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The SCRUM-Japan MONSTAR-SCREEN consortium is a nationwide molecular profiling project employing artificial intelligence-driven multi-omics analyses for patients with advanced malignancies, aiming to develop novel therapeutics and diagnostics and deliver effective drugs to patients. Concurrently, studies assessing molecular residual disease-based precision medicine for resectable solid tumors, including CIRCULATE-Japan, are ongoing. The substantial data generated by these platforms are stored within a state-of-the-art supercomputing infrastructure, VAPOR CONE. Since 2015, our project has registered over 24,000 patients as of December 2023. Among 16,144 patients with advanced solid tumors enrolled in MONSTAR-SCREEN projects, 5.0% participated in matched clinical trials, demonstrating a 29.2% objective response rate and 14.8-month median survival (95% confidence interval, 13.4-16.3), for patients treated in the matched clinical trials. Notably, patients who received matched therapy demonstrated significantly prolonged overall survival compared with those who did not (hazard ratio 0.77; 95% confidence interval, 0.71-0.83).
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In animal cells, vacuoles are absent, but can be induced by diseases and drugs. While phosphoinositides are critical for membrane trafficking, their role in the formation of these vacuoles remains unclear. The immunosuppressive KRP203/Mocravimod, which antagonizes sphingosine-1-phosphate receptors, has been identified as having novel multimodal activity against phosphoinositide kinases. However, the impact of this novel KRP203 activity is unknown. Here, we show that KRP203 disrupts the spatial organization of phosphoinositides and induces extensive vacuolization in tumor cells and immortalized fibroblasts. The KRP203-induced vacuoles are primarily from endosomes, and augmented by inhibition of PIKFYVE and VPS34. Conversely, overexpression of PTEN decreased KRP203-induced vacuole formation. Furthermore, V-ATPase inhibition completely blunted KRP203-induced vacuolization, pointing to a critical requirement of the endosomal maturation process. Importantly, nearly a half of KRP203-induced vacuoles are significantly decorated with PI4P, a phosphoinositide typically enriched at the plasma membrane and Golgi. These results suggest a model that noncanonical spatial reorganization of phosphoinositides by KRP203 alters the endosomal maturation process, leading to vacuolization. Taken together, this study reveals a previously unrecognized bioactivity of KRP203 as a vacuole-inducing agent and its unique mechanism of phosphoinositide modulation, providing a new insight of phosphoinositide regulation into vacuolization-associated diseases and their molecular pathologies.
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Endosomas , Fosfohidrolasa PTEN , Fosfatidilinositoles , Vacuolas , Vacuolas/metabolismo , Vacuolas/efectos de los fármacos , Endosomas/metabolismo , Endosomas/efectos de los fármacos , Humanos , Fosfatidilinositoles/metabolismo , Animales , Fosfohidrolasa PTEN/metabolismo , Fosfohidrolasa PTEN/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasas Clase III/metabolismo , Fosfatidilinositol 3-Quinasas Clase III/genética , Ratones , Morfolinas/farmacología , ATPasas de Translocación de Protón Vacuolares/metabolismo , ATPasas de Translocación de Protón Vacuolares/antagonistas & inhibidores , ATPasas de Translocación de Protón Vacuolares/genética , Citoplasma/metabolismo , Células HeLa , Aminopiridinas , Compuestos Heterocíclicos con 3 AnillosRESUMEN
BACKGROUND: We evaluated clinical and dosimetric outcomes of radiotherapy using two anterior oblique portals (AOP), to reduce the dose to the bilateral internal carotid arteries (CAs) and pharyngeal constrictor muscle (PCM) during early-stage glottic cancer (ESGC) treatment. METHODS: We identified patients with ESGC who underwent definitive radiotherapy between June 2014 and May 2020. RESULTS: Among the 66 patients, 32 (48%) underwent radiotherapy using AOP, and the remaining underwent typical radiotherapy using parallel opposed lateral portals (POLP). The median follow-up duration was 53 months. No significant differences were observed in the 5-year local failure (0%/9.4%), progression-free survival (90.6%/90.8%), and overall survival (90.6%/91.0%) rates between the two groups. The grade ≥2 acute mucositis incidence rate was significantly lower in the AOP group (44%/85%). Radiotherapy using AOP maintained an adequate dose coverage to the target while markedly reducing the CAs and PCM doses. CONCLUSION: Radiotherapy with AOP resulted in favorable clinical and dosimetric outcomes.
