Neutrophil-to-lymphocyte ratio and platelet-to-lymphocyte ratio: Markers predicting immune-checkpoint inhibitor efficacy and immune-related adverse events.

We conducted a comprehensive review of existing prediction models pertaining to the efficacy of immune-checkpoint inhibitor (ICI) and the occurrence of immune-related adverse events (irAEs). The predictive potential of neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) in determining ICI effectiveness has been extensively investigated, while limited research has been conducted on predicting irAEs. Furthermore, the combined model incorporating NLR and PLR, either with each other or in conjunction with additional markers such as carcinoembryonic antigen, exhibits superior predictive capabilities compared to individual markers alone. NLR and PLR are promising markers for clinical applications. Forthcoming models ought to incorporate established efficacious models and newly identified ones, thereby constituting a multifactor composite model. Furthermore, efforts should be made to explore effective clinical application approaches that enhance the predictive accuracy and efficiency.

Efficient gene delivery by multifunctional star poly (ß-amino ester)s into difficult-to-transfect macrophages for M1 polarization.

Gene delivery to macrophages holds great promise for cancer immunotherapy. However, traditional gene delivery methods exhibit low transfection efficiency in macrophages. The star-shaped topological structure of polymers is known to encapsulate genes inside their cores, thereby facilitating sustained release of the genetic material. Herein, combining the structural advantages of star polymers and the transfection advantages of poly (ß-amino ester)s (PAEs), we developed a novel linear oligomer grafting-onto strategy to synthesize a library of multi-terminal star structured PAEs (SPAEs), and evaluated their gene delivery efficiency in various tissue cells. The transfection with human hepatocellular carcinoma cells (HepG2, HCC-LM3 cells and MHCC-97H cells), rat normal liver cells (BRL-3 A cells), human ovarian cancer cells (A2780 cells), African green monkey kidney cells (Vero cells), human cervical cancer cells (HeLa cells), human chondrosarcoma cells (SW1353 cells), and difficult-to-transfect human epidermal keratinocytes (HaCaT cells) and normal human fibroblast cells (NHF cells) showed that SPAEs exhibited superior transfection profile. The GFP transfection efficiency of top-performing SPAEs in HeLa cells (96.1%) was 2.1-fold, and 3.2-fold higher compared to jetPEI and Lipo3000, respectively, indicating that the star-shaped topological structure can significantly enhance the transfection efficiency of PAEs. More importantly, the top-performing SPAEs could efficiently deliver Nod2 DNA to difficult-to-transfect RAW264.7 macrophages, with a high transfection efficiency of 33.9%, which could promote macrophage M1 polarization and enhanced CD8+ T cell response in co-incubation experiments. This work advances gene therapy by targeting difficult-to-transfect macrophages and remodeling the tumor immune microenvironment.

Inhibiting PLA2G7 reverses the immunosuppressive function of intratumoral macrophages and augments immunotherapy response in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is an exceptionally immunosuppressive malignancy characterized by limited treatment options and a dismal prognosis. Macrophages constitute the primary and heterogeneous immune cell population within the HCC microenvironment. Our objective is to identify distinct subsets of macrophages implicated in the progression of HCC and their resistance to immunotherapy. METHODS: Intratumoral macrophage-specific marker genes were identified via single-cell RNA sequencing analyses. The clinical relevance of phospholipase A2 Group VII (PLA2G7), a pivotal enzyme in phospholipid metabolism, was assessed in patients with HCC through immunohistochemistry and immunofluorescence. Flow cytometry and an in vitro co-culture system were used to elucidate the specific role of PLA2G7 in macrophages. Orthotopic and subcutaneous HCC mouse models were employed to evaluate the potential of the PLA2G7 inhibitor in complementing immune checkpoint blockade (ICB) therapy. RESULTS: Single-cell RNA sequencing analyses disclosed predominant PLA2G7 expression in intratumoral macrophages within the HCC microenvironment. The macrophage-specific PLA2G7 was significantly correlated with poorer prognosis and immunotherapy resistance in patients with HCC. PLA2G7high macrophages represent a highly immunosuppressive subset and impede CD8 T-cell activation. Pharmacological inhibition of PLA2G7 by darapladib improved the therapeutic efficacy of anti-programmed cell death protein 1 antibodies in the HCC mouse models. CONCLUSIONS: Macrophage-specific PLA2G7 serves as a novel biomarker capable of prognosticating immunotherapy responsiveness and inhibiting PLA2G7 has the potential to enhance the efficacy of ICB therapy for HCC.

