Search

Immune checkpoint inhibitors (ICIs) have contributed significantly to the treatment of various types of cancer, including skin cancer. However, not all patients respond; some patients do not respond at all (primary resistance), while others experience recurrence after the initial response (acquired resistance). Therefore, overcoming ICI resistance is an urgent priority. Numerous ICI resistance mechanisms have been reported. They are seemingly quite complex, varying from patient to patient. However, most involve T‐cell activation processes, especially in the tumor microenvironment (TME). ICIs exert their effects in the TME by reactivating suppressed T cells through inhibition of immune checkpoint molecules, such as cytotoxic T‐lymphocyte antigen‐4 (CTLA‐4) and programmed cell death protein 1 (PD‐1). Thus, this review focuses on the resistance mechanisms based on the T‐cell activation process. Here, we classify the main mechanisms of ICI resistance into three categories based on (1) antigen recognition, (2) T‐cell migration and infiltration, and (3) effector functions of T cells. By identifying and understanding these resistance mechanisms individually, including unknown mechanisms, we seek to contribute to the development of novel treatments to overcome ICI resistance. [ABSTRACT FROM AUTHOR]

In T-cell biology, 'exhaustion' was initially described as a hyporesponsive state in CD8+ T cells during chronic infections. Recently, exhaustion has been recognized as a T-cell dysfunctional state in the tumor microenvironment (TME). The term 'exhaustion' is used mainly to refer to effector T cells with a reduced capacity to secrete cytokines and an increased expression of inhibitory receptors. The up-regulation of exhaustion-related inhibitory receptors, including programmed cell death protein 1 (PD-1), in such T cells has been associated with the development of tumors, prompting the development of immune checkpoint inhibitors. In addition to CD8+ T cells, CD4+ T cells, including the regulatory T (Treg) cell subset, perform a wide variety of functions within the adaptive immune system. Up-regulation of the same inhibitory receptors that are associated with CD8+ T-cell exhaustion has also been identified in CD4+ T cells in chronic infections and cancers, suggesting a similar CD4+ T-cell exhaustion phenotype. For instance, high expression of PD-1 has been observed in Treg cells in the TME, and such Treg cells can play an important role in the resistance to PD-1 blockade therapies. Furthermore, recent progress in single-cell RNA sequencing has shown that CD4+ T cells with cytotoxic activity are also vulnerable to exhaustion. In this review, we will discuss novel insights into various exhausted T-cell subsets, which could reveal novel therapeutic targets and strategies to induce a robust anti-tumor immune response. [ABSTRACT FROM AUTHOR]

Aging leads to numerous changes that affect many components of the immune system, called "immunosenescence". Indeed, elderly individuals exhibit dysregulated immune responses against pathogens, poor responses to vaccination, and increased susceptibility to many diseases including cancer, autoimmune disorders, and other chronic inflammatory diseases. Despite progressed understanding of immunosenescence, its detailed mechanisms are still not fully understood. With advances in medicine, the population of older cancer patients is expected to rapidly increase in the coming years. Cancer immunotherapies, including immune checkpoint inhibitors (ICIs), have been shown to be effective for multiple cancer types, whereas to date, few specific data for elderly individuals have been published. Some systemic reviews have demonstrated that ICIs exhibit similar efficacy in older cancer patients, but they seem to be less effective in very old patients. In addition, toxicities might be more frequently observed in such patients. Here, we provide a summary to better understand immunosenescence and an overview of its relationship with cancer and antitumor immunity, including the efficacy and toxicity of ICIs. [ABSTRACT FROM AUTHOR]

Single-cell level analysis is powerful tool to assess the heterogeneity of cellular components in tumor microenvironments (TME). In this study, we investigated immune-profiles using the single-cell analyses of endoscopically- or surgically-resected tumors, and peripheral blood mononuclear cells from gastric cancer patients. Furthermore, we technically characterized two distinct platforms of the single-cell analysis; RNA-seq-based analysis (scRNA-seq), and mass cytometry-based analysis (CyTOF), both of which are broadly embraced technologies. Our study revealed that the scRNA-seq analysis could cover a broader range of immune cells of TME in the biopsy-resected small samples of tumors, detecting even small subgroups of B cells or Treg cells in the tumors, although CyTOF could distinguish the specific populations in more depth. These findings demonstrate that scRNA-seq analysis is a highly-feasible platform for elucidating the complexity of TME in small biopsy tumors, which would provide a novel strategies to overcome a therapeutic difficulties against cancer heterogeneity in TME. [ABSTRACT FROM AUTHOR]

Biliary tract cancer (BTC) is an aggressive cancer with a poor prognosis partially due to the limited success in developing novel therapies, including molecularly targeted therapies and immunotherapies. Programmed cell death-1 (PD-1) blockade therapy is less effective against BTCs, necessitating further studies to understand the detailed immunological status of the tumor microenvironment (TME) in BTC. Here, we examined the immunological status of the TME in 37 BTCs with early- to late-stage disease, especially focusing on PD-1+CD8+ T cells. PD-1+CD8+ T cells, which are reportedly associated with the clinical response to PD-1 blockade therapy, were frequently observed in early-stage BTC and decreased with disease progression. Imaging mass cytometry for representative PD-1+CD8+TIL-high and -low patients demonstrated that tumor-infiltrating PD-1+CD8+ T cells were localized adjacent to tumor cells, whereas PD-1−CD8+ T cells were detected mainly in the stroma of the TME. In a mouse model, PD-1 expression by tumor-infiltrating CD8+ T cells was higher in smaller tumors and decreased with tumor growth. Consequently, large tumors became resistant to PD-1 blockade, while small tumors containing higher numbers of PD-1+CD8+ T cells were sensitive. We propose the important role of tumor-infiltrating PD-1+CD8+ T cells in anti-tumor immunity and the potential application of PD-1 blockade therapy for early-stage BTC. [ABSTRACT FROM AUTHOR]

Precursor exhausted T cells can differentiate into terminally exhausted cells, and the author outlines memory T-cell differentiation for comparison. Coexpression of PD-1, 2B4, CD160 and KLRG1 on exhausted HCV-specific CD8 SP + sp T cells is linked to antigen recognition and T cell differentiation. Within tumors, T cell (and NK cell) numbers and functions are enhanced, and intratumoral T cells have shown reduced markers of cellular exhaustion. Although T-cell exhaustion was first described in CD8 SP + sp T cells during chronic viral infections, it has become clear that the phenomenon is also a crucial feature of various T-cell subsets in the tumor microenvironment (TME). [Extracted from the article]

Background: Although colorectal mucinous adenocarcinomas (MCs) are conventionally regarded as exhibiting high-grade differentiation, they can be divided by differentiation into 2 groups according to the glandular appearance: low-grade mucinous adenocarcinoma (low-MC) and high-grade mucinous adenocarcinoma (high-MC). Methods: Patients with colorectal cancer (CRC) who underwent surgical resection between 2000 and 2012 were enrolled in this study. Among the cases with MC, the clinicopathological and genetic differences between low-MC and high-MC were investigated with next-generation sequencing. Results: A total of 1373 patients with CRC were analyzed. Forty patients (2.9%) had MC, and 13 patients had high-MC. Patients with MC had significantly shorter disease-free survival (DFS) and overall survival (OS) periods than those with nonmucinous carcinoma. When low-MC patients and high-MC patients were compared, those with high-MC had significantly shorter DFS and OS periods than those with low-MC. Multivariate analyses revealed that high-MC was significantly associated with both shorter DFS and shorter OS, but low-MC was not. A genome analysis revealed that low-MC had a considerably larger number of mutations than high-MC, and Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations and adenomatous polyposis coli mutations were particularly frequently found in low-MC. In contrast, SMAD family member 4 (SMAD4) mutations were frequently found in high-MC. Conclusions: High-MC is an independent prognostic factor in CRC (but low-MC is not), and it is genetically different from other CRCs, including low-MC. Both the clinicopathological differences and the genetic differences suggest that low-MC and high-MC should be distinguished in clinical settings. [ABSTRACT FROM AUTHOR]

