Background: PIK3CA mutation is one of the major oncogenic events in gastric cancer (GC); however, the molecular heterogeneity and therapeutic vulnerabilities associated with different mutation domains remain poorly understood. This study aimed to investigate the clinicopathological characteristics and organoid-based therapeutic responses of gastric cancers harboring distinct PIK3CA mutation domains. Methods: A total of 285 patients with primary gastric adenocarcinoma who underwent surgical resection between 2023 and 2025 at the First Affiliated Hospital of Anhui Medical University were retrospectively enrolled. PIK3CA hotspot mutations were analyzed using real-time quantitative PCR, and microsatellite instability (MSI) status was evaluated. Patient-derived organoid (PDO) models, including PIK3CA-mutant and wild-type organoids, were established to evaluate drug sensitivity to the PI3Kα inhibitor alpelisib and chemotherapeutic agents, as well as potential synergistic effects of combination therapy. Results: PIK3CA mutations were identified in 16 of 285 gastric cancers (5.6%). Mutations were predominantly clustered within the helical domain (E542K, E545K, and Q546R) and kinase domain (H1047R and H1047L). Compared with wild-type tumors, PIK3CA-mutant gastric cancers showed a significantly higher proportion of female patients (62.5% vs. 24.5%, P = 0.002) and increased MSI-H frequency (25.0% vs. 6.3%, P = 0.028). All helical-domain mutations occurred in microsatellite-stable tumors, whereas MSI-H status was predominantly associated with kinase-domain mutations, particularly H1047R. Pathologically, most PIK3CA-mutant tumors exhibited aggressive features, including deep invasion, lymph node metastasis, neural invasion, and lymphovascular invasion. PDO-based drug sensitivity assays demonstrated that PIK3CA-mutant organoids were markedly more sensitive to alpelisib than wild-type organoids (median GI50: 2.77 μM vs. 11.93 μM). Enhanced sensitivity to 5-fluorouracil and oxaliplatin was also observed in mutant PDOs. Combination treatment analysis revealed that alpelisib combined with paclitaxel exhibited the strongest synergistic effect in PIK3CA-mutant PDOs, particularly in the H1047R/MSI-H model. Conclusions: PIK3CA-mutant gastric cancers exhibit substantial domain-specific molecular heterogeneity. Kinase-domain mutations are strongly associated with MSI-H status and may confer increased sensitivity to PI3Kα inhibitor–based combination therapy. These findings provide additional evidence supporting molecular stratification and precision therapeutic strategies for PIK3CA-mutant gastric cancer.

Background Immune imbalance in tumor microenvironment is closely linked to GC progression. MDSCs primarily drive immune imbalance by suppressing the anti-tumor activity of CD8⁺T cells. Notably, no prior study has explored the combination of CD8⁺T cells and MDSCs to assess the clinical prognosis or immunotherapy response in GC. Methods PubMed and EMBASE were searched, literature screened, and 5 eligible studies included for this first meta-analysis. Multiplex immunofluorescence detected the expression of CD11B, CD33, CD8, and PD-L1 in 236 GC patients. R software was used to construct the Nomogram model. Receiver operating characteristic curves, calibration curves, and decision curve analysis were applied to evaluate the accuracy, reliability, and clinical utility of the prediction model. In vitro and in vivo experiments explored the impact of MDSCs on the migration and invasion of GC cells. Multicolor flow cytometry measured the infiltration of CD8⁺T cells and MDSCs in the peripheral blood of patients with GC and GCLM. Single-cell RNA sequencing data and spatial transcriptomics data were compared to further validate the results. Results Meta-analysis showed high MDSC infiltration correlated with poor prognosis and advanced stage in GC patients. The AUC of the CD8⁺T/MDSC ratio for predicting OS in GC patients was significantly superior to that of CD8⁺T cells alone or MDSCs alone. A low CD8⁺T/MDSC ratio (≤ 5.38) was closely associated with lymph node metastasis and advanced TNM stage in GC patients. In vitro and in vivo experiments demonstrated MDSCs significantly enhanced the proliferation, migration, invasion, and liver metastatic capabilities of GC cells, while MDSC inhibitors reversed this pro-tumor effect. Multicolor flow cytometry indicated significantly increased MDSC infiltration and a decreased CD8⁺T/MDSC ratio in patients with liver metastasis, and scRNA-seq and spatial transcriptomics results corroborated these findings. Among GC patients treated with sintilimab, the therapeutic response rate was significantly higher in the high CD8⁺T/MDSC ratio group than in the low ratio group, establishing this ratio as the first reported composite immune biomarker for predicting sintilimab response in GC patients. Conclusions This is the first study to validate the CD8⁺T/MDSC ratio as a novel, reliable predictive biomarker for GC prognosis, metastasis, and immunotherapy response.