Inactivated vaccines reduce the risk of liver function abnormality in NAFLD patients with COVID-19: a multi-center retrospective study.

BACKGROUND: Abnormal liver function was frequently observed in nonalcoholic fatty liver disease (NAFLD) patients infected with SARS-CoV-2. Our aim was to explore the effect of SARS-CoV-2 inactivated vaccines on liver function abnormality among NAFLD patients with COVID-19. METHODS: The multi-center retrospective cohort included 517 NAFLD patients with COVID-19 from 1 April to 30 June 2022. Participants who received 2 doses of the vaccine (n = 274) were propensity score matched (PSM) with 243 unvaccinated controls. The primary outcome was liver function abnormality and the secondary outcome was viral shedding duration. Logistic and Cox regression models were used to calculate the odds ratio (OR) and hazard ratio (HR) for the outcomes. Sensitivity analysis was conducted to assess robustness. FINDINGS: PSM identified 171 pairs of vaccinated and unvaccinated patients. Liver function abnormality was less frequent in the vaccinated group (adjusted OR, 0.556 [95% CI (confidence interval), 0.356-0.869], p = 0.010). Additionally, the vaccinated group demonstrated a lower incidence of abnormal bilirubin levels (total bilirubin: adjusted OR, 0.223 [95% CI, 0.072-0.690], p = 0.009; direct bilirubin: adjusted OR, 0.175 [95% CI, 0.080-0.384], p < 0.001) and shorter viral shedding duration (adjusted HR, 0.798 [95% CI, 0.641-0.994], p = 0.044) than the unvaccinated group. Further subgroup analysis revealed similar results, while the sensitivity analyses indicated consistent findings. INTERPRETATION: SARS-CoV-2 vaccination in patients with NAFLD may reduce the risk of liver dysfunction during COVID-19. Furthermore, vaccination demonstrated beneficial effects on viral shedding in the NAFLD population. FUNDING: 23XD1422700, Tszb2023-01, Zdzk2020-10, Zdxk2020-01, 2308085J27 and JLY20180124.

The Pyroptosis-Related Signature Composed of GSDMC Predicts Prognosis and Contributes to Growth and Metastasis of Hepatocellular Carcinoma.

BACKGROUND: Pyroptosis-related genes (PRG) are closely associated with the progression and metastasis of hepatocellular carcinoma (HCC). The predictive power of PRGs could be used to assess the clinical outcomes of HCC. METHODS: The Cancer Genome Atlas (TCGA) RNA-seq data and clinical information from patients with liver hepatocellular carcinoma (LIHC) were used to identify PRG with differentially expressed between HCC and normal samples. Univariate Cox regression, least absolute shrinkage and selection operator (LASSO) Cox method, and multivariate Cox regression analysis were used to develop a prognostic model that included three PRGs. Gene set enrichment analysis (GSEA) was performed to identify differential immune cells and their associated pathways. The expression of Gasdermin C (GSDMC) in the HCC samples was detected by western blotting, and the function of GSDMC in HCC proliferation and metastasis was detected by the Cell Counting Kit-8 (CCK-8), colony formation, cell invasion, and wound healing assays. RESULTS: Of 52 PRGs, GSDMC, Bcl-2 homologusantagonist/ killer 1 (BAK1), and NOD-like receptor thermal protein domain associated protein 6 (NLRP6) were selected to establish a prognostic model. The model successfully differentiated HCC patients with varied survival in the TCGA training and test cohorts, as well as the International Cancer Genome Consortium (ICGC) validation cohorts. The risk score was proven to be an independent prognostic factor. In addition, we also reported a marked upregulation of GSDMC in HCC tissues, which could be induced by CD274 (PD-L1). Overexpression of GSDMC contributes to HCC cells invasion, proliferation, and migration. CONCLUSIONS: The three PRGs signatures containing GSDMC independently predicted HCC prognosis. As a new driver molecule, GSDMC could play a tumor-promoting role by facilitating HCC growth and metastasis.