Background MET splice site mutations resulting in an exon 14 deletion have been reported to be present in about 3% of all lung adenocarcinomas. Patients with lung adenocarcinoma and a MET splice site mutation who have responded to MET inhibitors have been reported. The CRISPR/Cas9 system is a recently developed genome-engineering tool that can easily and rapidly cause small insertions or deletions. Materials and methods We created an in vitro model for MET exon 14 deletion using the CRISPR/Cas9 system and the HEK293 cell line. The phenotype, which included MET inhibitor sensitivity, was then investigated in vitro . Additionally, MET splice site mutations were analyzed in several cancers included in The Cancer Genome Atlas (TCGA) dataset. Results An HEK293 cell line with a MET exon 14 deletion was easily and rapidly created; this cell line had a higher MET protein expression level, enhanced MET phosphorylation, and prolonged MET activation. In addition, a direct comparison of phenotypes using this system demonstrated enhanced cellular growth, colony formation, and MET inhibitor sensitivity. In the TCGA dataset, lung adenocarcinomas had the highest incidence of MET exon 14 deletions, while other cancers rarely carried such mutations. Approximately 10% of the lung adenocarcinoma samples without any of driver gene alterations carried the MET exon 14 deletion. Conclusions These findings suggested that this system may be useful for experiments requiring the creation of specific mutations, and the present experimental findings encourage the development of MET-targeted therapy against lung cancer carrying the MET exon 14 deletion. [ABSTRACT FROM AUTHOR]

Background Some of patients with non-small cell lung cancer (NSCLC) harboring somatic activating mutations of the epidermal growth factor receptor gene ( EGFR mutations) show poor responses to EGFR-tyrosine kinase inhibitors (EGFR-TKIs) treatment. Cigarette smoking is the strongest documented risk factor for the development of lung cancer. Nicotine, while not carcinogenic by itself, has been shown to induce proliferation, angiogenesis, and the epithelial-mesenchymal transition; these effects might be associated with EGFR-TKI resistance. Materials and methods PC-9 and 11_18 cell lines ( EGFR -mutated NSCLC cell lines) were cultured with 1 μM nicotine for 3 months and were designated as PC-9/N and 11_18/N cell lines, respectively. The sensitivities of these cell lines to EGFR-TKI were then tested in vitro . Moreover, the association between the smoking status and the progression-free survival (PFS) period was investigated in patients with EGFR -mutated NSCLC who were treated with gefitinib. Results The PC-9/N and 11_18/N cell lines were resistant to EGFR-TKI, compared with controls. The phosphorylation of EGFR in these cell lines was reduced by EGFR-TKI to a smaller extent than that observed in controls, and a higher concentration of EGFR-TKI was capable of further decreasing the phosphorylation. Clinically, smoking history was an independent predictor of a poor PFS period on gefitinib treatment. Conclusions Chronic nicotine exposure because of cigarette smoking mediates resistance to EGFR-TKI via an EGFR signal. Smoking cessation is of great importance, while resistance may be overcome through the administration of high-dose EGFR-TKI. [ABSTRACT FROM AUTHOR]

We previously reported that activin produces a signal with a tumor suppressive role in pancreatic cancer (PC). Here, the association between plasma activin A and survival in patients with advanced PC was investigated. Contrary to our expectations, however, patients with high plasma activin A levels had a significantly shorter survival period than those with low levels (median survival, 314 days vs. 482 days, P  = 0.034). The cellular growth of the MIA PaCa-2 cell line was greatly enhanced by activin A via non-SMAD pathways. The cellular growth and colony formation of an INHBA (beta subunit of inhibin)-overexpressed cell line were also enhanced. In a xenograft study, INHBA- overexpressed cells tended to result in a larger tumor volume, compared with a control. The bodyweights of mice inoculated with INHBA -overexpressed cells decreased dramatically, and these mice all died at an early stage, suggesting the occurrence of activin-induced cachexia. Our findings indicated that the activin signal can promote cancer progression in a subset of PC and might be involved in cachexia. The activin signal might be a novel target for the treatment of PC. [ABSTRACT FROM AUTHOR]

Background Transforming growth factor, beta (TGFB) signal is considered to be a tumor suppressive pathway based on the frequent genomic deletion of the SMAD4 gene in pancreatic cancer (PC); however; the role of the activin signal, which also belongs to the TGFB superfamily, remains largely unclear. Methods and results We found a homozygous deletion of the activin A receptor, type IB (ACVR1B) gene in 2 out of 8 PC cell lines using array-comparative genomic hybridization, and the absence of ACVR1B mRNA and protein expression was confirmed in these 2 cell lines. Activin A stimulation inhibited cellular growth and increased the phosphorylation level of SMAD2 and the expression level of p21CIP1/WAF1 in the Sui66 cell line (wild-type ACVR1B and SMAD4 genes) but not in the Sui68 cell line (homozygous deletion of ACVR1B gene). Stable ACVR1B-knockdown using short hairpin RNA cancelled the effects of activin A on the cellular growth of the PC cell lines. In addition, ACVR1B-knockdown significantly enhanced the cellular growth and colony formation abilities, compared with controls. In a xenograft study, ACVR1B-knockdown resulted in a significantly elevated level of tumorigenesis and a larger tumor volume, compared with the control. Furthermore, in clinical samples, 6 of the 29 PC samples (20.7%) carried a deletion of the ACVR1B gene, while 10 of the 29 samples (34.5%) carried a deletion of the SMAD4 gene. Of note, 5 of the 6 samples with a deletion of the ACVR1B gene also had a deletion of the SMAD4 gene. Conclusion We identified a homozygous deletion of the ACVR1B gene in PC cell lines and clinical samples and proposed that the deletion of the ACVR1B gene may mediate an aggressive cancer phenotype in PC. Our findings provide novel insight into the role of the activin signal in PC. [ABSTRACT FROM AUTHOR]

Introduction: This study aimed to evaluate the association between BCL2 single-nucleotide polymorphisms and survival outcome in advanced non-small cell lung cancer (NSCLC). Methods: One hundred and sixty-eight patients with advanced NSCLC who were treated with anti-cancer drugs and could be evaluated for therapeutic response between April 2005 and March 2010 at Kyoto University Hospital were enrolled. DNA was extracted from peripheral blood samples. The BCL2 polymorphisms -938 C→A (rs2279115) and +21 A→G (rs1801018) were genotyped using the 5′-nuclease assay. The univariate relationship between each independent clinicopathologic variable and BCL2 genotype was examined using Fisher's exact test. To evaluate risk factors associated with prognosis, a Cox proportional hazards regression model with a step-down procedure was used. Results: The median survival time of patients with the -938 AA and AC genotypes were significantly shorter than those with the -938 CC genotype (p = 0.027 by log-rank test). Based on multivariate analysis, poor performance status [hazard ratio (HR) 2.424, 95% confidence interval (CI) 1.727-3.262; p < 0.0001], non-adenocarcinoma histology (HR 1.512, 95% CI 1.167-1.938; p = 0.0048) and the BCL2 -938 AA + AC genotype (HR 1.219, 95% CI, 1.024-1.456; p = 0.0256) were significant independent prognostic factors for survival. Conclusions: Polymorphisms in BCL2 may be associated with survival in advanced-stage NSCLC patients who received chemotherapy. Copyright © 2013 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Purpose: Several cases have been reported in which central nervous system (CNS) metastases of non-small cell lung cancer (NSCLC) resistant to gefitinib were improved by erlotinib. However, there has been no study in which cerebrospinal fluid (CSF) concentrations of gefitinib and erlotinib are directly compared. Thus, we aimed to compare them. Methods: We examined 15 Japanese patients with NSCLC and CNS metastases with epidermal growth factor receptor gene mutations who received CSF examinations during epidermal growth factor receptor-tyrosine kinase inhibitors treatment (250 mg daily gefitinib or 150 mg daily erlotinib). Plasma and CSF concentrations were determined using high-performance liquid chromatography with tandem mass spectrometry. Results: The concentration and penetration rate of gefitinib (mean ± standard deviation) in the CSF were 3.7 ± 1.9 ng/mL (8.2 ± 4.3 nM) and 1.13 ± 0.36 %, respectively. The concentration and penetration rate of erlotinib in the CSF were 28.7 ± 16.8 ng/mL (66.9 ± 39.0 nM) and 2.77 ± 0.45 %, respectively. The CSF concentration and penetration rate of erlotinib were significantly higher than those of gefitinib ( P = 0.0008 and <0.0001, respectively). The CNS response rates of patients with erlotinib treatment were preferentially (but not significantly) higher than those with gefitinib treatment. (1/3 vs. 4/7, respectively). Leptomeningeal metastases in one patient, which were refractory to gefitinib, dramatically responded to erlotinib. Conclusions: This study suggested that higher CSF concentration could be achieved with erlotinib and that erlotinib could be more effective for the treatment for CNS metastases, especially leptomeningeal metastases, than gefitinib. [ABSTRACT FROM AUTHOR]