Abstract Background Adenocarcinomas often exhibit a poor response to immunotherapy due to their "cold tumor" nature and immunosuppressive microenvironment. Cancer-testis antigens (CTAs), which are selectively expressed in tumors but absent in most normal tissues, represent ideal targets for tumor-specific therapies. Despite promising individual CTAs like MAGE-A, MSLN, and PRAME, a systematic analysis of their role across various adenocarcinomas is lacking. Methods In this study, we explored the expression profiles, prognostic significance, and immunomodulatory roles of cancer-testis antigens (CTAs) across various types of adenocarcinomas. By integrating data from The Cancer Genome Atlas (TCGA) with a validation cohort of 198 patients, comprising cases of colon, rectal, and lung adenocarcinomas, we analyzed six CTAs (CT83, MAGEA1, MAGEA4, MAGEA10, MSLN, and PRAME) utilizing survival analysis and immunohistochemistry techniques. Results Our findings revealed that CTAs exhibit tumor type-specific expression profiles. Notably, MSLN was highly expressed in colon adenocarcinoma (COAD, 73%), rectal adenocarcinoma (READ, 69%), and lung adenocarcinoma (LUAD, 72%), whereas PRAME was exclusively expressed in LUAD (24%). Elevated levels of CT83 ( p = 0.026) and MAGEA10 ( p = 0.036) in COAD, MAGEA1 ( p = 0.028) in READ, MAGEA1 ( p = 0.008) and MAGEA4 ( p = 0.009) in LUAD, were significantly associated with shorter progression-free survival (PFS). Additionally, exploratory CTA-count analysis showed that the prognostically relevant cutoff differed across tumor types, with more than one CTA-positive marker associated with poorer progression-free survival in COAD and more than two CTA-positive markers associated with poorer progression-free survival in READ and LUAD (COAD: p = 0.008; READ: p = 0.012; LUAD: p = 0.045). In LUAD, CT83 and MAGEA1 expression were significantly associated with lower infiltration of B cells, CD4 + T cells, and dendritic cells (all p < 0.01), and with higher infiltration of immunosuppressive myeloid-derived suppressor cells (MDSCs, p < 0.001) and FOXP3 + regulatory T cells (Tregs, p < 0.05). These effects were corroborated in the validation cohort of COAD (MAGEA4-Tregs, p < 0.05) and LUAD (CT83-Tregs, p < 0.05). Conclusions We found that CTA expression was highly tumor-type-specific in adenocarcinomas. These findings provide preliminary evidence for the potential relevance of CTA-based stratification in adenocarcinomas and may inform future mechanistic studies and prospective evaluation of precision immunotherapeutic strategies.

Submucosal gland differentiation (SGD), including acinar and ductal differentiation, has been detected in different subtypes of esophageal carcinoma, complicating tumor classification and diagnosis. This study aims to delineate the histomorphologic characteristics and to reveal the genetic alterations and precursor squamous dysplasia of the esophageal carcinomas with SGD. A panel of immunohistochemical markers was used to recognize and subcategorize the SGD. Next-generation sequencing (NGS) was performed on selected cases. Group 1 retrospectively included 50 pure type BSCCs with SGD. Group 2 included 25 carcinomas with ductal differentiation. In group 1, the integral branching morphology was observed in 20

Non-small cell lung carcinomas showing co-expression of TTF-1 and p40 in sampled tumor tissue are rare, diagnostically challenging, and not formally recognized in the current WHO classification of thoracic tumors. We analyzed 15 pulmonary carcinomas with TTF-1/p40 co-expression to characterize their clinicopathologic features and molecular alterations. The cohort consisted predominantly of male patients and was composed mainly of high-grade, poorly differentiated tumors. All cases demonstrated diffuse nuclear co-expression of TTF-1 and p40 by immunohistochemistry. Whole-exome sequencing, performed in 12 cases, revealed frequent TP53 alterations (9/12), occasional concurrent RB1 alterations, and a rare FGFR3::TACC3 fusion, which was further validated by FISH. Two tumors with prominent lymphoid stroma were positive for Epstein-Barr virus-encoded RNA and lacked TP53 alterations, suggesting additional biological diversity within this phenotype. Follow-up data were available for 13 patients (range, 1-29 months); 4 died of disease and 9 were alive at last follow-up. Two additional patients were lost to follow-up. Collectively, these findings suggest that pulmonary carcinomas showing TTF-1/p40 co-expression in sampled tissue comprise a heterogeneous clinicopathologic and molecular group rather than a currently definable distinct entity. Despite the small sample size and the inclusion of several biopsy-only cases, this study provides a descriptive basis for future investigation of this unusual immunophenotypic pattern.

