American Association for Cancer Research (AACR)Acta Medica Okayama2767-9764622026Clinical Characteristics and Spatial Transcriptome Analysis of Non–Small Cell Lung Cancers Exhibiting Early Alectinib Resistance: A Retrospective OLCSG Study284293ENTadahiroKuribayashiDepartment of Hematology, Oncology and Respiratory Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesGoMakimotoDepartment of Respiratory Medicine, Okayama University HospitalKadoakiOhashiDepartment of Respiratory Medicine, Okayama University HospitalShutaTomidaCenter for Comprehensive Genomic Medicine, Okayama University HospitalHirofumiInoueCenter for Comprehensive Genomic Medicine, Okayama University HospitalToshihideYokoyamaDepartment of Respiratory Medicine, Ohara Healthcare Foundation, Kurashiki Central HospitalShoichiKuyamaDepartment of Respiratory Medicine, NHO Iwakuni Clinical CenterYukaKatoDepartment of Thoracic Oncology and Medicine, National Hospital Organization, Shikoku Cancer CenterKenichiroKudoDepartment of Respiratory Medicine, National Hospital Organization Okayama Medical CenterNaokatsuHoritaDepartment of Respiratory Medicine, Kure Kyosai HospitalHiroeKayataniDepartment of Respiratory Medicine, Japanese Red Cross Okayama HospitalMasaakiInoueDepartment of Chest Surgery, Shimonoseki City HospitalKeisukeSugimotoDepartment of Respiratory Medicine, Japanese Red Cross Kobe HospitalKiichiroNinomiyaCenter for Comprehensive Genomic Medicine, Okayama University HospitalYoshinobuMaedaDepartment of Hematology, Oncology and Respiratory Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYosukeTogashiDepartment of Respiratory Medicine, Okayama University HospitalKatsuyukiHottaCenter for Innovative Clinical Medicine, Okayama University HospitalSome anaplastic lymphoma kinase (ALK) gene rearrangement–positive lung cancers show early resistance, within 3 months, to alectinib. This study investigated the clinical and molecular characteristics of these patients. We analyzed patients with unresectable stage III/IV disease without indications for radical radiotherapy and recurrent ALK-positive lung cancer who received alectinib as the primary ALK tyrosine kinase inhibitor between 2013 and 2021 at nine hospitals. In total, 103 patients were included. The median age was 65 years; 44 were male and 22 had brain metastases. The median progression-free survival and overall survival (OS) were 28.7 and 80.6 months. Nineteen patients treated for ≤3 months and 84 treated for >3 months were categorized into the early resistance and responder groups, respectively. The early resistance group had significantly shorter OS (8.4 months vs. not estimable, P < 0.001) and was significantly more likely to have brain metastases (42% vs. 17%, P = 0.027). They also showed elevated inflammatory markers, including neutrophil-to-lymphocyte ratio (NLR). Univariate analysis identified brain metastases and high NLR as significant predictors of early resistance. Spatial transcriptome analysis and immunohistochemical staining revealed upregulation of annexin A1 (ANXA1), a calcium-dependent phospholipid-binding protein involved in inflammation and cancer progression, in the early resistance group. Interleukin 6 stimulation, prompted by elevated inflammatory markers, increased ANXA1 expression and reduced alectinib sensitivity. Knockdown of ANXA1 improved alectinib sensitivity in alectinib-resistant cells. In conclusion, brain metastases and high NLR are associated with early resistance. ANXA1 may play an important role in mediating early resistance. New treatment options for the early resistance group are required.No potential conflict of interest relevant to this article was reported.Elsevier BVActa Medica Okayama2211-12474512026Immunopeptidomics combined with full-length transcriptomics uncovers diverse neoantigens116781ENTakamasaIshinoDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesTomofumiWatanabeDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesSerinaTokitaDivision of Cancer Immunology, Graduate School of Medical and Dental Sciences, Niigata UniversityYoukiUedaDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesKatsushigeKawaseDivision of Cell Therapy, Chiba Cancer Center Research InstituteYukaTakanoDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesYin MinThuDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesYutaSuzukiDepartment of Computational Biology and Medical Sciences, The University of TokyoChieOwaDepartment of Computational Biology and Medical Sciences, The University of TokyoTakashiInozumeDepartment of Dermatology, Chiba University Graduate School of MedicineWenhaoZhouDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesJojiNagasakiDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesVitalyKochinDepartment of Immunology, Nagoya University Graduate School of MedicineToshihideUenoDivision of Cellular Signaling, National Cancer Center Research InstituteShinyaKojimaDivision of Cellular Signaling, National Cancer Center Research InstituteAkikoHonobe-TabuchiDepartment of Dermatology, University of YamanashiTatsuyoshiKawamuraDepartment of Dermatology, University of YamanashiTakehiroOhnumaDepartment of Dermatology, Kumamoto Kenhoku HospitalTakamitsuMatsuzawaDepartment of Dermatology, Chiba University Graduate School of MedicineYuKawaharaDepartment of Dermatology, Chiba University Graduate School of MedicineKazuoYamashitaKOTAI Biotechnologies, IncJasonLinDivision of Cell Therapy, Chiba Cancer Center Research InstituteJunKosekiDivision of Systems Biology, Nagoya University Graduate School of MedicineHiroyoshiNishikawaDepartment of Immunology, Nagoya University Graduate School of MedicineMotooArakiDepartment of Urology, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesNaoyaKatoDepartment of Gastroenterology, Graduate School of Medicine, Chiba UniversityTeppeiShimamuraDivision of Systems Biology, Nagoya University Graduate School of MedicineShinichiMorishitaDepartment of Computational Biology and Medical Sciences, The University of TokyoYutakaSuzukiDepartment of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of TokyoHiroyukiManoDivision of Cellular Signaling, National Cancer Center Research InstituteToshihikoTorigoeTakayukiKanasekiDivision of Cancer Immunology, Graduate School of Medical and Dental Sciences, Niigata UniversityMasahitoKawazuDivision of Cell Therapy, Chiba Cancer Center Research InstituteYosukeTogashiDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesNeoantigens are crucial for antitumor immunity and immune checkpoint inhibitor (ICI) efficacy by triggering strong immune responses. However, conventional methods for identifying neoantigens, such as whole-exon sequencing and short-read RNA sequencing (RNA-seq), appear to be insufficient, and the tumor mutational burden cannot sufficiently predict ICI efficacy. In this study, we employed a proteogenomic approach using long-read RNA-seq with Pacific Biosciences Single-Molecule Real-Time Sequencing technology to analyze full-length transcripts in combination with the human leukocyte antigen ligandome. As a result, many neoantigen candidates were identified, which were unregistered in a comprehensive database, including those from non-coding regions. Additionally, we validated the responses of specific T cell receptors (TCRs) to these candidates and identified several pairs of TCRs and neoantigens. These findings highlight the presence of more diverse neoantigens than expected that cannot be identified by conventional methods.