Investigation of lethal thresholds of nanosecond pulsed electric field in rabbit VX2 hepatic tumors through finite element analysis and verification with a single-needle bipolar electrode: A prospective strategy employing three-dimensional comparisons.

Pulsed electric field has emerged as a promising modality for the solid tumor ablation with the advantage in treatment planning, however, the accurate prediction of the lesion margin requires the determination of the lethal electric field (E) thresholds. Herein we employ the highly repetitive nanosecond pulsed electric field (RnsPEF) to ablate the normal and VX2 tumor-bearing livers of rabbits. The ultrasound-guided surgery is operated using the conventional double- and newly devised single-needle bipolar electrodes. Finite element analysis is also introduced to simulate the E distribution in the practical treatments. Two- and three-dimensional investigations are performed on the image measurements and reconstructed calcification models on micro-CT, respectively. Specially, an algorithm considering the model surface, volume and shape is employed to compare the similarities between the simulative and experimental models. Blood vessel injury, temperature and synergistic efficacy with doxorubicin (DOX) are also investigated. According to the three-dimensional calculation, the overall E threshold is 4536.4 ± 618.2 V/cm and the single-needle bipolar electrode is verified to be effective in tissue ablation. Vessels are well preserved and the increment of temperature is limited. Synergy of RnsPEF and DOX shows increased apoptosis and improved long-term tumor survival. Our study presents a prospective strategy for the evaluation of the lethal E threshold, which can be considered to guide the future clinical treatment planning for RnsPEF.

Blocking MARCO+ tumor-associated macrophages improves anti-PD-L1 therapy of hepatocellular carcinoma by promoting the activation of STING-IFN type I pathway.

The PD-L1/PD-1 axis is a classic immunotherapy target. However, anti-PD-L1/PD-1 therapy alone can not achieve satisfactory results in solid tumors, especially liver cancer. Among the several factors involved in tumor anti-PD-L1/PD-1 treatment resistance, tumor-associated macrophages (TAMs) have attracted attention because of their immunosuppressive ability. TAMs with a macrophage receptor with a collagenous structure (MARCO) are a macrophage subset group with strong immunosuppressive abilities. Clinical specimens and animal experiments revealed a negative correlation between MARCO + TAMs and patient prognosis with liver cancer. Transcriptional data and in vitro and in vivo experiments revealed that MARCO + TAM immunosuppressive ability was related to secretion. MARCO suppressed IFN-ß secretion from TAMs, reducing antigen presentation molecule expression, infiltration, and CD8+T cell dysfunction, thus producing an immunosuppressive microenvironment in liver cancer. MARCO can promote dying tumor cell clearance by macrophages, reducing tumor-derived cGAMP and ATP accumulation in the tumor microenvironment and inhibiting sting-IFN-ß pathway activation mediated by P2X7R in MARCO+TAMs. Animal experiments revealed that the MARCO and PD-L1 monoclonal antibody combination could significantly inhibit liver cancer growth. Conclusively, targeting MARCO+TAMs can significantly improve anti-PD-L1 resistance in liver cancer, making it a potential novel immune target for liver cancer therapy.

Nanosecond pulsed electric field stimulates CD103+ DC accumulation in tumor microenvironment via NK-CD103+ DC crosstalk.

CD103+ DC is crucial for antitumor immune response. As a promising local therapy on cancers, nanosecond pulsed electric field (nsPEF) has been widely reported to stimulate anti-tumor immune response, but the underlying relationship between intratumoral CD103+ DC and nsPEF treatment remains enigmatic. Here, we focused on the behavior of CD103+ DC in response to nsPEF treatment and explored the underlying mechanism. We found that the nsPEF treatment led to the activation and accumulation of CD103+ DC in tumor. Depletion of CD103+ DC via Batf3-/- mice demonstrated CD103+ DC was necessary for intratumoral CD8+ T cell infiltration and activation in response to nsPEF treatment. Notably, NK cells recruited CD103+ DC into nsPEF-treated tumor through CCL5. Inflammatory array revealed CD103+ DC-derived IL-12 mediated the CCL5 secretion in NK cells. In addition, the boosted activation and infiltration of intratumoral CD103+ DC were abolished by cGAS-STING pathway inhibition, following IL-12 and CCL5 decreasing. Furthermore, nsPEF treatment promoting CD103+ DC-mediated antitumor response enhanced the effects of CD47 blockade strategy. Together, this study uncovers an unprecedented role for CD103+ DC in nsPEF treatment-elicited antitumor immune response and elucidates the underlying mechanisms.

