induces sunburn, and neutrophils are pivotal in this inflammation. In this study, we examined the potential involvement of neutrophil extracellular traps (NETs) in ultraviolet B (UVB)-induced skin inflammation, correlating the skin inflammation-mitigating effects of Hochu-ekki-to on UV-B irradiation and NETs. To elucidate NET distribution in the dorsal skin, male ICR mice, exposed to UVB irradiation, were immunohistologically analyzed to detect citrullinated histone H3 (citH3) and peptidylarginine deiminase 4 (PAD4). Reactive oxygen species (ROS) production in the bloodstream was analyzed. To establish the involvement of NET-released DNA in this inflammatory response, mice were UV-B irradiated following the intraperitoneal administration of DNase I. In vitro experiments were performed to scrutinize the impact of Hochu-ekki-to on A23187-induced NETs in neutrophil-like HL-60 cells. UV-B irradiation induced dorsal skin inflammation, coinciding with a significant increase in citH3 and PAD4 expression. Administration of DNase I attenuated UV-B-induced skin inflammation, whereas Hochu-ekki-to administration considerably suppressed the inflammation, correlating with diminished levels of citH3 and PAD4 in the dorsal skin. UV-B irradiation conspicuously augmented ROS and hydrogen peroxide (H2O2) production in the blood. Hochu-ekki-to significantly inhibited ROS and H2O2 generation. In vitro experiments demonstrated that Hochu-ekki-to notably inhibited A23187-induced NETs in differentiated neutrophil-like cells. Hence, NETs have been implicated in UV-B-induced skin inflammation, and their inhibition reduces cutaneous inflammation. Additionally, Hochu-ekki-to mitigated skin inflammation by impeding neutrophil infiltration and NETs in the dorsal skin of mice.

Massive frozen stocked allogeneic bone grafts are often used to reconstruct large bone defects caused by trauma or tumor resections. However, the long-term failure rate of such massive allografts was reported to be 25% because of infection, fracture, and nonunion. In this study, we evaluated the ability of a recombinant human bone morphogenetic protein (rhBMP)-2-retaining paste to promote the osteogenic potential of frozen stocked allogeneic bone grafts to repair intercalated femoral shaft defects in a rat model.After confirming the transplantation intolerance between two rat strains (Wistar and Lewis) by skin transplantation from Lewis rats to Wistar rats, an 8-mm-long bone segment was removed from the Wistar rats, and a frozen stocked allograft coated with the rhBMP-2-retaining paste from the Lewis rats was placed into the defect and subjected to intramedullary fixation with an 18-gauge injection needle pin. The allografted femurs were evaluated by radiographic, histologic, and biomechanical examinations at specified time points.The results revealed successful repair of critical-size cortical bone defects by implanting frozen stocked allografts coated with the rhBMP-2-retaining synthetic biodegradable carrier paste from an immunologically intolerant host.This experimental study suggest that allogeneic bone grafting in combination with rhBMP-2 and its local delivery system may represent an innovative approach to the reconstruction of bone defects.

To investigate the prevalence and associated factors of severe low back pain (LBP) among patients with rheumatoid arthritis (RA).This cross-sectional study included 201 patients with RA without prior spinal surgery. Severe LBP was defined as that with a visual analog scale (VAS) score of ≥ 50 mm within the previous 4 weeks. Lumbar lesions, sagittal alignment, and disc degeneration were evaluated by plain standing X-rays and magnetic resonance imaging. Associated factors of severe LBP were evaluated using multiple logistic regression analysis.Forty-eight patients (23.8%) had LBP with a VAS score of ≥ 50 mm. Multivariate analysis indicated that the associated factors for severe LBP were female, smoking, and moderate and high disease activity on the Disease Activity Score in 28 joints-erythrocyte sedimentation rate (DAS28-ESR). There was no relationship between severe LBP and any radiological findings. Among DAS28-ESR subscores, patients with severe LBP had significantly higher tender joint counts and VAS scores for general health.The prevalence of severe LBP was relatively high in patients with RA. The factor most closely associated with severe LBP was Disease Activity Score, but not radiological findings. Severe LBP was related to the tender joint count or subjective complaints of RA.

