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Riki Toita,1,2 Masaharu Murata,1–3 Kana Abe,3 Sayoko Narahara,1 Jing Shu Piao,2 Jeong-Hun Kang,4 Kenoki Ohuchida,2,5 Makoto Hashizume1–31Innovation Center for Medical Redox Navigation, Kyushu University, 2Department of Advanced Medical Initiatives, Kyushu University, 3Center for Advanced Medical Innovation, Kyushu University, Fukuoka, 4Department of Biomedical Engineering, National Cerebral and Cardiovascular Center Research Institutes, Suita, 5Department of Surgery and Oncology, Kyushu University, Fukuoka, JapanAbstract: This study describes the applications of a naturally occurring small heat shock protein (Hsp) that forms a cage-like structure to act as a drug carrier. Mutant Hsp cages (HspG41C) were expressed in Escherichia coli by substituting glycine 41 located inside the cage with a cysteine residue to allow conjugation with a fluorophore or a drug. The HspG41C cages were taken up by various cancer cell lines, mainly through clathrin-mediated endocytosis. The cages were detected in acidic organelles (endosomes/lysosomes) for at least 48 hours, but none were detected in the mitochondria or nuclei. To generate HspG41C cages carrying doxorubicin (DOX), an anticancer agent, the HspG41C cages and DOX were conjugated using acid-labile hydrazone linkers. The release of DOX from HspG41C cages was accelerated at pH 5.0, but was negligible at pH 7.2. The cytotoxic effects of HspG41C–DOX against Suit-2 and HepG2 cells were slightly weaker than those of free DOX, but the effects were almost identical in Huh-7 cells. Considering the relatively low release of DOX from HspG41C–DOX, HspG41C–DOX exhibited comparable activity towards HepG2 and Suit-2 cells and slightly stronger cytotoxicity towards Huh-7 cells than free DOX. Hsp cages offer good biocompatibility, are easy to prepare, and are easy to modify; these properties facilitate their use as nanoplatforms in drug delivery systems and in other biomedical applications.Keywords: anticancer activity, biomedical application, doxorubicin, drug delivery system, protein nanocapsules, small heat shock protein

Masaharu Murata,1,2 Sayoko Narahara,1,2 Kaori Umezaki,1 Riki Toita,1,2 Shigekazu Tabata,1 Jing Shu Piao,1 Kana Abe,1 Jeong-Hun Kang,3 Kenoki Ohuchida,1,4 Lin Cui,4 Makoto Hashizume1,21Department of Advanced Medical Initiatives, Faculty of Medical Science, Kyushu University, Fukuoka, Japan; 2Innovation Center for Medical Redox Navigation, Kyushu University, Fukuoka, Japan; 3Department of Biomedical Engineering, National Cerebral and Cardiovascular Center Research Institute, Osaka, Japan; 4Department of Surgery and Oncology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, JapanAbstract: Protein nanocages are self-organized complexes of oligomers whose three-dimensional architecture can been determined in detail. These structures possess nanoscale inner cavities into which a variety of molecules, including therapeutic or diagnostic agents, can be encapsulated. These properties yield these particles suitable for a new class of drug delivery carrier, or as a bioimaging reagent that might respond to biochemical signals in many different cellular processes. We report here the design, synthesis, and biological characterization of a hepatocyte-specific nanocage carrying small heat-shock protein. These nanoscale protein cages, with a targeting peptide composed of a preS1 derivative from the hepatitis B virus on their surfaces, were prepared by genetic engineering techniques. PreS1-carrying nanocages showed lower cytotoxicity and significantly higher specificity for human hepatocyte cell lines than other cell lines in vitro. These results suggested that small heat-shock protein-based nanocages present great potential for the development of effective targeted delivery of various agents to specific cells.Keywords: protein nanocages, drug delivery system, hepatocyte cell lines specific, hepatitis B virus

Abstract Background Recently, the microRNA-200 family was reported to affect cancer biology by regulating epithelial to mesenchymal transition (EMT). Especially, the expression of miR-200c has been shown to be associated with upregulating the expression of E-cadherin, a gene known to be involved in pancreatic cancer behavior. However, the significance of miR-200c in pancreatic cancer is unknown. Methods In the present study, we investigated the relationship between E-cadherin and miR-200c expression in a panel of 14 pancreatic cancer cell lines and in macro-dissected formalin-fixed paraffin-embedded (FFPE) tissue samples obtained from 99 patients who underwent pancreatectomy for pancreatic cancer. We also investigated the effects of miR-200c on the proliferation and invasion of pancreatic cancer cells. Results We found that patients with high levels of miR-200c expression had significantly better survival rates than those with low levels of miR-200c expression. We also found a remarkably strong correlation between the levels of miR-200c and E-cadherin expression. Conclusions These data indicate that miR-200c may play a role in the pancreatic cancer biology and may be a novel marker for the prognosis of pancreatic cancer.

Abstract Background LIM-only 4 (LMO4), a member of the LIM-only (LMO) subfamily of LIM domain-containing transcription factors, was initially reported to have an oncogenic role in breast cancer. We hypothesized that LMO4 may be related to pancreatic carcinogenesis as it is in breast carcinogenesis. If so, this could result in a better understanding of tumorigenesis in pancreatic cancer. Methods We measured LMO4 mRNA levels in cultured cells, pancreatic bulk tissues and microdissected target cells (normal ductal cells; pancreatic intraepithelial neoplasia-1B [PanIN-1B] cells; PanIN-2 cells; invasive ductal carcinoma [IDC] cells; intraductal papillary-mucinous adenoma [IPMA] cells; IPM borderline [IPMB] cells; and invasive and non-invasive IPM carcinoma [IPMC]) by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Results 9 of 14 pancreatic cancer cell lines expressed higher levels of LMO4 mRNA than did the human pancreatic ductal epithelial cell line (HPDE). In bulk tissue samples, expression of LMO4 was higher in pancreatic carcinoma than in intraductal papillary-mucinous neoplasm (IPMN) or non-neoplastic pancreas (p < 0.0001 for both). We carried out microdissection-based analyses. IDC cells expressed significantly higher levels of LMO4 than did normal ductal epithelia or PanIN-1B cells (p < 0.001 for both) or PanIN-2 cells (p = 0.014). IPMC cells expressed significantly higher levels of LMO4 than did normal ductal epithelia (p < 0.001), IPMA (p < 0.001) and IPMB cells (p = 0.003). Conclusion Pancreatic carcinomas (both IDC and IPMC) expressed significantly higher levels of LMO4 mRNA than did normal ductal epithelia, PanIN-1B, PanIN-2, IPMA and IPMB. These results suggested that LMO4 is overexpressed at late stages in carcinogenesis of pancreatic cancer.

