Clonal Origin and Lineage Ambiguity in Mixed Neuroendocrine Carcinoma of the Uterine Cervix.

Small-cell neuroendocrine carcinoma (SCNEC) of the cervix is a rare disease characterized by a high incidence of mixed tumors with other types of cancer. The mechanism underlying this mixed phenotype is not well understood. This study established a panel of organoid lines from patients with SCNEC of the cervix and ultimately focused on one line, which retained a mixed tumor phenotype, both in vitro and in vivo. Histologically, both organoids and xenograft tumors showed distinct differentiation into either SCNEC or adenocarcinoma in some regions and ambiguous differentiation in others. Tracking single cells indicated the existence of cells with bipotential differentiation toward SCNEC and adenocarcinomas. Single-cell transcriptional analysis identified three distinct clusters: SCNEC-like, adenocarcinoma-like, and a cluster lacking specific differentiation markers. The expression of neuroendocrine markers was enriched in the SCNEC-like cluster but not exclusively. Human papillomavirus 18 E6 was enriched in the SCNEC-like cluster, which showed higher proliferation and lower levels of the p53 pathway. After treatment with anticancer drugs, the expression of adenocarcinoma markers increased, whereas that of SCNEC decreased. Using a reporter system for keratin 19 expression, changes in the differentiation of each cell were shown to be associated with the shift in differentiation induced by drug treatment. These data suggest that mixed SCNEC/cervical tumors have a clonal origin and are characterized by an ambiguous and flexible differentiation state.

The hepatic niche leads to aggressive natural killer cell leukemia proliferation through the transferrin-transferrin receptor 1 axis.

Aggressive natural killer cell leukemia (ANKL) is a rare lymphoid neoplasm frequently associated with Epstein-Barr virus, with a disastrously poor prognosis. Owing to the lack of samples from patients with ANKL and relevant murine models, comprehensive investigation of its pathogenesis including the tumor microenvironment (TME) has been hindered. Here we established 3 xenograft mice derived from patients with ANKL (PDXs), which enabled extensive analysis of tumor cells and their TME. ANKL cells primarily engrafted and proliferated in the hepatic sinusoid. Hepatic ANKL cells were characterized by an enriched Myc-pathway and proliferated faster than those in other organs. Interactome analyses and in vivo CRISPR-Cas9 analyses revealed transferrin (Tf)-transferrin receptor 1 (TfR1) axis as a potential molecular interaction between the liver and ANKL. ANKL cells were rather vulnerable to iron deprivation. PPMX-T003, a humanized anti-TfR1 monoclonal antibody, showed remarkable therapeutic efficacy in a preclinical setting using ANKL-PDXs. These findings indicate that the liver, a noncanonical hematopoietic organ in adults, serves as a principal niche for ANKL and the inhibition of the Tf-TfR1 axis is a promising therapeutic strategy for ANKL.

MCAM+CD161- Th17 Subset Expressing CD83 Enhances Tc17 Response in Psoriasis.

Recent studies have highlighted the pathogenic roles of IL-17-producing CD8+ T cells (T-cytotoxic 17 [Tc17]) in psoriasis. However, the underlying mechanisms of Tc17 induction remain unclear. In this study, we focused on the pathogenic subsets of Th17 and their mechanism of promotion of Tc17 responses. We determined that the pathogenic Th17-enriched fraction expressed melanoma cell adhesion molecule (MCAM) and CCR6, but not CD161, because this subset produced IL-17A abundantly and the presence of these cells in the peripheral blood of patients has been correlated with the severity of psoriasis. Intriguingly, the serial analysis of gene expression revealed that CCR6+MCAM+CD161-CD4+ T cells displayed the gene profile for adaptive immune responses, including CD83, which is an activator for CD8+ T cells. Coculture assay with or without intercellular contact between CD4+ and CD8+ T cells showed that CCR6+MCAM+CD161-CD4+ T cells induced the proliferation of CD8+ T cells in a CD83-dependent manner. However, the production of IL-17A by CD8+ T cells required exogenous IL-17A, suggesting that intercellular contact via CD83 and the production of IL-17A from activated CD4+ T cells elicit Tc17 responses. Intriguingly, the CD83 expression was enhanced in the presence of IL-15, and CD83+ cells stimulated with IL-1ß, IL-23, IL-15, and IL-15Rα did not express FOXP3. Furthermore, CCR6+MCAM+CD161-CD4+ T cells expressing CD83 were increased in the peripheral blood of patients, and the CD83+ Th17-type cells accumulated in the lesional skin of psoriasis. In conclusion, pathogenic MCAM+CD161- Th17 cells may be involved in the Tc17 responses via IL-17A and CD83 in psoriasis.