Galectin-1: A Traditionally Immunosuppressive Protein Displays Context-Dependent Capacities.

Galectin-Carbohydrate interactions are indispensable to pathogen recognition and immune response. Galectin-1, a ubiquitously expressed 14-kDa protein with an evolutionarily conserved ß-galactoside binding site, translates glycoconjugate recognition into function. That galectin-1 is demonstrated to induce T cell apoptosis has led to substantial attention to the immunosuppressive properties of this protein, such as inducing naive immune cells to suppressive phenotypes, promoting recruitment of immunosuppressing cells as well as impairing functions of cytotoxic leukocytes. However, only in recent years have studies shown that galectin-1 appears to perform a pro-inflammatory role in certain diseases. In this review, we describe the anti-inflammatory function of galectin-1 and its possible mechanisms and summarize the existing therapies and preclinical efficacy relating to these agents. In the meantime, we also discuss the potential causal factors by which galectin-1 promotes the progression of inflammation.

Targeting neuropilin-1 abolishes anti-PD-1-upregulated regulatory T cells and synergizes with 4-1BB agonist for liver cancer treatment.

BACKGROUND AIMS: Regulatory T cells (Tregs) are an obstacle to PD-1 blockade-mediated antitumor efficacy. However, the behaviors of Tregs response to anti-PD-1 in HCC and the characteristics of Tregs tissue adaptation from peripheral lymphoid tissues to the tumor are still unclear. APPROACH RESULTS: Here, we determine that PD-1 monotherapy potentially augments the accumulation of tumor CD4 + Tregs. Mechanistically, anti-PD-1 mediates Tregs proliferation in lymphoid tissues rather than in the tumor. Increased peripheral Tregs burden replenishes intratumoral Tregs, raising the ratio of intratumoral CD4 + Tregs to CD8 + T cells. Subsequently, single-cell transcriptomics revealed that neuropilin-1 (Nrp-1) supports Tregs migration behavior, and the genes of Crem and Tnfrsf9 regulate the behaviors of the terminal suppressive Tregs. Nrp-1 + 4-1BB - Tregs stepwise develop to the Nrp-1 - 4-1BB + Tregs from lymphoid tissues into the tumor. Moreover, Treg-restricted Nrp1 depletion abolishes anti-PD-1-upregulated intratumoral Tregs burden and synergizes with the 4-1BB agonist to enhance the antitumor response. Finally, a combination of the Nrp-1 inhibitor and the 4-1BB agonist in humanized HCC models showed a favorable and safe outcome and evoked the antitumor effect of the PD-1 blockade. CONCLUSION: Our findings elucidate the potential mechanism of anti-PD-1-mediated intratumoral Tregs accumulation in HCC and uncover the tissue adaptation characteristics of Tregs and identify the therapeutic potential of targeting Nrp-1 and 4-1BB for reprogramming the HCC microenvironment.

Disulfiram/Copper Induces Immunogenic Cell Death and Enhances CD47 Blockade in Hepatocellular Carcinoma.