Eleven metabolism­related genes composed of Stard5 predict prognosis and contribute to EMT phenotype in HCC.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, with a high mortality and poor survival rate. Abnormal tumor metabolism is considered a hallmark of HCC and is a potential therapeutic target. This study aimed to identify metabolism-related biomarkers to evaluate the prognosis of patients with HCC. METHOD: The Cancer Genome Atlas (TCGA) database was used to explore differential metabolic pathways based on high and low epithelial-mesenchymal transition (EMT) groupings. Genes in differential metabolic pathways were obtained for HCC metabolism-related molecular subtype analysis. Differentially expressed genes (DEGs) from the three subtypes were subjected to Lasso Cox regression analysis to construct prognostic risk models. Stard5 expression in HCC patients was detected by western blot and immunohistochemistry (IHC), and the role of Stard5 in the metastasis of HCC was investigated by cytological experiments. RESULTS: Unsupervised clustering analysis based on metabolism-related genes revealed three subtypes in HCC with differential prognosis. A risk prognostic model was constructed based on 11 genes (STARD5, FTCD, SCN4A, ADH4, CFHR3, CYP2C9, CCL14, GADD45G, SOX11, SCIN, and SLC2A1) obtained by LASSO Cox regression analysis of the three subtypes of DEGs. We validated that the model had a good predictive power. In addition, we found that the high-risk group had a poor prognosis, higher proportion of Tregs, and responded poorly to chemotherapy. We also found that Stard5 expression was markedly decreased in HCC tissues, which was associated with poor prognosis and EMT. Knockdown of Stard5 contributed to the invasion and migration of HCC cells. Overexpression of Stard5 inhibited EMT in HCC cells. CONCLUSION: We developed a new model based on 11 metabolism-related genes, which predicted the prognosis and response to chemotherapy or immunotherapy for HCC. Notably, we demonstrated for the first time that Stard5 acted as a tumor suppressor by inhibiting metastasis in HCC.

CD36-BATF2\MYB Axis Predicts Anti-PD-1 Immunotherapy Response in Gastric Cancer.

Despite the utilization of anti-PD-1 therapy in gastric cancer (GC), the absence of a reliable predictive biomarker continues to pose a challenge. In this study, we utilized bioinformatic analysis and immunohistochemistry to develop a prediction model for activated CD4+ memory T cells, considering both mRNA and protein levels. An elevation of activated CD4+ memory T cells in GC was noted, which exhibited a strong association with the patients' overall survival. By utilizing WGCNA and DEG analysis, we discovered that BATF2, MYB, and CD36 are genes that exhibit differential expression and are linked to activated CD4+ memory T cells. Afterwards, a forecast model was built utilizing Stepwise regression and immunohistochemistry relying on the three genes. The model's high-risk score showed significant associations with a suppressive immune microenvironment. Moreover, our model exhibited encouraging prognostic value and superior performance in predicting response to immune checkpoint blockade therapy compared with the conventional CD8+PD-L1 model. In terms of mechanism, CD36 could function as a receptor upstream that identifies Helicobacter pylori and fatty acids. This recognition then results in the reduction of the BATF2-MYB protein complex and subsequent alterations in the transcription of genes associated with classical T cell activation. As a result, the activation state of CD4+ memory T cells is ultimately suppressed. The CD36-BATF2/MYB signature serves as a robust predictor of anti-PD-1 immunotherapy response in GC.

Six immune-related promising biomarkers may promote hepatocellular carcinoma prognosis: a bioinformatics analysis and experimental validation.