Abstract: The most serious adverse reaction associated with treatment with epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) is drug-induced interstitial lung disease (ILD). Because EGFR-TKIs are key drugs for patients with non-small cell lung cancer who have somatic activating mutations of the epidermal growth factor receptor gene (EGFR mutations), several cases of retreatment with EGFR-TKIs after ILD induced by these drugs have been reported. Here, we present a 68-year-old man with lung adenocarcinoma and leptomeningeal metastases having an EGFR mutation who was retreated with erlotinib after erlotinib-induced ILD. He suffered no ILD recurrence and his leptomeningeal metastases dramatically improved. In addition to the present case, reports of nine patients who were retreated with EGFR-TKIs after ILD were found in the literature. Only one patient had recurrence of ILD (although seven were retreated at a reduced dose of EGFR-TKIs, including the patient with recurrence). In contrast, three patients had no recurrence of ILD even without dose-reduction. These reports suggest that dose-reduction plays a limited role in preventing recurrence. Many patients received corticosteroids during retreatment, but not the one with recurrence of ILD. This may suggest that corticosteroids can prevent recurrence due to their antiinflammatory properties. [Copyright &y& Elsevier]

Abstract: Purpose: The maximum tolerated dose (MTD) of erlotinib (150mg) is the approved daily dose. In contrast, the approved daily dose of gefitinib (250mg) is only one-third of its MTD. Significantly different adverse events have been associated with gefitinib and erlotinib. Experimental design: A retrospective investigation examining the adverse events and tolerances of 250mg daily gefitinib and 150mg daily erlotinib in Japanese patients with non-small cell lung cancer (NSCLC) was performed. Adverse events were assessed according to Common Terminology Criteria for Adverse Events version 3.0. To determine tolerance for each agent, failure was defined as dose reduction or discontinuation of the drug due to adverse events, and early failure as dose reduction or discontinuation due to adverse events before the first evaluation of response. Results: More adverse events including skin disorders, diarrhea, oral mucositis, asthenic conditions, anorexia, nausea, vomiting, and gastrointestinal bleeding were observed in the erlotinib group. Liver function test abnormalities and pneumonitis did not differ between the two groups. Based on multivariate analysis, failure, early failure, and discontinuation due to adverse events were independently associated with erlotinib use. Conclusion: Our data show that 150mg daily erlotinib was associated with more toxicity and less tolerability than 250mg daily gefitinib. [Copyright &y& Elsevier]

Purpose: Recent reports indicate that refractory central nervous system (CNS) metastases of non-small cell lung cancer (NSCLC) are improved by high-dose gefitinib or erlotinib administration. We describe a Japanese woman with NSCLC and CNS metastases who was resistant to 75 mg daily erlotinib, but the metastases were improved by 150 mg daily erlotinib. We investigated the plasma and CSF concentrations of erlotinib at each dose as well as the correlation between the plasma and CSF concentrations of erlotinib. Methods: Including this patient, we administered 150 mg erlotinib daily to nine NSCLC patients with CNS metastases and measured the plasma and CSF concentrations just before administration on day 8. The concentrations were determined using high-performance liquid chromatography with ultraviolet detection. Results: The plasma and CSF concentrations of erlotinib at a dose of 75 mg were 433 and 14 nM, respectively. The plasma and CSF concentrations of erlotinib at a dose of 150 mg were increased to 1,117 and 44 nM, respectively. The mean ± standard deviation of CSF concentrations and penetration rates were 106 ± 59 nM and 4.5 ± 1.5%, respectively. There was a good correlation ( R = 0.84) between plasma and CSF concentrations ( P = 0.0005). Conclusions: This study indicates that CSF concentrations of erlotinib depend on its plasma concentration. As seen in this patient, high CSF concentrations of erlotinib can be achieved by high-dose administration, and this finding suggests the efficacy of high-dose administration, especially to refractory CNS metastases of NSCLC patients. [ABSTRACT FROM AUTHOR]

BACKGROUND: Although the association of multiple pulmonary metastases, and particularly miliary metastases, with response to gefitinib treatment in patients with nonsmall cell lung cancer has been reported, the association of miliary pulmonary metastases with epidermal growth factor receptor gene (EGFR) mutations remains unclear. METHODS: The authors retrospectively investigated the association of diffuse, random pulmonary metastases in patients with lung adenocarcinoma. The study included 163 Japanese patients who had unresectable, advanced lung adenocarcinoma diagnosed between April 2003 and March 2010. Computed tomography scans that were obtained at the time of diagnosis were analyzed by 2 investigators. For the purposes of this study, diffuse, random pulmonary metastases were defined as multiple nodules (n = 50; ≤3 cm in greatest dimension) distributed diffusely and randomly throughout the lungs. RESULTS: Of 163 patients, 55 had pulmonary metastases, and EGFR mutations were detected in 22 of those 55 patients. The mutations were identified preferentially among women (P = .15) and were identified significantly among patients who had a smoking history of <10 pack-years (P = .0057). Diffuse, random pulmonary metastases were identified in 11 of 22 patients who had EGFR mutations and in 4 of 33 patients who had the wildtype EGFR (P = .0043). On the basis of multivariate analyses, EGFR mutations were associated independently with a smoking history of <10 pack-years (P = .026) and with diffuse, random pulmonary metastases (P = .012). CONCLUSIONS: When patients with lung adenocarcinomas who had EGFR mutations developed pulmonary metastases, they tended to be diffuse and random, including military metastases. However, such metastases were much less common in patients who had lung adenocarcinomas with wild-type EGFR. Cancer 2011;117:819-25. [ABSTRACT FROM AUTHOR]

Internal jugular vein thrombosis is much less common than deep venous thrombosis of lower limbs and is generally caused by an indwelling venous catheter or otological infection. Several cases of internal jugular vein thrombosis associated with malignancy have been also reported. Bevacizumab, a monoclonal neutralizing antibody against vascular endothelial growth factor, has shown benefits in the treatment of many types of malignancy and its use is increasing. Serious adverse effects, however, are associated with the use of bevacizumab, including venous thromboembolism. In this article, we present a rare case of non-small cell lung cancer complicated by pulmonary embolism due to internal jugular vein thrombosis associated with bevacizumab. [ABSTRACT FROM AUTHOR]