Patients with colorectal cancer (CRC) following renal transplantation require long-term immunosuppressants to prevent graft rejection. However, the impact of these immunosuppressants on the tumor immune microenvironment and the roles of immune cells within it remain poorly understood. We conducted comprehensive single-cell RNA sequencing on tumor and normal tissues from four CRC patients post renal transplantation and compared these with published data from 23 non-transplant CRC patients. We set four groups for detailed comparative analysis based on the renal transplantation status and tissue origin: non-renal transplantation normal (nRT&#95;Normal), non-renal transplantation tumor (nRT&#95;Tumor), renal transplantation normal (RT&#95;Normal), renal transplantation tumor (RT&#95;Tumor). Our analysis revealed significant tumor immune microenvironment landscape alterations in the transplantation group. CD8 + effector T cells of RT&#95;Tumor showed significantly diminished cytotoxicity and tumor neoantigen recognition (p < 0.0001), while CD4 + FOXP3 regulatory T cells of RT&#95;Tumor displayed a higher inhibitory score (p < 0.05), indicating preserved immunomodulatory potential compared with non-transplant CRC. Notably, significantly increased CTLA4 expression in T cells of RT&#95;Tumor was found and testified (p < 0.05). Our findings provide novel mechanistic insights for understanding the immune landscape in renal transplant recipients with CRC and pave the way for potential immunotherapeutic strategies that may improve survival and quality of life for this patient population., (© 2024 The Author(s). Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Purpose: Pancreatic ductal adenocarcinoma (PDAC) is considered a "cold" tumor because the tumor immune microenvironment (TIME) exhibits poor intratumoral T-cell infiltration. This study aimed to identify the marker genes associated with induction of cold TIME in PDAC cells., Methods: We orthotopically transplanted 10 primary cultures of PDAC derived from KrasG12D/+; Trp53R172H/+; Pdx-1-Cre (KPC) mice into immunocompetent mice and evaluated TIME by immunohistochemistry (IHC) staining of CD8. We divided primary cultures into two groups: cold TIME group with low CD8 + T-cell infiltration and a hot TIME group with high infiltration. RNA sequencing was performed to identify specific genes in the cold TIME group, and single-cell RNA sequencing (scRNA-seq) data was used for validation. IHC was performed to evaluate expressions in human PDAC samples., Results: We identified six genes specific in PDAC cells to the cold TIME group by RNA sequencing; these were defined as "cold tumor induction-related genes". Human PDAC scRNA-seq data revealed that cold tumor induction-related genes were significantly and negatively correlated with the number of CD8 + T-cells (p = 0.0341). These genes included protocadherin 7 (PCDH7). High expression of PCDH7 significantly and negatively correlated with the number of CD8 + T-cells in scRNA-seq (p = 0.0474) and IHC (p = 0.0110) data using human PDAC samples. PCDH7 was an independent factor for poor prognosis in PDAC (overall survival: hazard ratio = 2.07, p = 0.0367)., Conclusion: PCDH7 is a prognostic marker associated with CD8 + T-cell infiltration for PDAC patients., Competing Interests: Declarations. Ethical approval: This study was approved by the Ethics Committee of Kyushu University (approval number: 22002-00). All animal experiments were conducted following the guidelines of the institutional animal committee of Kyushu University (approval number: A22-128 and A23-026). Consent to participate: Written informed consent was waived owing to the retrospective analysis of the study. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: Gastric cancer (GC) shows limited response to immune checkpoint inhibitors due to its complex tumor immune microenvironment (TIME). This study explores the functions of various immune cells in the complex TIME in GC., Methods: We assessed CD8 + T-cell infiltration of GC tissues by immunohistochemistry, and performed single-cell RNA sequencing (scRNA-seq) of tumor and normal tissues from 34 patients with GC., Results: We categorized 157 GC patients into LOW, MID, and HIGH groups based on their CD8 + T-cell infiltration. Overall survival was notably lower for the HIGH and LOW groups compared with the MID group. Our scRNA-seq data analysis showed that CD8 + T-cell activity markers in the HIGH group were expressed at lower levels than in normal tissue, but the T-cell-attracting chemokine CCL5 was expressed at a higher level. Notably, CD8 + T-cells in the HIGH group displayed lower PD1 expression and higher CTLA4 expression. TCR repertoire analysis using only Epstein-Barr virus-negative cases showed that CD8 + T-cell receptor clonality was lower in the HIGH group than in the MID group. Furthermore, in the HIGH group, the antigen-presenting capacity of type 1 conventional dendritic cells was lower, the immunosuppressive capacity of myeloid-derived suppressor cells was higher, and the expression of CTLA4 in regulatory T-cells was higher., Conclusion: The present data suggest that the infiltration of inactive CD8 + T-cells with low clonality is induced by chemotaxis in the HIGH group, possibly leading to a poor prognosis for patients with GC., Competing Interests: Declarations. Ethics approval and consent to participation: For sample collection for scRNA-seq, written consent was obtained from all patients who participated in this study. After approval by the Ethics Committee of Kyushu University (approval number: #2023–79), we complied with the “Ethical Guidelines for Life Sciences and Medical Research Involving Human Subjects” and the principles of the Declaration of Helsinki. The immunohistochemistry and immunofluorescence samples were used according to a protocol approved by the Kyushu University Clinical Trials Accreditation Committee (#22002–01)., (© 2024. The Author(s).)

The effect of immune checkpoint inhibitors is extremely limited in patients with pancreatic ductal adenocarcinoma (PDAC) due to the suppressive tumor immune microenvironment. Autophagy, which has been shown to play a role in antitumor immunity, has been proposed as a therapeutic target for PDAC. In this study, single-cell RNA sequencing of autophagy-deficient murine PDAC tumors revealed that autophagy inhibition in cancer cells induced dendritic cell (DC) activation. Analysis of human PDAC tumors substantiated a negative correlation between autophagy and DC activation signatures. Mechanistically, autophagy inhibition increased the intracellular accumulation of tumor antigens, which could activate DCs. Administration of chloroquine, an autophagy inhibitor, in combination with Flt3 ligand-induced DC infiltration inhibited tumor growth and increased tumor-infiltrating T lymphocytes. However, autophagy inhibition in cancer cells also induced CD8+ T-cell exhaustion with high expression of immune checkpoint LAG3. A triple-therapy comprising chloroquine, Flt3 ligand, and an anti-LAG3 antibody markedly reduced tumor growth in orthotopic syngeneic PDAC mouse models. Thus, targeting autophagy in cancer cells and activating DCs sensitize PDAC tumors to immune checkpoint inhibitor therapy, warranting further development of this treatment approach to overcome immunosuppression in pancreatic cancer. Significance: Inhibiting autophagy in pancreatic cancer cells enhances intracellular accumulation of tumor antigens to induce dendritic cell activation and synergizes with immunotherapy to markedly inhibit the growth of pancreatic ductal adenocarcinoma., (©2024 The Authors; Published by the American Association for Cancer Research.)

Background: Pancreatic ductal adenocarcinoma tumors exhibit resistance to chemotherapy, targeted therapies, and even immunotherapy. Dendritic cells use glucose to support their effector functions and play a key role in anti-tumor immunity by promoting cytotoxic CD8 + T cell activity. However, the effects of glucose and lactate levels on dendritic cells in pancreatic ductal adenocarcinoma are unclear. In this study, we aimed to clarify how glucose and lactate can impact the dendritic cell antigen-presenting function and elucidate the relevant mechanisms., Methods: Glycolytic activity and immune cell infiltration in pancreatic ductal adenocarcinoma were evaluated using patient-derived organoids and resected specimens. Cell lines with increased or decreased glycolysis were established from KPC mice. Flow cytometry and single-cell RNA sequencing were used to evaluate the impacts on the tumor microenvironment. The effects of glucose and lactate on the bone marrow-derived dendritic cell antigen-presenting function were detected by flow cytometry., Results: The pancreatic ductal adenocarcinoma tumor microenvironment exhibited low glucose and high lactate concentrations from varying levels of glycolytic activity in cancer cells. In mouse transplantation models, tumors with increased glycolysis showed enhanced myeloid-derived suppressor cell infiltration and reduced dendritic cell and CD8 + T cell infiltration, whereas tumors with decreased glycolysis displayed the opposite trends. In three-dimensional co-culture, increased glycolysis in cancer cells suppressed the antigen-presenting function of bone marrow-derived dendritic cells. In addition, low-glucose and high-lactate media inhibited the antigen-presenting and mitochondrial functions of bone marrow-derived dendritic cells., Conclusions: Our study demonstrates the impact of dynamic glycolytic reprogramming on the composition of immune cells in the tumor microenvironment of pancreatic ductal adenocarcinoma, especially on the antigen-presenting function of dendritic cells., (© 2024. The Author(s).)

