Identification and in vitro and in vivo validation of the key role of GSDME in pyroptosis-related genes signature in hepatocellular carcinoma.

We used pyroptosis-related genes to establish a risk-score model for prognostic prediction of liver hepatocellular carcinoma (LIHC) patients. A total of 52 pyroptosis-associated genes were identified. Then, data for 374 LIHC patients and 50 normal individuals were acquired from the TCGA database. Through gene expression analyses, differentially expressed genes (DEGs) were determined. The 13 pyroptosis-related genes (PRGs) confirmed as potential prognostic factors through univariate Cox regression analysis were entered into Lasso and multivariate Cox regression to build a PRGs prognostic signature, containing four PRGs (BAK1, GSDME, NLRP6, and NOD2) determined as independent prognostic factors. mRNA levels were evaluated by qRT-PCR, while overall survival (OS) rates were assessed by the Kaplan-Meier method. Enrichment analyses were done to establish the mechanisms associated with differential survival status of LIHC patients from a tumor immunology perspective. Additionally, a risk score determined by the prognostic model could divide LIHC patients into low- and high-risk groups using median risk score as cut-off. A prognostic nomogram, derived from the prognostic model and integrating clinical characteristics of patients, was constructed. The prognostic function of the model was also validated using GEO, ICGC cohorts, and online databases Kaplan-Meier Plotter. Small interfering RNA-mediated knockdown of GSDME, as well as lentivirus-mediated GSDME knockdown, were performed to validate that knockdown of GSDME markedly suppressed growth of HCC cells both in vivo and in vitro. Collectively, our study demonstrated a PRGs prognostic signature that had great clinical value in prognosis assessment.

Autophagy gene Atg7 regulates the development of radiation-induced skin injury and fibrosis of skin.

BACKGROUND: Radiation-induced skin injury, which may progress to fibrosis, is a severe side effect of radiotherapy in patients with cancer. However, currently, there is a lack of preventive or curative treatments for this injury. Meanwhile, the mechanisms underlying this injury remain poorly understood. Here, we elucidated whether autophagy is essential for the development of radiation-induced skin injury and the potential molecular pathways and mechanisms involved. METHODS AND RESULTS: We used the myofibroblast-specific Atg7 knockout (namely, conditional Atg7 knockout) mice irradiated with a single electron beam irradiation dose of 30 Gy. Vaseline-based 0.2% rapamycin ointment was topically applied once daily from the day of irradiation for 30 days. On day 30 post irradiation, skin tissues were harvested for further analysis. In vitro, human foreskin fibroblast cells were treated with rapamycin (100 nM) for 24 h and pretreated with 3-MA (5 mM) for 12 h. Macroscopic skin manifestations, histological changes, and fibrosis markers at the mRNA and protein expression levels were measured. Post irradiation, the myofibroblast-specific autophagy-deficient (Atg7Flox/Flox Cre+ ) mice had increased fibrosis marker (COL1A1, CTGF, TGF-ß1, and α-SMA) levels in the irradiated area and had more severe macroscopic skin manifestations than the control group (Atg7Flox/Flox Cre- ) mice. Treatment with an autophagy agonist rapamycin attenuated macroscopic skin injury scores and skin fibrosis marker levels with decreased epidermal thickness and dermal collagen deposition in Atg7Flox/Flox Cre+ mice compared with the vehicle control. Moreover, in vitro experiment results were consistent with the in vivo results. Together with studies at the molecular level, we found that these changes involved the Akt/mTOR pathway. In addition, this phenomenon might also relate to Nrf2-autophagy signaling pathway under oxidative stress conditions. CONCLUSION: In conclusion, Atg7 and autophagy-related mechanisms confer radioprotection, and reactivation of the autophagy process can be a novel therapeutic strategy to reduce and prevent the occurrence of radiodermatitis, particularly skin fibrosis, in patients with cancer.

Comprehensive Analysis of N6-Methyladenosine-Related IncRNAs Prognostic Signature and the Associated Immune Infiltrates in Kidney Renal Clear Cell Carcinoma.