BACKGROUND: Abnormal miRNA and mRNA expression and dysregulated immune microenvironment have been found to frequently induce the progression of hepatocellular carcinoma (HCC) in recent reports. In particular, the immune-related competing endogenous RNAs (ceRNA) mechanism plays a crucial role in HCC progression. However, the underlying mechanisms remain unclear. METHODS: Differentially expressed immune-related genes were obtained from the Immport, GEO, and TCGA databases. The mRNA and protein expression levels in HCC tissues and adjacent normal tissues were confirmed, and we further investigated the methylation levels of these biomarkers to explore their function. Then, the TIMER and TISCH databases were used to assess the relationship between immune infiltration and hub genes. Survival analysis and univariate and multivariate Cox models were used to evaluate the association between hub genes and HCC diagnosis. Hub gene expression was experimentally validated in six HCC cell lines and 15 HCC samples using qRT-PCR and immunohistochemistry. The hub genes were uploaded to DSigDB for drug prediction enrichment analysis. RESULTS: We identified that patients with abnormal miRNAs (hsa-miR-125b-5p and hsa-miR-21-5p) and their targeted genes (NTF3, PSMD14, CD320, and SORT1) had a worse prognosis. Methylation analysis of miRNA-targeted genes suggested that alteration of methylation levels is also a factor in the induction of tumorigenesis. We also found that the development of HCC progression caused by miRNA-mRNA interactions may be closely correlated with the infiltration of immunocytes. Moreover, the GSEA, GO, and KEGG analysis suggested that several common immune-related biological processes and pathways were related to miRNA-targeted genes. The results of qRT-PCR, immunohistochemistry, and western blotting were consistent with our bioinformatics results, suggesting that abnormal miRNAs and their targeted genes may affect HCC progression. CONCLUSIONS: Briefly, our study systematically describes the mechanisms of miRNA-mRNA interactions in HCC and predicts promising biomarkers that are associated with immune filtration for HCC progression.

circRanGAP1/miR-27b-3p/NRAS Axis may promote the progression of hepatocellular Carcinoma.

BACKGROUND: Though circular RNAs (circRNAs) are the key regulators in tumor carcinogenesis, they remain largely unexplored in hepatocellular carcinoma (HCC). METHODS: The expression of RanGAP1-derived circRNAs (circ_0063531, circ_0063534, circ_0063513, circ_0063518, circ_0063507, circ_0063723) were evaluated in eight paired HCC and normal tissues, and the correlation between circRanGAP1 (circ_0063531) expression and clinicopathological characteristics in 40 HCC patients was determined. The association between miR-27b-3p and circRanGAP1 or NRAS was predicted using bioinformatics analysis. The expression of circRanGAP1, miR-27b-3p, and NRAS were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The potential oncogenic role of circ-RanGAP1 was assessed using CCK-8, colony formation, transwell assays in vitro, subcutaneous tumor mouse model, vein tail metastatic model, and orthotopically implanted intrahepatic HCC model in vivo. Luciferase reporter and RNA immunoprecipitation (RIP) assays were used to explore the binding site between miR-27b-3p and circ-RanGAP1 or NRAS. Protein expression was detected using western blotting. The localization of miR-27b-3p and circ-RanGAP1 was investigated using fluorescence in situ hybridization (FISH). The level of immune infiltration was assessed by bioinformatics analysis, flow cytometry, and orthotopically implanted intrahepatic HCC models. RESULTS: Here, we found elevated circRanGAP1 in the cells and clinical tissues of patients with HCC. Increased circRanGAP1 levels are associated with enlarged tumors and the advanced stage of TNM. CircRanGAP1 promotes the growth, migration, and HCC cell invasion, concurrently with the growth and metastasis of tumors in-vivo. Moreover, circRanGAP1 is mainly located inside the cytoplasm. Mechanistically, circRanGAP1 as an oncogene promotes HCC progression by miR-27b-3p/NRAS/ERK axis, furthermore, affects the infiltration level of tumor-associated macrophages probably by sponging miR-27b-3p. Immune infiltration analysis shows that NRAS is positively correlated with the levels of CD68+ tumor-associated macrophages in HCC samples and that NRAS and CD68 are related to the poor outcome of HCC. CONCLUSION: These results reveal that circRanGAP1 is a HCC oncogene that function by the miR-27b-3p/NRAS/ERK axis and regulates the infiltration levels of tumor-associated macrophages by sponging miR-27b-3p. Therefore, circRANGAP1/ NRAS axis may be an important potential treatment target against HCC.

Identification of prognostic miRNA-mRNA regulatory network in the progression of HCV-associated cirrhosis to hepatocellular carcinoma.