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama2399-3642812025Single-cell and spatial transcriptomic characterization of pulmonary pleomorphic carcinoma1773ENAtsushiMatsuokaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKazuhikoShienDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesShutaTomidaMasayoshiOhkiDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKazuyaHisamatsuDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesRyotaFujiwaraDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKoseiIshimuraDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesRyunosukeFujiiDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesTomoakiHigashiharaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNaohiroHayashiDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKazuhiroOkadaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesRyoYoshichikaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesFumiakiMukoharaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMaoYoshikawaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYumaFukumotoDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKenSuzawaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYasuakiTomiokaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesShinTanakaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKentarohMiyoshiDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMikioOkazakiDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesSeiichiroSugimotoDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYusukeOtaniDepartment of Pathology, Beth Israel Deaconess Medical CenterAtsushiTanakaDepartment of Pathology, Beth Israel Deaconess Medical CenterHirofumiInoueCenter for Comprehensive Genomic Medicine, Okayama University HospitalYosukeTogashiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHidetakaYamamotoDepartment of Pathology and Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesDaisukeEnnishiCenter for Comprehensive Genomic Medicine, Okayama University HospitalShinichiToyookaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesPulmonary pleomorphic carcinoma (PPC) is a rare subtype of lung cancer that comprises both epithelial and sarcomatoid components. The molecular basis of PPC, including the cellular dynamics of its components, remains largely unknown. To elucidate potential therapeutic targets for PPC, we perform a multi-omics analysis incorporating digital spatial profiling and single-cell RNA sequencing (scRNA-seq). PPC exhibits diverse driver gene alterations, including MET exon 14 skipping mutation (METex14) and ALK fusion. In spatial transcriptomics, MET gene and protein are overexpressed exclusively within the epithelial component and not in the sarcomatoid component, even in patients harboring METex14. Epithelial-mesenchymal transition (EMT)-related transcriptional changes, along with extracellular matrix (ECM) remodeling between the epithelial and sarcomatoid components, are observed. scRNA-seq identifies cell populations within the epithelial component that contribute to the malignant transformation and differentiation of the sarcomatoid component. They are characterized by an intermediate EMT state with ECM remodeling signature, suggesting their potential as novel therapeutic targets for PPC.No potential conflict of interest relevant to this article was reported.Japanese Society of Internal MedicineActa Medica Okayama0918-291864232025A Prompt Diagnosis of Ascites and Dramatic Effect of Alectinib for Advanced Lung Adenocarcinoma Harboring EML4-ALK Fusion34133418ENTakahiroBabaDepartment of Respiratory Medicine, Okayama University HospitalHirofumiInoueDepartment of Medical Support, Okayama University HospitalHiromiMatsuokaDepartment of Medical Support, Okayama University HospitalMioKyakunoDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of MedicineYusukeYoshinagaDepartment of Respiratory Medicine, Okayama University HospitalTetsuyaTakeguchiDepartment of Respiratory Medicine, Okayama University HospitalMihoFujiwaraDepartment of Respiratory Medicine, Okayama University HospitalKotaroYamadaDepartment of Respiratory Medicine, Okayama University HospitalEriNakamuraDepartment of Respiratory Medicine, Okayama University HospitalAyakoMoritaDepartment of Respiratory Medicine, Okayama University HospitalNaofumiHaraDepartment of Respiratory Medicine, Okayama University HospitalKiichiroNinomiyaCenter for Comprehensive Genomic Medicine, Okayama University HospitalHisaoHigoDepartment of Respiratory Medicine, Okayama University HospitalMasanoriFujiiDepartment of Geriatric Medicine, Okayama University HospitalEikiIchiharaCenter for Clinical Oncology, Okayama University HospitalKammeiRaiCenter for Innovative Clinical Medicine, Okayama University HospitalKadoakiOhashiDepartment of Respiratory Medicine, Okayama University HospitalKatsuyukiHottaCenter for Innovative Clinical Medicine, Okayama University HospitalMasahiroTabataCenter for Clinical Oncology, Okayama University HospitalYoshinobuMaedaDepartment of Hematology, Oncology, and Respiratory Medicine, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesYosukeTogashiDepartment of Respiratory Medicine, Okayama University HospitalGoMakimotoDepartment of Respiratory Medicine, Okayama University HospitalA 75-year-old never-smoker woman presented with dyspnea and loss of appetite. A mass was identified in the left upper lobe of the lung, and the patient was referred to our hospital. Despite the diagnosis of lung adenocarcinoma via bronchoscopy, anaplastic lymphoma kinase (ALK) immunostaining was negative. Rapid weight gain and abdominal distension caused by ascites prompted fluid testing using the AmoyDx® Pan Lung Cancer PCR Panel. EML4-ALK fusion was confirmed, and alectinib therapy was initiated immediately. The tumor size had decreased