BACKGROUND:Renal neuroendocrine tumors (RenNETs) are exceptionally rare neoplasms, accounting for less than 1 % of all renal tumors. Their clinicopathologic features, molecular characteristics, and biological behavior remain inadequately understood. This study aims to provide a comprehensive clinicopathologic and molecular analysis of RenNETs to distinguish them from other renal and neuroendocrine tumors. METHODS:A total of seven cases of RenNETs diagnosed between January 2017 and June 2023 were retrospectively analyzed. Clinical and pathologic features were reviewed, and immunohistochemical staining was performed. High-throughput next-generation sequencing (NGS) targeting 425 cancer-related genes was conducted on formalin-fixed, paraffin-embedded tumor samples RESULTS: Among the seven cases, six were non-functional RenNETs and one was associated with Cushing syndrome. Histologically, the tumors exhibited trabecular and solid growth patterns, with neuroendocrine features evident in the tumor cells. Immunohistochemically, all cases were positive for synaptophysin, and a subset showed expression of chromogranin A, INSM1, and β-catenin. NGS revealed 57 distinct somatic mutations, with TP53 (57.1 %) and APC (42.9 %) being the most frequently mutated genes. Additional recurrent alterations included POLE, ERCC6, ABCC2, and ALK. MCL1 amplification was observed in two cases, suggesting activation of anti-apoptotic pathways CONCLUSIONS: RenNETs represent a biologically distinct subtype of neuroendocrine tumors, with unique molecular features such as TP53 and APC mutations and frequent alterations in DNA repair pathways. These findings underscore the necessity for molecular profiling in guiding the diagnosis and potential therapeutic strategies for RenNETs. Further studies are warranted to explore the mechanisms underlying their pathogenesis and clinical behavior.

Gastric and gastroesophageal junction adenocarcinoma (GC/GEJC) poses a substantial clinical burden in China. This updated guideline introduces a refined human epidermal growth factor receptor 2 (HER2) classification system (high, intermediate, low, absent) to redefine patient stratification for targeted therapy. It integrates cutting-edge evidence on standardized detection-including immunohistochemistry, in situ hybridization, and liquid biopsy-to overcome spatial-temporal heterogeneity. Recent progress in the use of HER2-targeted agents, including monoclonal antibodies, antibody-drug conjugates, tyrosine kinase inhibitors, and bispecific antibodies, is systematically reviewed and presented to inform evidence-based clinical practice. These updates aim to optimize personalized HER2-targeted therapy and improve outcomes in GC/GEJC patients.

Esophageal basaloid squamous cell carcinoma (EBSCC) is a rare subtype of esophageal squamous cell carcinoma (ESCC) that might be misdiagnosed or missed in clinical practice. Through RNA sequencing on 20 pure EBSCC and 11 poorly differentiated ESCC samples, we identified CSPG4 as a potential marker for EBSCC at the mRNA level, and verified with CSPG4 immunohistochemical staining in these cases. Then we assessed the value of CSPG4 expression in differential diagnosis and prognosis in EBSCC cases (n = 360) and conventional ESCC cases (n = 160) from 11 different institutions with whole-tissue sections. CSPG4 had acceptable discriminatory capacity for EBSCC, with an AUC ROC of 0.891. The optimal cutoff value for the H-score determined by ROC curve analysis was 77.5, with 82.7% sensitivity and 79.9% specificity. In well-moderately differentiated ESCC with CSPG4 expression, staining was mostly observed on the edge of the tumor nest or infiltration front, with different expression pattern from EBSCC. The H-scores of CSPG4 expression for the EBSCC component were higher than those for other components in the same tissue section (P < 0.05). The expression level of CSPG4 was similar in groups with different percentages of the EBSCC component (P > 0.05). In additional 505 ESCC patients, patients with high CSPG4 expression had decreased DFS and OS, especially in stage I-II disease (P < 0.001). The similar prognostic significance was also found in EBSCC. Our data indicate that CSPG4 is not only a sensitive but also a specific marker for the diagnosis of EBSCC. CSPG4 might also be a prognostic marker for ESCC.