Inhibition of transforming growth factor-ß signals suppresses tumor formation by regulation of tumor microenvironment networks.

The tumor microenvironment (TME) consists of cancer cells surrounded by stromal components including tumor vessels. Transforming growth factor-ß (TGF-ß) promotes tumor progression by inducing epithelial-mesenchymal transition (EMT) in cancer cells and stimulating tumor angiogenesis in the tumor stroma. We previously developed an Fc chimeric TGF-ß receptor containing both TGF-ß type I (TßRI) and type II (TßRII) receptors (TßRI-TßRII-Fc), which trapped all TGF-ß isoforms and suppressed tumor growth. However, the precise mechanisms underlying this action have not yet been elucidated. In the present study, we showed that the recombinant TßRI-TßRII-Fc protein effectively suppressed in vitro EMT of oral cancer cells and in vivo tumor growth in a human oral cancer cell xenograft mouse model. Tumor cell proliferation and angiogenesis were suppressed in tumors treated with TßRI-TßRII-Fc. Molecular profiling of human cancer cells and mouse stroma revealed that K-Ras signaling and angiogenesis were suppressed. Administration of TßRI-TßRII-Fc protein decreased the expression of heparin-binding epidermal growth factor-like growth factor (HB-EGF), interleukin-1ß (IL-1ß) and epiregulin (EREG) in the TME of oral cancer tumor xenografts. HB-EGF increased proliferation of human oral cancer cells and mouse endothelial cells by activating ERK1/2 phosphorylation. HB-EGF also promoted oral cancer cell-derived tumor formation by enhancing cancer cell proliferation and tumor angiogenesis. In addition, increased expressions of IL-1ß and EREG in oral cancer cells significantly enhanced tumor formation. These results suggest that TGF-ß signaling in the TME controls cancer cell proliferation and angiogenesis by activating HB-EGF/IL-1ß/EREG pathways and that TßRI-TßRII-Fc protein is a promising tool for targeting the TME networks.

Fusion with type 2 macrophages induces melanoma cell heterogeneity that potentiates immunological escape from cytotoxic T lymphocytes.

Evasion from immunity is a major obstacle to the achievement of successful cancer immunotherapy. Hybrids derived from cell-cell fusion are theoretically associated with tumor heterogeneity and progression by conferring novel properties on tumor cells, including drug resistance and metastatic capacity; however, their impact on immune evasion remains unknown. Here, we investigated the potency of tumor-macrophage hybrids in immune evasion. Hybrids were established by co-culture of a melanoma cell line (A375 cells) and type 2 macrophages. The hybrids showed greater migration ability and greater tumorigenicity than the parental melanoma cells. The hybrids showed heterogeneous sensitivity to New York esophageal squamous cell carcinoma-1 (NY-ESO-1)-specific T-cell receptor-transduced T (TCR-T) cells and two out of four hybrid clones showed less sensitivity to TCR-T compared with the parental cells. An in vitro tumor heterogeneity model revealed that the TCR-T cells preferentially killed the parental cells compared with the hybrids and the survival rate of the hybrids was higher than that of the parental cells, indicating that the hybrids evade killing by TCR-T cells efficiently. Analysis of a single-cell RNA sequencing dataset of patients with melanoma revealed that a few macrophages expressed RNA encoding melanoma differentiation antigens including melan A, tyrosinase, and premelanosome protein, which indicated the presence of hybrids in primary melanoma. In addition, the number of potential hybrids was correlated with a poorer response to immune checkpoint blockade. These results provide evidence that melanoma-macrophage fusion has a role in tumor heterogeneity and immune evasion. © 2023 The Pathological Society of Great Britain and Ireland.