Some chemotherapeutic agents have been found to enhance antitumor immunity by inducing immunogenic cell death (ICD). The combination of disulfiram (DSF) and copper (Cu) has demonstrated anti-tumor effects in a range of malignancies including hepatocellular carcinoma (HCC). However, the potential of DSF/Cu as an ICD inducer and whether it can enhance the efficacy of the immune checkpoint blockade in HCC remains unknown. Here, we showed that DSF/Cu-treated HCC cells exhibited characteristics of ICD in vitro, such as calreticulin (CRT) exposure, ATP secretion, and high mobility group box 1 (HMGB1) release. DSF/Cu-treated HCC cells elicited significant immune memory in a vaccination assay. DSF/Cu treatment promoted dendritic cell activation and maturation. The combination of DSF/Cu and CD47 blockade further facilitated DC maturation and subsequently enhanced CD8+ T cell cytotoxicity. Mechanically, DSF/Cu promoted the nuclear accumulation and aggregation of nuclear protein localization protein 4 (NPL4) to inhibit the ubiquitin-proteasome system; thus, inducing endoplasmic reticulum (ER) stress. The inhibition of NPL4 induced ICD-associated damage-associated molecular patterns. Collectively, our findings demonstrated that DSF/Cu-induced ICD-mediated immune activation in HCC enhanced the efficacy of CD47 blockade.

Blocking CD47 promotes antitumour immunity through CD103+ dendritic cell-NK cell axis in murine hepatocellular carcinoma model.

BACKGROUND & AIMS: The CD47-signal regulatory protein α (SIRPα) axis inhibits dendritic cell (DC) phagocytosis and contributes to immune evasion. However, the behaviour of DCs and the potential crosstalk between DCs and natural killer (NK) cells in the hepatocellular carcinoma (HCC) microenvironment after CD47 blockade remain unclear. METHODS: The infiltration of CD103+ DCs and NK cells were analysed by immunohistochemistry and immunofluorescence in both human and murine HCC specimens. An orthotopic liver tumour model was used to evaluate the function of the CD103+ DC-NK cell axis after CD47 blockade in vivo in wild-type, Rag1-/-, Batf3-/-, and STING1-/- mice. Phagocytosis assays were performed in CD103+ DC and HCC cell lines. CD103+ DC-derived cytokines were analysed by chemokine array. Spleen-derived NK cells in C57BL/6J mice were used to evaluate cytotoxic functions in vitro. RESULTS: Higher CD47 expression was associated with worse prognosis in patients with HCC. CD47 blockade enhanced antitumour efficacy by stimulating the CD103+ DC-NK cell axis. The hypoxic microenvironment promoted CD47 blockade-induced tumour DNA phagocytosis by CD103+ DCs. By releasing IL-12 and CXCL9, activated CD103+ DCs induced the recruitment of NK cells with upregulated expression of granzyme B, NKG2D, interferon-γ, and tumour necrosis factor-α and downregulated expression of NKG2A. The antitumour effects of CD47 blockade could be abolished by cyclic GMP-AMP synthase (cGAS)-STING pathway inhibition. CONCLUSIONS: In addition to the classical DC-T cell axis, CD47 blockade significantly enhanced the ability of CD103+ DCs to take up tumour DNA, resulting in the stimulation of the cGAS-STING pathway, which promoted the infiltration and activation of NK cells in liver cancer. LAY SUMMARY: Hypoxia (low oxygen levels) is prevalent in the hepatocellular carcinoma microenvironment and promotes the phagocytosis (ingestion and elimination) of tumour DNA by CD103+ dendritic cells (a type of immune cell). Blockade of the cell surface protein CD47 resulted in activation of CD103+ dendritic cells which led to the recruitment and activation of natural killer cells (a different immune cell). When activated, these cells exhibit an antitumour effect.

A pan-cancer analysis of the oncogenic role of Holliday junction recognition protein in human tumors.