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Neoplasias Laríngeas , Radioterapia Conformacional , Radioterapia de Intensidad Modulada , Humanos , Arteria Carótida Interna , Neoplasias Laríngeas/radioterapia , Neoplasias Laríngeas/etiología , Radioterapia de Intensidad Modulada/métodos , Planificación de la Radioterapia Asistida por Computador/métodos , Radioterapia Conformacional/efectos adversos , Radioterapia Conformacional/métodos , Músculos , Dosificación RadioterapéuticaRESUMEN
BACKGROUNDS: We aimed to clarify the outcomes of postoperative radiotherapy (PORT) after salvage neck dissection for cervical lymph node (LN) recurrence in oral cavity cancer. METHODS: We retrospectively evaluated overall survival (OS), recurrence-free survival (RFS), recurrence patterns, and adverse events of 51 patients with high-risk features receiving PORT after salvage neck dissection between 2009 and 2019. RESULTS: After a median follow-up of 7.4 years from PORT initiation, the 7-year OS and RFS rates were 66.3% (95% CI: 54.0-81.3) and 54.6% (95% CI: 42.1-70.9), respectively. Age <70 years and isolated LN recurrence were significantly associated with longer OS and RFS. Among the 22 patients who experienced recurrence, 14 experienced recurrence within the radiation field. PORT-related grade 3 acute mucositis (35%) and late adverse events (osteoradionecrosis [4%] and laryngeal stenosis [2%]) were observed. CONCLUSIONS: PORT after salvage neck dissection for cervical LN recurrence achieved good survival with acceptable toxicity.
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Neoplasias de la Boca , Disección del Cuello , Humanos , Anciano , Estudios Retrospectivos , Ganglios Linfáticos/cirugía , Ganglios Linfáticos/patología , Neoplasias de la Boca/radioterapia , Neoplasias de la Boca/cirugía , Recurrencia Local de Neoplasia/radioterapia , Recurrencia Local de Neoplasia/cirugía , Recurrencia Local de Neoplasia/patología , Terapia Recuperativa , Escisión del Ganglio LinfáticoRESUMEN
Radiotherapy (RT) combined with immunotherapy is promising; however, the immune response signature in the clinical setting after RT remains unclear. Here, by integrative spatial and single-cell analyses using multiplex immunostaining (CODEX), spatial transcriptome (VISIUM), and single-cell RNA sequencing, we substantiated the infiltration of immune cells into tumors with dynamic changes in immunostimulatory and immunosuppressive gene expression after RT. In addition, our comprehensive analysis uncovered time- and cell type-dependent alterations in the gene expression profile after RT. Furthermore, myeloid cells showed prominent up-regulation of immune response-associated genes after RT. Notably, a subset of infiltrating tumor-associated myeloid cells showing PD-L1 positivity exhibited significant up-regulation of immunostimulatory (HMGB1 and ISG15), immunosuppressive (SIRPA and IDO1), and protumor genes (CXCL8, CCL3, IL-6, and IL-1AB), which can be targets of immunotherapy in combination with PD-L1. These datasets will provide information on the RT-induced gene signature to seek an appropriate target for personalized immunotherapy combined with RT and guide the timing of combination therapy.