Kidney renal clear cell carcinoma (KIRC) is the most common and aggressive subtype of renal cell carcinoma. N6-methyladenosine (m6A) RNA methylation is the most prevalent epigenetic RNA modification. Long non-coding RNAs (lncRNAs) have emerged as a key role in regulating cancer progression. However, little has been learned about the molecular functions of m6A-related lncRNAs in KIRC. The prognostic value of m6A-related ln-cRNAs was investigated in KIRC samples downloaded from The Cancer Genome Atlas (TCGA) dataset. The m6A-related lncRNAs were further screen out by Pearson correlation test. Then, 27 m6A-related lncRNAs were confirmed as potential prognostic factors through univariate Cox regression analysis. They were entered into Lasso and multivariate Cox regression to build a m6A-related lncRNA prognostic signature, including 14 m6A-related lncRNAs determined as independent prognostic factors. Additionally, a risk score calculated according to the prognostic model could divide KIRC patients into low- and high-risk groups depending on median risk score as cut-off. A prognostic nomogram, derived from the prognostic model and integrating clinical characteristics of patients, was constructed. Three distinct clusters were identified with different immune signatures through consensus clustering analysis according to the expression pattern of m6A-related lncRNAs. Twenty-seven prognostic m6A-related lncRNAs were determined as prognostic lncRNAs from TCGA-KIRC cohort. The m6A-related lncRNA prognostic signature containing 14 independent prognostic lncRNAs exhibited good accuracy in predicting overall survival of KIRC patients. We correlated the three distinct clusters with immune infiltration signature of KIRC for the first time. We found that the worse prognosis of cluster2 was probably mediated by immune evasion. In summary, our study identified a m6A-related lncRNAs prognostic signature which had great clinical value in prognosis assessment. We classified TCGA-KIRC samples into three clusters with distinct immune signatures, which could be considered as potential targets of immunotherapy for KIRC treatment in the future.

Cell-Derived Biogenetic Gold Nanoparticles for Sensitizing Radiotherapy and Boosting Immune Response against Cancer.

The biosynthesis of nanomedicine has gained enormous attention and exhibited promising prospects, while the underlying mechanism and advantage remain not fully understood. Here, a cell-reactor based on tumor cells is developed to obtain biogenetic gold nanoparticles (Au@MC38) for sensitizing radiotherapy and boosting immune responses. It demonstrates that the intracellular biomineralization and exocytosis process of Au@MC38 can be regulated by the cellular metabolites level and other factors, such as glutathione and reactive oxygen species (ROS), autophagy, and UV irradiation. The elucidation of mechanisms may promote the understanding of interaction principles between nanoparticles and biosystems in the process of biosynthesis. Combined with radiotherapy, Au@MC38 strengthens the radiation-induced DNA damage and ROS generation, thus aggravating cell apoptosis and necrosis. Benefiting from homologous targeting and transcytosis effect, Au@MC38 demonstrates good tumor distribution. Local radiation-induced immunogenic cell death initiates an effective immune response. Especially, CD8a+ dendritic cells are significantly increased in mice that received combinatorial treatment. This radio-sensitization strategy has demonstrated the effective inhibition on primary and metastatic tumors, and achieved satisfactory survival benefit in combinatorial with immune checkpoint blockade. Thus, this bio-inspired synthetic strategy may impulse the development of biosynthesis and its therapeutic applications, contributing to a non-invasive and efficient modality for nanomedicine exploitation.

Advancements in osteosarcoma management: integrating immune microenvironment insights with immunotherapeutic strategies.

Osteosarcoma, a malignant bone tumor predominantly affecting children and adolescents, presents significant therapeutic challenges, particularly in metastatic or recurrent cases. Conventional surgical and chemotherapeutic approaches have achieved partial therapeutic efficacy; however, the prognosis for long-term survival remains bleak. Recent studies have highlighted the imperative for a comprehensive exploration of the osteosarcoma immune microenvironment, focusing on the integration of diverse immunotherapeutic strategies-including immune checkpoint inhibitors, tumor microenvironment modulators, cytokine therapies, tumor antigen-specific interventions, cancer vaccines, cellular therapies, and antibody-based treatments-that are directly pertinent to modulating this intricate microenvironment. By targeting tumor cells, modulating the tumor microenvironment, and activating host immune responses, these innovative approaches have demonstrated substantial potential in enhancing the effectiveness of osteosarcoma treatments. Although most of these novel strategies are still in research or clinical trial phases, they have already demonstrated significant potential for individuals with osteosarcoma, suggesting the possibility of developing new, more personalized and effective treatment options. This review aims to provide a comprehensive overview of the current advancements in osteosarcoma immunotherapy, emphasizing the significance of integrating various immunotherapeutic methods to optimize therapeutic outcomes. Additionally, it underscores the imperative for subsequent research to further investigate the intricate interactions between the tumor microenvironment and the immune system, aiming to devise more effective treatment strategies. The present review comprehensively addresses the landscape of osteosarcoma immunotherapy, delineating crucial scientific concerns and clinical challenges, thereby outlining potential research directions.