Background: Tumor-associated neutrophils (TANs) constitute an abundant component among tumor-infiltrating immune cells and have recently emerged as a critical player in pancreatic ductal adenocarcinoma (PDAC) progression. This study aimed to elucidate the pro-tumor mechanisms of TAN and identify a novel target for effective immunotherapy against PDAC., Methods: Microarray and cytokine array analyses were performed to identify the mechanisms underlying the function of TANs. Human and mouse TANs were obtained from differentiated HL-60 cells and orthotopically transplanted PDAC tumors, respectively. The interactions of TANs with cancer and cytotoxic T-cells were evaluated through in vitro co-culture and in vivo orthotopic or subcutaneous models. Single-cell transcriptomes from patients with PDAC were analyzed to validate the cellular findings., Results: Increased neutrophil infiltration in the tumor microenvironment was associated with poor survival in patients with PDAC. TANs secreted abundant amounts of chemokine ligand 5 (CCL5), subsequently enhancing cancer cell migration and invasion. TANs subpopulations negatively correlated with cytotoxic CD8 + T-cell infiltration in PDAC and promoted T-cell dysfunction. TANs upregulated the membranous expression of Nectin2, which contributed to CD8 + T-cell exhaustion. Blocking Nectin2 improved CD8 + T-cell function and suppressed tumor progression in the mouse model. Single-cell analysis of human PDAC revealed two immunosuppressive TANs phenotypes: Nectin2 + TANs and OLR1 + TANs. Endoplasmic reticulum stress regulated the protumor activities in TANs., Conclusions: TANs enhance PDAC progression by secreting CCL5 and upregulating Nectin2. Targeting the immune checkpoint Nectin2 could represent a novel strategy to enhance immunotherapy efficacy in PDAC., (© 2024. The Author(s).)

Background: Preoperative vascular embolization is an effective strategy for managing meningiomas, neck paragangliomas, renal cell carcinomas, and bone metastasis by reducing the intraoperative bleeding volume and operation time. Although hypervascular tumors also occur in the pancreas, preoperative embolization for these tumors is not commonly practiced. We herein present a case of a giant serous cystic neoplasm (SCN) of the pancreas with significant arterial vascularity that was managed with preoperative interventional radiology and subsequently resected via pancreaticoduodenectomy., Case Presentation: A 60-year-old man presented with an 8-cm hypervascular tumor located at the head of the pancreas, identified as an SCN on pathologic examination. The tumor had increased by 13 mm over 5 years, necessitating surgical intervention. Computed tomography revealed a substantial blood supply to the tumor from the dorsal pancreatic artery and gastroduodenal artery, both branches of the superior mesenteric artery. To mitigate the risk of severe intraoperative bleeding from this giant hypervascular tumor, branches of the dorsal pancreatic artery and gastroduodenal artery were embolized using metallic coils and further secured using a gelatin sponge 1 day prior to pancreatectomy. During the laparotomy, the tumor appeared to have decreased in size, likely because of reduced distension and congestion. Despite significant adhesions to surrounding tissues secondary to prolonged compression and inflammation, the pancreaticoduodenectomy was completed successfully in 5 h and 15 min with blood loss of 763 mL. The patient was discharged on postoperative day 15 without complications., Conclusions: Preoperative arterial embolization for hypervascular pancreatic tumors might control the risk of massive intraoperative bleeding, contributing to a favorable postoperative outcome. Utilizing interventional radiology for preoperative inflow control is one of the beneficial strategies for pancreatectomy in patients with a giant SCN., (© 2024. The Author(s).)

Background: Although immune checkpoint blockade (ICB) therapy has proven to be extremely effective at managing certain cancers, its efficacy in treating pancreatic ductal adenocarcinoma (PDAC) has been limited. Therefore, enhancing the effect of ICB could improve the prognosis of PDAC. In this study, we focused on the histamine receptor H1 (HRH1) and investigated its impact on ICB therapy for PDAC., Methods: We assessed HRH1 expression in pancreatic cancer cell (PCC) specimens from PDAC patients through public data analysis and immunohistochemical (IHC) staining. The impact of HRH1 in PCCs was evaluated using HRH1 antagonists and small hairpin RNA (shRNA). Techniques including Western blot, flow cytometry, quantitative reverse transcription polymerase chain reaction (RT-PCR), and microarray analyses were performed to identify the relationships between HRH1 and major histocompatibility complex class I (MHC-I) expression in cancer cells. We combined HRH1 antagonism or knockdown with anti-programmed death receptor 1 (αPD-1) therapy in orthotopic models, employing IHC, immunofluorescence, and hematoxylin and eosin staining for assessment., Results: HRH1 expression in cancer cells was negatively correlated with HLA-ABC expression, CD8 + T cells, and cytotoxic CD8 + T cells. Our findings indicate that HRH1 blockade upregulates MHC-I expression in PCCs via cholesterol biosynthesis signaling. In the orthotopic model, the combined inhibition of HRH1 and αPD-1 blockade enhanced cytotoxic CD8 + T cell penetration and efficacy, overcoming resistance to ICB therapy., Conclusions: HRH1 plays an immunosuppressive role in cancer cells. Consequently, HRH1 intervention may be a promising method to amplify the responsiveness of PDAC to immunotherapy., (© 2024. The Author(s).)

Background: Pancreatic ductal adenocarcinoma (PDAC) has a high mortality rate owing to its late diagnosis and aggression. In addition, there are relatively few minimally invasive screening methods for the early detection of PDAC, making the identification of biomarkers for this disease a critical priority. Recent studies have reported that microRNAs in extracellular vesicles (EV-miRs) from bodily fluids can be useful for the diagnosis of PDACs. Given this, we designed this study to evaluate the utility of cancer EVs extracted from duodenal fluid (DF) and their resident EV-miRs as potential biomarkers for the detection of PDAC., Methods: EV-miRs were evaluated and identified in the supernatants of various pancreatic cancer cell lines (Panc-1, SUIT2, and MIAPaca2), human pancreatic duct epithelial cells, and the DF from patients with PDAC and healthy controls. EVs were extracted using ultracentrifugation and the relative expression of EV-miR-20a was quantified., Results: We collected a total of 34 DF samples (27 PDAC patients and seven controls) for evaluation and our data suggest that the relative expression levels of EV-miR-20a were significantly higher in patients with PDAC than in controls ( p = 0.0025). In addition, EV-miR-20a expression could discriminate PDAC from control patients regardless of the location of the tumor with an area under the curve values of 0.88 and 0.88, respectively., Conclusions: We confirmed the presence of EVs in the DF and suggest that the expression of EV-miR-20a in these samples may act as a potential diagnostic biomarker for PDAC., Competing Interests: None., (© 2024 The Authors. DEN Open published by John Wiley & Sons Australia, Ltd on behalf of Japan Gastroenterological Endoscopy Society.)

Background: Gastric cancer (GC) is characterized by an immunosuppressive and treatment-resistant tumor immune microenvironment (TIME). Here, we investigated the roles of different immunosuppressive cell types in the development of the GC TIME., Methods: Single-cell RNA sequencing (scRNA-seq) and multiplex immunostaining of samples from untreated or immune checkpoint inhibitor (ICI)-resistant GC patients were used to examine the correlation between certain immunosuppressive cells and the prognosis of GC patients., Results: The results of the scRNA-seq analysis revealed that tumor-infiltrating monocytic myeloid-derived suppressor cells (TI-M-MDSCs) expressed higher levels of genes with immunosuppressive functions than other immunosuppressive cell types. Additionally, M-MDSCs in GC tissues expressed significantly higher levels of these markers than adjacent normal tissues. The M-MDSCs were most enriched in GC tissues relative to adjacent normal tissues. Among the immunosuppressive cell types assessed, the M-MDSCs were most enriched in GC tissues relative to adjacent normal tissues; moreover, their presence was most strongly associated with a poor prognosis. Immediate early response 3 (IER3), which we identified as a differentially expressed gene between M-MDSCs of GC and adjacent normal tissues, was an independent poor prognostic factor in GC patients (P = 0.0003). IER3 + M-MDSCs expressed higher levels of genes with immunosuppressive functions than IER3 - M-MDSCs and were abundant in treatment-resistant GC patients., Conclusions: The present study suggests that TI-M-MDSCs, especially IER3 + ones, may play a predominant role in the development of the immunosuppressive and ICI-resistant GC TIME., (© 2024. The Author(s) under exclusive licence to The International Gastric Cancer Association and The Japanese Gastric Cancer Association.)