Nationwide multicenter study on adverse events associated with a patency capsule: Additional survey of appropriate use of patency capsule study.

BACKGROUND AND AIM: The PillCam patency capsule (PC) without a radio frequency identification tag was released to preclude retention of the small bowel capsule endoscope (CE) in Japan in 2012. We conducted a multicenter study to determine tag-less PC-related adverse events (AEs). METHODS: We first conducted a retrospective survey using a standardized data collection sheet for the clinical characteristics of PC-related AEs among 1096 patients collected in a prospective survey conducted between January 2013 and May 2014 (Cohort 1). Next, we retrospectively investigated additional AEs that occurred before and after Cohort 1 within the period June 2012 and December 2014 among 1482 patients (Cohort 2). RESULTS: Of the 2578 patients who underwent PC examinations from both cohorts, 74 AEs occurred among 61 patients (2.37%). The main AEs were residual parylene coating in 25 events (0.97%), PC-induced small bowel obstruction, suspicious of impaction, in 23 events (0.89%), and CE retention even after patency confirmation in 10 events (0.39%). Residual parylene coating was significantly associated with Crohn's disease (P < 0.01). Small bowel obstruction was significantly associated with physicians with less than 1 year of experience handling the PC and previous history of postprandial abdominal pain (P < 0.01 and P < 0.03, respectively). CE retention was ascribed to erroneous judgment of PC localization in all cases. CONCLUSIONS: This large-scale multicenter study provides evidence supporting the safety and efficiency of a PC to preclude CE retention. Accurate PC localization in patients without excretion and confirmation of previous history of postprandial abdominal pain before PC examinations is warranted (UMIN000010513).

Treatment of Inflammatory Bowel Disease with Biologics in Japan: A Single-Center, Retrospective Pharmacoeconomic Study.

Biologics are essential for treating inflammatory bowel disease (IBD); however, only a few studies have validated cost-effective treatment options and patient factors for biologic use using real-world data from Japanese patients with IBD. Here, we aimed to provide pharmacoeconomic evidence to support clinical decisions for IBD treatment using biologics. We assessed 183 cases (127 patients) of IBD treated with biologics between November 2004 and September 2021. Data on patient background, treatment other than biologics, treatment-related medical costs, and effectiveness index (ratio of the C-reactive protein-negative period to drug survival time) were analyzed using univariate and multivariate logistic regression analyses. Drug survival was determined using Kaplan-Meier survival curve analysis. The outcomes were to validate a novel assessment index and elucidate the following aspects using this index: the effectiveness-cost relationship of long-term biologic use in IBD and cost-effectiveness-associated patient factors. Body mass index ≥25 kg/m2 and duration of hypoalbuminemia during drug survival correlated significantly with the therapeutic effectiveness of biologics. There were no significant differences in surgical, granulocyte apheresis, or adverse-event costs per drug survival time. Biologic costs were significantly higher in the group showing lower effectiveness than in the group showing higher effectiveness. These findings hold major pharmacoeconomic implications for not only improving therapeutic outcomes through the amelioration of low albumin levels and obesity but also potentially reducing healthcare expenditure related to the use of biotherapeutics. To our knowledge, this is the first pharmacoeconomic study based on real-world data from Japanese patients with IBD receiving long-term biologic therapy.

Eribulin is an immune potentiator in breast cancer that upregulates human leukocyte antigen class I expression via the induction of NOD-like receptor family CARD domain-containing 5.