Background: Long-term hepatitis C virus (HCV) infection is strongly associated with hepatocellular carcinoma (HCC), yet the mechanisms of the progression process remain unclear. The research is aiming to establish a crucial prognostic model that indicates the risk of HCV-associated cirrhosis evolving into HCC. Methods: Differentially expressed microRNAs (DE-miRNAs) and differentially expressed genes (DEGs) between HCV-associated cirrhosis and HCC were screened from the GSE40744 and GSE6764 datasets, respectively. Downstream target genes of DE-miRNAs were predicted by the miRNet tool and then overlapped with the DEGs to select intersection genes. The GSE15654 was downloaded to establish a prognostic model. Expression levels of risk genes and their corresponding miRNAs were measured in liver tissues of clinical patients. HCC cell lines with UHRF1 knockdown or overexpression were assayed for cell proliferation and migration. Results: Thirty-nine DE-miRNAs and 796 DEGs are identified between HCV-associated cirrhosis and HCC. Main intersection genes and their corresponding miRNAs constitute a miRNA-mRNA regulatory network. PABPC1 (Polyadenylate-binding protein 1), SLC2A9 (solute carrier gene family 2, member 9), and UHRF1 (ubiquitin-like with PHD and ring finger domains 1) form a prognostic model indicating the risk of HCC development among HCV-associated cirrhosis. The genetic mutations of PABPC1, SLC2A9, and UHRF1 in HCC patients are 9%, 0.8%, and 0.6%, respectively. Compared to that in HCV-associated cirrhosis, the expression levels of PABPC1 and UHRF1 are higher while the expression level of SLC2A9 is lower in clinical HCV-associated HCC samples. UHRF1 enhances the proliferation and migration ability of HCC cells. Conclusions: PABPC1, SLC2A9, and UHRF1 and their corresponding miRNAs are involved in the evolution process of HCV-associated cirrhosis into malignant HCC. UHRF1 serves as an oncogene that promotes the proliferation and migration of HCC cells.

Future therapies for pancreatic carcinoma: Insights into cancer precision medicine.

Pancreatic carcinoma (PC) has one of the highest rates of cancer-related death worldwide. Except for surgery, adjuvant chemotherapy, chemoradiotherapy, and immunotherapy have shown various efficacies depending on the stage of the patient. We read the review "Current and emerging therapeutic strategies in pancreatic cancer: Challenges and opportunities" and offer some opinions that may improve its precision and completeness. This review presents a map of appropriate therapies for PC at different stages. Based on the clinical trial outcomes mentioned in the review, we evaluated the potential therapeutic options for PC and helped explain the contradictory efficacy between different programmed cell death protein 1/programmed cell death ligand 1 clinical trials, which may have resulted from the unique features of PC. Although R0 resection and adjuvant chemotherapy are still the gold standards for PC, new modalities, with or without clinical validation, are needed to establish more specific and precise treatments for PC.

Gut Microbiota and Immune Checkpoint Inhibitors-Based Immunotherapy.

Application of immune checkpoint inhibitors (ICIs) is a major breakthrough in the field of cancer therapy, which has displayed tremendous potential in various types of malignancies. However, their response rates range widely in different cancer types and a significant number of patients experience immune-related adverse effects (irAEs) induced by these drugs, limiting the proportion of patients who can truly benefit from ICIs. Gut microbiota has gained increasing attention due to its emerging role in regulating the immune system. In recent years, numerous studies have shown that gut microbiota can modulate antitumor response, as well as decrease the risk of colitis due to ICIs in patients receiving immunotherapy. The present review analyzed recent progress of relevant basic and clinical studies in this area and explored new perspectives to enhance the efficacy of ICIs and alleviate associated irAEs via manipulation of the gut microbiota.

The long non-coding RNA DKFZp434J0226 regulates the alternative splicing process through phosphorylation of SF3B6 in PDAC.