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Carcinoma de Células Escamosas , Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Humanos , Carcinoma de Células Escamosas/patología , Antígeno B7-H1/metabolismo , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patología , Macrófagos/metabolismo , InmunosupresoresRESUMEN
Serine/threonine kinase, cell division cycle 7 (CDC7) is critical for initiating DNA replication. TAK-931 is a specific CDC7 inhibitor, which is a next-generation replication stress (RS) inducer. This study preclinically investigates TAK-931 antitumor efficacy and immunity regulation. TAK-931 induce RS, generating senescence-like aneuploid cells, which highly expressed inflammatory cytokines and chemokines (senescence-associated secretory phenotype, SASP). In vivo multilayer-omics analyses in gene expression panel, immune panel, immunohistochemistry, RNA sequencing, and single-cell RNA sequencing reveal that the RS-mediated aneuploid cells generated by TAK-931 intensively activate inflammatory-related and senescence-associated pathways, resulting in accumulation of tumor-infiltrating immune cells and potent antitumor immunity and efficacy. Finally, the combination of TAK-931 and immune checkpoint inhibitors profoundly enhance antiproliferative activities. These findings suggest that TAK-931 has therapeutic antitumor properties and improved clinical benefits in combination with conventional immunotherapy.
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Proteínas de Ciclo Celular , Neoplasias , Humanos , Proteínas de Ciclo Celular/metabolismo , Inhibidores de Puntos de Control Inmunológico , Proteínas Serina-Treonina Quinasas/metabolismo , Aneuploidia , Neoplasias/tratamiento farmacológico , Neoplasias/genéticaRESUMEN
Radiotherapy (RT) plus immunotherapy is a promising modality; however, the therapeutic effects are insufficient, and the molecular mechanism requires clarification to further develop combination therapies. Here, we found that the RNA virus sensor pathway dominantly regulates the cellular immune response in NSCLC and ESCC cell lines. Notably, transposable elements (TEs), especially long terminal repeats (LTRs), functioned as key ligands for the RNA virus sensor RIG-I, and the mTOR-LTR-RIG-I axis induced the cellular immune response and dendritic cell and macrophage infiltration after irradiation. Moreover, RIG-I-dependent immune activation was observed in ESCC patient tissue. scRNA sequencing and spatial transcriptome analysis revealed that radiotherapy induced the expression of LTRs, and the RNA virus sensor pathway in immune and cancer cells; this pathway was also found to mediate tumour conversion to an immunological hot state. Here, we report the upstream and ligand of the RNA virus sensor pathway functions in irradiated cancer tissues.
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Elementos Transponibles de ADN , Macrófagos , Humanos , Línea Celular , Proteína 58 DEAD Box/genética , Proteína 58 DEAD Box/metabolismo , Macrófagos/metabolismoRESUMEN
The prognosis and prognostic factors of patients receiving whole-brain radiotherapy (WBRT) for leptomeningeal metastasis (LM) from lung adenocarcinoma have not been established. Particularly, the impact of EGFR mutations and ALK rearrangements on survival remains unclear. This retrospective study evaluated the prognosis and prognostic factors of patients receiving WBRT for LM. We evaluated overall survival (OS) from WBRT initiation and clinical variables in 80 consecutive patients receiving WBRT for LM from lung adenocarcinoma at our institution between June 2013 and June 2021. After a median follow-up of 5.2 (range 0.5-56.5) months, the median OS was 6.2 months (95% CI 4.4-12.4). Of the 80 patients, 51 were classified as EGFR/ALK mutant (EGFR: 44; ALK: 6; both: 1) and 29 as wild-type. The median OS was 10.4 (95% CI 5.9-20.9) versus 3.8 (95% CI 2.5-7.7) months in the EGFR/ALK-mutant versus wild-type patients (HR = 0.49, P = 0.0063). Multivariate analysis indicated that EGFR/ALK alterations (HR = 0.54, P = 0.021) and Eastern Cooperative Oncology Group performance status (ECOG PS) of 0-1 (HR = 0.25, P < 0.001) were independent factors associated with favorable OS. Among the patients who underwent brain MRI before and after WBRT, intracranial progression-free survival was longer in the 26 EGFR/ALK-mutant than 13 wild-type patients (HR = 0.31, P = 0.0039). Although the prognosis of patients receiving WBRT for LM remains poor, EGFR/ALK alterations and good ECOG PS may positively impact OS in those eligible for WBRT.