Eribulin inhibits microtubule polymerization and improves the overall survival of patients with recurrent metastatic breast cancer. A subgroup analysis revealed a low neutrophil to lymphocyte ratio (NLR) (<3) to be a prognostic factor of eribulin treatment. We thus hypothesized that eribulin might be related to the immune response for breast cancer cells and we analyzed the effects of eribulin on the immune system. Immunohistochemical staining revealed that human leukocyte antigen (HLA) class I expression was increased in clinical samples after eribulin treatment. In vitro assays revealed that eribulin treatment increased HLA class I expression in breast cancer line cells. RNA-sequencing demonstrated that eribulin treatment increased the expression of the NOD-like family CARD domain-containing 5 (NLRC5), a master regulator of HLA class I expression. Eribulin treatment increased the NY-ESO-1-specific T-cell receptor (TCR) transduced T (TCR-T) cell response for New York oesophageal squamous cell carcinoma 1 (NY-ESO-1) overexpressed breast cancer cells. The eribulin and TCR-T combined therapy model revealed that eribulin and immunotherapy using TCR-T cells has a synergistic effect. In summary, eribulin increases the expression of HLA class 1 via HLA class 1 transactivatior NLRC5 and eribulin combination with immunotherapy can be effective for the treatment of breast cancer.

Cisplatin resistance driver claspin is a target for immunotherapy in urothelial carcinoma.

Bladder cancer is a major and fatal urological disease. Cisplatin is a key drug for the treatment of bladder cancer, especially in muscle-invasive cases. In most cases of bladder cancer, cisplatin is effective; however, resistance to cisplatin has a significant negative impact on prognosis. Thus, a treatment strategy for cisplatin-resistant bladder cancer is essential to improve the prognosis. In this study, we established a cisplatin-resistant (CR) bladder cancer cell line using an urothelial carcinoma cell lines (UM-UC-3 and J82). We screened for potential targets in CR cells and found that claspin (CLSPN) was overexpressed. CLSPN mRNA knockdown revealed that CLSPN had a role in cisplatin resistance in CR cells. In our previous study, we identified human leukocyte antigen (HLA)-A*02:01-restricted CLSPN peptide by HLA ligandome analysis. Thus, we generated a CLSPN peptide-specific cytotoxic T lymphocyte clone that recognized CR cells at a higher level than wild-type UM-UC-3 cells. These findings indicate that CLSPN is a driver of cisplatin resistance and CLSPN peptide-specific immunotherapy may be effective for cisplatin-resistant cases.

Cooperation between artificial intelligence and endoscopists for diagnosing invasion depth of early gastric cancer.

BACKGROUND AND STUDY AIMS: The diagnostic ability of endoscopists to determine invasion depth of early gastric cancer is not favorable. We designed an artificial intelligence (AI) classifier for differentiating intramucosal and submucosal gastric cancers and examined it to establish a diagnostic method based on cooperation between AI and endoscopists. PATIENTS AND METHODS: We prepared 500 training images using cases of mainly depressed-type early gastric cancer from 250 intramucosal cancers and 250 submucosal cancers. We also prepared 200 test images each of 100 cancers from another institution. We designed an AI classifier to differentiate between intramucosal and submucosal cancers by deep learning. We examined the performance of the AI classifier and the majority vote of the endoscopists as high confidence and low confidence diagnostic probability, respectively, and cooperatively combined them to establish a diagnostic method providing high accuracy. RESULTS: Internal evaluation of the training images showed that accuracy, sensitivity, specificity, and F1 measure by the AI classifier were 77%, 76%, 78%, and 0.768, and those of the majority vote of the endoscopists were 72.6%, 53.6%, 91.6%, and 0.662, respectively. A diagnostic method based on cooperation between AI and the endoscopists showed that the respective values were 78.0%, 76.0%, 80.0%, and 0.776 for the test images. The value of F1 measure was especially higher than those by AI or the endoscopists alone. CONCLUSIONS: Cooperation between AI and endoscopists improved the diagnostic ability to determine invasion depth of early gastric cancer.

Lactoferrin and its digestive peptides induce interferon-α production and activate plasmacytoid dendritic cells ex vivo.