Only a small percentage of patients afflicted with gastric cancer (GC) respond to immune checkpoint blockade (ICB). To study the mechanisms underlying this resistance, we examined the immune landscape of GC. A subset of these tumors was characterized by high frequencies of regulatory T (Treg) cells and low numbers of effector T cells. Genomic analyses revealed that these tumors bore mutations in RHOA that are known to drive tumor progression. RHOA mutations in cancer cells activated the PI3K-AKT-mTOR signaling pathway, increasing production of free fatty acids that are more effectively consumed by Treg cells than effector T cells. RHOA mutant tumors were resistant to PD-1 blockade but responded to combination of PD-1 blockade with inhibitors of the PI3K pathway or therapies targeting Treg cells. We propose that the metabolic advantage conferred by RHOA mutations enables Treg cell accumulation within GC tumors, generating an immunosuppressive TME that underlies resistance to ICB. • RHOA Y42-mutated GC harbors abundant Treg cell and low CD8+ T cell infiltration in TME • RHOA Y42 mutations reduce CXCL10 and CXCL11 production via activated PI3K-AKT pathways • PI3K pathways activated by RHOA Y42 mutations provide a metabolic advantage for Treg cells • A PI3K inhibitor overcomes resistance to PD-1 blockade in RHOA Y42-mutated tumors Kumagai et al. find that a subset of gastric cancer (GC) tumors are characterized by high frequencies of regulatory T (Treg) cells and low numbers of effector T cells; these tumors bear mutations in RHOA. RHOA mutations activated the PI3K-AKT-mTOR pathway, increasing production of free fatty acids that are more effectively consumed by Treg cells than effector T cells. [ABSTRACT FROM AUTHOR]

During the replication of viral genomes, RNA viruses produce double‐stranded RNA (dsRNA), through the activity of their RNA‐dependent RNA polymerases (RdRps) as viral replication intermediates. Recognition of viral dsRNA by host pattern recognition receptors – such as retinoic acid‐induced gene‐I (RIG‐I)‐like receptors and Toll‐like receptor 3 – triggers the production of interferon (IFN)‐β via the activation of IFN regulatory factor (IRF)‐3. It has been proposed that, during the replication of viral genomes, each of RIG‐I and melanoma differentiation‐associated gene 5 (MDA5) form homodimers for the efficient activation of a downstream signalling pathway in host cells. We previously reported that, in the non‐neoplastic human hepatocyte line PH5CH8, the RdRp NS5B derived from hepatitis C virus (HCV) could induce IFN‐β expression by its RdRp activity without the actual replication of viral genomes. However, the exact mechanism by which HCV NS5B produced IFN‐β remained unknown. In the present study, we first showed that NS5B derived from another Flaviviridae family member, GB virus B (GBV‐B), also possessed the ability to induce IFN‐β in PH5CH8 cells. Similarly, HCV NS5B, but not its G317V mutant, which lacks RdRp activity, induced the dimerization of MDA5 and subsequently the activation of IRF‐3. Interestingly, immunofluorescence analysis showed that HCV NS5B produced dsRNA. Like HCV NS5B, GBV‐B NS5B also triggered the production of dsRNA and subsequently the dimerization of MDA5. Taken together, our results show that HCV NS5B triggers an MDA5‐mediated innate immune response by producing dsRNA without the replication of viral genomes in human hepatocytes. [ABSTRACT FROM AUTHOR]

Background: Immune checkpoint inhibitors may enhance the efficacy of radiotherapy (RT) in cancer treatment but the effect remains unknown in metastatic gastric cancer (mGC). This study aimed to compare the tumor shrinkage by palliative RT for mGC patients with or without previous exposure to anti-PD-1 therapy. Methods: Data of 36 mGC patients who had received palliative RT from April 2013 to May 2019 were analyzed. Primary tumor responses were evaluated through a volumetric measurement-based method using computed tomography (CT) and endoscopic responses were evaluated in patients who underwent endoscopy before and after RT. Tumor microenvironment (TME) immune status was investigated by analyzing tumor-infiltrating lymphocytes by flow cytometry. Results: Among 36 patients, 18 had previous exposure to anti-PD-1 before RT showing no significant differences in baseline characteristics with the other 18 patients without exposure to anti-PD-1 treatment. Tumor responses were observed in 28% (5/18) and none (0/18) in the anti-PD-1-exposed vs. naïve group, respectively (P = 0.045). Five out of eight patients in the anti-PD-1-exposed group, who underwent endoscopy after RT showed partial response, but none in the anti-PD-1-naïve patients showed response (P = 0.026). Increase in the CD8+ T cell/effector regulatory T cell ratio in TILs after anti-PD-1 therapy was noted in three responders to RT, but not in the other three non-responders. Conclusions: Prior exposure to anti-PD-1 therapy increases tumor response to RT. Immune profiling suggests that anti-PD-1 therapy may enhance the efficacy of RT by immunoactivation in the TME. [ABSTRACT FROM AUTHOR]

Dear Editor, Congenital melanocytic nevi (CMN) is known to develop into melanoma at various frequencies. The risk of melanoma development is low for a small single CMN (<1.5 cm projected adult size), whereas large CMN (>40 cm projected adult size) accompanied by multiple CMN have a lifetime melanoma incidence risk of 10 %-5 % [[1]]. Copy number gains of GNAS and MYC were commonly detected in the CMN and metastatic melanoma, and those of BCL11B were commonly detected in the CMN and primary melanoma (Fig. [Extracted from the article]

Background: Identifying biomarkers to predict immune checkpoint inhibitor (ICI) efficacy is warranted. Considering that somatic mutation-derived neoantigens induce strong immune responses, patients with a high tumour mutational burden reportedly tend to respond to ICIs. However, there are several conflicting data. Therefore, we focused on the original function of neoantigenic mutations and their impact on the tumour microenvironment (TME). Methods: We evaluated 88 high-frequency microsatellite instability (MSI-H) colorectal cancers and analysed the function of the identified neoantigenic mutations and their influence on programmed cell death 1 (PD-1) blockade efficacy. The results were validated using The Cancer Genome Atlas (TCGA) datasets. Results: We identified frameshift mutations in RNF43 as a common neoantigenic gene mutation in MSI-H tumours. However, loss-of-function RNF43 mutations induced noninflamed TME by activating the WNT/β-catenin signalling pathway. In addition, loss of RNF43 function induced resistance to PD-1 blockade even in neoantigen-rich tumours. TCGA dataset analyses demonstrated that passenger rather than driver gene mutations were related to the inflamed TME in diverse cancer types. Conclusions: We propose a novel concept of "paradoxical neoantigenic mutations" that can induce noninflamed TME through their original gene functions, despite deriving neoantigens, suggesting the significance of qualities as well as quantities in neoantigenic mutations. [ABSTRACT FROM AUTHOR]

Comprehensive genomic profiling for non–small cell lung cancer (NSCLC) is likely to identify more patients with rare genetic alterations including uncommon epidermal growth factor receptor gene ( EGFR ) mutations. It remains unclear how such patients should be treated, however. We here report a case of NSCLC positive for two uncommon mutations of EGFR and a KRAS mutation, including its treatment with the second-generation EGFR tyrosine kinase inhibitor (TKI) afatinib. Tumor specimen obtained by a NSCLC patient with no smoking history was analyzed by next-generation sequencing. Comprehensive genomic profiling revealed that the patient harbored the EGFR mutations G719C and S768I as well as the E49K mutation of KRAS . Treatment with afatinib was clinically effective as confirmed by PET-CT scans of bone metastases and by a marked decrease in the serum concentration of carcinoembryonic antigen. Afatinib was the most effective among seven EGFR-TKIs tested in inhibiting the growth of Ba/F3 cells expressing EGFR(S768I), showing an efficacy similar to that apparent with cells expressing the common EGFR mutant L858R, whereas first- and third-generation EGFR-TKIs were markedly less effective against EGFR(S768I) than against EGFR(L858R). These data suggest that EGFR-TKIs differ in their activity toward cells expressing EGFR(S768I) in vitro. Consistently, afatinib was clinically effective for the treatment of NSCLC harboring G719C and S768I mutations of EGFR . Further studies are warranted to determine the most appropriate EGFR-TKI for treatment of NSCLC harboring uncommon EGFR mutations. [ABSTRACT FROM AUTHOR]