MUC1 and MUC16: critical for immune modulation in cancer therapeutics.

The Mucin (MUC) family, a range of highly glycosylated macromolecules, is ubiquitously expressed in mammalian epithelial cells. Such molecules are pivotal in establishing protective mucosal barriers, serving as defenses against pathogenic assaults. Intriguingly, the aberrant expression of specific MUC proteins, notably Mucin 1 (MUC1) and Mucin 16 (MUC16), within tumor cells, is intimately associated with oncogenesis, proliferation, and metastasis. This association involves various mechanisms, including cellular proliferation, viability, apoptosis resistance, chemotherapeutic resilience, metabolic shifts, and immune surveillance evasion. Due to their distinctive biological roles and structural features in oncology, MUC proteins have attracted considerable attention as prospective targets and biomarkers in cancer therapy. The current review offers an exhaustive exploration of the roles of MUC1 and MUC16 in the context of cancer biomarkers, elucidating their critical contributions to the mechanisms of cellular signal transduction, regulation of immune responses, and the modulation of the tumor microenvironment. Additionally, the article evaluates the latest advances in therapeutic strategies targeting these mucins, focusing on innovations in immunotherapies and targeted drugs, aiming to enhance customization and accuracy in cancer treatments.

Mendelian randomization unraveled: gender-specific insights into obesity-related phenotypes and colorectal cancer susceptibility.

Objective: Evidence has been increasingly pointing towards a potential link between phenotypes related to obesity and the incidence of colorectal cancer. However, confirming this as a direct causal connection remains elusive. This investigation aims to elucidate the causative links between obesity-associated phenotypes and the incidence of colorectal cancer. Methods: Employing the Two Sample Mendelian Randomization (TwoSampleMR) R package, analyses were conducted using Mendelian randomization (MR) to discern potential causative links between obesity categories sourced from both the Institute for Education and University (IEU) Open GWAS Project and Zenodo, and colorectal tumors (data obtained from IEU Open GWAS and FinnGen). For primary evaluations, the study utilized the Wald ratio and the Inverse Variance Weighting (IVW) methods, while the MR-Egger approach was integrated for sensitivity assessment. Bidirectional Mendelian Randomization (Bidirectional MR), as well as Linkage Disequilibrium (LD) Score Regression with well-imputed HapMap3 single nucleotide polymorphisms (SNPs), were additionally executed. Sensitivity assessments entailed IVW, MR-Egger methodologies to assess heterogeneity and pleiotropy, along with a leave-one-out strategy. Instrumental variables were chosen judiciously based on predetermined P-value thresholds and F-statistics. Results: Results from MR evaluations did not identify a clear causative link between BMI and colorectal malignancy. Conversely, both measures of obesity, the Waist-Hip Ratio (WHR) and its adjusted form for BMI (WHRadjBMI), displayed a connection to increased risk of colorectal cancer, especially prominent among female subjects. Reverse MR analyses dismissed potential reverse causality between colorectal malignancies and obesity. A significant genetic interplay was observed between WHR, WHRadjBMI, and colorectal cancer instances. Ensuing MR probes spotlighted inflammatory bowel ailment as a protective factor, while salad intake was indicated as a potential risk concerning colorectal malignancies. Sensitivity reviews, which included tests for both pleiotropy and heterogeneity, validated the robustness of the MR findings. Conclusion: Findings from this research indicate that specific obesity-related parameters, notably WHR and WHRadjBMI, carry a causal relationship with an elevated colorectal cancer risk. The impact is distinctly more evident among females. Such insights might be pivotal for public health deliberations, hinting that individuals boasting a high WHR might necessitate intensified colorectal cancer screenings.