We identified transmembrane protease , serine 4 ( TMPRSS4 ) as a putative, druggable target by screening surgically resected samples from 90 Japanese non‐small‐cell lung cancer (NSCLC) patients using cDNA microarray. TMPRSS4 has two druggable domains and was upregulated in 94.5% of the lung cancer specimens. Interestingly, we found that TMPRSS4 expression was associated with tissue factor pathway inhibitor 2 ( TFPI‐2 ) expression in these clinical samples. In contrast to TMPRSS4, TFPI‐2 expression was downregulated in NSCLC samples. The in vitro induction of TFPI‐2 in lung cancer cell lines decreased the expression of TMPRSS4 mRNA levels. Reporter assay showed that TFPI‐2 inhibited transcription of TMPRSS4 , although partially. Knockdown of TMPRSS4 reduced the proliferation rate in several lung cancer cell lines. When lung cancer cell lines were treated with 5‐aza‐2′‐deoxycytidine or trichostatin A, their proliferation rate and TMPRSS4 mRNA expression levels were also reduced through the upregulation of TFPI‐2 by decreasing its methylation in vitro . The TFPI‐2 methylation level in the low TMPRSS4 group appeared to be significantly low in NSCLC samples ( P = 0.02). We found a novel molecular mechanism that TFPI‐2 negatively regulates cell growth by inhibiting transcription of TMPRSS4 . We suggest that TMPRSS4 is upregulated by silencing of TFPI‐2 through aberrant DNA methylation and contributes to oncogenesis in NSCLC.

Anaplastic lymphoma kinase (ALK) rearrangements are present in an important subset of non-small-cell lung cancer (NSCLC) and predict for response to the tyrosine kinase inhibitor crizotinib. In this study, we evaluated the yet unknown frequency and functional role of ALK splicing isoforms in NSCLC.We analyzed 270 cases of NSCLC for ALK kinase domain splicing aberrations and in addition generated constructs with full-length echinoderm microtubule-associated protein-like 4 (EML4)-ALK (E13;A20) and a splicing isoform.Splicing isoforms of the kinase domain of ALK-including complete skipping of exon 23 (ALKdel23, ALK p.I1171fs*42) and exon 27 (ALKdel27, ALK p.T1312fs*0)-were identified in 11.1% (30 of 270 cases) of NSCLC, and these changes coexisted with ALK rearrangements, KRAS mutations, and EGFR mutations. ALK splicing isoforms were observed with full-length EML4-ALK in crizotinib-naive and treated NSCLCs. ALK T1312fs*0 was unable to render cells solely dependent on ALK signaling. Unlike EML4-ALK and EML4-ALK p.L1196M, EML4-ALK T1312fs*0 did not autophosphorylate ALK or other phosphotyrosine sites. Coexpression of equal amounts of EML4-ALK T1312fs*0 and EML4-ALK did not result in resistance to crizotinib, whereas coexpression of EML4-ALK L1196M with EML4-ALK resulted in resistance to inhibition of ALK by crizotinib.ALK kinase splicing isoforms were present in NSCLC and even if translated seemed to be nonfunctional variants of ALK.

Abstract Vemurafenib improves survival of melanoma patients. However, cutaneous side‐effects commonly occur in them. Nivolumab and ipilimumab are monoclonal antibodies against programmed death 1 and cytotoxic T‐lymphocyte‐associated antigen 4, both of which regulate excessive T‐cell activation. Although these agents induce antitumor immunity against melanoma, the modified immune condition may result in an unexpected adverse reaction which has not been observed previously. Herein, we report a case who manifested severe erythema multiforme‐like eruption with mucosal involvement associated with vemurafenib following nivolumab. The patient also subsequently suffered from ipilimumab‐induced interstitial pneumonia with refractory course. Such a case has never been reported. This case suggested that dermatologists should pay special attention to unexpected adverse events of these drugs, and carefully observe cutaneous and respiratory status of patients during the treatment of melanoma.

Lung cancer and chronic obstructive pulmonary disease are leading causes of morbidity and mortality worldwide, and cigarette smoking is a main risk factor for both. The presence of emphysema, an irreversible lung disease, further raises the risk of lung cancer in patients with chronic obstructive pulmonary disease. The mechanisms involved in smoke-induced tumorigenesis and emphysema are not fully understood, attributable to a lack of appropriate animal models. Here, we optimized a model of cigarette smoke (CS)–induced lung cancer and emphysema in A/J mice treated with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, a potent carcinogen. We investigated whether variations in CS exposure patterns with the same total amount and duration of exposure affect tumorigenesis and/or development of emphysema. Continuous CS exposure for 3 months significantly suppressed 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone–induced development of adenomas and adenocarcinomas; however, emphysema independently developed during this period. Surprisingly, intermittent CS exposure increased the severity of emphysema and resulted in a higher incidence of adenocarcinomas. Furthermore, intermittent CS exposure elicited a marked increase in M2-polarized macrophages within and near the developed tumors. By employing a CS exposure protocol with repeated cycles of cessation and relapse, we provide evidence that intermittent CS exposure enhances tumorigenesis and emphysema progression more than that of continuous CS exposure.