Plasmacytoid dendritic cells (pDCs) recognise viral single-stranded RNA (ssRNA) or CpG DNA via Toll-like receptor (TLR)-7 and TLR9, and produce interferon (IFN)-α. Activated pDCs upregulate human leukocyte antigen (HLA)-DR and CD86 expression levels. Ingestion of bovine lactoferrin (LF) activates pDCs, but little is known about its effects. In this study, the effects of LF and its pepsin hydrolysate (LFH) on the production of IFN-α from peripheral blood mononuclear cells (PBMCs) and pDCs were examined. PBMCs were prepared from peripheral blood of healthy adults and incubated with LF, LFH, or lactoferricin (LFcin) in the absence or presence of ssRNA derived from human immunodeficiency virus. The concentration of IFN-α in the supernatant and the expression levels of IFN-α, HLA-DR, and CD86 in pDCs were quantified by enzyme-linked immunosorbent assay and flow cytometry. In the absence of ssRNA, the concentration of IFN-α was negligible and LF had no effect on it. In the presence of ssRNA, IFN-α was detected at a certain level, and LF and LFH significantly increased its concentration. The increase caused by LFH and LFcin were comparable. In addition, LF significantly upregulated the expression levels of IFN-α, HLA-DR, and CD86 in pDCs. LF and its digestive peptides induced IFN-α production and activated pDCs in the presence of ssRNA, suggesting that LF modulates the immune system by promoting pDC activation upon viral recognition.

Effects of Sprayable, Highly Adhesive Hydrophobized Gelatin Microparticles on Endoscopic Submucosal Dissection: A Swine Model.

INTRODUCTION: Sprayable wound dressings containing hydrophobized microparticles (hMPs) are characterized by strong adhesiveness. We examined the effect of hMPs derived from Alaska pollock gelatin on endoscopic submucosal dissection (ESD) ulcers. METHODS: (1) In an in vivo model of miniature swine gastric ESD, gastric ulcers were created by ESD and then sprayed with hMPs or untreated followed by microscopic examination. (2) In an ex vivo ESD model of resected stomach, a pinhole-shaped perforation was created on the ESD ulcer of resected stomach; hMPs were then sprayed on the perforation; and air leakage and intragastric pressure were measured. (3) In an in vivo duodenal ESD model of miniature swine, duodenal artificial ESD ulcers with pinhole-shaped perforation were examined; ulcers were classified into hMPs-sprayed and nonsprayed groups, and inflammation in the intrinsic muscle layer and serosa were compared between the groups. RESULTS: (1) Histological observation of submucosal tissues showed a decreased number of invading inflammatory cells in hMP-sprayed tissues compared with the control in miniature swine gastric ESD (p < 0.05). In addition, the rates of anti-alpha smooth muscle actin and type I collagen positivity were significantly lower in the hMPs group than in the control group (p < 0.05). (2) Intragastric pressure could not be measured in the nonsprayed group, whereas no air leakage was observed in the sprayed group when pressurized up to 26 mm Hg in the resected stomach model. (3) The sprayed group showed suppressed inflammation of the intrinsic muscular layer and serosa in both cases compared with the nonsprayed group in miniature swine duodenal ESD (p < 0.05). CONCLUSIONS: Sprayable, tissue-adhesive hMPs are a promising medical material for intraoperative and postoperative treatment of ESD-induced wound via anti-inflammation and strong adhesiveness.

Usefulness of the aldehyde breath test for predicting metachronous recurrence in patients with esophageal squamous cell carcinoma and hypopharyngeal squamous cell carcinoma.