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Adenocarcinoma del Pulmón , Neoplasias Encefálicas , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Carcinomatosis Meníngea , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/radioterapia , Neoplasias Pulmonares/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Pronóstico , Estudios Retrospectivos , Receptores ErbB/genética , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/radioterapia , Adenocarcinoma del Pulmón/tratamiento farmacológico , Carcinomatosis Meníngea/genética , Carcinomatosis Meníngea/radioterapia , Mutación , Encéfalo/patología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/radioterapia , Inhibidores de Proteínas Quinasas/uso terapéuticoRESUMEN
NRF2 is a transcription factor responsible for antioxidant stress responses that is usually regulated in a redox-dependent manner. p62 bodies formed by liquid-liquid phase separation contain Ser349-phosphorylated p62, which participates in the redox-independent activation of NRF2. However, the regulatory mechanism and physiological significance of p62 phosphorylation remain unclear. Here, we identify ULK1 as a kinase responsible for the phosphorylation of p62. ULK1 colocalizes with p62 bodies, directly interacting with p62. ULK1-dependent phosphorylation of p62 allows KEAP1 to be retained within p62 bodies, thus activating NRF2. p62S351E/+ mice are phosphomimetic knock-in mice in which Ser351, corresponding to human Ser349, is replaced by Glu. These mice, but not their phosphodefective p62S351A/S351A counterparts, exhibit NRF2 hyperactivation and growth retardation. This retardation is caused by malnutrition and dehydration due to obstruction of the esophagus and forestomach secondary to hyperkeratosis, a phenotype also observed in systemic Keap1-knockout mice. Our results expand our understanding of the physiological importance of the redox-independent NRF2 activation pathway and provide new insights into the role of phase separation in this process.
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Factor 2 Relacionado con NF-E2 , Estrés Oxidativo , Humanos , Animales , Ratones , Proteína 1 Asociada A ECH Tipo Kelch/genética , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Fosforilación , Proteína Sequestosoma-1/genética , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Oxidación-Reducción , Autofagia/fisiología , Homólogo de la Proteína 1 Relacionada con la Autofagia/genética , Homólogo de la Proteína 1 Relacionada con la Autofagia/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismoRESUMEN
In addition to membranous organelles, autophagy selectively degrades biomolecular condensates, in particular p62/SQSTM1 bodies, to prevent diseases including cancer. Evidence is growing regarding the mechanisms by which autophagy degrades p62 bodies, but little is known about their constituents. Here, we established a fluorescence-activated-particle-sorting-based purification method for p62 bodies using human cell lines and determined their constituents by mass spectrometry. Combined with mass spectrometry of selective-autophagy-defective mouse tissues, we identified vault, a large supramolecular complex, as a cargo within p62 bodies. Mechanistically, major vault protein directly interacts with NBR1, a p62-interacting protein, to recruit vault into p62 bodies for efficient degradation. This process, named vault-phagy, regulates homeostatic vault levels in vivo, and its impairment may be associated with non-alcoholic-steatohepatitis-derived hepatocellular carcinoma. Our study provides an approach to identifying phase-separation-mediated selective autophagy cargoes, expanding our understanding of the role of phase separation in proteostasis.