Cancers, including lung cancer, are a leading cause of death worldwide. To overcome this deadly disease, multiple modality inhibitors have been developed. These include cytotoxic agents, molecular targeted small molecules, such as tyrosine kinase inhibitors, and neutralizing antibodies. An aptamer is a short single-stranded nucleic acid molecule that is selected in vitro from a large random sequence library based on its high and specific affinity to a target molecule. Aptamers can be applied to therapeutics of various types of diseases, including cancer, due to their strong and specific neutralizing activities. However, the efficacy of aptamer-based therapy for cancer cells is not well characterized. In this study, we aimed to show that the FGF2 aptamer is effective for the treatment of FGF2 dependent lung cancer cells. We previously developed PC9GR lung cancer cells, whose proliferation is dependent on EGFR and FGF2-FGFR pathways in a cell autonomous manner. Using PC9GR cells, we demonstrate that the addition of the FGF2 aptamer induces more significant inhibition of PC9GR cell proliferation than does the addition of EGFR inhibitor alone. Furthermore, the addition of the FGF2 aptamer more significantly inhibits the downstream signals and induces apoptosis to a higher extent than does the addition of EGFR inhibitor alone. Our results show that the FGF2 aptamer inhibits the growth of FGF2-FGFR pathway-dependent lung cancer cells. The findings provide preclinical evidence that aptamers can be useful for cancer treatment.

Osteoprotic vertebral fractures (OVFs) are common in elderly people. The association between back pain due to OVF with magnetic resonance imaging (MRI) signal change is unclear. In this study we hypothesized that MRI findings would be a predictive factor for back pain measured by visual analogue scale (VAS) at 6 months follow-up.The aim was to study the MRI findings that predict back pain after OVF and the association between radiological findings and scores of back pain.Multicenter prospective cohort study.A total of 153 OVF patients.The outcome measures were VAS back pain and MRI signal change.This study was performed from 2012 to 2015. Consecutive patients with less than 2-week-old OVFs at 11 institutions were enrolled prospectively. MRI was performed at enrollment and at 1, 3, and 6 months follow-up. T1- and T2-weighted images (T1WI and T2W1) were obtained at each time point and their association with VAS scores of back pain at 6 months were investigated. Anterior compression ratio, posterior compression ratio, and angular motion of vertebral bodies were also measured on X-rays at each follow-up. This research had no financial support. There are no conflicts of interest.The 6 months follow-up was completed by 153 patients. At enrollment, the average VAS score of back pain was 75 mm, and it had improved at the 6-month follow-up to an average score of 20 mm. There was a significant correlation between T1 diffuse low signal change and VAS scores at the 6-month follow-up (p<.01). T2 high signal changes (odds ratio; 4.01, p<.01) and old vertebral fractures (odds ratio; 2.47, p=.04) were independent risk factors for back pain. The correlation between angular motion of vertebrae on X-rays and the VAS score of back pain was significant at all time points.This study demonstrates the radiological factors associated with persistent back pain after an OVF and the association between the VAS score of back pain and radiological findings. In addition, T2 high signal changes in acute phase and old vertebral fractures were independent risk factors for residual back pain.

Abstract Balloon kyphoplasty (BKP) sometimes fails to improve patients’ outcomes, with revision surgery, using anterior or posterior reconstruction, being required. The purpose of this study was to investigate the radiological risk factors of failure after BKP in the treatment of osteoporotic vertebral fractures (OVFs). This case-control study included 105 patients treated with single BKP and 14 patients who required revision BKP. We evaluated radiological findings differentiating both groups, using plain radiography and computed tomography, before BKP. Angular flexion-extension motion was significantly greater in the revision than BKP group. While the frequency of pedicle fracture and posterior wall injury was not different between the groups, a split type fracture was more frequent in the revision group. Split type fracture had the highest adjusted odds ratio (OR) for revision (16.5, p = 0.018). Angular motion ≥14° increased the risk for revision surgery by 6-fold (p = 0.013), with endplate deficit having an OR of revision of 5.0 (p = 0.032). The revision rate after BKP was 3.8%, with split type fracture, greater angular motion and large endplate deficit being risk factors for revision. Treatment strategies for patients with these risk factors should be carefully evaluated, considering the inherent difficulties in performing revision surgery after BKP.