RUNX transcription factors: biological functions and implications in cancer.

Runt-related transcription factors (RUNX) are a family of transcription factors that are essential for normal and malignant hematopoietic processes. Their most widely recognized role in malignancy is to promote the occurrence and development of acute myeloid leukemia. However, it is worth noting that during the last decade, studies of RUNX proteins in solid tumors have made considerable progress, suggesting that these proteins are directly involved in different stages of tumor development, including tumor initiation, progression, and invasion. RUNX proteins also play a role in tumor angiogenesis, the maintenance of tumor cell stemness, and resistance to antitumor drugs. These findings have led to the consideration of RUNX as a tumor biomarker. All RUNX proteins are involved in the occurrence and development of solid tumors, but the role of each RUNX protein in different tumors and the major signaling pathways involved are complicated by tumor heterogeneity and the interacting tumor microenvironment. Understanding how the dysregulation of RUNX in tumors affects normal biological processes is important to elucidate the molecular mechanisms by which RUNX affects malignant tumors.

Nintedanib Mitigates Radiation-Induced Pulmonary Fibrosis by Suppressing Epithelial Cell Inflammatory Response and Inhibiting Fibroblast-to-Myofibroblast Transition.

Radiation-induced pulmonary fibrosis (RIPF) represents a serious complication observed in individuals undergoing thoracic radiation therapy. Currently, effective interventions for RIPF are unavailable. Prior research has demonstrated that nintedanib, a Food and Drug Administration (FDA)-approved anti-fibrotic agent for idiopathic pulmonary fibrosis, exerts therapeutic effects on chronic fibrosing interstitial lung disease. This research aimed to investigate the anti-fibrotic influences of nintedanib on RIPF and reveal the fundamental mechanisms. To assess its therapeutic impact, a mouse model of RIPF was established. The process involved nintedanib administration at various time points, both prior to and following thoracic radiation. In the RIPF mouse model, an assessment was conducted on survival rates, body weight, computed tomography features, histological parameters, and changes in gene expression. In vitro experiments were performed to discover the mechanism underlying the therapeutic impact of nintedanib on RIPF. Treatment with nintedanib, administered either two days prior or four weeks after thoracic radiation, significantly alleviated lung pathological changes, suppressed collagen deposition, and improved the overall health status of the mice. Additionally, nintedanib demonstrated significant mitigation of radiation-induced inflammatory responses in epithelial cells by inhibiting the PI3K/AKT and MAPK signaling pathways. Furthermore, nintedanib substantially inhibited fibroblast-to-myofibroblast transition by suppressing the TGF-ß/Smad and PI3K/AKT/mTOR signaling pathways. These findings suggest that nintedanib exerts preventive and therapeutic effects on RIPF by modulating multiple targets instead of a single anti-fibrotic pathway and encourage the further clinical trials to determine the efficacy of nintedanib in patients with RIPF.

ATM inhibition enhance immunotherapy by activating STING signaling and augmenting MHC Class I.

Accumulating evidence supports the concept that DNA damage response targeted therapies can improve antitumor immune response by increasing the immunogenicity of tumor cells and improving the tumor immune microenvironment. Ataxia telangiectasia mutated (ATM) is a core component of the DNA repair system. Although the ATM gene has a significant mutation rate in many human cancers, including colorectal, prostate, lung, and breast, it remains understudied compared with other DDR-involved molecules such as PARP and ATR. Here, we found that either gene knockout or drug intervention, ATM inhibition activated the cGAS/STING pathway and augmented MHC class I in CRC cells, and these effects could be amplified by radiation. Furthermore, we found that MHC class I upregulation induced by ATM inhibition is dependent on the activation of the NFκB/IRF1/NLRC5 pathway and independent of STING. Animal experiments have shown increasing infiltration and cytotoxic function of T cells and better survival in ATM-deficient tumors. This work indicated that ATM nonsense mutation predicted the clinical benefits of radiotherapy combined with immune checkpoint blockade for patients with CRC. It also provides a molecular mechanism rationale for ATM-targeted agents for patients with CRC.

Integrated bioinformatics and validation reveal SOX12 as potential biomarker in colon adenocarcinoma based on an immune infiltration-related ceRNA network.