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Neoplasias Hepáticas , Proteómica , Animales , Humanos , Ratones , Proteína Sequestosoma-1/metabolismo , Autofagia , Orgánulos/metabolismoAsunto(s)
Carcinoma de Pulmón de Células no Pequeñas , ADN Tumoral Circulante , Neoplasias Pulmonares , Terapia de Protones , Carcinoma Pulmonar de Células Pequeñas , Humanos , Carcinoma de Pulmón de Células no Pequeñas/radioterapia , Carcinoma de Pulmón de Células no Pequeñas/patología , Neoplasias Pulmonares/radioterapia , Neoplasias Pulmonares/patología , Carcinoma Pulmonar de Células Pequeñas/patología , Recurrencia Local de Neoplasia/patología , Estadificación de NeoplasiasRESUMEN
Human cathelicidin LL-37 is a multifunctional antimicrobial peptide that exhibits antimicrobial and immunomodulatory activities. LL-37 regulates skin barrier function and was recently reported to activate autophagy in macrophages. Because autophagy deficiency is associated with skin diseases characterized by a dysfunctional epidermal barrier, we hypothesized that LL-37 might regulate the skin barrier through autophagy modulation. We showed that LL-37 activated autophagy in human keratinocytes and three-dimensional skin equivalent models as indicated by increases in LC3 puncta formation, decreases in p62, and autophagosome and autolysosome formation. LL-37âinduced autophagy was suppressed by P2X7 receptor, adenosine monophosphateâactivated protein kinase, and unc-51-like kinase 1 inhibitors, suggesting that the P2X7, adenosine monophosphateâactivated protein kinase, and unc-51-like kinase 1 pathways are involved. Moreover, LL-37 enhanced the phosphorylation of adenosine monophosphateâactivated protein kinase and unc-51-like kinase 1. In addition, LL-37âmediated autophagy involves the mechanistic target of rapamycin and MAPK pathways. Interestingly, the LL-37âinduced distribution of tight junction proteins and improvement in the tight junction barrier were inhibited in autophagy-deficient keratinocytes and keratinocytes and skin models treated with autophagy inhibitors, indicating that the LL-37âmediated tight junction barrier is associated with autophagy activation. Collectively, these findings suggest that LL-37 is a potential therapeutic target for skin diseases characterized by dysfunctional autophagy and skin barriers.
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Péptidos Catiónicos Antimicrobianos , Catelicidinas , Humanos , Adenosina Monofosfato/metabolismo , Péptidos Catiónicos Antimicrobianos/farmacología , Péptidos Catiónicos Antimicrobianos/metabolismo , Autofagia , Homólogo de la Proteína 1 Relacionada con la Autofagia/metabolismo , Catelicidinas/farmacología , Catelicidinas/metabolismo , Queratinocitos/metabolismo , Sirolimus , Transducción de SeñalRESUMEN
Colorectal cancer (CRC) is one of the most common malignant diseases. Generally, stoma construction is performed following surgery for the resection of the primary tumor in patients with CRC. The association of CRC with the gut microbiota has been widely reported, and the gut microbiota is known to play an important role in the carcinogenesis, progression, and treatment of CRC. In this study, we compared the microbiota of patients with CRC between with and without a stoma using fecal metagenomic sequencing data from SCRUM-Japan MONSTAR-SCREEN, a joint industry-academia cancer research project in Japan. We found that the composition of anaerobes was reduced in patients with a stoma. In particular, the abundance of Alistipes, Akkermansia, Intestinimonas, and methane-producing archaea decreased. We also compared gene function (e.g., KEGG Orthology and KEGG pathway) and found that gene function for methane and short-chain fatty acids (SCFAs) production was underrepresented in patients with a stoma. Furthermore, a stoma decreased Shannon diversity based on taxonomic composition but increased that of the KEGG pathway. These results suggest that the feces of patients with a stoma have a reduced abundance of favorable microbes for cancer immunotherapy. In conclusion, we showed that a stoma alters the taxonomic and functional profiles in feces and may be a confounding factor in fecal microbiota analysis.
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Neoplasias Colorrectales , Microbiota , Bacterias Anaerobias/genética , Bacterias Anaerobias/metabolismo , Neoplasias Colorrectales/microbiología , Neoplasias Colorrectales/cirugía , Ácidos Grasos Volátiles/metabolismo , Heces/microbiología , Humanos , Metano , ARN Ribosómico 16S/genéticaRESUMEN
Human ß-defensin-3 (hBD-3) exhibits antimicrobial and immunomodulatory activities; however, its contribution to autophagy regulation remains unclear, and the role of autophagy in the regulation of the epidermal barrier in atopic dermatitis (AD) is poorly understood. Here, keratinocyte autophagy was restrained in the skin lesions of patients with AD and murine models of AD. Interestingly, hBD-3 alleviated the IL-4- and IL-13-mediated impairment of the tight junction (TJ) barrier through keratinocyte autophagy activation, which involved aryl hydrocarbon receptor (AhR) signaling. While autophagy deficiency impaired the epidermal barrier and exacerbated inflammation, hBD-3 attenuated skin inflammation and enhanced the TJ barrier in AD. Importantly, hBD-3-mediated improvement of the TJ barrier was abolished in autophagy-deficient AD mice and in AhR-suppressed AD mice, suggesting a role for hBD-3-mediated autophagy in the regulation of the epidermal barrier and inflammation in AD. Thus, autophagy contributes to the pathogenesis of AD, and hBD-3 could be used for therapeutic purposes.