Purpose Bronchoscopic microsampling (BMS) is a novel and direct method with which to obtain epithelial lining fluid (ELF) from the lungs. Analysis of DNA hypermethylation of tumor suppressor genes (TSGs) is expected to be a sensitive tool for the early detection of lung cancer. It has been reported that the existence of EGFR mutations and EML4-ALK gene rearrangements are related to the sensitivity of corresponding kinase inhibitors. We aimed to evaluate the suitability of ELF as a sample for analyzing molecular changes specific for lung cancer. Patients and methods We collected ELF from 61 lung cancer patients by BMS from the airway close to the peripheral lung nodule and purified the nucleic acids. We performed methylation specific PCR in each ELF as well as matched serum and tumor tissue for TSGs for DNA methylation analysis. We also examined EGFR mutations and EML4-ALK rearrangement. Results The sensitivity for detecting DNA hypermethylation in ELF vs serum was 74.1% vs 18.5%. We found 60.1% of patients had at least one hypermethylation in ELF, while only 27.9% had it in serum. Of note, DNA hypermethylation was detected even in stage I patients (60.0%) and the detection rate was almost the same level in each stage. We also found the sensitivity for detecting EGFR mutation in ELF vs serum was 58.3% vs 8.3%. We detected an EML4-ALK fusion gene using ELF in one patient. Conclusions BMS is an alternative method to detect cancer specific genetic and epigenetic alterations and will be a useful complementary diagnostic tool for lung cancer. Summary Investigation of genetic and epigenetic changes associated with lung cancer has clinical importance for its diagnosis and management. The clinical usefulness of bronchoscopic microsampling (BMS) in lung cancer has not yet been evaluated. This study demonstrates that BMS could be useful for detecting lung cancer specific molecular changes and valuable for early diagnosis and determination of treatment options for lung cancer.

Cancer cells are surrounded by the extracellular fluid, matrix, and stroma cells. Little is known about how extracellular components such as growth factor ligands affect the biology of lung cancer cells. The objective of this study was to determine whether extracellular fibroblast growth factors (FGFs) can affect the biology of lung cancer cells and to understand how extracellular FGFs affect the biology of lung cancer cells, including non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) cells. Out of the 23 reported FGFs, we focused on FGF2, FGF9 and FGF10. We examined the effect of FGFs on proliferation, treatment sensitivity, and apoptosis of NSCLC (PC9) and SCLC (H69, H82 and H146) cells in vitro. To determine which FGF was the most clinically relevant, we also examined FGF2 and FGF9 concentrations in the serum of patients with lung cancer. We found that extracellular FGFs can affect proliferation, treatment sensitivity, and apoptosis of lung cancer cells in a cell-specific manner. Our results indicate that extracellular FGFs affect the biology of lung cancer cells through multiple functions.

It has been thought that corrective posterior surgery for adolescent idiopathic scoliosis (AIS) should be started on the concave side because initial convex manipulation would increase the risk of vertebral malrotation, worsening the rib hump. With the many new materials, implants, and manipulation techniques (e.g., direct vertebral rotation) now available, we hypothesized that manipulating the convex side first is no longer taboo. Our technique has two major facets. (1) Curve correction is started from the convex side with a derotation maneuver and in situ bending followed by concave rod application. (2) A 6.35 mm diameter pure titanium rod is used on the convex side and a 6.35 mm diameter titanium alloy rod on the concave side. Altogether, 52 patients were divided into two groups. Group N included 40 patients (3 male, 37 female; average age 15.9 years) of Lenke type 1 (23 patients), type 2 (2), type 3 (3), type 5 (10), type 6 (2). They were treated with a new technique using 6.35 mm diameter different-stiffness titanium rods. Group C included 12 patients (all female, average age 18.8 years) of Lenke type 1 (6 patients), type 2 (3), type 3 (1), type 5 (1), type 6 (1). They were treated with conventional methods using 5.5 mm diameter titanium alloy rods. Radiographic parameters (Cobb angle/thoracic kyphosis/correction rates) and perioperative data were retrospectively collected and analyzed. Preoperative main Cobb angles (groups N/C) were 56.8°/60.0°, which had improved to 15.2°/17.1° at the latest follow-up. Thoracic kyphosis increased from 16.8° to 21.3° in group N and from 16.0° to 23.4° in group C. Correction rates were 73.2% in group N and 71.7% in group C. There were no significant differences for either parameter. Mean operating time, however, was significantly shorter in group N (364 min) than in group C (456 min). We developed a new corrective surgical technique for AIS using a 6.35 mm diameter pure titanium rod initially on the convex side. Correction rates in the coronal, sagittal, and axial planes were the same as those achieved with conventional methods, but the operation time was significantly shorter.