Preoperative treatment is commonly carried out for borderline resectable pancreatic ductal adenocarcinoma (PDAC). However, the relationship between the combination of immune cells in the tumor microenvironment and their intratumoral heterogeneity along with their association with histological findings remains unclear, especially in patients receiving preoperative chemotherapy. We aimed to explore the therapeutic strategies for patients with PDAC with poor prognosis after receiving chemotherapy based on histological and immunological microenvironmental classifications. We investigated the correlation between the prognosis and histological immune microenvironmental factors of patients who initially underwent surgery (n = 100) and were receiving gemcitabine plus nab-paclitaxel (GEM + nabPTX) as preoperative chemotherapy (n = 103). Immune profiles were generated based on immune cell infiltration into the tumor, and their correlation with patient outcomes and histological features was analyzed. Tumor-infiltrating neutrophils (TINs) were identified as independent poor prognostic factors using multivariate analysis in both surgery-first and preoperative chemotherapy groups. The patients were further classified into four groups based on immune cell infiltration into the tumor. Patients with high CD15 infiltration into the tumor and immature stroma at the cancer margins showed the worst prognosis in the preoperative chemotherapy group. The analysis of mRNA expression and immunohistochemical features revealed that CXCR2, the receptor for CXCL8, was correlated with disease-free and overall survival. We inferred that patients with immature stroma at the margins and high infiltration of CD15 + neutrophils within the tumor showed the worst prognosis and they could particularly benefit from treatment with inhibitors targeting CXCR2 or CXCL8., (© 2024 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Purpose: Cholelithiasis occurs often after gastrectomy. However, no consensus has been established regarding the difference in the incidence of postgastrectomy cholelithiasis with different reconstruction methods. In this study, we examined the frequency of cholelithiasis after two major reconstruction methods, namely Billroth-I (B-I) and Roux-en-Y (R-Y) following laparoscopic distal gastrectomy (LDG) for gastric cancer., Methods: Among 696 gastric cancer patients who underwent LDG between April 2000 and March 2017, after applying the exclusion criteria, 284 patients who underwent B-I and 310 who underwent R-Y were examined retrospectively. The estimated incidence of cholelithiasis was compared between the methods, and factors associated with the development of cholelithiasis in the gallbladder and/or common bile duct were investigated., Results: During the median follow-up of 61.2 months, 52 patients (8.8%) developed cholelithiasis postgastrectomy; 12 patients (4.2%) after B-I and 40 (12.9%) after R-Y (p = 0.0002). Among them, choledocholithiasis was more frequent in patients who underwent R-Y (n = 11, 27.5%) vs. B-I (n = 1, 8.3%) (p = 0.0056). Univariate and multivariate analyses revealed that male sex, body mass index > 22.5 kg/m 2 , and R-Y reconstruction were significant predictors of the development of postLDG cholelithiasis., Conclusion: Regarding cholelithiasis development, B-I reconstruction should be preferred whenever possible during distal gastrectomy., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Pancreatic cancer is one of the most adverse diseases and it is very difficult to treat because the cancer cells formed in the pancreas intertwine themselves with nearby blood vessels and connective tissue. Hence, the surgical procedure of treatment becomes complicated and it does not always lead to a cure. Histopathological diagnosis is the usual approach for cancer diagnosis. However, the pancreas remains so deep inside the body that experts sometimes struggle to detect cancer in it. Computer-aided diagnosis can come to the aid of pathologists in this scenario. It assists experts by supporting their diagnostic decisions. In this research, we carried out a deep learning-based approach to analyze histopathology images. We collected whole-slide images of KPC mice to implement this work. The pancreatic abnormalities observed in KPC mice develop similar histological features to human beings. We created random patches from whole-slide images. Then, a convolutional autoencoder framework was used to embed these patches into an integrated latent space. We applied 'information maximization', a deep learning clustering technique to cluster the identical patches in an unsupervised manner since our dataset does not have annotation. Moreover, Uniform manifold approximation and projection, a nonlinear dimension reduction technique was utilized to visualize the embedded patches in a 2-dimensional space. Finally, we calculated a few internal cluster validation metrics to determine the optimal cluster set. Our work concentrated on patch-based anomaly detection in the whole slide histopathology images of KPC mice., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Rumman et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: There are several options for the treatment of gastrointestinal stricture, including endoscopic stent placement and bypass surgery. However, a benign stricture is difficult to manage in a reconstructed gastric tube in the thoracic cavity owing to the technical difficulty of bypass surgery, and the possibility of stent migration., Case Presentation: A 78-year-old woman was admitted to our hospital for treatment for her inability to eat. She had undergone video-assisted subtotal esophagectomy with retromediastinal gastric tube reconstruction 7 years earlier. At the current admission, there was a severely dilated gastric tube in the thoracic cavity with a soft stricture immediately anterior to the spine. Conservative therapy was ineffective; therefore, endoscopic stenting was performed. However, the stent migrated to the upper side of the stricture because the stricture was mild, and the stent was not fixed in the gastric tube. Next, endoscopic stent placement followed by laparoscopic stent fixation was performed. The stent was patent and worked well, and the patient's body weight increased. However, the stent collapsed 2 years later, with recurrence of symptoms. Stent-in-stent placement with an over-the-scope clip was performed, and the second stent was also patent and worked well., Conclusions: Laparoscopic stent fixation with endoscopic stent placement could be an effective option for patients with a benign stricture in the reconstructed gastric tube., (© 2023. The Author(s).)

Background: Tertiary lymphoid structures (TLSs) are associated with a favorable prognosis in several cancers. However, the correlation between TLSs and outcomes of esophageal squamous cell carcinoma (ESCC) and the impact of TLSs on the tumor immune microenvironment (TIME) remain unknown., Methods: We pathologically evaluated the significance of TLSs in ESCC focusing on TLS maturation using 180 ESCC specimens and performed single-cell RNA sequencing (scRNA-seq) using 14 ESCC tissues to investigate functional differences of immune cells according to TLS presence., Results: TLS+ cases had better recurrence-free-survival (RFS) (p < 0.0001) and overall survival (OS) (p = 0.0016) compared with TLS- cases. Additionally, mature TLS+ cases had better RFS and OS compared with immature TLS+ cases (p = 0.019 and p = 0.015) and TLS- cases (p < 0.0001 and p = 0.0002). The scRNA-seq showed that CD8 + T cells in TLS+ tumors expressed high levels of cytotoxic signatures and antigen-presentation of dendritic cells (DCs) was enhanced in TLS+ tumors. Immunohistochemistry showed that the densities of tumor-infiltrating CD8 + T cells and DCs were significantly higher in TLS+ tumors than those in TLS- tumors., Conclusions: These data suggest the prognostic and functional significance of TLSs in ESCC and provides new insights into TLSs on the TIME., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Intratumor bacteria modify the tumor immune microenvironment and influence outcomes of various tumors. Periodontal pathogen Fusobacterium nucleatum has been detected in pancreatic cancer tissues and is associated with poor prognosis. However, it remains unclear how F. nucleatum affects pancreatic cancer. Here, we compared clinical features with F. nucleatum colonization in pancreatic cancer tissues. F. nucleatum was detected in 15.5% (13/84) of pancreatic cancer patients. The tumor size was significantly larger in the F. nucleatum-positive group than in the negative group. To clarify the biological effect of intratumor F. nucleatum on pancreatic cancer progression, we performed migration/invasion assays and cytokine array analysis of cancer cells cocultured with F. nucleatum. F. nucleatum promoted CXCL1 secretion from pancreatic cancer cells, leading to cancer progression through autocrine signaling. Intratumor F. nucleatum suppressed tumor-infiltrating CD8 + T cells by recruiting myeloid-derived suppressor cells (MDSCs) to the tumor in an F. nucleatum-injected subcutaneous pancreatic cancer mouse model, resulting in tumor progression. Furthermore, tumor growth accelerated by F. nucleatum was suppressed by MDSC depletion or cytokine inhibitors. Intratumor F. nucleatum promoted pancreatic cancer progression through autocrine and paracrine mechanisms of the CXCL1-CXCR2 axis. Blockade of the CXCL1-CXCR2 axis may be a novel therapeutic approach for patients with intratumor F. nucleatum-positive pancreatic cancer., (© 2023 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Background: Malnutrition impacts the clinical course of Crohn's disease; however, there is little evidence of its influence on perioperative adverse events. We assessed whether nutritional indicators are associated with postoperative complications in surgical treatment of Crohn's disease., Methods: 137 patients with Crohn's disease who underwent surgical treatment between January 2011 and December 2020 were included. Skeletal muscle index was calculated by a single CT slice. We analyzed the risk factors for adverse events., Results: 37 % of patients had postoperative complications. Adverse events occurred more frequently in patients with high serum C-reactive protein, low serum albumin, prognostic nutritional index <38.3, skeletal muscle index <38.9 cm 2 /m 2 , abdominoperineal resection, long surgical duration, and mass hemorrhage. Among patients with skeletal muscle index <38.9 cm 2 /m 2 , patients who experienced adverse events had higher visceral fat index compared with those who did not (0.85 vs. 0.45, P  = 0.04). Multivariate analysis revealed that skeletal muscle index <38.9 cm 2 /m 2 and low serum albumin were the independent risk factors for postoperative complications (Odds ratio, 2.85; 95 % confidence interval, 1.13-7.16; P  = 0.03, 2.62; 1.09-6.26; P  = 0.03, respectively). Separated by sex, low serum albumin (<3.5 and <2.8 g/dL, male and female, respectively) and skeletal muscle index (<38.9 and <36.6 cm 2 /m 2 , male and female, respectively) were statistically related to postoperative complications., Conclusions: Skeletal muscle index is the most useful nutritional predictor of postoperative complications in Crohn's disease patients among other nutritional indices. We believe that these patients are at high risk of postoperative complications and need appropriate nutritional support in the perioperative period., Competing Interests: The authors have declared no conflicts of interest., (© 2023 The Authors.)