Although transcriptome alteration is an essential driver of carcinogenesis, the effects of chromosomal structural alterations on the cancer transcriptome are not yet fully understood. Short-read transcript sequencing has prevented researchers from directly exploring full-length transcripts, forcing them to focus on individual splice sites. Here, we develop a pipeline for Multi-Sample long-read Transcriptome Assembly (MuSTA), which enables construction of a transcriptome from long-read sequence data. Using the constructed transcriptome as a reference, we analyze RNA extracted from 22 clinical breast cancer specimens. We identify a comprehensive set of subtype-specific and differentially used isoforms, which extended our knowledge of isoform regulation to unannotated isoforms including a short form TNS3. We also find that the exon–intron structure of fusion transcripts depends on their genomic context, and we identify double-hop fusion transcripts that are transcribed from complex structural rearrangements. For example, a double-hop fusion results in aberrant expression of an endogenous retroviral gene, ERVFRD-1, which is normally expressed exclusively in placenta and is thought to protect fetus from maternal rejection; expression is elevated in several TCGA samples with ERVFRD-1 fusions. Our analyses provide direct evidence that full-length transcript sequencing of clinical samples can add to our understanding of cancer biology and genomics in general. Namba et al develop a new pipeline called MuSTA to enable the efficient assembly of transcriptome from long-read sequencing data of breast cancer samples. This method enables the authors to discover subtype-specific isoforms, find that fusion transcript structures depend on their genomic context and identify a double-hop fusion that results in aberrant expression of an endogenous retroviral gene. [ABSTRACT FROM AUTHOR]

Background: Erlotinib shows large inter-patient pharmacokinetic variability, but the impact of early drug exposure and genetic variations on the clinical outcomes of erlotinib remains fully investigated. The primary objective of this study was to clarify the population pharmacokinetics/pharmacodynamics of erlotinib in Japanese patients with non-small cell lung cancer (NSCLC). The secondary objective was to identify genetic determinant(s) for the cerebrospinal fluid (CSF) permeability of erlotinib and its active metabolite OSI-420. Methods: A total of 88 patients treated with erlotinib (150 mg/day) were enrolled, and CSF samples were available from 23 of these patients with leptomeningeal metastases. Plasma and CSF concentrations of erlotinib and OSI-420 were measured by high-performance liquid chromatography with UV detection. Population pharmacokinetic analysis was performed with the nonlinear mixed-effects modelling program NONMEM. Germline mutations including ABCB1 (1236C>T, 2677G>T/A, 3435C>T), ABCG2 (421C>A), and CYP3A5 (6986A>G) polymorphisms, as well as somatic EGFR activating mutations if available, were examined. Early exposure to erlotinib and its safety/efficacy relationship were evaluated. Results: The apparent clearance of erlotinib and OSI-420 were significantly decreased by 24 and 35 % in patients with the ABCG2 421A allele, respectively ( p < 0.001), while ABCB1 and CYP3A5 polymorphisms did not affect their apparent clearance. The ABCG2 421A allele was significantly associated with increased CSF penetration for both erlotinib and OSI-420 ( p < 0.05). Furthermore, the incidence of grade ≥2 diarrhea was significantly higher in patients harboring this mutant allele ( p = 0.035). A multivariate logistic regression model showed that erlotinib trough (C) levels on day 8 were an independent risk factor for the development of grade ≥2 diarrhea ( p = 0.037) and skin rash ( p = 0.031). Interstitial lung disease (ILD)-like events occurred in 3 patients (3.4 %), and the median value of erlotinib C levels adjacent to these events was approximately 3 times higher than that in patients who did not develop ILD (3253 versus 1107 ng/mL; p = 0.014). The objective response rate in the EGFR wild-type group was marginally higher in patients achieving higher erlotinib C levels (≥1711 ng/mL) than that in patients having lower erlotinib C levels (38 versus 5 %; p = 0.058), whereas no greater response was observed in the higher group (67 %) versus the lower group (77 %) within EGFR mutation-positive patients ( p = 0.62). Conclusions: ABCG2 can influence the apparent clearance of erlotinib and OSI-420, and their CSF permeabilities in patients with NSCLC. Our preliminary findings indicate that early exposure to erlotinib may be associated with the development of adverse events and that increased erlotinib exposure may be relevant to the antitumor effects in EGFR wild-type patients while having less of an impact on the tumor response in EGFR mutation-positive patients. [ABSTRACT FROM AUTHOR]

Background: Subgroup analyses of randomized studies have consistently shown that pemetrexed is exclusively effective in non-small-cell lung cancer (NSCLC) other than squamous cell carcinoma and the combination of pemetrexed and platinum agents is recommended for first-line chemotherapy in advanced non-squamous NSCLC; however, there have been few prospective studies of a selected population. Patients and methods: This was a single-arm phase II study of carboplatin and pemetrexed in Japanese patients with chemo-naive advanced non-squamous NSCLC. Patients received six cycles of pemetrexed (500 mg/m) combined with carboplatin (area under the curve: AUC 6) every 3 weeks. Maintenance chemotherapy with pemetrexed was permitted in patients whose disease did not progress after combination chemotherapy. The primary endpoint was the response rate, and secondary endpoints were safety and survival. Results: Fifty-one patients were enrolled between November 2009 and March 2011, and 49 patients were evaluable for both safety and efficacy. All but one patient had adenocarcinoma histology. Forty-four (90 %) patients completed four cycles, and 33 (67 %) completed six cycles of chemotherapy. Partial response was achieved in 25 patients (response rate: 51 %) and stable disease in 18 patients (37 %). Median progression-free survival (PFS) and overall survival (OS) were 6.3 months and 24.3 months, respectively. The median PFS and OS were 7.9 months and 24.3 months in patients with epidermal growth factor receptor (EGFR) mutation, and 6.3 months and 21.0 months in patients with EGFR wild type or unknown. There were no statistical differences between EGFR mutants and non-mutants for both PFS ( p = 0.09) and OS ( p = 0.23). Grade 3/4 neutropenia and thrombocytopenia were observed in 16 (33 %) and 9 (18 %) patients, respectively. Non-hematologic toxicities were generally mild, and there were no treatment-related deaths. Conclusions: The combination of carboplatin and pemetrexed was safe and effective in advanced non-squamous NSCLC. Although the sample size was small, our results indicate that pemetrexed is a key drug for advanced non-squamous NSCLC, irrespective of the EGFR mutation status (UMIN-CTR number 000002451). [ABSTRACT FROM AUTHOR]

Purpose: We examined patients with advanced nonsquamous, non-small cell lung cancer (NSCLC) to evaluate epidermal growth factor receptor (EGFR) mutation status and serum C-reactive protein (CRP) for their associations with response to gefitinib therapy and for prognostic impacts. Methods: Serum levels of CRP from 79 Japanese patients with advanced nonsquamous NSCLC were measured before the start of gefitinib. We used the peptic nucleic acid-locked nucleic acid clamp method to determine their EGFR somatic mutation status. We evaluated the relationship between each independent clinicopathological variable and the response to gefitinib therapy and the risk factors associated with prognosis. Results: Having CRP-positive serum and having wild-type EGFR were both independent negative predictive factors for the response to gefitinib treatment by multivariate logistic regression model analysis. Having CRP-positive serum and having wild-type EGFR were significant independent negative prognostic factors for survival based on multivariate analysis. Conclusions: Having CRP-positive serum predicted a lack of response to gefitinib therapy independent of EGFR mutational status. Both CRP-positive serum and wild-type EGFR were independent poor prognostic factors in patients with nonsquamous NSCLC who received gefitinib therapy. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