Aldehyde dehydrogenases (ALDHs) play a major role in detoxification of aldehydes. High expression of ALDHs is a marker for stem cells of many organs including the lungs. A common polymorphism in ALDH2 gene (ALDH2*2) results in inactivation of the enzyme and is associated with alcohol flushing syndrome and increased risk for cardiovascular and Alzheimer's diseases and some cancers. The effect of this ALDH2 polymorphism on the lung and its stem cells has not been thoroughly examined.We examined the association between the ALDH2*2 allele and lung function parameters in a population of healthy individuals. We also examined its association with the incidence of asthma and COPD in patient cohorts. We used the in vitro colony forming assay to detect the effect of the polymorphism on lung epithelial stem cells from both primary human surgical samples and Aldh2*2 transgenic (Tg) and Aldh2 -/- mice. Response to acute and chronic lung injuries was compared between wild type (WT), Aldh2*2 Tg and Aldh2 -/- mice.In humans, the ALDH2*2 allele was associated with lower FEV1/FVC in the general population, but not with the development of asthma or COPD. Both the bronchial and lung epithelium carrying the ALDH2*2 allele showed a tendency for lower colony forming efficiency (CFE) compared to ALDH2 allele. In mice, the tracheal epithelial thickness, nuclear density, and number of basal stem cells were significantly lower in Aldh2 -/- and Aldh2*2 Tg adult mice than in WT. Electron microscopy showed significantly increased number of morphologically abnormal mitochondria in the trachea of Aldh2 -/- mice. Aldh2 -/- tracheal and lung cells showed higher ROS levels and fewer functional mitochondria than those from WT mice. No significant differences were detected when tracheal and lung epithelial stem cells were examined for their in vitro CFE. When exposed to chronic cigarette smoke, Aldh2*2 Tg mice were resistant to emphysema development, whereas influenza infection caused more epithelial damage in Aldh2 -/- mice than in WT mice.ALDH2 polymorphism has several subtle effects on the lungs, some of which are similar to changes observed during normal aging, suggesting a "premature lung aging" effect.

Background: Both advanced cancer patients and their family caregivers experience distress and have a range of concerns after cancer diagnosis. However, longitudinal studies on this topic have been lacking. Aim: To investigate concerns in both patients with advanced lung cancer and their family caregivers longitudinally from diagnosis. Design: A multi-center prospective questionnaire-based study. Setting/participants: We recruited patients with newly diagnosed advanced lung cancer and their family caregivers at 16 hospitals in Japan. We prospectively assessed the prevalence of their concerns using the Concerns Checklist and investigated the associations between their concerns and mental status as well as quality of life until 24 months after diagnosis. Results: A total of 248 patients and their 232 family caregivers were enrolled. The prevalence of serious concerns was highest at diagnosis (patients: 68.3%, family caregivers: 65.3%). The most common serious concern was concern about the future in both groups at diagnosis (38.2% and 40.5%, respectively) and this remained high in prevalence over time, while the high prevalence of concern about lack of information improved 3 months after diagnosis in both groups. Approximately one-third of patient-family caregiver dyads had discrepant reports of serious concerns. The presence of serious concerns was significantly associated with anxiety and depression continuously in both groups. Conclusions: The majority of advanced lung cancer patients and their family caregivers have serious concerns from diagnosis, which is associated with their psychological distress. The spectrum of concerns alters over the disease trajectory, warranting efficient tailored care and support for both groups immediately after diagnosis.