BACKGROUND: The rate of metachronous recurrence after endoscopic submucosal dissection for early-stage esophageal squamous cell carcinoma and hypopharynx squamous cell carcinoma is as high (10-15%). The acetaldehyde breath test may detect acetaldehyde dehydrogenase 2 gene polymorphisms. Therefore, we evaluated its usefulness in assessing metachronous recurrence in patients with esophageal squamous cell carcinoma and hypopharynx squamous cell carcinoma. METHODS: A total of 76 patients underwent endoscopic submucosal dissection for esophageal squamous cell carcinoma and hypopharynx squamous cell carcinoma and were followed up for at least 3 years (non-recurrence group: 52 patients; recurrence group: 24 patients). The risk factors for carcinogenesis were compared between the recurrence and non-recurrence groups, and the acetaldehyde-to-ethanol ratio was assessed. The cutoff acetaldehyde-to-ethanol ratio that correlated with recurrence was established, and the cumulative recurrence rate was evaluated. RESULTS: The recurrence group had a higher acetaldehyde-to-ethanol ratio, daily alcohol consumption, and Lugol-voiding lesion grade than the non-recurrence group in the univariate analysis. The cutoff acetaldehyde-to-ethanol ratio for recurrence was 28.1 based on the receiver operating characteristic curve. The multivariate analysis revealed an acetaldehyde-to-ethanol ratio of > 28.1 and a Lugol-voiding lesion grade associated with carcinogenesis. Patients with an acetaldehyde-to-ethanol ratio of ≥ 28.1 had a significantly high recurrence rate using the Kaplan-Meier method. CONCLUSIONS: The acetaldehyde-to-ethanol ratio detected using the acetaldehyde breath test could be a novel biomarker of metachronous recurrence after endoscopic submucosal dissection in patients with esophageal squamous cell carcinoma and hypopharynx squamous cell carcinoma. TRIAL REGISTRATION NUMBER: UMIN000040615.

IFN-α/ß-mediated NK2R expression is related to the malignancy of colon cancer cells.

Neurokinin 2 receptor (NK2R), a G protein-coupled receptor for neurokinin A (NKA), a tachykinin family member, regulates various physiological functions including pain response, relaxation of smooth muscle, dilation of blood vessels, and vascular permeability. However, the precise role and regulation of NK2R expression in cancer cells have not been fully elucidated. In this study, we found that high NK2R gene expression was correlated with the poor survival of colorectal cancer patients, and Interferon (IFN-α/ß) stimulation significantly enhanced NK2R gene expression level of colon cancer cells in a Janus kinas 1/2 (JAK 1/2)-dependent manner. NKA stimulation augmented viability/proliferation and phosphorylation of Extracellular-signal-regulated kinase 1/2 (ERK1/2) levels of IFN-α/ß-treated colon cancer cells and NK2R blockade by using a selective antagonist reduced the proliferation in vitro. Administration of an NK2R antagonist alone or combined with polyinosinic-polycytidylic acid, a synthetic analog of double-stranded RNA, to CT26-bearing mice significantly suppressed tumorigenesis. NK2R-overexpressing CT26 cells showed enhanced tumorigenesis and metastatic colonization in both lung and liver after the inoculation into mice. These findings indicate that IFN-α/ß-mediated NK2R expression is related to the malignancy of colon cancer cells, suggesting that NK2R blockade may be a promising strategy for colon cancers.

High aldehyde dehydrogenase 1 activity is related to radiation resistance due to activation of AKT signaling after insulin stimulation in prostate cancer.

The association between type 2 diabetes mellitus and prostate cancer is still under investigation, and the relationship between hyperinsulinemia and prostate cancer stem-like cells (CSCs) is elusive. Here, we investigated the function of insulin/AKT signaling in prostate CSCs. We isolated prostate CSCs as aldehyde dehydrogenase 1-high (ALDH1high) cells from the human prostate cancer 22Rv1 cell line using an ALDEFLUOR assay and established several ALDH1high and ALDH1low clones. ALDH1high clones showed high ALDH1 expression which is a putative CSC marker; however, they showed heterogeneity regarding tumorigenicity and resistance to radiation and chemotherapy. Interestingly, all ALDH1high clones showed lower phosphorylated AKT (Ser473) (pAKT) levels than the ALDH1low clones. PI3K/AKT signaling is a key cell survival pathway and we analyzed radiation resistance under AKT signaling activation by insulin. Insulin increased pAKT levels in ALDH1high and ALDH1low cells; the fold increase rate of pAKT was higher in ALDH1high cells than in ALDH1low cells. Insulin induced resistance to radiation and chemotherapy in ALDH1high cells, and the increased levels of pAKT induced by insulin were significantly related to radiation resistance. These results suggest that ALDH1 suppresses baseline pAKT levels, but AKT can be activated by insulin, leading to treatment resistance.