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Dermatitis Atópica , beta-Defensinas , Animales , Autofagia , Dermatitis Atópica/tratamiento farmacológico , Dermatitis Atópica/genética , Humanos , Inflamación/genética , Inflamación/metabolismo , Queratinocitos/patología , Ratones , Receptores de Hidrocarburo de Aril/genética , Receptores de Hidrocarburo de Aril/metabolismo , Transducción de Señal , beta-Defensinas/genética , beta-Defensinas/metabolismo , beta-Defensinas/uso terapéuticoRESUMEN
Radiotherapy (RT) combined with immune checkpoint inhibitors has recently produced outstanding results and is expected to be adaptable for various cancers. However, the precise molecular mechanism by which immune reactions are induced by fractionated RT is still controversial. We aimed to investigate the mechanism of the immune response regarding multifractionated, long-term radiation, which is most often combined with immunotherapy. Two human esophageal cancer cell lines, KYSE-450 and OE-21, were irradiated by fractionated irradiation (FIR) daily at a dose of 3 Gy in 5 d/wk for 2 weeks. Western blot analysis and RNA sequencing identified type I interferon (IFN) and the stimulator of IFN genes (STING) pathway as candidates that regulate immune response by FIR. We inhibited STING, IFNAR1, STAT1, and IFN regulatory factor 1 (IRF1) and investigated the effects on the immune response in cancer cells and the invasion of surrounding immune cells. We herein revealed type I IFN-dependent immune reactions and the positive feedback of STING, IRF1, and phosphorylated STAT1 induced by FIR. Knocking out STING, IFNAR1, STAT1, and IRF1 resulted in a poorer immunological response than that in WT cells. The STING-KO KYSE-450 cell line showed significantly less invasion of PBMCs than the WT cell line under FIR. In the analysis of STING-KO cells and migrated PBMCs, we confirmed the occurrence of STING-dependent immune activation under FIR. In conclusion, we identified that the STING-IFNAR1-STAT1-IRF1 axis regulates immune reactions in cancer cells triggered by FIR and that the STING pathway also contributes to immune cell invasion of cancer cells.
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Neoplasias Esofágicas , Inmunidad , Factor 1 Regulador del Interferón , Factor de Transcripción STAT1 , Línea Celular/efectos de la radiación , Neoplasias Esofágicas/genética , Humanos , Inmunidad/efectos de la radiación , Factor 1 Regulador del Interferón/genética , Factor 1 Regulador del Interferón/metabolismo , Factor 1 Regulador del Interferón/efectos de la radiación , Interferón Tipo I , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/efectos de la radiación , Receptor de Interferón alfa y beta/genética , Receptor de Interferón alfa y beta/metabolismo , Receptor de Interferón alfa y beta/efectos de la radiación , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT1/metabolismo , Factor de Transcripción STAT1/efectos de la radiaciónRESUMEN
BACKGROUND: Combination therapy based on radiotherapy and immune checkpoint inhibitors (ICIs) was recently reported as effective for various cancers. The radiation-induced immune response (RIIR) is an essential feature in ICI-combined radiotherapy; however, the effects of drugs used concomitantly with RIIR remain unclear. We screened for drugs that can modify RIIR to understand the mutual relationship between radiotherapy and combined drugs in ICI-combined radiotherapy. METHODS: We established a high-throughput system with reporter gene assays for evaluating RIIR, focusing on factors acting downstream of the STING-IRF pathway, which can stimulate cancer cells, T cells, and dendritic cells. We further quantified the effects of 2595 drugs, including those approved by the Food and Drug Administration, on RIIR in vitro. RESULTS: The reporter assay results correlated well with the expression of immune response proteins such as programmed death-ligand 1. This high-throughput system enabled the identification of drugs including cytotoxic agents, molecular-targeted agents, and other agents that activate or suppress RIIR. CONCLUSIONS: Our study provides an encyclopedic catalogue of clinically approved drugs based on their effect on RIIR. In ICIs combined radiotherapy, activation of STING-IFN may improve the therapeutic effect and our result could form a biological basis for further clinical trials combining radiotherapy with ICIs.