As a powerful bone-inducing cytokine, rhBMP-2 has been used as a bone graft substitute in combination with animal-derived collagen to achieve interbody or posterolateral spinal fusion. Successful interspinous process fusion using rhBMP-2 in combination with synthetic carrier materials would offer a safe, minimally invasive spinal fusion option for the treatment of spinal disorders. The aims of the present study were to achieve interspinous process fusion by implanting rhBMP-2-retaining degradable material instead of bone grafting and to evaluate efficacy for vertebral stabilization.A polymer gel (200 mg), β-tricalcium phosphate powder (400 mg), and rhBMP-2 (0, 30, 60 or 120 μg) were mixed to generate a plastic implant, which was then placed during surgery to bridge the L5-6 interspinous processes of 58 rabbits. Control animals received implants either without rhBMP-2 or with autogenous bone chips from the iliac crest. L5-6 vertebrae were recovered 8 weeks postoperatively. Interspinous process fusion was evaluated by radiography, biomechanical bending test, intradiscal pressure (IDP) measurement, and histology.In bending tests, strength of fusion was significantly greater in BMP60 and BMP120 groups than in sham, BMP0, BMP30 or autogenous bone groups. IDP at L5-6 was significantly reduced in BMP60 and BMP120 groups compared to sham, BMP0, BMP30, and autograft groups. Histologically, coronal sections of the fusion mass showed a bone mass bridging both spinous processes.Solid interspinous process fusion was achieved in rabbit models by 8 weeks after implanting the biodegradable bone-inducing material. These results suggest a potential new less-invasive option without bone grafting for the treatment of lumbar disorders.

This study was designed to identify epigenetically regulated genes and to clarify the contribution of epige-netic alteration to acquired resistance to epidermal growth factor receptor‑tyrosine kinase inhibitors (EGFR‑TKIs). We established a gefitinib‑resistant lung cancer cell line, PC9, which was originally gefitinib‑sensitive, by serial long‑term exposure to gefitinib. RNA and DNA were collected from both gefitinib‑sensitive and ‑resistant PC9 cells, and comprehensive DNA methylation and mRNA expression analyses were performed using Infinium HumanMethylation27 Bead Arrays and Agilent SurePrint G3 Human Gene Expression 8x60K Array, respectively. DNA methylation was increased in 640 genes in gefitinib‑resistant cells compared to parental cells. Among them, we selected 29 candidate genes that presented a decrease in mRNA expression in resistant PC9. We further studied four of the selected genes (C10orf116, IGFBP3, KL, and S100P) and found that KL or S100P silencing by siRNA induced a decrease in gefitinib sensitivity compared to that in the negative control in PC9. In conclusion, KL and S100P could be potential targets to overcome resistance to EGFR‑TKIs.

Epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (EGFR-TKIs) are widely used for the treatment of non-small cell lung cancers (NSCLCs) harboring EGFR-activating mutations. However, lung cancer cells inevitably acquire resistance to these EGFR-TKIs. The majority of patients whose lung cancer acquires resistance to EGFR-TKIs are subjected to treatment using cytotoxic agents. The present study aimed to determine if lung cancer cells acquiring resistance to EGFR-TKIs also develop altered sensitivity to cytotoxic agents. It was revealed that lung cancer cells that had developed resistance to EGFR-TKIs had increased sensitivity to gemcitabine and vinorelbine compared with EGFR-TKI naïve cells. The expression levels of ATP-binding cassette (ABC) transporter genes, including ABCC3, ABCC5 and ABCG2, were observed to be commonly repressed in EGFR-TKI-resistant cells. In addition, two cases were identified in which gemcitabine and vinorelbine exerted marked responses to lung cancers that had acquired resistance to EGFR-TKIs, even with late-line treatment. Therefore, it was proposed that gemcitabine and vinorelbine may be effective agents for patients with lung cancer previously treated with EGFR-TKIs.

Epigenetic gene regulation plays essential roles in differentiation of embryonic and tissue stem cells. In these benign undifferentiated cells, some polycomb targeted genes are kept in a state of DNA hypomethylation and they have a distinct chromatin signature termed bivalent chromatin structure to maintain their plasticity. We hypothesized that cancer stem cells (CSC), the malignant counterpart of these cells, are also under the control of epigenetics like benign stem cells. We compared the DNA methylation and chromatin structure in 10 tumor suppressor genes between CSC and differentiated cancer cells of MCF7 and Huh7 cells. We found that the level of DNA methylation was indeed significantly lower in CSC, while surprisingly, the bivalent chromatin structure was more ubiquitously seen in differentiated cancer cells compared to CSC. However, repressive marks of chromatin structure, namely H3K27me3 and EZH2, were significantly lower in CSC. As a consequence, CSC remained in a higher transcriptionally active chromatin state compared to differentiated cancer cells. We found that the differentiation of CSCs is also epigenetically regulated. These findings could help towards a comprehensive understanding of CSC, and also improve the development of eradicative therapies against human malignancies.