GRIK2 is a target for bladder cancer stem-like cell-targeting immunotherapy.

Recent studies have revealed that treatment-resistant cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) can be targeted by cytotoxic T lymphocytes (CTLs). CTLs recognize antigenic peptides derived from tumor-associated antigens; thus, the identification of tumor-associated antigens expressed by CSCs/CICs is essential. Human leucocyte antigen (HLA) ligandome analysis using mass spectrometry enables the analysis of naturally expressed antigenic peptides; however, HLA ligandome analysis requires a large number of cells and is challenging for CSCs/CICs. In this study, we established a novel bladder CSC/CIC model from a bladder cancer cell line (UM-UC-3 cells) using an ALDEFLUOR assay. CSCs/CICs were isolated as aldehyde dehydrogenase (ALDH)-high cells and several ALDHhigh clone cells were established. ALDHhigh clone cells were enriched with CSCs/CICs by sphere formation and tumorigenicity in immunodeficient mice. HLA ligandome analysis and cap analysis of gene expression using ALDHhigh clone cells revealed a distinctive antigenic peptide repertoire in bladder CSCs/CICs, and we found that a glutamate receptor, ionotropic, kainite 2 (GRIK2)-derived antigenic peptide (LMYDAVHVV) was specifically expressed by CSCs/CICs. A GRIK2 peptide-specific CTL clone recognized GRIK2-overexpressing UM-UC-3 cells and ALDHhigh clone cells, indicating that GRIK2 peptide can be a novel target for bladder CSC/CIC-targeting immunotherapy.

Tumor-infiltrating CD8+ T cells recognize a heterogeneously expressed functional neoantigen in clear cell renal cell carcinoma.

Immune checkpoint inhibitors (ICIs) are used in cancer immunotherapy to block programmed death-1 and cytotoxic T-lymphocyte antigen 4, but the response rate for ICIs is still low and tumor cell heterogeneity is considered to be responsible for resistance to immunotherapy. Tumor-infiltrating lymphocytes (TILs) have an essential role in the anti-tumor effect of cancer immunotherapy; however, the specificity of TILs in renal cell carcinoma (RCC) is elusive. In this study, we analyzed a 58-year-old case with clear cell RCC (ccRCC) with the tumor showing macroscopic and microscopic heterogeneity. The tumor was composed of low-grade and high-grade ccRCC. A tumor cell line (1226 RCC cells) and TILs were isolated from the high-grade ccRCC lesion, and a TIL clone recognized a novel neoantigen peptide (YVVPGSPCL) encoded by a missense mutation of the tensin 1 (TNS1) gene in a human leukocyte antigen-C*03:03-restricted fashion. The TNS1 gene mutation was not detected in the low-grade ccRCC lesion and the TIL clone did not recognized low-grade ccRCC cells. The missense mutation of TNS1 encoding the S1309Y mutation was found to be related to cell migration by gene over-expression. These findings suggest that macroscopically and microscopically heterogenous tumors might show heterogenous gene mutations and reactivity to TILs.

STAT1-mediated induction of Ly6c-expressing macrophages are involved in the pathogenesis of an acute colitis model.

BACKGROUND: The development of inflammatory bowel diseases is thought to be multifactorial, but the exact steps in pathogenesis are poorly understood. In this study, we investigated involvement of the activation of STAT1 signal pathway in the pathogenesis of an acute colitis model. METHODS: A dextran sulfate sodium-induced acute colitis model was established by using wild-type C57BL/6 mice and STAT1-deficient mice. Disease indicators such as body weight loss and clinical score, induction of cytokines, chemokines, and inflammatory cells were evaluated in the acute colitis model. RESULTS: Disease state was significantly improved in the acute colitis model using STAT1-deficient mice compared with wild-type mice. The induction of Ly6c-highly expressing cells in colorectal tissues was attenuated in STAT1-deficient mice. IL-6, CCL2, and CCR2 gene expressions in Ly6c-highly expressing cells accumulated in the inflamed colon tissues and were significantly higher than in Ly6c-intermediate-expressing cells, whereas TNF-α and IFN-α/ß gene expression was higher in Ly6c-intermediate-expressing cells. Blockade of CCR2-mediated signaling significantly reduced the disease state in the acute colitis model. CONCLUSIONS: Two different types of Ly6c-expressing macrophages are induced in the inflamed tissues through the IFN-α/ß-STAT1-mediated CCL2/CCR2 cascade and this is associated with the pathogenesis such as onset, exacerbation, and subsequent chronicity of acute colitis.