Purpose: This study aimed to demonstrate the involvement of angiogenesis in cancer-associated acinar-to-ductal metaplasia (CA-ADM) lesion of invasive front pancreatic ductal adenocarcinoma (PDAC) and investigate the possible mechanism., Methods: Tissue samples from 128 patients with PDAC and 36 LSL-Kras G12D/+ ; LSL-Trp53 R172H/+ ; Pdx-1-Cre mice were analyzed. Immunohistochemical assay was performed using HE, anti-CK19 and anti-amylase to confirm the presence of CA-ADM lesions, using anti-CD34 and anti-CD31 to measure microvessel density (MVD), and using anti-CD68, anti-CD163, anti-iNOS, or anti-MMP9 to evaluate the immune microenvironment. We performed multiplex immunohistochemical assay to detect the co-expression of MMP9 and CD68 on macrophage. We examined clinical outcomes and other clinicopathological factors to determine the significance of high-level MVD of CA-ADM on survival and liver metastasis. We performed tube formation assay to evaluate the effect of macrophage on angiogenic capacity in vitro., Results: Angiogenesis was significantly abundant in CA-ADM lesions compared with that in PDAC lesions in human and mouse tissues. High-level MVD in CA-ADM lesions was an independent predictor of poor prognosis (P = 0.0047) and the recurrence of liver metastasis (P = 0.0027). More CD68-positive and CD163-positive macrophages were detected in CA-ADM lesions than in PDAC. The percentage of CD68-positive macrophages was positively correlated with MVD in CA-ADM lesions. Multiplex-immunostaining revealed that MMP9 was expressed in CD68-positive macrophages of CA-ADM lesions. In CA-ADM lesions, the percentage of macrophages was positively correlated with MMP9 expression, which positively correlated with microvessel density., Conclusion: CA-ADM related angiogenesis is a promising predictive marker for poor prognosis of PDAC and may provide an attractive therapeutic target for PDAC., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Purpose: This study evaluated the reliability, validity, and responsiveness of the Japanese version of the European Organization for Research and Treatment of Cancer (EORTC) QLQ-ELD14 and measured the health-related quality of life (HRQOL) of elderly Japanese patients with cancer aged ≥ 60 and ≥ 70 years., Methods: The study recruited elderly Japanese patients with cancer aged ≥ 60 (≥ 70) years (n = 1803 [n = 1236]). The EORTC QLQ-ELD14 was evaluated for reliability, validity, responsiveness, and correlations of changes in score between the EORTC QLQ-ELD14 and the EORTC QLQ-C30 before and after the commencement of the COVID-19 pandemic., Results: In both age groups, the proportion of missing items was low (< 3%). Cronbach's α was good at ≥ 0.70, except for two of the seven items. All the intraclass coefficient constants were good at ≥ 0.70. The concurrent validity was good but correlation with the EORTC QLQ-C30 was not strong, except for the hypothesis items. Regarding the assessment of responsiveness, only one item ("maintaining purpose") of the EORTC QLQ-ELD14 worsened (- 6.14 ± 29.20, standard response of mean > 0.2) after the commencement of the COVID-19 pandemic. The changes in score between the EORTC QLQ-ELD14 and the "global health status/QOL" and "summary score" of the EORTC QLQ-C30 had moderate-to-high negative correlations for all items, except two. Hypotheses to evaluate construct validity were accepted at 90%, while responsiveness was accepted at 80%., Conclusion: The Japanese version of the EORTC QLQ-ELD14 questionnaire appears to have acceptable reliability, validity, and responsiveness to evaluate HRQOL in elderly Japanese people with cancer., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Although chemotherapy has been an essential treatment for cancer, the development of immune checkpoint blockade therapy was revolutionary, and a comprehensive understanding of the immunological tumor microenvironment (TME) has become crucial. Here, we investigated the impact of neoadjuvant chemotherapy (NAC) on immune cells in the TME of human esophageal squamous cell carcinoma using single cell RNA-sequencing. Analysis of 30 fresh samples revealed that CD8+/CD4+ T cells, dendritic cells (DCs), and macrophages in the TME of human esophageal squamous cell carcinoma showed higher levels of an anti-tumor immune response in the NAC(+) group than in the NAC(-) group. Furthermore, the immune cells of the NAC(+) group interacted with each other resulting in enhanced anti-tumor immune response via various cytokines, including IFNG in CD8+/CD4+ T cells, EBI3 in DCs, and NAMPT in macrophages. Our results suggest that NAC potentially enhances the anti-tumor immune response of immune cells in the TME., Competing Interests: The authors declare no competing interests., (© 2023 The Authors.)