Study Design. A propensity score matching study. Objective. The aim of this study was to assess the cost-effectiveness of balloon kyphoplasty (BKP) in Japan. Summary of Background Data. Osteoporotic vertebral fracture (OVF) is a common disease in elderly people. In Japan, the incidence of painful OVF in 2008 was estimated as 880,000, and approximately 40% of patients with painful OVF are hospitalized due to the severity of pain. Japan is the front runner among super-aged societies and rising health care costs are an economic problem. Methods. BKP and nonsurgical management (NSM) for acute/subacute OVF were performed in 116 and 420 cases, respectively. Quality-adjusted life years (QALY) and incremental costs were calculated on the basis of a propensity score matching study. QALY was evaluated using the SF-6D questionnaire. Finally, using a Markov model, incremental cost-effectiveness ratios (ICERs) were calculated for 71 matched cases. Results. In the comparison between BKP and NSM, mean patients age was 78.3 and 77.7 years, respectively ( P = 0.456). The BKP procedure cost 402,988 JPY more than NSM and the gains in QALY at the 6-month follow-up were 0.153 and 0.120, respectively (difference = 0.033). ICERs for 3 and 20 years were 4,404,158 JPY and 2,416,406 JPY, respectively. According to sensitivity analysis, ICERs ranged from 652,181 JPY to 4,896,645 JPY (4418–33,168 GBP). Conclusion. This study demonstrated that BKP is a cost-effective treatment option for OVF in Japan. However, the effect might be blunted in patients aged &gt; 80 years. Further research is necessary to elucidate the cost-effectiveness of BKP in this population. Level of Evidence: 4

Abstract Background Immune checkpoint inhibitors (ICIs) have demonstrated efficacy over previous cytotoxic chemotherapies in clinical trials among various tumors. Despite their favorable outcomes, they are associated with a unique set of toxicities termed as immune-related adverse events (irAEs). Among the toxicities, ICI-related pneumonitis has poor outcomes with little understanding of its risk factors. This retrospective study aimed to investigate whether pre-existing interstitial lung abnormality (ILA) is a potential risk factor for ICI-related pneumonitis. Materials and Methods Patients with non-small cell lung cancer, malignant melanoma, renal cell carcinoma, and gastric cancer, who was administered either nivolumab, pembrolizumab, or atezolizumab between September 2014 and January 2019 were retrospectively reviewed. Information on baseline characteristics, computed tomography findings before administration of ICIs, clinical outcomes, and irAEs were collected from their medical records. Pre-existing ILA was categorized based on previous studies. Results Two-hundred-nine patients with a median age of 68 years were included and 23 (11.0%) developed ICI-related pneumonitis. While smoking history and ICI agents were associated with ICI-related pneumonitis (P = .005 and .044, respectively), the categories of ILA were not associated with ICI-related pneumonitis (P = .428). None of the features of lung abnormalities were also associated with ICI-related pneumonitis. Multivariate logistic analysis indicated that smoking history was the only significant predictor of ICI-related pneumonitis (P = .028). Conclusion This retrospective study did not demonstrate statistically significant association between pre-existing ILA and ICI-related pneumonitis, nor an association between radiologic features of ILA and ICI-related pneumonitis. Smoking history was independently associated with ICI-related pneumonitis. Further research is warranted for further understanding of the risk factors of ICI-related pneumonitis.

The aim of this study was to assess the efficacy and safety of erlotinib, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), as second‑ or third‑line treatment for elderly Japanese patients with non‑small‑cell lung cancer (NSCLC). The patients eligible for this phase II trial were aged ≥70 years, had stage III/IV or recurrent NSCLC, and had previously received 1 or 2 chemotherapy regimens that did not include EGFR‑TKIs. The patients received erlotinib at a dose of 150 mg/day. The primary endpoint was overall response rate (ORR), and the secondary endpoints were progression‑free survival (PFS), overall survival (OS) and toxicity. A total of 38 patients with a median age of 76 years were enrolled. The majority of the patients were men (66%), had an Eastern Cooperative Oncology Group performance status of 1 (58%), stage IV disease (66%) and adenocarcinoma (74%). Of the 35 patients, 13 (34%) had tumors with EGFR mutations. The ORR was 26.3% (95% confidence interval: 12.1‑40.5%) and the disease control rate was 47.4%. The median PFS was 3.7 months and the median OS was 17.3 months. The grade 3 adverse events observed included rash (13%), diarrhea (5%), interstitial pneumonitis (5%), anorexia (3%) and gastrointestinal bleeding (3%). Grade 4 or 5 adverse events were not observed. The median OS did not differ significantly between patients aged <75 years (14.9 months) and those aged ≥75 years (19.0 months; P=0.226). Therefore, erlotinib was found to be effective and well‑tolerated in elderly patients with previously treated NSCLC.