Although cell-based or animal-based research evidence support the association of Holliday junction recognition protein (HJURP) with cancers, no pan-cancer investigation has been reported. The datasets of Gene Expression Omnibus database along with The Cancer Genome Atlas project were used to evaluate the expression of HJURP in various types of tumors. HJURP is overexpressed in a considerable number of cancers, and some changes in DNA methylation and genetic alterations are discovered in some types of tumors, such as kidney-related and adrenal gland-related tumors. Based on PrognoScan and gene expression profiling interactive analysis (GEPIA), the elevated expression of HJURP worsened the survival time of individuals with cancer. The biological general repository for interaction datasets (BioGRID) and The database for annotation, visualization and integrated discovery (DAVID) were used to establish the functional molecular network. It revealed that the cell cycle and p53 signaling pathway are the key molecular mechanisms that HJURP promotes carcinogenesis. The nomograms between HJURP and clinical pathological factors based on the Cox proportional hazards model showed a good prognostic performance in kidney carcinoma, hepatocellular carcinoma, and lung adenocarcinoma. Our first pan-cancer study provides a relatively profound insights into the oncogenic roles of HJURP across different tumors.

Integrative Network Analysis Revealed Genetic Impact of Pyruvate Kinase L/R on Hepatocyte Proliferation and Graft Survival after Liver Transplantation.

BACKGROUND: Pyruvate kinase L/R (PKLR) has been suggested to affect the proliferation of hepatocytes via regulation of the cell cycle and lipid metabolism. However, its impact on the global metabolome and its clinical implications remain unclear. AIMS: We aimed to clarify the genetic impact of PKLR on the metabolomic profiles of hepatoma cells and its potential effects on grafts for liver transplantation (LT). METHODS: Nontargeted and targeted metabolomic assays were performed in human hepatoma cells transfected with lentiviral vectors causing PKLR overexpression and silencing, respectively. We then constructed a molecular network based on integrative analysis of transcriptomic and metabolomic data. We also assessed the biological functions of PKLR in the global metabolome in LT grafts in patients via a weighted correlation network model. RESULTS: Multiomic analysis revealed that PKLR perturbations significantly affected the pyruvate, citrate, and glycerophospholipid metabolism pathways, as crucial steps in de novo lipogenesis (DNL). We also confirmed the importance of phosphatidylcholines (PC) and its derivative lyso-PC supply on improved survival of LT grafts in patients. Coexpression analysis revealed beneficial effects of PKLR overexpression on posttransplant prognosis by alleviating arachidonic acid metabolism of the grafts, independent of operational risk factors. CONCLUSION: This systems-level analysis indicated that PKLR affected hepatoma cell viability via impacts on the whole process of DNL, from glycolysis to final PC synthesis. PKLR also improved prognosis after LT, possibly via its impact on the increased genesis of beneficial glycerophospholipids.

Self-assembly nanovaccine containing TLR7/8 agonist and STAT3 inhibitor enhances tumor immunotherapy by augmenting tumor-specific immune response.

BACKGROUND: Cancer vaccines are a promising strategy for cancer immunotherapy. Cancer vaccines elicits a specific cytotoxic immune response to tumor antigens. However, the efficacy of traditional peptide-based cancer vaccines is limited due to the inefficient delivery of antigens and adjuvants to dendritic cells (DCs). Therefore, it is necessary to develop a novel rationally designed cancer vaccine to maximize its desired effects. METHODS: A Self-assembling Vehicle-free Multi-component Antitumor nanoVaccine (SVMAV) was constructed by using an unsaturated fatty acid docosahexaenoic acid (DHA)-conjugated antigen and R848 (a Toll-like receptor 7/8 agonist) to encapsulate stattic (a signal transducer and activator of transcription 3 inhibitor). The characteristics of SVMAV were investigated. The ability of SVMAV to promote DC functions was examined by in vitro analysis. The antitumor effects of SVMAV and its combination with antiprogrammed cell death protein 1 antibody (aPD-1) were also investigated in vivo. The potential application of SVMAV for neoantigen-targeted, personalized cancer vaccines was examined in an orthotopic hepatocellular carcinoma model. RESULTS: The obtained SVMAV efficiently migrated into lymph nodes and primed CD8+ T cells for exert neoantigen-specific killing by promoting the antigen uptake by DCs, stimulating DC maturation, and enhancing antigen cross-presentation, due to the simultaneous delivery of the antigen, R848 and stattic. SVMAV could not only yield a robust antitumor effect for primary melanoma allografts, but also exert a protective effect for lung metastases. Moreover, combination treatment of SVMAV and aPD-1 exerted synergistic antitumor activity and extended the survival duration of melanoma-bearing mice. Notably, a cell line-specific neoantigen-based SVMAV was designed according to predicted neoantigens for Hepa1-6 cells to examine the potential application of SVMAV for personalized cancer vaccine. Encouragingly, neoantigen-specific SVMAV achieved stronger antitumor activity than aPD-1 in an orthotopic hepatocellular cancer model established with Hepa1-6 cells. CONCLUSIONS: In summary, this study offers an efficient codelivery platform for neoantigens and immunoregulatory compounds to enhance immune responses during cancer immune therapy.