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 FROM AUTHOR]

• Not smoking history but smoking signature affects FOXP3 + T cell elevation. • Smoking signature can be a biomarker for immune microenvironment. • Smoking signature also associates with CD20 + B cell elevation. Mutational signatures associated with tobacco smoking (mutational smoking signatures: SS) are characterized mainly by C > A mutations. The aim of this study was to characterize the association between the tumor immune microenvironment and the SS in lung adenocarcinoma. Lung adenocarcinomas surgically resected from 96 patients, for which whole exome sequencing data was available, were included in the study. We extracted the SS from whole exome sequencing data, calculated the weights of SS using deconstructSigs, and compared the clinicopathological features of SS positive (SS+) and negative (SS-) adenocarcinomas. We selected 18 tumor pairs from SS + and SS- adenocarcinomas (sex, EGFR mutation, and tumor size-matched) and examined the expression of five immune markers (CD20, CD8, FOXP3, CD204, and PD-L1) by immunohistochemistry. Of 96 specimens, there were 33 (34 %) SS + adenocarcinoma tumors. The smoking index significantly correlated with the weight of the SS (R = 0.43). Between SS + and SS- tumors, there was no significant difference in clinicopathological factors excluding smoking history. Immunohistochemistry revealed that the number of FOXP3 + T cells in SS + adenocarcinomas was significantly higher than that in the SS- adenocarcinomas (median number 58 vs. 36, p < 0.01). Also, the number of CD20 + B cells in SS + adenocarcinomas was significantly higher than that in the SS- adenocarcinomas (median number 77 vs. 29, p < 0.01); however; these phenomena could not be confirmed when stratified by smoking history. In lung adenocarcinoma, SS is associated with an immunosuppressive tumor microenvironment. [ABSTRACT FROM AUTHOR]

Background: The standard treatment for patients with unresectable locally advanced esophageal squamous cell carcinoma (ESCC) is definitive chemoradiotherapy (CRT) using 5-FU plus cisplatin. However, complete response (CR) rates are low at 11-25%, resulting in 9-10 months of median overall survival (OS). An improved therapeutic efficacy by combining immunotherapy with radiation has been reported in patients with locally advanced non-small cell lung cancer. The results using ESCC cell lines suggest sequential treatment with anti-PD-L1 agents soon after completion of CRT is the most effective combination.Methods: TENERGY trial is a multicenter, phase II, proof-of-concept study to assess the efficacy and safety of atezolizumab following definitive CRT in patients with locally advanced ESCC. The main inclusion criteria are unresectable locally advanced ESCC without distant metastasis, completion of 60 Gy of radiation plus two concomitant cycles of chemotherapy (cisplatin 70 mg/m2 on day 1 and 5-FU 700 mg/m2 on days 1-4, every 28 days), and adequate organ function. Within 6 weeks after CRT, participants will start taking 1200 mg of atezolizumab every three weeks and continue until 12 months or disease progression. The primary endpoint is the confirmed CR rate by the investigator's assessment. Secondary endpoints include overall response rate, progression-free survival (PFS), OS, adverse events, and confirmed CR rate by central assessment. We will enroll 50 patients (40 with primary locally advanced ESCC and 10 with postoperative locoregionally recurrent ESCC). We will obtain biopsies from the primary site and will collect blood at 3 time points (before CRT, after CRT, and four weeks after the start of atezolizumab) for an exploratory biomarker study. We will analyze the phenotype of immune-competent cells, neoantigens, tumor mutational burden, PD-L1 status, and Human Leukocyte Antigen haplotyping.Discussion: The synergistic efficacies of the sequential combination of CRT and atezolizumab should improve the CR rate, resulting in survival improvement for patients with unresectable locally advanced ESCC. Because CRT is a standard treatment option for patients with early stage to locally advanced ESCC, the sequential combination of CRT and atezolizumab has the potential to change the standard ESCC treatments.Trial Registration: UMIN000034373, 10/04/2018 and EPOC1802. [ABSTRACT FROM AUTHOR]

Background: Recently, the U.S. Food and Drug Administration approved pembrolizumab for patients (pts) with PD-L1-positive metastatic gastric cancer (MGC) based on 22C3 immunohistochemistry (IHC) assay. However, little is known about detailed clinicopathological features of 22C3 PD-L1 expression in MGC.Patients and methods: Pts with histologically confirmed MGC were eligible for this prospective observational study. PD-L1 expression (22C3) on tumor cell (TC) or immune cell (IC) and mismatch repair (MMR) were analyzed by IHC. Epstein-Barr virus (EBV) was detected by in situ hybridization. The expressions of tyrosine kinase receptors (RTKs) and cancer genome alterations were evaluated by IHC or next-generation sequencing.Results: A total of 225 pts were analyzed in this study. PD-L1 expression on TC, PD-L1 on IC, MMR-deficient (D-MMR), and EBV positivity were identified in 8.4, 65.3, 6.2, and 6.2% cases, respectively. PD-L1 expression in TC was more frequently observed in pts with D-MMR (P < 0.001), PIK3CA mutation (P = 0.020), and KRAS mutation (P = 0.002), and PD-L1 on IC was associated with EBV positivity (P = 0.034), and lymph-node metastasis (P < 0.001). PD-L1 expression on either IC or TC was less frequently observed in pts with peritoneal metastasis and Borrmann Type 4. A significant association was not observed between PD-L1 expression and RTKs expression or presence of other gene alterations. PD-L1 expression on either TC or IC was not prognostic factor.Conclusions: 22C3 PD-L1 expression in MGC was associated with distinct clinicopathological features, but was not a prognostic factor. [ABSTRACT FROM AUTHOR]

Background: Non-small cell lung cancer (NSCLC) harboring common epidermal growth factor receptor (EGFR) gene mutations (exon 19 deletion or exon 21 L858R) respond to EGFR tyrosine kinase inhibitors (EGFR-TKIs). The secondary T790 M mutation in exon 20 of the EGFR gene is the most common type of acquired resistance mutation. Several reports have also shown that other secondary mutations (L747S, D761Y and T854A), while uncommon, can induce acquired resistance to first-generation EGFR-TKIs. However, little is known about the anticancer activities of second- or third-generation EGFR-TKIs. Methods: Uncommon secondary mutations were introduced into Ba/F3 cells along with the sensitive EGFR L858R mutation (Ba/F3-L858R/L747S, Ba/F3-L858R/D761Y, and Ba/F3-L858R/T854A), and the sensitivities to various EGFR-TKIs were then investigated. Results: Both the Ba/F3-L858R/L747S and Ba/F3-L858R/D761Y cell lines exhibited weak resistances to first-generation reversible EGFR-TKIs, while the Ba/F3-L858R/T854A cell line exhibited a strong resistance. In contrast, irreversible EGFR-TKIs, especially third-generation EGFR-TKIs, were capable of overcoming these resistances. Western blot analyses demonstrated that gefitinib (first-generation) inhibited the phosphorylation of EGFR to a lesser extent in cells with these secondary mutations than in cells with the sensitive L858R mutation alone. In contrast, afatinib and osimertinib (second- and thirdgeneration) inhibited the phosphorylation of EGFR in cells with these secondary mutations to a similar extent as that seen in cells with the sensitive L858R mutation alone. Conclusions: Our experimental findings suggest that irreversible EGFR-TKIs, especially third-generation EGFR-TKIs, can be effective against uncommon secondary mutations and that switching to third-generation EGFR-TKIs could be a promising treatment strategy for patients with acquired resistance because of these uncommon secondary mutations. [ABSTRACT FROM AUTHOR]