significantly, and the patient was discharged on day 34. This case highlights the necessity of multiplex genetic testing even when ALK immunostaining is negative.No potential conflict of interest relevant to this article was reported.American Chemical Society (ACS)Acta Medica Okayama0002-78632025Optogenetic Cancer Therapy Using the Light-Driven Outward Proton Pump Rhodopsin Archaerhodopsin-3 (AR3)ENShinNakaoGraduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKeiichiKojimaGraduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKeitaSatoGraduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityNaoyaKemmotsuGraduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHideyoOhuchiGraduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYosukeTogashiGraduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYukiSudoGraduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMedicines used for cancer treatment often cause serious side effects by damaging normal cells due to nonspecific diffusion. To address this issue, we previously developed an optical method to induce apoptotic cell death via intracellular pH alkalinization using the outward proton pump rhodopsin, Archaerhodopsin-3 (AR3) in various noncancer model cells in vitro and in vivo. In this study, we applied this method to cancer cells and tumors to evaluate its potential as an anticancer therapeutic strategy. First, we confirmed that AR3-expressing murine cancer cell lines (MC38, B16F10) showed apoptotic cell death upon green light irradiation, as indicated by increased levels of cell death and apoptosis-related markers. Next, we established stable AR3-expressing MC38 and B16F10 cells by using viral vectors. When these AR3-expressing cells were subcutaneously transplanted into C57BL/6 mice, the resulting tumors initially grew at a rate comparable to that of control tumors lacking AR3 expression or light stimulation. However, upon green light irradiation, AR3-expressing tumors exhibited either a marked reduction in size or significantly suppressed growth, accompanied by the induction of apoptosis signals and decreased proliferation signals. These results demonstrate that AR3-mediated cell death has potent antitumor effects both in vitro and in vivo. This optical method thus holds promise as a novel cancer therapy with potentially reduced side effects.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama0028-083663880492025Immune evasion through mitochondrial transfer in the tumour microenvironment225236ENHidekiIkedaDivision of Cell Therapy, Chiba Cancer Center Research InstituteKatsushigeKawaseDivision of Cell Therapy, Chiba Cancer Center Research InstituteTatsuyaNishiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTomofumiWatanabeDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKeizoTakenagaDivision of Innovative Cancer Therapeutics, Chiba Cancer Center Research InstituteTakashiInozumeDivision of Cell Therapy, Chiba Cancer Center Research InstituteTakamasaIshinoDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityShoAkiDivision of Nutriomics and Oncology, RCAST, The University of TokyoJasonLinDivision of Cell Therapy, Chiba Cancer Center Research InstituteShusukeKawashimaDivision of Cell Therapy, Chiba Cancer Center Research Institute, Chiba, Japan Department of Dermatology, Graduate School of Medicine, Chiba UniversityJojiNagasakiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYoukiUedaDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityShinichiroSuzukiDepartment of Medical Oncology, Kindai University Faculty of MedicineHidekiMakinoshimaTsuruoka Metabolomics Laboratory, National Cancer CenterMakikoItamiDepartment of Surgical Pathology, Chiba Cancer CenterYukiNakamuraDivision of Cell Therapy, Chiba Cancer Center Research InstituteYasutoshiTatsumiDivision of Cell Therapy, Chiba Cancer Center Research InstituteYusukeSuenagaLaboratory of Evolutionary Oncology, Chiba Cancer Center Research InstituteTakaoMorinagaDivision of Cell Therapy, Chiba Cancer Center Research InstituteAkikoHonobe-TabuchiDepartment of Dermatology, Faculty of Medicine, University of YamanashiTakehiroOhnumaDepartment of Dermatology, Faculty of Medicine, University of YamanashiTatsuyoshiKawamuraDepartment of Dermatology, Faculty of Medicine, University of YamanashiYoshiyasuUmedaDepartment of Skin Oncology/Dermatology, Saitama Medical University International Medical CenterYasuhiroNakamuraDepartment of Skin Oncology/Dermatology, Saitama Medical University International Medical CenterYukikoKiniwaDepartment of Dermatology, Shinshu University School of MedicineEikiIchiharaDepartment of Allergy and Respiratory Medicine, Okayama University HospitalHidetoshiHayashiDepartment of Medical Oncology, Kindai University Faculty of MedicineJun-ichiroIkedaDepartment of Diagnostic Pathology, Graduate School of Medicine, Chiba UniversityToyoyukiHanazawaDepartment of Otorhinolaryngology/Head and Neck Surgery, Chiba University Graduate School of MedicineShinichiToyookaDepartment of General Thoracic Surgery and Endocrinological Surgery, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiroyukiManoDivision of Cellular Signalling, National Cancer Center Research InstituteTakujiSuzukiDepartment of Respirology, Graduate School of Medicine, Chiba UniversityTsuyoshiOsawaDivision of Nutriomics and Oncology, RCAST, The University of TokyoMasahitoKawazuDivision of Cell Therapy, Chiba Cancer Center Research InstituteYosukeTogashiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityCancer cells in the tumour microenvironment use various mechanisms to evade the immune system, particularly T cell attack1. For example, metabolic reprogramming in the tumour microenvironment and mitochondrial dysfunction in tumour-infiltrating lymphocytes (TILs) impair antitumour immune responses2,3,4. However, detailed mechanisms of such processes remain unclear. Here we analyse clinical specimens and identify mitochondrial DNA (mtDNA) mutations in TILs that are shared with cancer cells. Moreover, mitochondria with mtDNA mutations from cancer cells are able to transfer to TILs. Typically, mitochondria in TILs readily undergo mitophagy through reactive oxygen species. However, mitochondria transferred from cancer cells do not undergo mitophagy, which we find is due to mitophagy-inhibitory molecules. These molecules attach to mitochondria and together are transferred to TILs, which results in homoplasmic replacement. T cells that acquire mtDNA mutations from cancer cells exhibit metabolic abnormalities and senescence, with defects in effector functions and memory formation. This in turn leads to impaired antitumour immunity