Purpose: A diagnostic and treatment strategy for appendiceal tumors (ATs) has not been established. We aimed to evaluate our treatment strategy in ATs, including laparoscopic surgery (LS), and to identify preoperative malignancy predictors., Methods: A total of 51 patients between 2011 and 2021 were retrospectively reviewed. Data, including tumor markers and imaging findings, were compared between carcinoma and non-carcinoma patients. Validity of planned operation was evaluated based on pathological diagnosis., Results: Twenty-five patients were diagnosed with carcinoma, 13 with low-grade mucinous neoplasm, and 13 with other diseases. Symptoms were more commonly present in carcinoma patients than in non-carcinoma patients (68.0% vs. 23.1%, p = 0.001). Elevated CEA and CA19-9 were more frequently observed in carcinoma patients than in non-carcinoma patients (p < 0.01 and p = 0.04, respectively). Five carcinoma patients had malignancy on biopsy, compared with zero non-carcinoma patients. Significant differences were noted in the percentages of carcinoma and non-carcinoma patients with solid enhanced mass (41.7% vs. 0%, p < 0.001) and tumor wall irregularity (16.7% vs. 0%, p = 0.03) on imaging. Although the sensitivity was not high, the specificity and positive predictive value of these findings were 100%. Forty-two patients (82.4%) underwent LS as minimally invasive exploratory and/or radical operation, of whom 2 were converted to open surgery for invasion of adjacent organ. No patients had intraoperative complications or postoperative mortality., Conclusion: Clinical symptoms, elevated tumor markers, and worrisome features of solid enhanced mass and tumor wall irregularity on imaging can be malignancy predictors. For management of ATs, LS is feasible and useful for diagnosis and treatment., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Purpose: To determine whether N3 nodal involvement predicts outcomes and whether its prognostic implications vary with tumor location in patients with Stage III colon cancer (CC)., Methods: We defined N3 as lymph node metastases near the bases of the major feeding arteries. We retrospectively examined recurrence rates and patterns by tumor location and sites of lymph node metastases in 57 patients with N3 CC who had undergone curative resections between January 2000 and March 2019. Survival analysis was performed to compare the prognoses of patients with and without N3 lymph node metastasis., Results: Most N3 patients had large tumors (T ≥ 3); five had T2 disease. Recurrence occurred quickly in one patient with T2N3M0 disease. Multivariate survival analysis demonstrated that N3 lymph node metastasis is an independent predictor of poor prognosis in Stage III CC patients (P < 0.001). Categorizing N3 patients according to UICC-TNM staging system does not stratify risk of recurrence (P = 0.970). To investigate the impact of tumor location on recurrence risk, we classified N3 CC into two subtypes according to tumor location: metastasis at the base of the superior mesenteric artery in right-sided CC and inferior mesenteric artery in left-sided CC. The former was found to have a statistically significant poorer prognosis than the latter (P = 0.091)., Conclusion: N3 is a robust prognostic marker in CC patients. Recurrence risk varies by tumor location. N3 right-sided CCs with lymph node metastasis at the base of the superior mesenteric artery have poorer prognoses than do N3 left-sided CCs., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Tumour immune microenvironment is related with carcinogenesis and efficacy of immunotherapy. B cells play major roles in humoral immunity, but detailed functions of tumour-infiltrating B lymphocytes (TIL-Bs) are unknown. Therefore, our aim was to investigate the functional heterogeneity of TIL-Bs in oesophageal squamous cell carcinoma (ESCC) and lymph nodes (LNs) during chemotherapy., Methods: Single-cell transcriptome analysis was performed on 23 specimens. We also performed immunohistochemical analysis of immunoglobulin κ C (IGKC), an antibody-secreting cell (ASC) marker, in 166 ESCC samples and evaluated the implication of IGKC in 2-year recurrence free survival (RFS) and 3-year overall survival (OS)., Results: A total of 81,246 cells were grouped into 24 clusters. We extracted B cell clusters based on canonical markers and identified 12 TIL-B subtypes in ESCC. We found that several functions, such as co-stimulation and CD40 signalling, were enhanced in TIL-Bs after chemotherapy. The proportion of naive B cells (NBCs) decreased and B cell activation genes were up-regulated in NBCs after chemotherapy. The proportion of ASCs in tumours increased with the loss of migratory abilities and antibody production in ASCs was promoted after chemotherapy. Differentially expressed genes up-regulated with chemotherapy in ASCs correlated with prolonged survival with oesophageal cancer (p = .028). In a metastatic LN, the ASC proportion increased and B cell differentiation was enhanced. In immunohistochemical analysis, RFS and OS of high IGKC expression cases were significantly better than those of low IGKC expression cases (RFS: p < .0001, OS: p < .0001). And in multivariable analysis, the expression of IGKC was an independent favourable prognostic factor for RFS (hazard ratio (HR): 0.23, 95% confidence interval (CI): 0.12-0.45, p < .0001) and OS (HR: 0.20, 95% CI: 0.086-0.47, p = .0002) in ESCC., Conclusions: Our findings provide novel insights for the heterogeneity of TIL-Bs during chemotherapy and will be useful to understand the clinical importance of TIL-Bs., (© 2023 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Purpose: Preoperative sarcopenia worsens postoperative outcomes in various cancer types including colorectal cancer. However, we often experienced postoperative anastomotic leakage in muscular male patients such as Judo players, especially in rectal cancer surgery with lower anastomosis. It is controversial whether the whole skeletal muscle mass impacts the potential for anastomotic failure in male rectal cancer patients. Thus, the purpose of this study was to clarify whether skeletal muscle mass impacts anastomotic leakage in rectal cancer in men., Methods: We reviewed the medical charts of male patients suffering from rectal cancer who underwent colo-procto anastomosis below the peritoneal reflection without a protective diverting stoma. We measured the psoas muscle area and calculated the psoas muscle index., Results: One hundred ninety-seven male rectal cancer patients were enrolled in this study. The psoas muscle index was significantly higher in patients with anastomotic leakage (P<0.001). Receiver operating characteristic curve determined the optimal cut-off value of the psoas muscle index for predicting anastomotic leakage as 812.67 cm2/m2 (sensitivity of 60% and specificity of 74.3%). Multivariate analysis revealed that high psoas muscle index (risk ratio [RR], 3.933; P<0.001; 95% confidence interval [CI], 1.917-8.070) and super low anastomosis (RR, 2.792; P=0.015; 95% CI, 1.221-6.384) were independent predictive factors of anastomotic leakage., Conclusion: This study showed that male rectal cancer patients with a large psoas muscle mass who underwent lower anastomosis had a higher rate of postoperative anastomotic leakage.

Background: Loss of E-cadherin expression is frequently observed in signet ring carcinoma (SRCC). People with germline mutations in CDH1, which encodes E-cadherin, develop diffuse gastric cancer at a higher rate. Loss of E-cadherin expression is thus assumed to trigger oncogenic development., Methods: To investigate novel therapeutic targets for gastric SRCC, we engineered an E-cadherin-deficient SRCC model in vitro using a human gastric organoid (hGO) with CDH1 knockout (KO)., Results: CDH1 KO hGO cells demonstrated distinctive morphological changes similar to SRCC and high cell motility. RNA-sequencing revealed up-regulation of matrix metalloproteinase (MMP) genes in CDH1 KO hGO cells compared to wild type. MMP inhibitors suppressed cell motility of CDH1 KO hGO cells and SRCC cell lines in vitro. Immunofluorescent analysis with 95 clinical gastric cancer tissues revealed that MMP-3 was specifically abundant in E-cadherin-aberrant SRCC. In addition, CXCR4 molecules translocated onto the cell membrane after CDH1 KO. Addition of CXCL12, a ligand of CXCR4, to the culture medium prolonged cell survival of CDH1 KO hGO cells and was abolished by the inhibitor, AMD3100. In clinical SRCC samples, CXCL12-secreting fibroblasts showed marked infiltration into the cancer area., Conclusions: E-cadherin deficient SRCCs might gain cell motility through upregulation of MMPs. CXCL12-positive cancer-associated fibroblasts could serve to maintain cancer-cell survival as a niche. MMPs and the CXCL12/CXCR4 axis represent promising candidates as novel therapeutic targets for E-cadherin-deficient SRCC., (© 2022. The Author(s) under exclusive licence to The International Gastric Cancer Association and The Japanese Gastric Cancer Association.)

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers, with a 5-year survival rate of 9%. Cancer-associated fibroblasts (CAFs) have historically been considered tumor-promoting. However, multiple studies reporting that suppression of CAFs in PDAC mouse models resulted in more aggressive tumors and worse prognosis have suggested the existence of a tumor-suppressive population within CAFs, leading to further research on heterogeneity within CAFs. In recent years, the benefits of cancer immunotherapy have been reported in various carcinomas. Unfortunately, the efficacy of immunotherapies in PDAC has been limited, and the CAF-driven cancer immunosuppressive microenvironment has been suggested as the cause. Thus, clarification of heterogeneity within the tumor microenvironment, including CAFs and tumor immunity, is urgently needed to establish effective therapeutic strategies for PDAC. In this review, we report the latest findings on the heterogeneity of CAFs and the functions of each major CAF subtype, which have been revealed by single-cell RNA sequencing in recent years. We also describe reports of tumor-suppressive CAF subtypes and the existence of CAFs that maintain a differentiated PDAC phenotype and review the potential for targeted therapy.

For stage II and III esophageal squamous cell carcinoma (ESCC), neoadjuvant chemotherapy (NAC) followed by esophagectomy is recommended in the Japanese guidelines for the diagnosis and treatment of esophageal cancer. However, recurrence of ESCC is common regardless of the NAC regimen and surgical method, and NAC demonstrates limited efficacy against recurrence. Therefore, the present study was conducted to identify risk factors of recurrence of ESCC with surgery after NAC. The outcomes of 51 patients who underwent esophagectomy for ESCC after NAC from 2010 to 2017 at Kyushu University Hospital were retrospectively analyzed. A total of 52 patients with ESCC without NAC followed by esophagectomy from 2001 to 2017 were selected for comparison. Among patients who underwent NAC followed by surgery, only lymphatic invasion (LY; hazard ratio, 2.761; 95% CI, 1.86-6.43, P=0.018) was an independent factor significantly associated with 3-year recurrence-free survival in the multivariate analysis. In patients with pathologic lymph node metastasis (pN) and no LY after NAC, there was significantly less recurrence compared with patients with pN and LY (P=0.0085), whereas in patients without LY after NAC, the presence of pN was not significantly associated with recurrence (P=0.2401). There were significantly fewer LY (+) patients in the NAC (+) group (P=0.0158) compared with those in the NAC (-) group. The presence of LY was an independent risk factor for recurrence of ESCC after esophagectomy following NAC. Overall, adjuvant treatment after surgery may be required in cases with remnant LY after NAC., Competing Interests: The authors declare that they have no competing interests., (Copyright: © Okuda et al.)