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Anticuerpos Monoclonales , Neoplasias , Anticuerpos Monoclonales/uso terapéutico , Humanos , Inhibidores de Puntos de Control Inmunológico , Inmunidad , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/radioterapia , Preparaciones FarmacéuticasRESUMEN
A germ line copy number duplication of chromosome 14q32, which contains ATG2B and GSKIP, was identified in families with myeloproliferative neoplasm (MPN). Here, we show that mice lacking both Atg2b and Gskip, but not either alone, exhibited decreased hematopoiesis, resulting in death in utero accompanied by anemia. In marked contrast to MPN patients with duplication of ATG2B and GSKIP, the number of hematopoietic stem cells (HSCs), in particular long-term HSCs, in double-knockout fetal livers was significantly decreased due to increased cell death. Although the remaining HSCs still had the ability to differentiate into hematopoietic progenitor cells, the differentiation efficiency was quite low. Remarkably, mice with knockout of Atg2b or Gskip alone did not show any hematopoietic abnormality. Mechanistically, while loss of both genes had no effect on autophagy, it increased the expression of genes encoding enzymes involved in oxidative phosphorylation. Taken together, our results indicate that Atg2b and Gskip play a synergistic effect in maintaining the pool size of HSCs.
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Proteínas Relacionadas con la Autofagia/genética , Hematopoyesis/genética , Células Madre Hematopoyéticas/metabolismo , Proteínas Represoras/genética , Proteínas de Transporte Vesicular/genética , Animales , Autofagia/fisiología , Proteínas Relacionadas con la Autofagia/metabolismo , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Cromosomas/genética , Hematopoyesis/fisiología , Ratones , Proteínas Represoras/metabolismo , Proteínas de Transporte Vesicular/metabolismoRESUMEN
Radiotherapy (RT) is an effective treatment option for cancer; however, its efficacy remains less than optimal in locally advanced cancer. Immune checkpoint inhibitor-based therapy, including the administration of anti-PD-L1 antibodies, is a promising approach that works synergistically with RT. Proton beam therapy and carbon-ion therapy are common options for patients with cancer. Proton and carbon ions are reported to induce an immune reaction in cancer cells; however, the underlying mechanisms remain unclear. Here, we aimed to compare the immune responses after irradiation (IR) with X-ray, protons, and carbon ions in an oesophageal cancer cell line and the underlying mechanisms. An oesophageal cancer cell line, KYSE450, was irradiated with 1 fraction/15 GyE (Gy equivalent) of X-ray, proton, or carbon-ion beams, and then, the cells were harvested for RNA sequencing and gene enrichment analysis. We also knocked out STING and STAT1 in the quest for mechanistic insights. RNA sequencing data revealed that gene expression signatures and biological processes were different in KYSE450 irradiated with X-ray, proton, and carbon-ion beams 6-24 h after IR. However, after 3 days, a common gene expression signature was detected, associated with biological pathways involved in innate immune responses. Gene knock-out experiments revealed that the STING-STAT1 axis underlies the immune reactions after IR. X-Ray, proton, and carbon-ion IRs induced similar immune responses, regulated by the STING-STAT1 axis.