Somatic mutations in KRAS, NRAS, and BRAF genes are related to resistance to anti-EGFR antibodies in colorectal cancer. We have established an extended RAS and BRAF mutation assay using a next-generation sequencer to analyze these mutations. Multiplexed deep sequencing was performed to detect somatic mutations within KRAS, NRAS, and BRAF, including minor mutated components. We first validated the technical performance of the multiplexed deep sequencing using 10 normal DNA and 20 formalin-fixed, paraffin-embedded (FFPE) tumor samples. To demonstrate the potential clinical utility of our assay, we profiled 100 FFPE tumor samples and 15 plasma samples obtained from colorectal cancer patients. We used a variant calling approach based on a Poisson distribution. The distribution of the mutation-positive population was hypothesized to follow a Poisson distribution, and a mutation-positive status was defined as a value greater than the significance level of the error rate (α = 2 x 10-5). The cut-off value was determined to be the average error rate plus 7 standard deviations. Mutation analysis of 100 clinical FFPE tumor specimens was performed without any invalid cases. Mutations were detected at a frequency of 59% (59/100). KRAS mutation concordance between this assay and Scorpion-ARMS was 92% (92/100). DNA obtained from 15 plasma samples was also analyzed. KRAS and BRAF mutations were identified in both the plasma and tissue samples of 6 patients. The genetic screening assay using next-generation sequencer was validated for the detection of clinically relevant RAS and BRAF mutations using FFPE and liquid samples. [ABSTRACT FROM AUTHOR]

Intra- and inter-tumor heterogeneity may hinder personalized molecular-target treatment that depends on the somatic mutation profiles. We performed mutation profiling of formalin-fixed paraffin embedded tumors of multi-regional colon cancer and characterized the consequences of intra- and inter-tumor heterogeneity and metastasis using targeted re-sequencing. We performed targeted re-sequencing on multiple spatially separated samples obtained from multi-regional primary colon carcinoma and associated metastatic sites in two patients using next-generation sequencing. In Patient 1 with four primary tumors (P1-1, P1-2, P1-3, and P1-4) and one liver metastasis (H1), mutually exclusive pattern of mutations was observed in four primary tumors. Mutations in primary tumors were identified in three regions; KARS ( G13D ) and APC ( R876* ) in P1-2, TP53 ( A161S ) in P1-3, and KRAS ( G12D ), PIK3CA ( Q546R ), and ERBB4 ( T272A ) in P1-4. Similar combinatorial mutations were observed between P1-4 and H1. The ERBB4 ( T272A ) mutation observed in P1-4, however, disappeared in H1. In Patient 2 with two primary tumors (P2-1 and P2-2) and one liver metastasis (H2), mutually exclusive pattern of mutations were observed in two primary tumors. We identified mutations; KRAS ( G12V ), SMAD4 ( N129K , R445* , and G508D ), TP53 ( R175H ), and FGFR3 ( R805W ) in P2-1, and NRAS ( Q61K ) and FBXW7 ( R425C ) in P2-2. Similar combinatorial mutations were observed between P2-1 and H2. The SMAD4 ( N129K and G508D ) mutations observed in P2-1, however, were nor detected in H2. These results suggested that different clones existed in primary tumors and metastatic tumor in Patient 1 and 2 likely originated from P1-4 and P2-1, respectively. In conclusion, we detected the muti-clonalities between intra- and inter-tumors based on mutational profiling in multi-regional colon cancer using next-generation sequencing. Primary region from which metastasis originated could be speculated by mutation profile. Characterization of inter- and inter-tumor heterogeneity can lead to underestimation of the tumor genomics landscape and treatment strategy of personal medicine. [ABSTRACT FROM AUTHOR]

The balance of programmed death-1 (PD-1)-expressing CD8+ T cells and regulatory T (Treg) cells in the tumor microenvironment (TME) determines the clinical efficacy of PD-1 blockade therapy through the competition of their reactivation. However, factors that determine this balance remain unknown. Here, we show that Treg cells gain higher PD-1 expression than effector T cells in highly glycolytic tumors, including MYC-amplified tumors and liver tumors. Under low-glucose environments via glucose consumption by tumor cells, Treg cells actively absorbed lactic acid (LA) through monocarboxylate transporter 1 (MCT1), promoting NFAT1 translocation into the nucleus, thereby enhancing the expression of PD-1, whereas PD-1 expression by effector T cells was dampened. PD-1 blockade invigorated the PD-1-expressing Treg cells, resulting in treatment failure. We propose that LA in the highly glycolytic TME is an active checkpoint for the function of Treg cells in the TME via upregulation of PD-1 expression. [Display omitted] • LA induces PD-1 expression by Treg cells in highly glycolytic tumors • LA absorbed through MCT1 is a metabolic checkpoint of immune responses • MYC -amplified or liver metastatic tumors augment PD-1+ Treg cells with abundant LA • MCT1 highly expressed by Treg cells provides therapeutic target for immunotherapy Kumagai et al. show that Treg cells uptake lactic acid in the highly glycolytic tumor microenvironment via MCT1 and robustly express PD-1, resulting in the impairment of PD-1 blockade therapy. [ABSTRACT FROM AUTHOR]

We evaluated the potential of a linear β-1,3-glucan (curdlan) as a starting material to access C6-modified glucose derivatives and found that 6-bromo-6-deoxyglucose, 6-azide-6-deoxyglucose, and 6-acetamido-6-deoxyglucose could be readily prepared from curdlan through its C6-selective and quantitative modifications and subsequent acid-catalyzed hydrolysis. [ABSTRACT FROM AUTHOR]

The tissue distribution and function of hemoglobin or myoglobin are well known; however, a newly found cytoglobin (CYGB), which also belongs to the globin family, remains to be characterized. To assess its expression in human malignancies, we sought to screen a number of cell lines originated from many tissues using northern blotting and real time PCR techniques. Unexpectedly, we found that several, but not all, melanoma cell lines expressed CYGB mRNA and protein at much higher levels than cells of other origins. Melanocytes, the primary origin of melanoma, also expressed CYGB at a high level. To verify these observations, immunostaining and immunoblotting using anti-CYGB antibody were also performed. Bisulfite-modified genomic sequencing revealed that several melanoma cell lines that abrogated CYGB expression were found to be epigenetically regulated by hypermethylation in the promoter region of CYGB gene. The RNA interference-mediated knockdown of the CYGB transcript in CYGB expression-positive melanoma cell lines resulted in increased proliferation in vitro and in vivo. Flow cytometric analysis using 2′-, 7′-dichlorofluorescein diacetate (DCFH-DA), an indicator of reactive oxygen species (ROS), revealed that the cellular ROS level may be involved in the proliferative effect of CYGB. Thus, CYGB appears to play a tumor suppressive role as a ROS regulator, and its epigenetic silencing, as observed in CYGB expression-negative melanoma cell lines, might function as an alternative pathway in the melanocyte-to-melanoma transition. [ABSTRACT FROM AUTHOR]

Abstract: The glycopolymer composed of an inulin scaffold and pendent β-lactosides was developed from commercially available inulin through sequential chemical modification processes composed of tosylation, azidation, and the subsequent Huisgen cyclocoupling with an alkyne-terminated β-lactoside. The resultant inulin-based glycopolymer has unique dual affinity towards β-galactoside and α-glucoside specific lectins which is attributable to its pendent β-lactosides and terminal α-glucoside. Its gellation property was also accessed to find that the inulin-based glycopolymer forms hydrogels whose critical gellation concentration (CGC) was lower than that required for hydrogels made from native inulin. Drug release properties of the inulin-based glycopolymer were also discussed in this paper. [Copyright &y& Elsevier]