Background: Pancreatic ductal adenocarcinoma (PDAC) is characterized by abundant stroma in which microenvironmental (niche) factors promote PDAC progression. In mouse models, reduction of the stroma increased the proportion of poorly differentiated PDAC with a worse prognosis. Here, we aimed to clarify the effects of stroma on PDAC that may define the PDAC phenotype and induce distinct therapeutic responses., Methods: The molecular features of PDAC based on differentiation grade were clarified by genome and transcriptome analysis using PDAC organoids (PDOs). We identified the dependency on niche factors that might regulate the differentiation grade. A three-dimensional co-culture model with cancer-associated fibroblasts (CAFs) was generated to determine whether CAFs provide niche factors essential for differentiated PDAC. PDOs were subtyped based on niche factor dependency, and the therapeutic responses for each subtype were compared., Results: The expression profiles of PDOs differed depending on the differentiation grade. Consistent with the distinct profiles, well differentiated types showed high niche dependency, while poorly differentiated types showed low niche dependency. The three-dimensional co-culture model revealed that well differentiated PDOs were strongly dependent on CAFs for growth, and moderately differentiated PDOs showed plasticity to change morphology depending on CAFs. Differentiated PDOs upregulated the expression of mevalonate pathway-related genes correlated with the niche dependency and were more sensitive to simvastatin than poorly differentiated PDOs., Conclusions: Our findings suggest that CAFs maintain the differentiated PDAC phenotype through secreting niche factors and induce distinct drug responses. These results may lead to the development of novel subtype-based therapeutic strategies., (© 2022. The Author(s).)

Purpose: T1 gastric cancer (GC) with seven or more metastatic lymph nodes is extremely rare, and very few clinical studies have been conducted to evaluate the clinicopathological features of their recurrence., Methods: We retrospectively analyzed the outcomes of T1 GC and T2-4 GC patients who had multiple nodal metastases after radical surgery from 2006 to 2020. Propensity score matching was performed to compare the two groups of patients., Results: After propensity score matching, 18 of 22 patients in the T1 group and 36 of 144 patients in the T2-4 group were selected. Recurrence occurred in six patients (33.3%) in the T1 group. In the T1 group, the most common site of initial recurrence was bone (15.0%). The prevalence of bone recurrence was significantly higher in the T1 group than in the T2-4 group (P = 0.02). The median interval time between radical surgery and bone recurrence was 24 months, and the median survival time after bone recurrence was 14 months., Conclusion: Bone recurrence was more frequently identified as an initial recurrence site in T1 GC cases with multiple metastases after radical surgery compared with that in T2-4 GC cases. Careful attention should be paid to postoperative bone recurrence in the long-term postoperative course of these patients., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Pancreatic cancer cells (PCCs) are surrounded by an abundant stroma, which is produced by pancreatic stellate cells (PSCs). PSCs promote tumor cell proliferation and invasion. The objective of the current study was to identify compounds that suppress PSC activation. Gene expression profiles of cancer-derived fibroblasts and normal fibroblasts were used, and the pathway analysis suggested altered pathways that were chosen for validation. It was found that the 'neuroactive ligand-receptor interaction' pathway from the Kyoto Encyclopedia of Genes and Genomes pathway analysis was one of the altered pathways. Several compounds related with this pathway were chosen, and changes in PSC activity were investigated using fluorescence staining of lipid droplets, reverse transcription-quantitative PCR, western blotting, and invasion and migration assays. Among these candidates, duloxetine, a serotonin-noradrenaline reuptake inhibitor, was found to suppress PSC activation and disrupt tumor-stromal interaction. Thus, duloxetine may be a potential drug for suppressing PSC activation and pancreatic cancer growth., Competing Interests: The authors declare that they have no competing interests., (Copyright: © Sagara et al.)

Pancreatic ductal adenocarcinoma (PDAC) is characterized by notorious resistance to current therapies attributed to inherent tumor heterogeneity and highly desmoplastic and immunosuppressive tumor microenvironment (TME). Unique proline isomerase Pin1 regulates multiple cancer pathways, but its role in the TME and cancer immunotherapy is unknown. Here, we find that Pin1 is overexpressed both in cancer cells and cancer-associated fibroblasts (CAFs) and correlates with poor survival in PDAC patients. Targeting Pin1 using clinically available drugs induces complete elimination or sustained remissions of aggressive PDAC by synergizing with anti-PD-1 and gemcitabine in diverse model systems. Mechanistically, Pin1 drives the desmoplastic and immunosuppressive TME by acting on CAFs and induces lysosomal degradation of the PD-1 ligand PD-L1 and the gemcitabine transporter ENT1 in cancer cells, besides activating multiple cancer pathways. Thus, Pin1 inhibition simultaneously blocks multiple cancer pathways, disrupts the desmoplastic and immunosuppressive TME, and upregulates PD-L1 and ENT1, rendering PDAC eradicable by immunochemotherapy., Competing Interests: Declaration of interests K.K., G.M.W., T.D.M., B.J.P., N.L., N.S.G., X.Z.Z., and/or K.P.L. are inventors of a number of issued patents and/or pending patent applications on Pin1, Pin1 biomarkers, Pin1 inhibitors, and Pin1 inhibitor combination to treat human diseases; X.Z.Z. and K.P.L. are inventors of cis P-tau antibody technology, which was licensed by BIDMC to the startup Pinteon Therapeutics. X.Z.Z. and K.P.L. are the scientific founders and former scientific advisors of and own equity in Pinteon. Their interests were reviewed and are managed by BIDMC in accordance with its conflict of interest policy. G.M.W. reports research funding from Glaxo Smith Kline (institutional funding). N.S.G. is a founder, science advisory board member (SAB), and equity holder in Gatekeeper, Syros, C4, Allorion, Jengu, Inception, B2S, EoCys, Larkspur, and Soltego (board member). The Gray lab receives or has received research funding from Novartis, Takeda, Astellas, Taiho, Jansen, Kinogen, Arbella, Deerfield, and Sanofi. All other authors do not have any competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Pancreatic stellate cells (PSCs) play a key role in desmoplastic stroma, which is a characteristic of pancreatic ductal adenocarcinoma (PDAC), and they also enhance the malignancy of pancreatic cancer cells. Our previous study reported chloroquine's mitigating effects on PSC activation; however, the drug is known to induce adverse effects in clinical practice. The present study aimed to reduce chloroquine doses and develop a useful pre-treatment that targets PSCs using nanoparticles. Poly lactic-co-glycolic acid (PLGA) nanoparticles were used as carriers and loaded with indocyanine green (Nano-ICG) or chloroquine (Nano-CQ). Tumor accumulation of Nano-ICG was evaluated using an in vivo imaging system. The effects of chloroquine, Nano-CQ and/or chemotherapy drug gemcitabine were investigated in an orthotopic xenograft mouse model. Nano-ICG selectively accumulated in pancreatic tumors and persisted therein for over 7 days after administration. Additionally, Nano-ICG accumulated in the peritoneal metastasized regions, but not in the liver, kidney and normal pancreatic tissues. Nano-CQ reduced the density of activated PSCs at lower chloroquine doses and significantly restrained tumor progression in combination with gemcitabine. In conclusion, the PLGA nanosystem successfully delivered the drug to pancreatic tumors. Nano-CQ efficiently reduced PSC activation and may be a promising novel pre-treatment strategy for PDAC., Competing Interests: The authors declare that they have no competing interests., (Copyright: © Matsumoto et al.)