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Carbono , Neoplasias Esofágicas/genética , Regulación Neoplásica de la Expresión Génica/efectos de la radiación , Inmunidad/efectos de la radiación , Protones , Transcriptoma/efectos de la radiación , Rayos X , Línea Celular Tumoral , Neoplasias Esofágicas/inmunología , Neoplasias Esofágicas/patología , Perfilación de la Expresión Génica/métodos , Regulación Neoplásica de la Expresión Génica/inmunología , Ontología de Genes , Humanos , Inmunidad/genética , Iones , RNA-Seq/métodos , Radiación/clasificación , Transducción de Señal/genética , Transducción de Señal/inmunología , Transducción de Señal/efectos de la radiación , Transcriptoma/inmunologíaRESUMEN
Several amyotrophic lateral sclerosis (ALS)-related proteins such as FUS, TDP-43, and hnRNPA1 demonstrate liquid-liquid phase separation, and their disease-related mutations correlate with a transition of their liquid droplet form into aggregates. Missense mutations in SQSTM1/p62, which have been identified throughout the gene, are associated with ALS, frontotemporal degeneration (FTD), and Paget's disease of bone. SQSTM1/p62 protein forms liquid droplets through interaction with ubiquitinated proteins, and these droplets serve as a platform for autophagosome formation and the antioxidative stress response via the LC3-interacting region (LIR) and KEAP1-interacting region (KIR) of p62, respectively. However, it remains unclear whether ALS/FTD-related p62 mutations in the LIR and KIR disrupt liquid droplet formation leading to defects in autophagy, the stress response, or both. To evaluate the effects of ALS/FTD-related p62 mutations in the LIR and KIR on a major oxidative stress system, the Keap1-Nrf2 pathway, as well as on autophagic turnover, we developed systems to monitor each of these with high sensitivity. These methods such as intracellular protein-protein interaction assay, doxycycline-inducible gene expression system, and gene expression into primary cultured cells with recombinant adenovirus revealed that some mutants, but not all, caused reduced NRF2 activation and delayed autophagic cargo turnover. In contrast, while all p62 mutants demonstrated sufficient ability to form liquid droplets, all of these droplets also exhibited reduced inner fluidity. These results indicate that like other ALS-related mutant proteins, p62 missense mutations result in a primary defect in ALS/FTD via a qualitative change in p62 liquid droplet fluidity.
Asunto(s)
Esclerosis Amiotrófica Lateral/metabolismo , Proteína Sequestosoma-1/metabolismo , Esclerosis Amiotrófica Lateral/genética , Animales , Células HEK293 , Humanos , Proteína 1 Asociada A ECH Tipo Kelch/genética , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Ratones , Mutación Missense , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Proteína Sequestosoma-1/genéticaRESUMEN
Lung cancer constitutes a threat to human health. BHLHE41 plays important roles in circadian rhythm and cell differentiation as a negative regulatory transcription factor. This study investigates the role of BHLHE41 in lung cancer progression. We analyzed BHLHE41 function via in silico and immunohistochemical studies of 177 surgically resected non-small cell lung cancer (NSCLC) samples and 18 early lung squamous cell carcinoma (LUSC) cases. We also examined doxycycline (DOX)-inducible BHLHE41-expressing A549 and H2030 adenocarcinoma cells. BHLHE41 expression was higher in normal lung than in lung adenocarcinoma (LUAD) tissues and was associated with better prognosis for the overall survival (OS) of patients. In total, 15 of 132 LUAD tissues expressed BHLHE41 in normal lung epithelial cells. Staining was mainly observed in adenocarcinoma in situ and the lepidic growth part of invasive cancer tissue. BHLHE41 expression constituted a favorable prognostic factor for OS (p = 0.049) and cause-specific survival (p = 0.042) in patients with LUAD. During early LUSC, 7 of 18 cases expressed BHLHE41, and this expression was inversely correlated with the depth of invasion. DOX suppressed cell proliferation and increased the autophagy protein LC3, while chloroquine enhanced LC3 accumulation and suppressed cell death. In a xenograft model, DOX suppressed tumor growth. Our results indicate that BHLHE41 expression prevents early lung tumor malignant progression by inducing autophagic cell death in NSCLC.