BACKGROUND: Long noncoding RNAs (lncRNAs), a type of pervasive genes that regulates various biological processes, are differentially expressed in different types of malignant tumors. The role of lncRNAs in the carcinogenesis of pancreatic ductal adenocarcinoma (PDAC) remains unclear. Here, we investigated the role of the lncRNA DKFZp434J0226 in PDAC. METHODS: Aberrantly expressed mRNAs and lncRNAs among six PDAC and paired non-tumorous tissues were profiled using microarray analysis. Quantitative real-time polymerase chain reaction was used to evaluate DKFZp434J0226 expression in PDAC tissues. CCK-8 assay, wound-healing assay, soft agar colony formation assay, and transwell assay were performed to assess the invasiveness and proliferation of PDAC cells. Furthermore, RNA pull-down, immunofluorescence, RNA immunoprecipitation, and western blotting assays were performed to investigate the association between DKFZp434J0226 and SF3B6. Tumor xenografts in mice were used to test for tumor formation in vivo. RESULTS: In our study, 222 mRNAs and 128 lncRNAs were aberrantly expressed (≥ twofold change). Of these, 66 mRNAs and 53 lncRNAs were upregulated, while 75 lncRNAs and 156 mRNAs were downregulated. KEGG pathway analysis and the Gene ontology category indicated that these genes were associated with the regulation of mRNA alternative splicing and metabolic balance. Clinical analyses revealed that overexpression of DKFZp434J0226 was associated with worse tumor grading, frequent perineural invasion, advanced tumor-node-metastasis stage, and decreased overall survival and time to progression. Functional assays demonstrated that DKFZp434J0226 promoted PDAC cell migration, invasion, and growth in vitro and accelerated tumor proliferation in vivo. Mechanistically, DKFZp434J0226 interacted with the splicing factor SF3B6 and promoted its phosphorylation, which further regulated the alternative splicing of pre-mRNA. CONCLUSIONS: This study indicates that DKFZp434J0226 regulates alternative splicing through phosphorylation of SF3B6 in PDAC and leads to an oncogenic phenotype in PDAC.

Activation of Platelet NLRP3 Inflammasome in Crohn's Disease.

Patients with Crohn's disease (CD) are inclined to have platelet hyperactivity and an increased risk of intestinal micro-thrombosis. However, the mechanisms underlying platelet hyperactivity in CD are not well understood. We investigated the assembly of platelet NLRP3 inflammasome in patients with active CD and its correlation with platelet hyperactivity. In this study, Real-time PCR and western blotting analyses uncovered that ASC, NLRP3, and active caspase-1 were significantly upregulated in platelets from patients with active CD compared with healthy subjects. As revealed by flow cytometry (FCM) and ELISA analyses, the levels of interleukin-1ß in both serum and isolated platelets were elevated in patients with active CD. Co-immunoprecipitation and immunofluorescence experiments revealed an increased assembly of NLRP3 inflammasome in platelets from patients with active CD. In addition, higher levels of intracellular reactive oxygen species (ROS) were observed in these platelets by FCM. Furthermore, elevated levels of platelet P-selectin exposure and fibrinogen binding were demonstrated in patients with active CD by FCM. They were positively correlated with the protein levels of NLRP3 inflammasome components. Collectively, our results indicate that the ROS-NLRP3 inflammasome-interleukin-1ß axis may contribute to platelet hyperactivity in active CD.

The role of Sphingomyelin synthase 2 (SMS2) in platelet activation and its clinical significance.

BACKGROUND: Sphingomyelin (SM) is an essential component of biological lipid rafts, and it plays an indispensable role in maintaining plasma membrane stability and in mediating signal transduction. The ultimate biosynthesis of SM is catalyzed by two sphingomyelin synthases (SMSs) namely SMS1 and SMS2, which are selectively distributed in the trans-Golgi apparatus and the plasma membrane. It has been demonstrated that SMS2 acts as an irreplaceable molecule in the regulation of transmembrane signaling, and loss of SMS2 has been reported to worsen atherosclerosis and liver steatosis. However, the function of SMS2 in platelet activation and its association with the pathological process of thrombosis in acute coronary syndrome (ACS) and portal hypertension (PH) remain unclear. METHODS: In this study, we tested the role of SMS2 in platelet activation and thrombosis using SMS2 knockout (SMS2 -/-) mice and SMS2-specific inhibitor, D609. Furthermore, we detected SMS2 expression in patients with ACS and PH. RESULTS: SMS2 -/- platelets showed significant reduction in platelet aggregation, spreading, clot retraction and in vivo thrombosis. Similar inhibitory effects on platelet activation were detected in D609-treated wild-type platelets. PLCγ/PI3K/Akt signaling pathway was inhibited in SMS2 -/- platelets and D609-treated wild-type platelets. In addition, we discovered that platelet SMS2 expression was remarkably increased in patients with ACS and PH, compared with healthy subjects. CONCLUSIONS: Our study indicates that SMS2 acts as a positive regulator of platelet activation and thrombosis, and provides a theoretical basis for the potential use of D609 in anti-thrombosis treatment.

Hepatic NOD2 promotes hepatocarcinogenesis via a RIP2-mediated proinflammatory response and a novel nuclear autophagy-mediated DNA damage mechanism.