PURPOSE: The primary objective of this study was to construct competing endogenous RNA (ceRNA) networks and evaluate the prognostic significance of tumor-infiltrating immune cells (TIICs) and key biomarkers within the ceRNA networks in colon adenocarcinoma (COAD) patients. METHODS: Comprehensive bioinformatics tools were used to screen differentially expressed genes (DEGs), miRNAs (DEMs), and lncRNAs (DELs) related to COAD, leading to the creation of ceRNA networks. The CIBERSORT technique was employed to assess the significance of TIICs in COAD, and an immune-related prognosis prediction model was subsequently developed. Co-expression analyses were conducted to determine the relationship between key genes in ceRNA networks and immunologically significant TIICs. The study also utilized 5 GEO datasets and web-based databases to externally validate the findings. RESULTS: The study revealed a statistically significant relationship between key hub genes and immune cells, as determined through co-expression analysis. Two hub regulators (SOX12 and H19) demonstrated significant prognostic value in the ceRNA-related prognostic model, and their elevated expression levels were verified across multiple CRC cell lines. Additionally, the knockdown of SOX12 led to a suppression of proliferation, migration, and invasion in colon cancer cells. CONCLUSION: Through the construction of ceRNA networks and evaluation of TIICs, the study successfully established two risk score models and nomograms. These models serve as valuable tools for understanding the molecular processes and predicting the prognosis of COAD patients. Further validation of hub regulators SOX12 and H19 substantiates their potential role as key biomarkers in COAD.

The application and research progress of anti-angiogenesis therapy in tumor immunotherapy.

Tumor immunotherapy, as the focus of scientific research and clinical tumor treatment in recent years, has received extensive attention. Due to its remarkable curative effect and fewer side effects than traditional treatments, it has significant clinical benefits for the treatment of various advanced cancers and can improve cancer patient survival in the long term. Currently, most patients cannot benefit from immunotherapy, and some patients may experience tumor recurrence and drug resistance even if they achieve remission overcome. Numerous studies have shown that the abnormal angiogenesis state of tumors can lead to immunosuppressive tumor microenvironment, which affects the efficacy of immunotherapy. Actually, to improve the efficacy of immunotherapy, the application of anti-angiogenesis drugs to normalize abnormal tumor vessel has been widely confirmed in basic and clinical research. This review not only discusses the risk factors, mechanisms, and effects of abnormal and normalized tumor angiogenesis state on the immune environment, but summarizes the latest progress of immunotherapy combined with anti-angiogenic therapy. We hope this review provides an applied reference for anti-angiogenesis drugs and synergistic immunotherapy therapy.

Boosting B Cell and Macrophage-Mediated Humoral Immunity with Fusion Nanovesicles for Triple-Negative Breast Cancer Combined Therapy.

Cell-derived nanovesicles are widely utilized as therapeutic agents for cancer therapy. Current research mostly focuses on their ability to activate antitumor cellular immunity. However, whether they can activate and participate in antitumor humoral immunity is rarely studied. Here, doxorubicin-loaded hybrid cell nanovesicles (DNVs) are designed for boosting antitumor humoral and cellular immunity. The hybrid cell nanovesicles are generated through fusion of nanovesicles derived from M1-type macrophages and 4T1 tumor cells. It is found that DNVs can accumulate at tumor tissues and draining lymph nodes effectively, which results in the activation of antitumor immune response and significant inhibition of tumor progression. During this process, dendritic cells are effectively activated, subsequently inducing cytotoxicity T lymphocytes-mediated cellular immunity. Furthermore, DNVs elicit the antitumor humoral immunity through boosting T follicular helper cells and germinal center B cells. By analyzing the mechanism behind humoral immunity activation, it is found that M1-type macrophages repolarized by DNVs play an important role. In general, besides antitumor cellular immunity, the proposed hybrid nanovesicles provide a promising strategy for enhancing antitumor humoral immunity by macrophages repolarization and germinal center B cells activation.

PLGA Nanoparticles Loaded with Sorafenib Combined with Thermosensitive Hydrogel System and Microwave Hyperthermia for Multiple Sensitized Radiotherapy.