Abstract: 2,2-Bipyridines having β-lactoside, β-d-glucoside, β-d-galactoside, and N-acetyl-β-d-glucosaminide were prepared and then, complexed with ferrous ion to afford trivalent glycoclusters having tris-bipyridine ferrous complex cores. Each glycocluster provides a dynamic combinatorial library composed of four diastereomeric stereoisomers (Δmer, Δfac, Λmer, and Λfac) whose ratios depend on their relative stabilities. CD spectral analyses of these glycoclusters showed that various cations (Na+, Mg2+, K+ or Ca2+) enriched Δ-forms of the glycocluster having β-lactosides and N-acetyl-β-d-glucosaminides possibly by cations-induced intramolecular carbohydrate–carbohydrate interactions. [Copyright &y& Elsevier]

Purpose: The objectives of this phase I trial were to evaluate the toxicity of the nedaplatin/gemcitabine regimen, determine the maximum tolerated doses (MTDs) of these agents, and observe the anti-tumor effects of this regimen on advanced squamous cell lung cancer. Methods: Patients with previously untreated advanced squamous cell lung cancer were eligible if they had a performance status of 0 or 1 with adequate organ function. The doses of gemcitabine (days 1 and 8) and nedaplatin (day 8) studied were 800/70, 1,000/80, 1,000/90, and 1,000/100 (mg/m), repeated every 3 weeks. Results: Toxicity and response could be assessed in all 13 patients enrolled. The patients included 12 men and one woman with a median age of 69 years (range 57-81 years). Three patients had stage IIIB disease and 10 patients had stage IV disease. The MTDs were reached at 1,000 mg/m gemcitabine and 80 mg/m nedaplatin. The most frequent toxic effects were thrombocytopenia and neutropenia; grade 3 or 4 thrombocytopenia was observed in 23% of patients, and grade 3 or 4 neutropenia was seen in 46% of patients. Non-hematologic toxicities were mild. Grade 3 fatigue, nausea/vomiting, and appetite loss occurred in two patients. The overall response rate was 62%. Conclusions: We recommend doses of 800 mg/m gemcitabine and 70 mg/m nedaplatin for phase II study. This combination chemotherapeutic regimen is active and well tolerated. [ABSTRACT FROM AUTHOR]

Background Although several studies suggested that immune checkpoint inhibitors may enhance an efficacy of radiotherapy (RT), it remains unclear in patients (pts) with advanced gastric cancer (AGC). Methods Total 34 AGC pts treated with RT for the primary gastric lesion from April 2013 to March 2019 were analyzed in this study. Pts were divided into two groups according to whether they had received nivolumab before RT or not. Primary tumor responses were assessed according to volumetry on CT scan with tumor response as > 70% decrease in tumor volume. Endoscopic responses were evaluated in pts who underwent endoscopy after RT. Tumor infiltrating lymphocytes (TILs) were also analyzed to examine the tumor microenvironmental immune status in the pts. Results Among 34 pts, 16 pts had received nivolumab before RT. There were no significant differences in baseline characteristics between nivolumab-exposed and nivolumab-naive groups. 70% tumor reductions by RT were observed in 25% (4/16) in nivolumab-exposed group but no patients (0/18) in nivolumab-naive group (P = 0.039). Among pts who received endoscopy after RT, four of seven pts in nivolumab-exposed group experienced partial response, while no response was observed among 10 nivolumab-naive pts (P = 0.015). TILs were evaluated in pts whose tumor samples were available. CD8+ T cells/effector regulatory T cells (eTreg cells) ratio in TILs was markedly increased after nivolumab in pts who achieved response, but not observed in pts who had no tumor reduction. A significant increase of CD8+ T cells/eTreg cells ratio after nivolumab treatment was observed in nivolumab-exposed pts (n = 15), whereas such a tendency was not observed in nivolumab-naive pts (n = 10). Conclusions RT to primary gastric cancer significantly led to tumor response in nivolumab-exposed pts compared with nivolumab-naive pts. Immune profiling suggested that nivolumab might enhance the sensitivity to RT by immunoactivation via increasing CD8+ T cell/eTreg cell ratio. [ABSTRACT FROM AUTHOR]

Although several studies suggested that immune checkpoint inhibitors may enhance an efficacy of radiotherapy (RT), it remains unclear in patients (pts) with advanced gastric cancer (AGC). Total 34 AGC pts treated with RT for the primary gastric lesion from April 2013 to March 2019 were analyzed in this study. Pts were divided into two groups according to whether they had received nivolumab before RT or not. Primary tumor responses were assessed according to volumetry on CT scan with tumor response as > 70% decrease in tumor volume. Endoscopic responses were evaluated in pts who underwent endoscopy after RT. Tumor infiltrating lymphocytes (TILs) were also analyzed to examine the tumor microenvironmental immune status in the pts. Among 34 pts, 16 pts had received nivolumab before RT. There were no significant differences in baseline characteristics between nivolumab-exposed and nivolumab-naive groups. 70% tumor reductions by RT were observed in 25% (4/16) in nivolumab-exposed group but no patients (0/18) in nivolumab-naive group (P = 0.039). Among pts who received endoscopy after RT, four of seven pts in nivolumab-exposed group experienced partial response, while no response was observed among 10 nivolumab-naive pts (P = 0.015). TILs were evaluated in pts whose tumor samples were available. CD8+ T cells/effector regulatory T cells (eTreg cells) ratio in TILs was markedly increased after nivolumab in pts who achieved response, but not observed in pts who had no tumor reduction. A significant increase of CD8+ T cells/eTreg cells ratio after nivolumab treatment was observed in nivolumab-exposed pts (n = 15), whereas such a tendency was not observed in nivolumab-naive pts (n = 10). RT to primary gastric cancer significantly led to tumor response in nivolumab-exposed pts compared with nivolumab-naive pts. Immune profiling suggested that nivolumab might enhance the sensitivity to RT by immunoactivation via increasing CD8+ T cell/eTreg cell ratio. [ABSTRACT FROM AUTHOR]

Background: Immune checkpoint inhibitors (ICIs) confer a survival benefit in many cancer types. Given that the survival outcome for cancer of unknown primary site (CUP) remains poor, we investigated the potential of CUP for immunotherapy. Methods: A total of 164 patients with CUP (favorable subset, 34 patients; unfavorable subset, 130 patients) who were treated between January 2009 and March 2017 was identified from a review of medical records at Kindai University Hospital. They included 92 patients for whom pretreatment tumor tissue was available both for determination of programmed cell death–ligand 1 expression and tumor-infiltrating lymphocyte (TIL) density by immunohistochemistry (IHC) and for immune-related gene expression profiling (irGEP). The results of irGEP for CUP were compared with published data for ICI-treated solid cancers classified into progressive disease (PD) and non-PD subsets according to their best response to ICIs. Results: The median overall survival of all CUP patients was 29.3 months (95% confidence interval [CI], 15.7–not reached) and 7.1 months (95% CI, 5.0–9.4) for favorable and unfavorable subsets, respectively. IHC and irGEP revealed that pretreatment immune activity—including expression of immune checkpoint molecules—for CUP was similar to that for ICI-responsive malignancies (antitumor immune cell signatures: CUP versus PD, P = 0.002–0.067; CUP versus non-PD, P = 0.591–0.999), although VEGFA expression was associated with suppression of antitumor immunity in CUP (P = 0.008, false discovery rate = 0.010). In addition, one case of CUP in the unfavorable subset that was associated with prominent PD-L1 expression on TILs and showed a durable response to nivolumab is presented. Conclusions: The survival outcome of CUP remains unsatisfactory. However, our clinical and immune profiling of CUP has revealed a potential to benefit from immunotherapy, with ICIs thus being a potential option for CUP treatment. [ABSTRACT FROM AUTHOR]