BACKGROUND: Key hepatic molecules linking gut dysbiosis and hepatocarcinogenesis remain largely unknown. Gut-derived gut microbiota contains pathogen-associated molecular patterns (PAMPs) that may circulate into the liver and, consequently, be recognized by hepatic pattern recognition receptors (PRRs). NOD2, a general intracellular PRR, recognizes muramyl dipeptide (MDP), present in both gram (+) and gram (-) bacteria. Here, we investigated the role of NOD2 as a molecular sensor translating gut dysbiosis signaling into hepatocarcinogenesis. METHODS: NOD2 expression was measured in clinical hepatocellular carcinoma (HCC) samples using qPCR (80 pairs), western blotting (30 pairs) and immunostaining (141 pairs). The role of NOD2 in hepatocarcinogenesis was examined in the hepatocyte-specific Nod2-knockout (Nod2△hep), Rip2-knockout (Rip2△hep), Lamin A/C-knockout (Lamn△hep) and Rip2/Lamin A/C double-knockout (Rip2/Lamn△hep) mice models of diethylnitrosamine (DEN)/CCl4-induced HCC. RESULTS: NOD2 was upregulated and activated in HCC samples, and high NOD2 expression correlated with poor prognosis in HCC patients. Hepatic NOD2 deletion in vivo decreased DEN/CCl4-induced HCC by reducing the inflammatory response, DNA damage and genomic instability. NOD2 activation increased liver inflammation via RIP2-dependent activation of the MAPK, NF-κB and STAT3 pathways. Notably, a novel RIP2-independent mechanism was discovered, whereby NOD2 activation induces the nuclear autophagy pathway. We showed that NOD2 undergoes nuclear transport and directly binds to a component of nuclear laminae, lamin A/C, to promote its protein degradation, leading to impaired DNA damage repair and increased genomic instability. CONCLUSIONS: We reveal a novel bridge, bacterial sensor NOD2, linking gut-derived microbial metabolites to hepatocarcinogenesis via induction of the inflammatory response and nuclear autophagy. Thus, we propose hepatic NOD2 as a promising therapeutic target against HCC.

Expression and Clinical Prognostic Value of Platelet NLRP3 in Acute Coronary Syndrome.

PURPOSE: Little is known about the relationship between the level of platelet NOD-like receptor protein 3 (NLRP3) and the severity of acute coronary syndrome (ACS) or the prognostic value of platelet NLRP3 for percutaneous coronary intervention (PCI). METHODS: Platelets collected from 25 healthy subjects, 23 patients with stable angina pectoris (SAP), and 72 patients with ACS were analyzed by Western blotting and real-time fluorescence quantitative PCR (qPCR). A total of 152 patients with ACS who had undergone PCI were included in this study to evaluate the prognostic value of platelet NLRP3. RESULTS: The levels of platelet NLRP3 in both the healthy and SAP groups were clearly lower than in the ACS group (P<0.001). According to the Pearson correlation analysis, the expression of platelet NLRP3 was closely related to the mean platelet volume (MPV), left ventricular ejection fraction (LVEF), the Gensini score, and the Global Registry of Acute Coronary Events (GRACE) score (all P<0.001). Multivariate logistic regression analysis identified NLRP3 as an independent risk factor for adverse cardiovascular events (ACEs) after PCI (P=0.004). The proportion of patients with high NLPR3 expression (the NLRP3-high group) remaining free of adverse events for 3 years was remarkably lower than that in patients with low NLPR3 expression (the NLRP3-low group; P=0.024). The NLRP3-high group had a significantly higher proportion of patients with interleukin-1ß-expressing (20.4%±6.1%) platelets than the NLRP3-low group (10.7%±3.5%, P<0.001). Moreover, the NLRP3-high group exhibited higher platelet activity, as indicated by increased PAC-1 binding and CD62P expression, compared with the NLRP3-low group (P<0.001). CONCLUSION: These results indicated that platelet NLRP3 was a novel potential prognostic factor for patients with ACS that underwent PCI.

Tumor-associated tertiary lymphoid structure predicts postoperative outcomes in patients with primary gastrointestinal stromal tumors.