Hypoxia is typically the leading cause of radiotherapy (RT) resistance in solid tumors, and glutathione (GSH) overexpression in tumor cells is a potent antioxidant mechanism that protects tumor cells from radiation damage. Herein, we developed a sorafenib (SFN) loaded-PLGA hydrogel system (SPH) in combination with microwave (MW) hyperthermia for RT sensitization. SPH with stable properties was produced by combining SFN and PLGA in a specific ratio and encapsulating the mixture in agarose hydrogel. Intratumoral injection of SPH to mice combined with MW hyperthermia can not only directly cause thermal damage to tumor cells, but also increase blood oxygen delivery to the tumor site, thus overcoming the problem of intratumoral hypoxia and achieving "first layer" RT sensitization. Moreover, high temperatures can cause the hydrogel to disintegrate and release SFN. Not only can SFN inhibit tumor growth, but it can also achieve the "second layer" of RT sensitization by inhibiting glutathione (GSH) synthesis in cells and increasing reactive oxygen species (ROS) production. Experiments, both in vitro and in vivo, have indicated that SPH and MW hyperthermia can achieve a double RT sensitization effect and a significant tumor inhibition effect. In conclusion, combining our SPH nanosystem and thermoradiotherapy is a promising anti-tumor treatment.

Combining radiation and the ATR inhibitor berzosertib activates STING signaling and enhances immunotherapy via inhibiting SHP1 function in colorectal cancer.

BACKGROUND: Immune checkpoint inhibitors (ICIs) targeting programmed cell death protein 1 (PD-1) and programmed death-ligand 1 (PD-L1) have shown a moderate response in colorectal cancer (CRC) with deficient mismatch repair (dMMR) functions and poor response in patients with proficient MMR (pMMR). pMMR tumors are generally immunogenically "cold", emphasizing combination strategies to turn the "cold" tumor "hot" to enhance the efficacy of ICIs. ATR inhibitors (ATRi) have been proven to cooperate with radiation to promote antitumor immunity, but it is unclear whether ATRi could facilitate the efficacy of IR and ICI combinations in CRCs. This study aimed to investigate the efficacy of combining ATRi, irradiation (IR), and anti-PD-L1 antibodies in CRC mouse models with different microsatellite statuses. METHODS: The efficacy of combining ATRi, IR, and anti-PD-L1 antibodies was evaluated in CRC tumors. The tumor microenvironment and transcriptome signatures were investigated under different treatment regimens. The mechanisms were explored via cell viability assay, flow cytometry, immunofluorescence, immunoblotting, co-immunoprecipitation, and real-time quantitative PCR in multiple murine and human CRC cell lines. RESULTS: Combining ATRi berzosertib and IR enhanced CD8+ T cell infiltration and enhanced the efficacy of anti-PD-L1 therapy in mouse CRC models with different microsatellite statuses. The mechanistic study demonstrated that IR + ATRi could activate both the canonical cGAS-STING-pTBK1/pIRF3 axis by increasing cytosolic double-stranded DNA levels and the non-canonical STING signaling by attenuating SHP1-mediated inhibition of the TRAF6-STING-p65 axis, via promoting SUMOylation of SHP1 at lysine 127. By boosting the STING signaling, IR + ATRi induced type I interferon-related gene expression and strong innate immune activation and reinvigorated the cold tumor microenvironment, thus facilitating immunotherapy. CONCLUSIONS: The combination of ATRi and IR could facilitate anti-PD-L1 therapy by promoting STING signaling in CRC models with different microsatellite statuses. The new combination strategy raised by our study is worth investigating in the management of CRC.

MicroRNA-621 functions as a metastasis suppressor in colorectal cancer by directly targeting LEF1 and suppressing Wnt/ß-catenin signaling.