both in vitro and in vivo. Accordingly, the presence of an mtDNA mutation in tumour tissue is a poor prognostic factor for immune checkpoint inhibitors in patients with melanoma or non-small-cell lung cancer. These findings reveal a previously unknown mechanism of cancer immune evasion through mitochondrial transfer and can contribute to the development of future cancer immunotherapies.No potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama1347-903211652025High Antigenicity for Treg Cells Confers Resistance to PD-1 Blockade Therapy via High PD-1 Expression in Treg Cells12141226ENHiroakiMatsuuraDepartment of Tumor Microenvironment, Okayama UniversityTakamasaIshinoDepartment of Tumor Microenvironment, Okayama UniversityToshifumiNinomiyaDepartment of Tumor Microenvironment, Okayama UniversityKiichiroNinomiyaDepartment of Hematology, Oncology and Respiratory Medicine,Okayama UniversityKotaTachibanaDepartment of Dermatology, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityAkikoHonobe-TabuchiDepartment of Dermatology, University of YamanashiYoshinoriMutoDepartment of Dermatology, University of YamanashiTakashiInozumeDepartment of Dermatology, University of YamanashiYoukiUedaDepartment of Tumor Microenvironment, Okayama UniversityKadoakiOhashiDepartment of Hematology, Oncology and Respiratory Medicine,Okayama UniversityYoshinobuMaedaDepartment of Hematology, Oncology and Respiratory Medicine,Okayama UniversityJojiNagasakiDepartment of Tumor Microenvironment, Okayama UniversityYosukeTogashiDepartment of Tumor Microenvironment, Okayama UniversityRegulatory T (Treg) cells have an immunosuppressive function, and programmed death-1 (PD-1)-expressing Treg cells reportedly induce resistance to PD-1 blockade therapies through their reactivation. However, the effects of antigenicity on PD-1 expression in Treg cells and the resistance to PD-1 blockade therapy remain unclear. Here, we show that Treg cells gain high PD-1 expression through an antigen with high antigenicity. Additionally, tumors with high antigenicity for Treg cells were resistant to PD-1 blockade in vivo due to PD-1+ Treg-cell infiltration. Because such PD-1+ Treg cells have high cytotoxic T lymphocyte antigen (CTLA)-4 expression, resistance could be overcome by combination with an anti-CTLA-4 monoclonal antibody (mAb). Patients who responded to combination therapy with anti-PD-1 and anti-CTLA-4 mAbs sequentially after primary resistance to PD-1 blockade monotherapy showed high Treg cell infiltration. We propose that the high antigenicity of Treg cells confers resistance to PD-1 blockade therapy via high PD-1 expression in Treg cells, which can be overcome by combination therapy with an anti-CTLA-4 mAb.No potential conflict of interest relevant to this article was reported.Proceedings of the National Academy of SciencesActa Medica Okayama0027-8424121352024Somatic mutations in tumor-infiltrating lymphocytes impact on antitumor immunitye2320189121ENFumiakiMukoharaDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKazumaIwataDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakamasaIshinoDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakashiInozumeDepartment of Dermatology, Chiba University Graduate School of MedicineJojiNagasakiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYoukiUedaDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKenSuzawaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Dentistry and Pharmaceutical Sciences, Okayama University Graduate School of Medicine, Okayama UniversityToshihideUenoDivision of Cellular Signaling, National Cancer Center Research InstituteHidekiIkedaDivision of Cell Therapy, Chiba Cancer Research InstituteKatsushigeKawaseDivision of Cell Therapy, Chiba Cancer Research InstituteYukaSaekiDepartment of Dermatology, Chiba University Graduate School of MedicineShusukeKawashimaDepartment of Dermatology, Chiba University Graduate School of MedicineKazuoYamashitaKOTAI Biotechnologies, Inc.YuKawaharaDepartment of Dermatology, Chiba University Graduate School of MedicineYasuhiroNakamuraDepartment of Skin Oncology/Dermatology, Saitama Medical University International Medical CenterAkikoHonobe-TabuchiDepartment of Dermatology, University of YamanashiHirokoWatanabeDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiromichiDansakoDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTatsuyoshiKawamuraDepartment of Dermatology, University of YamanashiYutakaSuzukiDepartment of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, KashiwaHiroakiHondaDepartment of Pathology, Tokyo Women's Medical UniversityHiroyukiManoDivision of Cellular Signaling, National Cancer Center Research InstituteShinichiToyookaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Dentistry and Pharmaceutical Sciences, Okayama University Graduate School of Medicine, Okayama UniversityMasahitoKawazuDivision of Cell Therapy, Chiba Cancer Research InstituteYosukeTogashiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityImmune checkpoint inhibitors (ICIs) exert clinical efficacy against various types of cancers by reinvigorating exhausted CD8+ T cells that can expand and directly attack cancer cells (cancer-specific T cells) among tumor-infiltrating lymphocytes (TILs). Although some reports have identified somatic mutations in TILs, their effect on antitumor immunity remains unclear. In this study, we successfully established 18 cancer-specific T cell clones, which have an exhaustion phenotype, from the TILs of four patients with melanoma. We conducted whole-genome sequencing for these T cell clones and identified various somatic mutations in them with high clonality. Among the somatic mutations, an SH2D2A loss-of-function frameshift mutation and TNFAIP3 deletion could activate T cell effector functions in vitro. Furthermore, we generated CD8+ T cell–specific Tnfaip3 knockout mice and showed that Tnfaip3 function loss in CD8+ T cell increased antitumor immunity, leading to remarkable response to PD-1 blockade in vivo. In addition, we analyzed bulk CD3+ T cells from TILs in additional 12 patients and identified an SH2D2A mutation in one patient through amplicon sequencing. These findings suggest that somatic mutations in TILs can affect antitumor immunity and suggest unique biomarkers and therapeutic targets.No potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama1347-9032115102024Overcoming immunotherapy resistance and inducing abscopal effects with boron neutron immunotherapy (B-NIT)32313247ENTakuyaFujimotoDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesOsamuYamasakiDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesNoriyukiKanehiraDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesHirokazuMatsushitaDivision of Translational Oncoimmunology, Aichi Cancer Center Research InstituteYoshinoriSakuraiInstitute for Integrated Radiation and Nuclear Science, Kyoto UniversityNaoyaKenmotsuDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesRyoMizutaDepartment of Neurological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesNatsukoKondoInstitute for Integrated Radiation and Nuclear Science, Kyoto UniversityTakushiTakataInstitute for Integrated Radiation and Nuclear Science, Kyoto UniversityMizukiKitamatsuFaculty of Science and Engineering, Kindai UniversityKazuyoIgawaNeutron Therapy Research Center, Okayama UniversityAtsushiFujimuraNeutron Therapy Research Center, Okayama UniversityYoshihiroOtaniDepartment of Neurological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesMakotoShirakawaNeutron Therapy Research Center, Okayama UniversityKunitoshiShigeyasuDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesFuminoriTeraishiDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesYosukeTogashiDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesMinoruSuzukiInstitute for Integrated Radiation and Nuclear Science, Kyoto UniversityToshiyoshiFujiwaraDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical SciencesHiroyukiMichiueNeutron Therapy Research Center, Okayama UniversityImmune checkpoint inhibitors (ICIs) are effective against many advanced malignancies. However, many patients are nonresponders to immunotherapy, and overcoming this resistance to treatment is important. Boron neutron capture therapy (BNCT) is a local chemoradiation therapy with the combination of boron drugs that accumulate selectively in cancer and the neutron irradiation of the cancer site. Here, we report the first boron neutron immunotherapy (B-NIT), combining BNCT and ICI immunotherapy, which was performed on a radioresistant and immunotherapy-resistant advanced-stage B16F10 melanoma mouse model. The BNCT group showed localized tumor suppression, but the anti-PD-1 antibody immunotherapy group did not show tumor suppression. Only the B-NIT group showed strong tumor growth inhibition at both BNCT-treated and shielded distant sites. Intratumoral CD8+ T-cell infiltration and serum high mobility group box 1 (HMGB1) levels were higher in the B-NIT group. Analysis of CD8(+) T cells in tumor-infiltrating lymphocytes (TILs) showed that CD62L- CD44(+) effector memory T cells and CD69(+) early-activated T cells were predominantly increased in the B-NIT group. Administration of CD8-depleting mAb to the B-NIT group completely suppressed the augmented therapeutic effects. This indicated that B-NIT has a potent immune-induced abscopal effect, directly destroying tumors with BNCT, inducing antigen-spreading effects, and protecting normal tissue. B-NIT, immunotherapy combined with BNCT, is the first treatment to overcome immunotherapy resistance in malignant melanoma. In the future, as its therapeutic efficacy is demonstrated not only in melanoma but also in other immunotherapy-resistant malignancies, B-NIT can become a new treatment candidate for advanced-stage cancers.No potential conflict of interest relevant to this article was reported.Elsevier BVActa Medica Okayama2211-12474322024Stem-like progenitor and terminally differentiated TFH-like CD4+ T cell exhaustion in the tumor microenvironment113797ENWenhaoZhouDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesShusukeKawashimaDepartment of Dermatology, Chiba University Graduate School of MedicineTakamasaIshinoDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKatsushigeKawaseChiba Cancer Center, Research InstituteYoukiUedaDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKazuoYamashitaKOTAI Biotechnologies, Inc. TomofumiWatanabeDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMasahitoKawazuChiba Cancer Center, Research Institute, Division of Cell TherapyHiromichiDansakoDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYutakaSuzukiDepartment of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of TokyoHiroyoshiNishikawaDepartment of Immunology, Nagoya University Graduate School of MedicineTakashiInozumeDepartment of Dermatology, Chiba University Graduate School of MedicineJojiNagasakiDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYosukeTogashiDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesImmune checkpoint inhibitors exert clinical efficacy against various types of cancer through reinvigoration of exhausted CD8+ T cells that attack cancer cells directly in the tumor microenvironment (TME). Using single-cell sequencing and mouse models, we show that CXCL13, highly expressed in tumor-infiltrating exhausted CD8+ T cells, induces CD4+ follicular helper T (TFH) cell infiltration, contributing to anti-tumor immunity. Furthermore, a part of the TFH cells in the TME exhibits cytotoxicity and directly attacks major histocompatibility complex-II-expressing tumors. TFH-like cytotoxic CD4+ T cells have high LAG-3/BLIMP1 and low TCF1 expression without self-renewal ability, whereas non-cytotoxic TFH cells express low LAG-3/BLIMP1 and high TCF1 with self-renewal ability, closely resembling the relationship between terminally differentiated and stem-like progenitor exhaustion in CD8+ T cells, respectively. Our findings provide deep insights into TFH-like CD4+ T cell exhaustion with helper progenitor and cytotoxic differentiated functions, mediating anti-tumor immunity orchestrally with CD8+ T cells.No potential conflict of interest relevant to this article was reported.ElsevierActa Medica Okayama2666-36434102023Immunologic Significance of CD80/CD86 or Major Histocompatibility Complex-II Expression in Thymic Epithelial Tumors100573ENHidekiIkedaChiba Cancer Center, Research InstituteJojiNagasakiChiba Cancer Center, Research InstituteDaikiShimizuDivision of Thoracic Surgery, Chiba Cancer CenterYukiKatsuyaDepartment of Experimental Therapeutics, National Cancer Center HospitalHidehitoHorinouchiDepartment of Thoracic Oncology, National Cancer Center HospitalYukioHosomiDepartment of Thoracic Oncology and Respiratory Medicine, Tokyo Metropolitan Cancer and Infectious Diseases Center Komagome HospitalEtsukoTanjiChiba Cancer Center, Research InstituteTakekazuIwataDivision of Thoracic Surgery, Chiba Cancer CenterMakikoItamiDepartment of Surgical Pathology, Chiba Cancer CenterMasahitoKawazuChiba Cancer Center, Research InstituteYuichiroOheDepartment of Thoracic Oncology, National Cancer Center HospitalTakujiSuzukiDepartment of Respirology, Graduate School of Medicine, Chiba UniversityYosukeTogashiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityIntroduction: Unresectable or recurrent thymic epithelial tumors (TETs) have a poor prognosis, and treatment options are limited. This study aimed to investigate the immunologic significance of CD80/CD86 or major histocompatibility complex class II (MHC-II) expression in TETs, as potential predictive biomarkers for immune checkpoint inhibitors (ICIs).<br>