Tumor-infiltrating tertiary lymphoid structures (TLS) are thought to have anti-tumor activity and are believed to indicate a favorable prognosis in cancer patients. However, the prognostic value of TLS in gastrointestinal stromal tumors (GIST) is unknown. We evaluated the prognostic value of TLS using two independent GIST cohorts. Pathological examinations identified TLS in 44.9% of patients in our discovery cohort (DC). TLS was significantly associated with smaller tumor size (P = .011), relatively well morphological classification (P < .001), lower NIH classification (P < .001), lower recurrence (P = .005), longer survival time (P < .001) and lower imatinib resistance (P = .006). Kaplan-Meier curves showed that TLS was remarkably associated with favorable survival (P = .0002) and recurrence (P = .0015) time. In addition, the presence of KIT mutations and the absence of TLS suggested worst prognosis both in terms of overall survival (OS) (P = .0029) and time to recurrence (TTR) (P = .0150), while the presence of PDGFRA mutations and TLS suggested optimal prognosis for OS and TTR. Multivariate analyzes demonstrated that TLS was an independent prognostic factor for OS (HR:0.180, P = .002) and TTR (HR:0.412, P = .023). These results were confirmed using our validation cohort. Multiplexed immunohistochemistry staining was used to determine the composition of TLS. Therapies designed to target TLS may be a novel therapeutic strategy for GIST patients with imatinib resistance.

Comprehensive Characterization of Immunological Profiles and Clinical Significance in Hepatocellular Carcinoma.

BACKGROUND: Therapies targeting immune molecules have rapidly been adopted and advanced the treatment of hepatocellular carcinoma (HCC). Nonetheless, no studies have reported a systematic analysis between immunological profiles and clinical significance in HCC. METHODS: We comprehensively investigated immune patterns and systematically correlated 22 types of both adaptive and innate immune cells with genomic characteristics and clinical outcomes based on 370 HCC patients from The Cancer Genome Atlas (TCGA) database through a metagene approach (known as CIBERSORT). Based on the Quantitative Pathology Imaging and Analysis System coupled with integrated high-dimensional bioinformatics analysis, we further independently validated six immune subsets (CD4+ T cells, CD8+ T cells, CD20+ B cells, CD14+ monocytes, CD56+ NK cells, and CD68+ macrophages), and shortlisted three (CD4+ T cells, CD8+ T cells, and CD56+ NK cells) of which to investigate their association with clinical outcomes in two independent Zhongshan cohorts of HCC patients (n = 258 and n = 178). Patient prognosis was further evaluated by Kaplan-Meier analysis and univariate and multivariate regression analysis. RESULTS: By using the CIBERSORT method, the immunome landscape of HCC was constructed based on integrated transcriptomics analysis and multiplexed sequential immunohistochemistry. Further, the patients were categorized into four immune subgroups featured with distinct clinical outcomes. Strikingly, significant inter-tumoral and intra-tumoral immune heterogeneity was further identified according to the in-depth interrogation of the immune landscape. CONCLUSION: This work represents a potential useful resource for the immunoscore establishment for prognostic prediction in HCC patients.

Clinicopathological and prognostic implications of vessels encapsulate tumor clusters with PD-L1 in intrahepatic cholangiocarcinoma patients.

BACKGROUND: Frequently abnormal vascularization and immunologic derangement have been uncovered in malignant tumors. In present research, we evaluated prognostic characteristic and clinicopathological features of vessels encapsulate tumor clusters (VETC) and the immune checkpoint molecule, programmed cell death-ligand 1 (PD-L1) in patients diagnosed as intrahepatic cholangiocarcinoma (ICC). METHODS: VETC and PD-L1 were investigated in two cohort enrolling 412 ICC patients. VETC and PD-L1 was easily detectable in whole slides and tissue microarray (TMA). Prognostic analysis was performed through Kaplan-Meier cures, log-rank tests and nomograms. RESULTS: VETC+ was significantly associated with aggressive tumor features. VETC+ predicted a significantly unfavorable survival and higher metastasis and recurrence rates. Furthermore, nomograms integrated by the combination of VETC and PD-L1, that heralded better prognostic value compared to previous staging systems. CONCLUSIONS: Heterogeneous patterns of VETC phenotype and PD-L1 status were both illustrated to be an independent prognostic predictor for clinical outcomes. Therapies designed to target both vascularization and autoimmunity may open a novel direction for HCC. HCC should be replaced by ICC.