Crystal Structures and Conduction Paths of Sc-Doped CaTiO3 Fast Oxide-Ion Conductors with Orthorhombic Symmetry.

The crystal structures and electron density distributions (EDDs) of Ca-deficient Sc-doped CaTiO3 fast oxide-ion conductors, Ca0.97(Ti0.97Sc0.03)O3-δ (CTS3) and Ca0.96(Ti0.9Sc0.1)O3-δ (CTS10), were investigated in the temperature range of 298-1173 K in N2 to analyze the effect of composition on the perovskite structure and oxide-ion transport mechanism. CTS3 and CTS10 exhibited orthorhombic Pnma symmetry in temperature ranges of 298-1173 K and 973-1173 K, respectively, with CTS10 exhibiting lower symmetry and reduction in oxide-ion conductivity below 973 K. The EDDs of CTS3 and CTS10 at 1173 K indicated unique chemical bonds and conduction paths. CTS3 and CTS10 showed covalent bonds between (Ti,Sc) and O1 (or O2) sites. CTS3, with a lower oxide-ion conductivity than that of CTS10, exhibited pseudo-one-dimensional (1D) zig-zag curved conduction paths for oxide-ions along the a-axis, unlike previously reported curved migration paths along the b-axis in CaTiO3, and chemical bonds between Ca and O1 sites, indicating oxide-ion conduction suppression. In CTS10, additional conduction paths were observed along the a-axis, forming three-dimensional (3D) zig-zag curved conduction paths in the ac-plane and along the b-axis, with the weakening of the chemical bonds between the Ca and O1 sites. The oxide-ion conductivity and mobile oxide-ion concentration of CTS10 were 3.6 and 2.0 times those of CTS3, respectively, at 1173 K; the higher oxide-ion conductivity of CTS10 could be attributed to an increase in the mobile oxide-ion concentration and mobility with a 1D to 3D change in the conduction paths and a weakening of chemical bonds between the Ca and O1 sites.

Expression and localization of tight junction-related proteins in adult rat pituitary stem/progenitor cell niches.

Pituitary endocrine cells are supplied by Sox2-expressing stem/progenitor cells in the anterior lobe of the adult pituitary gland. These SOX2-positive cells are maintained in two types of microenvironments (niches): the marginal cell layer (MCL)-niche and the parenchymal-niche. Recently, we isolated dense SOX2-positive cell clusters from the parenchymal-niche by taking advantage of their resistance to protease treatment as parenchymal stem/progenitor cell (PS)-clusters. In the present study, by analyzing these isolated PS-clusters, we attempted to identify novel structural characteristics of pituitary stem/progenitor cell niches. Quantitative real-time PCR showed that tight junction-related genes were distinctly expressed in the isolated PS-clusters. Immunocytostaining showed that the tight junction molecules, ZO-1 and occludin, were localized in the apical membrane facing the pseudo-follicle-like structure of the isolated PS-clusters regardless of the expression of S100ß, which distinguishes the sub-population of SOX2-positive cells. Furthermore, immunohistochemistry of the pituitary glands of adult rats clearly demonstrated that ZO-1 and occludin were densely present in the parenchymal-niche encircling the pseudo-follicle, while they were observed in the apical membrane in the MCL-niche facing the residual lumen. Collectively, these tight junction-related proteins might be involved in the architecture and maintenance of the plasticity of pituitary stem/progenitor cell niches.