Methods: We analyzed CD80, CD86, MHC class I (MHC-I), and MHC-II expression in TETs using immunohistochemistry and investigated their association with T-cell infiltration or ICI efficacy. In addition, we generated CD80- or MHC-II–expressing mouse tumors, evaluated the effects of ICIs, and analyzed tumor-infiltrating lymphocytes. We also performed tumor-rechallenge experiments in vivo.<br>
Results: We found that approximately 50% and 30% of TETs had high expression of CD80/CD86 and MHC-II in tumor cells, respectively, and that this expression was related to T-cell infiltration in clinical samples. In mouse models, both CD80 and MHC-II increase the effects of ICIs. In addition, senescent T cells and long-lived memory precursor effector T cells were significantly decreased and increased, respectively, in tumor-infiltrating lymphocytes from CD80-expressing tumors, and rechallenged tumors were completely rejected after the initial eradication of CD80-expressing tumors by programmed cell death protein 1 blockade. Indeed, patients with CD80-high thymic carcinoma had longer progression-free survival with anti–programmed cell death protein 1 monoclonal antibody.<br>
Conclusions: Half of the TETs had high expression of CD80/CD86 or MHC-II with high T-cell infiltration. These molecules could potentially increase the effects of ICIs, particularly inducing a durable response. CD80/CD86 and MHC-II can be predictive biomarkers of ICIs in TETs, promoting the development of drugs for such TETs.No potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama1742-464X29162023Hepatitis C virus NS5B triggers an MDA5-mediated innate immune response by producing dsRNA without the replication of viral genomes11191130ENHiromichiDansakoDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMasanoriIkedaDivision of Biological Information Technology, Joint Research Center for Human Retrovirus Infection, Kagoshima UniversityYasuoAriumiManagement Department of Biosafety, Laboratory Animal, and Pathogen Bank, National Institute of Infectious DiseasesYosukeTogashiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityNobuyukiKatoDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityDuring 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.No potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama0020-713615412023Low frequency of intracranial progression in advanced NSCLC patients treated with cancer immunotherapies169179ENNaoyaKemmotsuDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKiichiroNinomiyaDepartment of Respiratory Medicine, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKeiKunimasaDepartment of Thoracic Oncology, Osaka International Cancer InstituteTakamasaIshinoDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityJojiNagasakiDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYoshihiroOtaniDepartment of Neurological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiroyukiMichiueDepartment of Neurological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityEikiIchiharaDepartment of Respiratory Medicine, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKadoakiOhashiDepartment of Respiratory Medicine, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakakoInoueDepartment of Thoracic Oncology, Osaka International Cancer InstituteMotohiroTamiyaDepartment of Thoracic Oncology, Osaka International Cancer InstituteKazukoSakaiDepartment of Genome Biology, Kindai University Faculty of MedicineYoukiUedaDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiromichiDansakoDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKazutoNishioDepartment of Genome Biology, Kindai University Faculty of MedicineKatsuyukiKiuraDepartment of Respiratory Medicine, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityIsaoDateDepartment of Neurological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYosukeTogashiDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityIntracranial metastases are common in nonsmall-cell lung cancer (NSCLC) patients, whose prognosis is very poor. In addition, intracranial progression is common during systemic treatments due to the inability to penetrate central nervous system (CNS) barriers, whereas the intracranial effects of cancer immunotherapies remain unclear. We analyzed clinical data to evaluate the frequency of intracranial progression in advanced NSCLC patients treated with PD-1 blockade therapies compared with those treated without PD-1 blockade therapies, and found that the frequency of intracranial progression in advanced NSCLC patients treated with PD-1 blockade therapies was significantly lower than that in patients treated with cytotoxic chemotherapies. In murine models, intracranial rechallenged tumors after initial rejection by PD-1 blockade were suppressed. Accordingly, long-lived memory precursor effector T cells and antigen-specific T cells were increased by PD-1 blockade in intracranial lesions. However, intracranial rechallenged different tumors are not suppressed. Our results indicate that cancer immunotherapies can prevent intracranial progression, maintaining long-term effects intracranially as well as systemically. If intracranial recurrence occurs during the treatment with PD-1 blockade therapies, aggressive local therapies could be worthwhile.No potential conflict of interest relevant to this article was reported.American Association for Cancer ResearchActa Medica Okayama2767-9764272022Mixed Response to Cancer Immunotherapy is Driven by Intratumor Heterogeneity and Differential Interlesion Immune Infiltration739753ENTakaoMorinagaChiba Cancer Center, Research InstituteTakashiInozumeChiba Cancer Center, Research InstituteMasahitoKawazuChiba Cancer Center, Research InstituteYoukiUedaDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine Dentistry and Pharmaceutical SciencesNicolasSaxKOTAI Biotechnologies IncKazuoYamashitaKOTAI Biotechnologies IncShusukeKawashimaChiba Cancer Center, Research InstituteJojiNagasakiDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine Dentistry and Pharmaceutical SciencesToshihideUenoDivision of Cellular Signaling, National Cancer Center Research InstituteJasonLinChiba Cancer Center, Research InstituteYuukiOharaDepartment of Pathology, National Cancer Center Hospital