Emerging Roles of Liquid-Liquid Phase Separation in Cancer: From Protein Aggregation to Immune-Associated Signaling.

Liquid-liquid Phase Separation (LLPS) of proteins and nucleic acids has emerged as a new paradigm in the study of cellular activities. It drives the formation of liquid-like condensates containing biomolecules in the absence of membrane structures in living cells. In addition, typical membrane-less condensates such as nuclear speckles, stress granules and cell signaling clusters play important roles in various cellular activities, including regulation of transcription, cellular stress response and signal transduction. Previous studies highlighted the biophysical and biochemical principles underlying the formation of these liquid condensates. The studies also showed how these principles determine the molecular properties, LLPS behavior, and composition of liquid condensates. While the basic rules driving LLPS are continuously being uncovered, their function in cellular activities is still unclear, especially within a pathological context. Therefore, the present review summarizes the recent progress made on the existing roles of LLPS in cancer, including cancer-related signaling pathways, transcription regulation and maintenance of genome stability. Additionally, the review briefly introduces the basic rules of LLPS, and cellular signaling that potentially plays a role in cancer, including pathways relevant to immune responses and autophagy.

Chemokine-Like Factor-Like MARVEL Transmembrane Domain-Containing Family in Hepatocellular Carcinoma: Latest Advances.

Chemokine-like factor (CKLF)-like MARVEL transmembrane domain-containing family (CMTMs) is a new gene family, consisting of CKLF and CMTM1 to CMTM8, which plays an important role in hematopoiesis system, autoimmune diseases, male reproduction etc. Abnormal expression of CMTMs is also associated with tumor genesis, development and metastasis. In this review, we briefly describe the characteristics of CMTM family, outline its functions in multiple kinds of carcinomas, and summarize the latest research on their roles in hepatocellular carcinoma which are mainly related to the expression, prognostic effect, potential functions, and mechanism of action. The CMTM family is expected to provide new ideas and targets for HCC diagnosis and treatment.

Syndecan-4 promotes vascular beds formation in tissue engineered liver via thrombospondin 1.

Instantaneous blood coagulation after bioengineered liver transplantation is a major issue, and the key process in its prevention is the construction of the endothelial vascular bed on biomimetic scaffolds. However, the specific molecules involved in the regulation of the vascular bed formation remain unclear. Syndecan-4 is a type I transmembrane glycoprotein commonly expressed in the human body; its receptor has been reported as critical for optimal cell adhesion and initiation of intracellular signaling, indicating its promising application in vascular bed formation. In the current study, bioinformatics analysis and in vitro experiments were performed to evaluate whether syndecan-4 promoted endothelial cell migration and functional activation. Exogenous syndecan-4-overexpressing endothelial cells were perfused into the decellularized liver scaffold, which was assessed by Masson's trichrome staining. Western blotting and qRT-PCR were used to evaluate the effects of syndecan-4 on the thrombospondin 1 (THBS1) stability. We found that syndecan-4 promoted the adhesion of vascular endothelial cells and facilitated cell migration and angiogenesis. Furthermore, syndecan-4 overexpression resulted in a well-aligned endothelium on the decellularized liver scaffolds. Mechanistically, syndecan-4 destabilized THBS1 at the protein level. Therefore, our data revealed that syndecan-4 promoted the biological activity of endothelial cells on the bionic liver vascular bed through THBS1. These findings provide scientific evidences for solving transient blood coagulation after bionic liver transplantation.