Accurate segmentation and classification of glomeruli are fundamental to histopathology slide analysis in renal pathology, which helps to characterize individual kidney disease. Accurate segmentation of glomeruli of different types faces two main challenges compared to traditional primitives segmentation in computational image analysis. Limited by small kernel size, traditional convolutional neural networks could hardly understand the complete context information of different glomeruli. Moreover, typical semantic segmentation networks lack adequate attention to difficult glomerular samples during the training process due to serious class imbalance between different glomeruli types. We propose a new deep learning approach, Glo-Net, which accurately segments and classifies glomeruli based on digitized pathology slides. Specifically, Glo-Net divides the traditional semantic segmentation network into two branches, i.e., segmentation and classification. While the segmentation branch specifically aims at localizing and delineating the boundary of individual glomerulus, the classification branch could focus on differentiating the glomerular types based on segmented pixels. In addition, an innovative loss function is added to the classification task to compensate for the class imbalance and minor types of glomeruli. The proposed network's average accuracy and F-score in classification tasks on the multi-institution datasets (including an external validation set) are 0.858 and 0.704, respectively. The average intersection over union (IoU) in segmentation tasks is 0.866. The Glo-Net demonstrates a 5 % improvement in classification accuracy, with up to 14 % increases for minor classes and an average 6 % IoU increase for segmentation tasks. Quantitative results show that our network achieves overall higher accuracy for segmentation and classification among nine subtypes of glomeruli compared to previous work with improved robustness and generalizability.

MiR-646, a small non-coding RNA, poorly expressed in a variety of tumors. This study aimed to clarify the role of miR-646 and its underlying mechanisms in glioblastoma (GBM). In our study, we found that miR-646 mRNA levels were lower in tumor tissues than in non-cancer tissues. The ability of glioma cells to proliferate, invade, and migrate is diminished by miR-646 overexpression in vitro and in vivo. Mechanistically, miR-646 targeted sequestosome 1 (p62) in the 3'UTR and affected the Keap1/Nrf2 pathway, thus attenuating the expression of the HO-1 gene. In conclusion, this study provided a novel finding that miR-646 tampered with gliomagenesis by regulating the p62/Keap1/Nrf2 axis, which provides a potential target for GBM therapy.

BACKGROUND:Glioma is the most prevalent and lethal tumor of the central nervous system. Routine treatment with temozolomide (TMZ) would unfortunately result in inevitable recurrence and therapy resistance, severely limiting therapeutic efficacy. Tumor-associated astrocytes (TAAs) are key components of the tumor microenvironment and increasing evidence has demonstrated that aberrant expression of connexin43 (Cx43) was closely associated with glioma progression and TMZ resistance. However, the specific role of Cx43 in mediating TMZ resistance through glioma and astrocyte interactions has not been fully explored. METHODS:The expression and prognostic value of Cx43 were evaluated in tumor samples and clinical databases. ShRNA-medicated knockdown and Gfap-Cre Cx43flox/flox gene mouse were used to assess the role and functional significance of Cx43 in vitro and in vivo. Moreover, we performed mass spectrometry analysis, chromatin immunoprecipitation, and other biochemical assays to define the molecular mechanisms by which Cx43 promotes TMZ resistance. RESULTS:We confirmed that the upregulation of Cx43 expression between TAAs and glioma cells contributed to TMZ resistance and tumor recurrence. Genetic knockdown or pharmacological inhibition of Cx43 enhanced TMZ-induced cytotoxicity. Mechanistically, elevated Cx43 expression induced β-catenin accumulation at the cell surface of glioma cells, suppressing T-cell factor/lymphoid enhancer-binding factor transcription. This led to impaired miR-205-5p expression and subsequent activation of the E2F1/ERCC1 axis, which eventually led to chemoresistance. CONCLUSIONS:Our study reveals a novel regulatory mechanism in which the Cx43/miR-205-5p/E2F1/ERCC1 axis contributes to TMZ resistance in glioma. These findings further highlight the potential of targeting Cx43 as a therapeutic strategy in glioma.