EastTakeshiKuwataDepartment of Genetic Medicineand Services, National Cancer Center Hospital EastHirokiYukamiDepartment of Gastroenterology and Gastrointestinal Oncology, National Cancer Center Hospital EastAkihitoKawazoeDepartment of Gastroenterology and Gastrointestinal Oncology, National Cancer Center Hospital EastKoheiShitaraDepartment of Gastroenterology and Gastrointestinal Oncology, National Cancer Center Hospital EastAkikoHonobe-TabuchiDepartment of Dermatology, University of YamanashiTakehiroOhnumaDepartment of Dermatology, University of YamanashiTatsuyoshiKawamuraDepartment of Dermatology, University of YamanashiYoshiyasuUmedaDepartment of Skin Oncology/Dermatology, Saitama Medical University International Medical CenterYuKawaharaDepartment of Skin Oncology/Dermatology, Saitama Medical University International Medical CenterYasuhiroNakamuraDepartment of Skin Oncology/Dermatology, Saitama Medical University International Medical CenterYukikoKiniwaDepartment of Dermatology, Shinshu University School of Medicine AyakoMoritaDepartment of Allergy and Respiratory Medicine, Okayama University HospitalEikiIchiharaDepartment of Allergy and Respiratory Medicine, Okayama University HospitalKatsuyukiKiuraDepartment of Allergy and Respiratory Medicine, Okayama University HospitalTomohiroEnokidaDepartment of Head and Neck Medical Oncology, National Cancer Center Hospital EastMakotoTaharaDepartment of Head and Neck Medical Oncology, National Cancer Center Hospital EastYoshinoriHasegawaDepartment of Applied Genomics, Kazusa DNA Research InstituteHiroyukiManoDivision of Cellular Signaling, National Cancer Center Research InstituteYutakaSuzukiDepartment of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of TokyoHiroyoshiNishikawaDivision of Cancer Immunology, Research Institute/Exploratory Oncology Research and Clinical Trial Center (EPOC), National Cancer CenterYosukeTogashiDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine Dentistry and Pharmaceutical SciencesSome patients experience mixed response to immunotherapy, whose biological mechanisms and clinical impact have been obscure. We obtained two tumor samples from lymph node (LN) metastatic lesions in a same patient. Whole exome sequencing for the both tumors and single-cell sequencing for the both tumor-infiltrating lymphocytes (TIL) demonstrated a significant difference in tumor clonality and TILs' characteristics, especially exhausted T-cell clonotypes, although a close relationship between the tumor cell and T-cell clones were observed as a response of an overlapped exhausted T-cell clone to an overlapped neoantigen. To mimic the clinical setting, we generated a mouse model of several clones from a same tumor cell line. Similarly, differential tumor clones harbored distinct TILs, and one responded to programmed cell death protein 1 (PD-1) blockade but the other did not in this model. We further conducted cohort study (n = 503) treated with PD-1 blockade monotherapies to investigate the outcome of mixed response. Patients with mixed responses to PD-1 blockade had a poor prognosis in our cohort. Particularly, there were significant differences in both tumor and T-cell clones between the primary and LN lesions in a patient who experienced tumor response to anti-PD-1 mAb followed by disease progression in only LN metastasis. Our results underscore that intertumoral heterogeneity alters characteristics of TILs even in the same patient, leading to mixed response to immunotherapy and significant difference in the outcome.<br>
Significance: Several patients experience mixed responses to immunotherapies, but the biological mechanisms and clinical significance remain unclear. Our results from clinical and mouse studies underscore that intertumoral heterogeneity alters characteristics of TILs even in the same patient, leading to mixed response to immunotherapy and significant difference in the outcome.No potential conflict of interest relevant to this article was reported.American Association for Cancer Research (AACR)Acta Medica Okayama2326-60661172023High Expression of MHC Class I Overcomes Cancer Immunotherapy Resistance Due to IFNγ Signaling Pathway Defects895908ENKatsushigeKawaseDivision of Cell Therapy, Chiba Cancer Center Research InstituteShusukeKawashimaDivision of Cell Therapy, Chiba Cancer Center Research InstituteJojiNagasakiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakashiInozumeDivision of Cell Therapy, Chiba Cancer Center Research InstituteEtsukoTanjiDivision of Cell Therapy, Chiba Cancer Center Research InstituteMasahitoKawazuDivision of Cell Therapy, Chiba Cancer Center Research InstituteToyoyukiHanazawaDepartment of Otorhinolaryngology/Head & Neck Surgery, Graduate School of Medicine, Chiba UniversityYosukeTogashiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityIFNγ signaling pathway defects are well-known mechanisms of resistance to immune checkpoint inhibitors. However, conflicting data have been reported, and the detailed mechanisms remain unclear. In this study, we have demonstrated that resistance to immune checkpoint inhibitors owing to IFNγ signaling pathway defects may be primarily caused by reduced MHC-I expression rather than by the loss of inhibitory effects on cellular proliferation or decreased chemokine production. In particular, we found that chemokines that recruit effector T cells were mainly produced by immune cells rather than cancer cells in the tumor microenvironment of a mouse model, with defects in IFNγ signaling pathways. Furthermore, we found a response to immune checkpoint inhibitors in a patient with JAK-negative head and neck squamous cell carcinoma whose HLA-I expression level was maintained. In addition, CRISPR screening to identify molecules associated with elevated MHC-I expression independent of IFNγ signaling pathways demonstrated that guanine nucleotide-binding protein subunit gamma 4 (GNG4) maintained MHC-I expression via the NF-κB signaling pathway. Our results indicate that patients with IFNγ signaling pathway defects are not always resistant to immune checkpoint inhibitors and highlight the importance of MHC-I expression among the pathways and the possibility of NF-κB–targeted therapies to overcome such resistance.No potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama1347-9032114102023Combination therapy with hydrogen peroxide and irradiation promotes an abscopal effect in mouse models38483856ENNaoyaKemmotsuDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityLiZhuDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityJojiNagasakiDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYoshihiroOtaniDepartment of Neurological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYoukiUedaDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiromichiDansakoDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYueFangDepartment of Microbial and Biochemical Pharmacy, School of Pharmacy, China Medical UniversityIsaoDateDepartment of Neurological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYosukeTogashiDepartment of Tumor Microenvironment, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHydrogen peroxide (H2O2) induces oxidative stress and cytotoxicity, and can be used for treating cancers in combination with radiotherapy. A product comprising H2O2 and sodium hyaluronate has been developed as a radiosensitizer. However, the effects of H2O2 on antitumor immunity remain unclear. To investigate the effects of H2O2, especially the abscopal effect when combined with radiotherapy (RT), we implanted murine tumor cells simultaneously in two locations in mouse models: the hind limb and back. H2O2 mixed with sodium hyaluronate was injected intratumorally, followed by irradiation only at the hind limb lesion. No treatment was administered to the back lesion. The H2O2/RT combination significantly reduced tumor growth at the noninjected/nonirradiated site in the back lesion, whereas H2O2 or RT individually did not reduce tumor growth. Flow cytometric analyses of the tumor-draining lymph nodes in the injected/irradiated areas showed that the number of dendritic cells increased significantly with maturation in the H2O2/RT combination group. In addition, analyses of tumor-infiltrating lymphocytes showed that the number of CD8+ (cluster of differentiation 8) T cells and the frequency of IFN-γ+ (interferon gamma) CD8+ T cells were higher in the noninjected/nonirradiated tumors in the H2O2/RT group compared to those in the other groups. PD-1 (programmed death receptor 1) blockade further increased the antitumor effect against noninjected/nonirradiated tumors in the H2O2/RT group. Intratumoral injection of H2O2 combined with RT therefore induces an abscopal effect by activating antitumor immunity, which can be further enhanced by PD-1 blockade. These findings promote the development of H2O2/RT therapy combined with cancer immunotherapies, even for advanced cancers.No potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama1347-90322023Activated CTLA-4-independent immunosuppression of Treg cells disturbs CTLA-4 blockade-mediated antitumor immunityENTomofumiWatanabeDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakamasaIshinoDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYoukiUedaDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityJojiNagasakiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakuyaSadahiraDepartment of Urology, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiromichiDansakoDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMotooArakiDepartment of Urology, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYosukeTogashiDepartment of Tumor Microenvironment, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityCombination therapy with anti-cytotoxic T lymphocyte-associated protein 4 (CTLA-4) and anti-programmed death-1 (PD-1) monoclonal antibodies (mAbs) has dramatically improved the prognosis of patients with multiple types of cancer, including renal cell carcinoma (RCC). However, more than half of RCC patients fail to respond to this therapy. Regulatory T cells (Treg cells) are a subset of highly immunosuppressive CD4(+) T cells that promote the immune escape of tumors by suppressing effector T cells in the tumor microenvironment (TME) through various mechanisms. CTLA-4 is constitutively expressed in Treg cells and is regarded as a key molecule for Treg-cell-mediated immunosuppressive functions, suppressing antigen-presenting cells by binding to CD80/CD86. Reducing Treg cells in the TME with an anti-CTLA-4 mAb with antibody-dependent cellular cytotoxicity (ADCC) activity is considered an essential mechanism to achieve tumor regression. In contrast, we demonstrated that CTLA-4 blockade without ADCC activity enhanced CD28 costimulatory signaling pathways in Treg cells and promoted Treg-cell proliferation in mouse models. CTLA-4 blockade also augmented CTLA-4-independent immunosuppressive functions, including cytokine production, leading to insufficient antitumor effects. Similar results were also observed in human peripheral blood lymphocytes and tumor-infiltrating lymphocytes from patients with RCC. Our findings highlight the importance of Treg-cell depletion to achieve tumor regression in response to CTLA-4 blockade therapies.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama0007-09202023Somatic mutations can induce a noninflamed tumour microenvironment via their original gene functions, despite deriving neoantigensENTakamasaIshinoDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesShusukeKawashimaDivision of Cell Therapy, Chiba Cancer Center Research InstituteEtsukoTanjiDivision of Cell Therapy, Chiba Cancer Center Research InstituteToshihideUenoDivision of Cellular Signaling, National Cancer Center Research InstituteYoukiUedaDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesSadahisaOgasawaraDepartment of Gastroenterology, Graduate School of Medicine, Chiba UniversityKazuhitoSatoDepartment of Surgical Oncology, Graduate School of Medicine, The University of TokyoHiroyukiManoDivision of Cellular Signaling, National Cancer Center Research InstituteSoichiroIshiharaDepartment of Surgical Oncology, Graduate School of Medicine, The University of TokyoNaoyaKatoDepartment of Gastroenterology, Graduate School of Medicine, Chiba UniversityMasahitoKawazuDivision of Cell Therapy, Chiba Cancer Center Research InstituteYosukeTogashiDepartment of Tumor Microenvironment, Okayama University, Graduate School of Medicine Dentistry and Pharmaceutical SciencesBackground<br>
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).<br>
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Methods<br>
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.<br>
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Results<br>
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.<br>
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Conclusions<br>
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.No potential conflict of interest relevant to this article was reported.岡山医学会Acta Medica Okayama0030-155813332021腫瘍微小環境とがん免疫療法のバイオマーカー151157ENYosukeTogashiDepartment of Tumor Microenvironment, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNo potential conflict of interest relevant to this article was reported.