Integrated analysis of microbiome and host transcriptome reveals correlations between gut microbiota and clinical outcomes in HBV-related hepatocellular carcinoma.

BACKGROUND: The gut-liver axis plays a pivotal role in the pathogenesis of hepatocellular carcinoma (HCC). However, the correlations between the gut microbiome and the liver tumor transcriptome in patients with HCC and the impact of the gut microbiota on clinical outcome are less well-understood. METHODS: Fecal samples collected from HBV-related HCC patients (n = 113) and healthy volunteers (n = 100) were subjected to 16S rRNA sequencing of the microbiome. After a rigorous selection process, 32 paired tumor and adjacent non-tumor liver tissues from the HCC group were subjected to next-generation sequencing (NGS) RNA-seq. The datasets were analyzed individually and integrated with clinical characteristics for combined analysis using bioinformatics approaches. We further verified the potential of the gut microbiota to predict clinical outcome by a random forest model and a support vector machine model. RESULTS: We found that Bacteroides, Lachnospiracea incertae sedis, and Clostridium XIVa were enriched in HCC patients with a high tumor burden. By integrating the microbiome and transcriptome, we identified 31 robust associations between the above three genera and well-characterized genes, indicating possible mechanistic relationships in tumor immune microenvironment. Clinical characteristics and database analysis suggested that serum bile acids may be important communication mediators between these three genera and the host transcriptome. Finally, among these three genera, six important microbial markers associated with tumor immune microenvironment or bile acid metabolism showed the potential to predict clinical outcome (AUC = 81%). CONCLUSIONS: This study revealed that changes in tumor immune microenvironment caused by the gut microbiota via serum bile acids may be important factors associated with tumor burden and adverse clinical outcome. Gut microbes can be used as biomarkers of clinical features and outcomes, and the microbe-associated transcripts of host tumors can partly explain how gut microbiota promotes HCC pathogenesis.

Metabonomic Profile of Macrosteatotic Allografts for Orthotopic Liver Transplantation in Patients With Initial Poor Function: Mechanistic Investigation and Prognostic Prediction.

BACKGROUND: Our previous study revealled amplified hazardous effects of macrosteatosis (MaS) on graft failure (GF) in recipients with severe liver damage in short post-operative days, with vague mechanism inside. AIM: We aimed to uncover the molecular mechanism of donor MaS on GF, and construct the predictive model to monitor post-transplant prognosis based on "omics" perspective. METHODS: Ultra-performance liquid chromatography coupled to mass spectrometry metabolomic analysis was performed in allograft tissues from 82 patients with initial poor function (IPF) from multi-liver transplant (LT) centers. Pathway analysis was performed by on-line toolkit Metaboanalyst (v 3.0). Predictive model was constructed based on combinative metabonomic and clinical data extracted by stepwised cox proportional analysis. RESULTS: Principle component analysis (PCA) analysis revealled stratification on metabolic feature in organs classified by MaS status. Differential metabolits both associated with MaS and GF were significantly enriched on pathway of glycerophospholipid metabolism (P < 0.05). Phosphatidylcholine (PC) and phosphatidylethanolamine (PE) involved in glycerophospholipid metabolism was significantly decreased in cases with MaS donors and GF (P < 0.05). Better prediction was observed on graft survival by combinative model (area under the curve = 0.91) and confirmed by internal validation. CONCLUSION: Metabonomic features of allografts can be clearly distinguished by MaS status in patients with IPF. Dysfunction on glycerophospholipid metabolism was culprit to link donor MaS and final GF. Decrement on PC and PE exerted the fatal effects of MaS on organ failure. Metabonomic data might help for monitoring long-term graft survival after LT.

Blocking exposed PD-L1 elicited by nanosecond pulsed electric field reverses dysfunction of CD8+ T cells in liver cancer.