AIM:Microsatellite instability (MSI) as a result of deficient deoxyribonucleic acid (DNA) mismatch repair (dMMR) is a key contributor to the development of tumors with a high mutation rate and cancer-specific neoantigens. dMMR identification can be beneficial for selection of immune checkpoint inhibitor (ICI) therapy-eligible patients. While multiple studies have focused on dMMR prevalence in colorectal cancer (CRC), fewer investigate the prevalence of dMMR in tumor types besides CRC, especially in Chinese patients. In this study, we aimed to determine the prevalence of dMMR in China across five gastrointestinal and gynecological tumor types. METHODS:Tissue samples from Chinese patients with advanced endometrial, ovarian, cervical, biliary tract, or gastric metastatic or unresectable solid tumors were tested for dMMR status using immunohistochemistry with the Ventana MMR RxDx panel. Data were analyzed to determine the prevalence of dMMR for each tumor type. RESULTS:A total of 748 patients were included in the study, representing five tumor types. Prevalence of dMMR varied across tumor types, with an overall prevalence of 9.4%. Patients with endometrial tumors had the highest proportion of patients with dMMR at 49/164 (29.9%). Patients with cervical tumors had the lowest prevalence of dMMR with 6/221 (2.7%) patients. The prevalence of dMMR was similar across most demographic characteristics. In the dMMR population, co-occurring MLH1 and PMS2 protein loss across all tumor types was observed most commonly, in 48/70 (68.6%) patients. CONCLUSIONS:These data highlight the importance of dMMR testing in patients with advanced solid tumors in China to optimize biomarker testing and treatment decisions.

AimsCurrently, the clinicopathological characteristics of gastric cancer (GC) with oncogenic NTRK alterations are not well known. Although NTRK fusion has been identified as prevalent in DNA mismatch repair protein deficient (dMMR) colorectal cancer (CRC), the relationship between NTRK alterations and dMMR protein expression in GC has not been previously explored. MethodsOur study comprised 51 cases of EBV(Epstein-barr virus)-associated gastric carcinomas, 94 cases of dMMR GC, 90 cases of gastric adenocarcinoma with hepatoid or enteroblastic differentiation (GAHED) and 256 cases of conventional GC. Furthermore, to investigate the connection between NTRK fusion and dMMR proteins, we collected dMMR tumours of various types, including 21 cases of duodenal adenocarcinomas, 46 endometrioid carcinomas and 82 CRCs. NTRK fusion and amplification were screened in GC and various types of dMMR tumours using fluorescence in situ hybridisation (FISH), while cases positive for FISH translocation underwent next-generation sequencing testing. ResultsOur findings revealed the existence of two cases each of NTRK fusions and NTRK amplifications, which were all enriched in case of GAHED. Additionally, following an analysis of several types of cancers, we discovered that NTRK gene alterations were only present in dMMR CRC. ConclusionsOur results indicate that NTRK gene alterations are not enriched in GC with dMMR but are specifically enriched in cases of GAHED.

肝未分化胚胎性肉瘤（undifferentiated embryonal sarcoma of the liver，UESL）是一种侵袭性恶性肿瘤，主要发生于儿童，偶有发生于成人的病例报道且预后极差。由于临床症状和影像学表现的非特异性，术前诊断的准确率低，主要依靠病理组织学及免疫组织化学检查明确诊断。本文报道1例成人多发性肝未分化胚胎性肉瘤，重点从病理检查、免疫组织化学及鉴别诊断等方面进行介绍，以提高对成人UESL的认识。

Esophageal basaloid squamous cell carcinoma (BSCC) is a heterogenous entity with multilineage differentiation. It lacks systematical analysis on submucosal gland differentiation (SGD) due to the histological diversity and low incidence of esophageal BSCC. This study aims to find the correlation of SGD and clinicopathological features. A total of 152 esophageal BSCCs were separated into three histological groups: pure, mixed, and borderline group. The clinicopathological features were compared between different groups. The prevalence of SGD was also compared between cases of different groups. A panel of antibodies were used to identify SGD. The pure group differed from the mixed and borderline groups in many aspects, lymph node metastasis (LNM), cancer embolus, perineural invasion, and advanced stage occurred less frequently in the pure group ( P <0.01). The pure group had a better but statistically insignificant overall survival ( P =0.097). The squamous cell carcinoma (SCC) component or focal squamous differentiation was present in metastatic lymph nodes in almost all mixed BSCCs (95.7%, 22/23) with LNM. The LNM rate of superficial (T1b) BSCCs (17.6%, 6/34) was comparable to that of superficial (T1b) SCCs (18.5%, 57/308). However, LNM exclusively occurred in superficial mixed (3/5) and borderline (3/10) BSCCs. The IHC results demonstrated a prevalence of SGD in pure group (77%, 43/56). SGD is considered to be a favorable factor, while the squamous differentiation or invasive SCC component is an adverse factor in esophageal BSCCs. Refinement of classification is a promising way to improve patient management.