AIMS: Colorectal liver metastasis (CRLM) is the leading death-causing among colorectal cancer (CRC) patients. Recently, a novel tumor-related microRNA, miR-621, has been identified as a tumor suppressor in diverse tumor types, but its role in CRLM remains unclear and requires further investigation. MAIN METHODS: To elucidate novel regulators of CRLM progression, we used a well-established CRLM animal model. After serially transplanting human colon carcinoma cell lines Caco-2 into the liver, we obtained liver metastatic variants that exhibited a strong ability for invasion and metastasis. High-throughput sequencing was conducted on these newly established cell lines. After comparison and prediction between the two cell lines: parental Caco-2 (hereafter referred to as F0) and F3, miR-621 was identified as a candidate regulator for lymphoid enhancer-binding factor 1 (LEF1) expression. Further validation was achieved with dual-luciferase reporter assay. KEY FINDINGS: The gain- and loss-of-function validation showed that miR-621 inhibits cell viability, cell cycle progression, colony formation, and proliferation in vitro. Meanwhile, miR-621 could reverse EMT malignant phenotype. LEF1, an important downstream mediator of activated Wnt/ß-catenin signaling pathway, was validated as the direct functional target of miR-621. miR-621 interacts directly with the LEF1 3'-UTR and post-transcriptionally suppresses LEF1 expression. Moreover, LEF1 overexpression reversed the effect of miR-621. LEF1 silencing counteracted miR-621 down-regulation-induced effects. Further in vivo experiments revealed that miR-621 over-expression suppressed CRLM, but LEF1 abrogated the inhibitory effect of miR-621. SIGNIFICANCE: MiR-621 is a vital tumor suppressor in CRC and could be a promising anti-cancer therapeutic target.

Analysis of Ferroptosis-Related LncRNAs Signatures Associated with Tumor Immune Infiltration and Experimental Validation in Clear Cell Renal Cell Carcinoma.

Background: Clear cell renal cell carcinoma (ccRCC) is the most aggressive subtype of renal cell carcinoma. Ferroptosis is an iron-dependent programmed cell death. Long non-coding RNAs (lncRNAs) emerge as a critical role in regulating cancer progression. Objective: This study aimed to identify molecular regulation of ferroptosis-related lncRNAs (FRLs) in ccRCC. Methods: The prognostic value of FRLs was investigated in ccRCC samples downloaded from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) dataset. The FRLs were screened out by Pearson correlation test. The 465 FRLs confirmed as potential prognostic factors through univariate Cox regression analysis were entered into Lasso and multivariate Cox regression to build a FRLs prognostic signature. A risk score based on the prognostic model divided ccRCC patients into low- and high-risk groups. A prognostic nomogram, derived from the prognostic signature and integrating clinical characteristics, was constructed. Gene set enrichment analysis (GSEA) revealed the immune- and tumor-associated pathways. Two distinct clusters were identified with different immune signatures through consensus clustering analysis. The prognostic value of some hub FRLs was externally validated via three GEO datasets (GSE46699, GSE53757 and GSE66272) and online databases. Finally, the three FRLs (LINC00460, LINC00941 and LINC02027) were verified through in vitro experiments. Results: The FRLs prognostic signature, including 7 independent prognostic lncRNAs, exhibited good accuracy in predicting overall survival (OS) of ccRCC patients. This signature was correlated with immune infiltration and immune checkpoint blockade (ICB). We correlated two distinct clusters with immune infiltration signature of ccRCC. The worse prognosis of cluster 2 was probably mediated by immune evasion. We also found that the expression levels of LINC00460 and LINC00941 in ccRCC cell lines were higher than those in HK-2 cells, but LINC02027 showed the inverse trend. Conclusion: Collectively, our study demonstrated a FRLs prognostic signature which had great clinical value in prognosis assessment.

Comprehensive and Integrative Analysis of Two Novel SARS-CoV-2 Entry Associated Proteases CTSB and CTSL in Healthy Individuals and Cancer Patients.

More than 200 million people have been infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and 4 million deaths have been reported worldwide to date. Cathepsin B/cathepsin L (CTSB/L) are SARS-CoV-2 entry-associated proteases and facilitate SARS-CoV-2 to infect host cells. However, the expressions of CTSB/L in healthy individuals and cancer patients remain not fully elucidated yet. Here, we comprehensively profiled the expressions and distributions of CTSB/L in human normal tissues, cancer tissues, and cell lines. Moreover, we compared CTSB/L expressions between various cancers and matched normal tissues, and investigated their genetic alteration and prognostic values in pan-cancer. Finally, we also explored the correlation between CTSB/L expressions and immune infiltration. We found that CTSB was highly expressed in most tissues, and CTSL was highly expressed predominantly in the digestive, urinary, and respiratory systems, such as the lungs, liver and gallbladder, and kidney tissues in the translational level. Moreover, cancer patients may be more susceptible to SARS-CoV-2 infection. Our data suggested that CTSB/L are overexpressed in aerodigestive and genitourinary cancers when compared with that in matched normal tissues, and their expressions were closely related to the prognosis of some cancer types. Interestingly, CTSB/L expressions were significantly correlated with immune cell infiltration in manifold cancer tissues and their corresponding normal tissues. In conclusion, our study shows a comprehensive bioinformatic analysis of two important SARS-CoV-2 entry-related proteases, which could provide a potential indication on prevention of SARS-CoV-2 infection.