Postoperative adjuvant chemotherapy for patients with stage III colon cancer (CC) is regarded as the standard treatment worldwide for outcome improvement and relapse prevention. Similarly, high-risk stage II CC requires adjuvant chemotherapy because of its high recurrence rate. Previous randomized controlled trials showed that oxaliplatin (OX), in addition to fluorinated pyrimidine-based therapy for patients with stage II/III CC, significantly improves cancer survival but it remains controversial as to which patient groups should receive OX-containing regimens. Among 1,150 consecutive patients who underwent curative resection for stage II/III CC between 2009 and 2016 at two tertiary hospitals, 349 patients treated with only peroral (PO) fluorinated pyrimidine-based chemotherapy and 149 patients who received fluorinated pyrimidine-based chemotherapy with OX as adjuvant chemotherapy were retrospectively reviewed. The primary outcome was recurrence-free survival (RFS). Clinicopathological factors were more advanced in patients treated with OX than in patients treated only with PO fluorinated pyrimidine agents. Multivariate analysis for 5-year RFS showed that T4 [hazard ratio (HR), 2.947; P=0.0001], N2 (HR, 2.704; P=0.0075), vessel or lymphatic invasion (HR, 1.675; P=0.0437) and high cancer antigen (CA)19-9 (HR 3.367, P=0.0002) levels were independent risk factors of cancer relapse. Propensity score matching analysis was performed to match clinicopathological differences between the PO and OX groups. After matching, subgroup analysis of the patients showed that greater effects of OX on cancer survival were observed in patients in the OX group with high CA19-9 levels and tended to be associated with T4 and N2 compared with the PO group. Thus, OX-containing regimens should be recommended for patients with CC with these factors in an adjuvant setting., Competing Interests: The authors declare that they have no competing interests., (Copyright © 2020, Spandidos Publications.)

Background: Pancreatic stellate cells (PSCs) occupy the majority of the pancreatic cancer microenvironment, contributing to aggressive behavior of pancreatic cancer cells (PCCs). Recently, anti-fibrotic agents have proven to be an effective strategy against cancer, but clinical trials have shown little efficacy, and the driving mechanism remains unknown. N-acetyl-cysteine (NAC) is often used for pulmonary cystic fibrosis. Pioglitazone, an agonist of peroxisome proliferator-activated receptor gamma, was habitually used for type II diabetes, but recently reported to inhibit metastasis of PCCs. However, few studies have focused on the effects of these two agents on cancer-stromal interactions., Method: We evaluated the expression of α-smooth muscle actin (α-SMA) and the number of lipid droplets in PSCs cultured with or without NAC. We also evaluated changes in invasiveness, viability, and oxidative level in PSCs and PCCs after NAC treatment. Using an indirect co-culture system, we investigated changes in viability, invasiveness, and migration of PSCs and PCCs. Combined treatment effects of NAC and Pioglitazone were evaluated in PSCs and PCCs. In vivo, we co-transplanted KPC-derived organoids and PSCs to evaluate the effects of NAC and Pioglitazone's combination therapy on subcutaneous tumor formation and splenic xenografted mouse models., Results: In vitro, NAC inhibited the viability, invasiveness, and migration of PSCs at a low concentration, but not those of PCCs. NAC treatment significantly reduced oxidative stress level and expression of α-SMA, collagen type I in PSCs, which apparently present a quiescent-like state with a high number of lipid droplets. Co-cultured PSCs and PCCs mutually promoted the viability, invasiveness, and migration of each other. However, these promotion effects were attenuated by NAC treatment. Pioglitazone maintained the NAC-induced quiescent-like state of PSCs, which were reactivated by PCC-supernatant, and enhanced chemosensitivity of PCCs. In vivo, NAC and Pioglitazone's combination suppressed tumor growth and liver metastasis with fewer stromal components and oxidative stress level., Conclusion: NAC suppressed activated PSCs and attenuated cancer-stromal interactions. NAC induces quiescent-like PSCs that were maintained in this state by pioglitazone treatment.

Introduction: Pancreatic adenocarcinoma (PAAD) is an aggressive malignancy, with a major mortality resulting from the rapid progression of metastasis. Unfortunately, no effective treatment strategy has been developed for PAAD metastasis to date. Thus, unraveling the mechanisms involved in PAAD metastatic phenotype may facilitate the treatment for PAAD patients., Objectives: PIK3CB is an oncogene implicated in cancer development and progression but less is known about whether PIK3CB participates in PAAD metastasis. Therefore, the objective of this study is to explore the mechanism(s) of PIK3CB in PAAD metastasis., Methods: In our study, we examined the PIK3CB expression pattern using bioinformatic analysis and clinical material derived from patients with PAAD. Subsequently, a series of biochemical experiments were conducted to investigate the role of PIK3CB as potential mechanism(s) underlying PAAD metastasis in vivo using nude mice and in vitro using cell lines., Results: We observed that PIK3CB was involved in PAAD progression. Notably, we identified that PIK3CB was involved in PAAD metastasis. Downregulation of PIK3CB significantly reduced PAAD metastatic potential in vivo. Furthermore, a series of bioinformatic analyses showed that PIK3CB was involved in cell adhesion in PAAD. Notably, PIK3CB depletion inhibited invasion potential specifically via suppressing cell adhesion to collagen I in PAAD cells., Conclusion: Collectively, our findings indicate that PIK3CB is involved in PAAD metastasis through cell-matrix adhesion. We proposed that PIK3CB is a potential therapeutic target for PAAD therapy., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Authors. Published by Elsevier B.V. on behalf of Cairo University. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Robotic surgery is expanding in the minimally invasive treatment of gastrointestinal cancer. In the field of gastrointestinal cancer, robotic surgery is performed using a robot-assisted surgery system. In this system, the robot does not operate automatically but is controlled by the surgeon. The surgery assistant robot currently used in clinical practice worldwide is the leader-follower type, including the da Vinci® Surgical System (Intuitive Surgical). This review describes the current state of robotic surgery in the treatment of gastrointestinal cancer and discusses the future development of robotic systems in gastrointestinal surgery., Competing Interests: Kenoki Ohuchida has no conflict of interest or financial ties to disclose., (Copyright © 2020 Kenoki Ohuchida.)

Background: Metastatic cancer to the stomach is relatively rare. Prostate-specific antigen (PSA) is a reliable biomarker used in the screening and management of patients with prostate cancer. However, it is difficult to definitively diagnose a PSA-negative metastatic gastric tumor of prostate cancer because the cancer sometimes resembles primary gastric cancer in clinical images. It is also difficult to distinguish metastatic cancer from primary cancer even in the pathological examination of biopsy samples when the lesion is poorly differentiated adenocarcinoma. There is a possibility that the characteristics of the cancer are changed during treatment such as chemotherapy or radiation therapy. Therefore, careful consideration is required for surgical indication., Case Presentation: A 60-year-old male underwent radical prostatectomy and subsequent radiation therapy for advanced prostate cancer (pT3N1M0) 10 years previously, and hormone therapy was started for metachronous multiple bone metastasis 10 months before. Upper gastrointestinal endoscopy revealed an irregular depressed lesion with a convergence of folds at the greater curvature of the upper gastric body. Biopsy showed poorly differentiated adenocarcinoma that was negative for PSA upon immunohistochemistry. He had high serum carcinoembryonic antigen (CEA) (946.1 ng/ml) and carbohydrate antigen 19-9 (CA19-9) (465.1 U/ml) levels with no elevation of PSA (0.152 ng/ml). The tumor was diagnosed as primary gastric cancer based on the clinical imaging and pathological examination of the biopsy sample including the PSA staining. Based on the diagnosis, laparoscopic proximal gastrectomy with lymphadenectomy was performed. However, pathological examination of the resected specimen revealed poorly differentiated adenocarcinoma that was positive for other prostate markers such as androgen receptor. Thus, the patient was diagnosed with metastasized prostate cancer to the stomach., Conclusions: We report a case of metastatic gastric cancer of prostate cancer 10 years after radical prostatectomy. In the present case, it was difficult to diagnose a metastatic gastric tumor of prostate cancer preoperatively, because of its resemblance to primary gastric cancer without PSA expression and no serum PSA elevation. Although a rare case entity, it is important to consider the possibility of a metastatic gastric tumor when the surgical indication is determined in cases with another co-existing cancer.