A 69-year-old man with post-operative recurrence of lung adenocarcinoma was treated with multiple chemotherapies, including epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors. A second biopsy revealed an EGFR T790M mutation. As 10th-line chemotherapy, osimertinib was initiated. After 24 weeks, chest computed tomography (CT) revealed asymptomatic ground-glass opacities in both lobes. After four weeks of osimertinib discontinuation, imaging revealed rapid lung cancer progression. Osimertinib was resumed. After 11 weeks, CT revealed decreased lung nodules with no exacerbation of interstitial lung disease. We describe a patient who experienced transient asymptomatic pulmonary opacities during treatment with osimertinib, which was successfully managed by a "stop-and-go" approach.

Objective:Targeted therapy-induced facial skin toxicities may reduce overall quality of life (QoL) in cancer patients. We investigated whether facial skin toxicities affect QoL and attempted to identify factors related to QoL in patients with advanced/recurrent cancer.Methods:We performed a cross-sectional study in 34 outpatients with advanced/recurrent cancer showing targeted therapy-induced facial skin toxicities in Japan between November 2016 and February 2017. For measurement, we used the Kessler Psychological Distress Scale (K6), Mental Adjustment to Cancer (MAC) Scale, and Dermatology Life Quality Index (DLQI). Data were analyzed using Spearman's rank correlation coefficient.Results:Mean DLQI score in 34 patients was 4.59 (standard deviation ± 4.70), which was interpreted as a small effect on a patient's life. Acneiform rash was the most common skin condition noted, followed by xerosis, pruritus, and erythema. Analysis of DLQI scores revealed that symptoms and feelings was the domain most commonly affected among different domains constituting the DLQI. MAC analysis revealed that the fighting spirit score was the highest among MAC scales. We found that age, K6, and fatalism construct in MAC were significantly correlated with total DLQI scores (age: Spearman's ρ= −0.48, P = 0.004; K6: ρ= 0.58, P < 0.001; fatalism; ρ= −0.39, P = 0.025).Conclusions:This is the first study investigating targeted therapy-induced facial skin toxicities in cancer patients. Our results suggest potential negative effects of facial skin toxicities on overall QoL in patients with advanced/recurrent cancer in middle and early old age. Targeted therapy-induced facial skin toxicities may reduce overall quality of life (QoL) in cancer patients. We investigated whether facial skin toxicities affect QoL and attempted to identify factors related to QoL in patients with advanced/recurrent cancer. We performed a cross-sectional study in 34 outpatients with advanced/recurrent cancer showing targeted therapy-induced facial skin toxicities in Japan between November 2016 and February 2017. For measurement, we used the Kessler Psychological Distress Scale (K6), Mental Adjustment to Cancer (MAC) Scale, and Dermatology Life Quality Index (DLQI). Data were analyzed using Spearman's rank correlation coefficient. Mean DLQI score in 34 patients was 4.59 (standard deviation ± 4.70), which was interpreted as a small effect on a patient's life. Acneiform rash was the most common skin condition noted, followed by xerosis, pruritus, and erythema. Analysis of DLQI scores revealed that symptoms and feelings was the domain most commonly affected among different domains constituting the DLQI. MAC analysis revealed that the fighting spirit score was the highest among MAC scales. We found that age, K6, and fatalism construct in MAC were significantly correlated with total DLQI scores (age: Spearman's ρ= −0.48, P = 0.004; K6: ρ= 0.58, P < 0.001; fatalism; ρ= −0.39, P = 0.025). This is the first study investigating targeted therapy-induced facial skin toxicities in cancer patients. Our results suggest potential negative effects of facial skin toxicities on overall QoL in patients with advanced/recurrent cancer in middle and early old age.