As a promising method for local tumor treatment, nanosecond pulsed electric field (nsPEF) ablation elicits a potent anti-tumor immune response. However, the mechanism of the nsPEF-mediated anti-tumor immune response and its effects on the tumor microenvironment remains unclear. Here, we demonstrated that nsPEF treatment increased the level of membrane PD-L1 in liver cancer cells. Furthermore, nsPEF induced the release of PD-L1-associated extra-cellular vesicles, leading to the dysfunction of CD8+ T cells, which could potentially be reversed by PD-L1 blockade. Biological and functional assays also demonstrated that nsPEF treatment resulted in the increased PD-L1 level and dysfunction of infiltrated CD8+ T cells in tumor tissues in vivo, indicating the long term antitumor efficacy of nsPEF treatment. A combination of nsPEF treatment and PD-L1 blockade effectively inhibited tumor growth and improved the survival of the tumor-bearing mouse. In conclusion, nsPEF treatment induced the translocation and release of PD-L1 and contributed to the dysfunction of infiltrated CD8+ T cells, resulting in tumor progression at later stages. The combination of nsPEF treatment and PD-L1 blockade is a promising therapeutic strategy for liver cancer.

Mechanisms of RNA N6-Methyladenosine in Hepatocellular Carcinoma: From the Perspectives of Etiology.

N6-Methyladenosine (m6A) is the most common RNA internal modification in eukaryotic cells. Its regulatory effects at the post-transcriptional level on both messenger RNAs (mRNAs) and noncoding RNAs have been widely studied; these include alternative splicing, stability, translation efficiency, nucleus export, and degradation. m6A modification is implicated in a series of physiological and pathological activities, such as embryonic stem cell differentiation, immunoregulation, adipogenesis, and cancer development. Recently, the significance of m6A methylation has been identified in both viral hepatitis and non-alcohol fatty liver disease (NAFLD), which are major risk factors in the development of hepatocellular carcinoma (HCC). Given the high incidence and mortality rate of HCC worldwide, it is of great importance to elucidate the mechanisms underlying HCC initiation and progression. m6A as an emerging research focus has great potential to facilitate the understanding of HCC, particularly from an etiological perspective. Thus, in this review, we summarize recent progress in understanding m6A modification related to viral hepatitis, NAFLD, and HCC, including their mechanisms and clinical applications.

Systematic Analysis of Alternative Splicing Landscape in Pancreatic Adenocarcinoma Reveals Regulatory Network Associated with Tumorigenesis and Immune Response.

BACKGROUND Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive gastrointestinal tumors and has an extremely high mortality rate. Recent studies indicate that alternative splicing (AS), a common post-transcriptional process, has important roles in tumor biological behaviors and may provide novel immunotherapeutic targets. This study systematically analyzes AS profiles in PDAC and reveals their potential regulatory effects on cancer immune response. MATERIAL AND METHODS AS event, RNA sequencing, and splicing factor (SF) data were extracted from SpliceSeq, The Cancer Genome Atlas, and SpliceAid2, respectively. Overall survival (OS)-associated AS events and SFs were identified with univariate analysis. The LASSO method and multivariate Cox regression analysis were used to construct predictive signatures for the prediction of patient prognosis. The proportions of immune cells within PDAC samples were evaluated using the CIBERSORT algorithm. The correlations among AS events, SFs, and immune cell proportions were calculated using Spearman correlation analysis. Consensus clustering and immune classification were performed on the PDAC cohort. RESULTS A total of 4812 OS-related AS events from 3341 parent genes were identified, and 8 AS-based predictive models were constructed for PDAC. An OS-related SF-AS regulatory network was constructed. The AS events regulated by ELAVL4 exhibited strong correlations with CD8 T cells and regulatory T cells. In addition, AS-based clusters demonstrated distinct OS outcomes and immune features. CONCLUSIONS AS-based predictive models with high accuracy were constructed to facilitate prognosis prediction and treatment of PDAC. An SF-AS regulatory network was constructed, revealing the potential relationships among SF, AS, and immune response.