Nintedanib enhances the efficacy of PD-L1 blockade by upregulating MHC-I and PD-L1 expression in tumor cells.

Background: Immune checkpoint inhibitors (ICIs), such as programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1), have been widely applied in clinical and scientific research. Despite their effective antitumor effects in clinical tumor therapy, most tumors are still resistant to ICIs and long-term benefits are lacking. In addition, tumor patients complicated with interstitial lung disease limit the application of ICI therapy. Therefore, for these cases, there is an urgent need to develop new methods to relieve lung complications and enhance the efficacy of ICI therapy. Nintedanib, a potent triple angiokinase inhibitor approved for the treatment of progressive fibrotic interstitial lung disease. However, its immunotherapy synergy properties and mechanism are still pending further exploration. Methods: To explore the therapeutic potential of nintedanib and αPD-L1 combination therapy, MC38, LLC, and 4T1 tumor models were used to investigate antitumor and antimetastatic activities in vivo. An idiopathic pulmonary fibrosis-tumor bearing model was used to evaluate the effect of the synergy therapy on tumor model complicated with lung disease. Moreover, RNA-seq, immunohistochemistry, and flow cytometry were utilized to analyze the effect of combination treatment on the tumor microenvironment. The bioactivity following different treatments was determined by western blotting, CCK-8, and flow cytometry. Results: In this study, nintedanib and αPD-L1 synergy therapy exhibited significant antitumor, antimetastatic and anti-pulmonary fibrosis effects. Both in vitro and in vivo experiments revealed that these effects included promoting vessel normalization, increasing infiltration and activation of immune cells in tumors, enhancing the response of interferon-gamma, and activating the MHC class I-mediated antigen presentation process. Moreover, our results showed an increased expression of PD-L1 and promoted phosphorylation of STAT3 after nintedanib (1 µM) treatment. Conclusion: The combination of nintedanib and αPD-L1 increased ICI therapy responses, relieved lung complications and further activated the tumor immune microenvironment; thus, exhibiting a notable antitumor effect. Accordingly, the nintedanib synergy strategy is expected to be a promising candidate therapy for tumor patients complicated with interstitial lung disease in clinical practice.

Combination of Chidamide-Mediated Epigenetic Modulation with Immunotherapy: Boosting Tumor Immunogenicity and Response to PD-1/PD-L1 Blockade.

Improving tumor immunogenicity is critical for increasing the responsiveness of triple-negative breast cancer (TNBC) to anti-PD-(L)1 treatment. Here, we verified that chidamide (CHI), an epigenetic modulator, could elicit immunogenic cell death within TNBC to enhance cancer immunogenicity and elicit an antitumor immune response. Additionally, CHI increased the expression level of PD-L1, MHC I, and MHC II on cancer cells, which contributed to T-cell recognition and PD-1/PD-L1 blockade therapy response. The synergistic antitumor efficacy of CHI and PD-L1 blockade therapy was further explored through liposomes co-delivering CHI and BMS-202 (a small-molecule PD-L1 inhibitor). The liposomes possessed good biocompatibility, security, and controllable drug release and endowed therapeutics drugs with favorable tumor accumulation. Furthermore, the drug-loaded liposomes could obviously boost the antitumor immunity of TNBC through CHI-enhanced tumor immunogenicity and BMS-202-mediated PD-L1 blockade, thereby effectively inhibiting the growth of primary and metastatic tumors with an inhibitory rate of metastasis of up to 96%. In summary, this work provided a referable and optional approach for clinical antitumor therapy based on the combination of an epigenetic modulator and PD-1/PD-L1 blockade therapy.