Oncolytic adenovirus encoding apolipoprotein A1 suppresses metastasis of triple-negative breast cancer in mice.

BACKGROUND: Dysregulation of cholesterol metabolism is associated with the metastasis of triple-negative breast cancer (TNBC). Apolipoprotein A1 (ApoA1) is widely recognized for its pivotal role in regulating cholesterol efflux and maintaining cellular cholesterol homeostasis. However, further exploration is needed to determine whether it inhibits TNBC metastasis by affecting cholesterol metabolism. Additionally, it is necessary to investigate whether ApoA1-based oncolytic virus therapy can be used to treat TNBC. METHODS: In vitro experiments and mouse breast cancer models were utilized to evaluate the molecular mechanism of ApoA1 in regulating cholesterol efflux and inhibiting breast cancer progression and metastasis. The gene encoding ApoA1 was inserted into the adenovirus genome to construct a recombinant adenovirus (ADV-ApoA1). Subsequently, the efficacy of ADV-ApoA1 in inhibiting the growth and metastasis of TNBC was evaluated in several mouse models, including orthotopic breast cancer, spontaneous breast cancer, and human xenografts. In addition, a comprehensive safety assessment of Syrian hamsters and rhesus monkeys injected with oncolytic adenovirus was conducted. RESULTS: This study found that dysregulation of cholesterol homeostasis is critical for the progression and metastasis of TNBC. In a mouse orthotopic model of TNBC, a high-cholesterol diet promoted lung and liver metastasis, which was associated with keratin 14 (KRT14), a protein responsible for TNBC metastasis. Furthermore, studies have shown that ApoA1, a cholesterol reverse transporter, inhibits TNBC metastasis by regulating the cholesterol/IKBKB/FOXO3a/KRT14 axis. Moreover, ADV-ApoA1 was found to promote cholesterol efflux, inhibit tumor growth, reduce lung metastasis, and prolonged the survival of mice with TNBC. Importantly, high doses of ADV-ApoA1 administered intravenously and subcutaneously were well tolerated in rhesus monkeys and Syrian hamsters. CONCLUSIONS: This study provides a promising oncolytic virus treatment strategy for TNBC based on targeting dysregulated cholesterol metabolism. It also establishes a basis for subsequent clinical trials of ADV-ApoA1 in the treatment of TNBC.

Expanding the ubiquitin code in pancreatic cancer.

The ubiquitin-proteasome system (UPS) is a fundamental regulatory mechanism in cells, vital for maintaining cellular homeostasis, compiling signaling transduction, and determining cell fates. These biological processes require the coordinated signal cascades of UPS members, including ubiquitin ligases, ubiquitin-conjugating enzymes, deubiquitinases, and proteasomes, to ubiquitination and de-ubiquitination on substrates. Recent studies indicate that ubiquitination code rewriting is particularly prominent in pancreatic cancer. High frequency mutation or aberrant hyperexpression of UPS members dysregulates ferroptosis, tumor microenvironment, and metabolic rewiring processes and contribute to tumor growth, metastasis, immune evasion, and acquired drug resistance. We conduct an in-depth overview of ubiquitination process in pancreatic cancer, highlighting the role of ubiquitin code in tumor-promoting and tumor-suppressor pathways. Furthermore, we review current UPS modulators and analyze the potential of UPS modulators as cancer therapy.

Oncolytic viruses engineered to enforce cholesterol efflux restore tumor-associated macrophage phagocytosis and anti-tumor immunity in glioblastoma.

The codependency of cholesterol metabolism sustains the malignant progression of glioblastoma (GBM) and effective therapeutics remain scarce. In orthotopic GBM models in male mice, we identify that codependent cholesterol metabolism in tumors induces phagocytic dysfunction in monocyte-derived tumor-associated macrophages (TAMs), resulting in disease progression. Manipulating cholesterol efflux with apolipoprotein A1 (ApoA1), a cholesterol reverse transporter, restores TAM phagocytosis and reactivates TAM-T cell antitumor immunity. Cholesterol metabolomics analysis of in vivo-sorted TAMs further reveals that ApoA1 mediates lipid-related metabolic remodeling and lowers 7-ketocholesterol levels, which directly inhibits tumor necrosis factor signaling in TAMs through mitochondrial translation inhibition. An ApoA1-armed oncolytic adenovirus is also developed, which restores antitumor immunity and elicits long-term tumor-specific immune surveillance. Our findings provide insight into the mechanisms by which cholesterol metabolism impairs antitumor immunity in GBM and offer an immunometabolic approach to target cholesterol disturbances in GBM.

Metabolic-related gene pairs signature analysis identifies ABCA1 expression levels on tumor-associated macrophages as a prognostic biomarker in primary IDHWT glioblastoma.

Background: Although isocitrate dehydrogenase (IDH) mutation serves as a prognostic signature for routine clinical management of glioma, nearly 90% of glioblastomas (GBM) patients have a wild-type IDH genotype (IDHWT) and lack reliable signatures to identify distinct entities. Methods: To develop a robust prognostic signature for IDHWT GBM patients, we retrospectively analyzed 4 public datasets of 377 primary frozen tumor tissue transcriptome profiling and clinical follow-up data. Samples were divided into a training dataset (204 samples) and a validation (173 samples) dataset. A prognostic signature consisting of 21 metabolism-related gene pairs (MRGPs) was developed based on the relative ranking of single-sample gene expression levels. GSEA and immune subtype analyses were performed to reveal differences in biological processes between MRGP risk groups. The single-cell RNA-seq dataset was used to examine the expression distribution of each MRG constituting the signature in tumor tissue subsets. Finally, the association of MRGs with tumor progression was biologically validated in orthotopic GBM models. Results: The metabolic signature remained an independent prognostic factor (hazard ratio, 5.71 [3.542-9.218], P < 0.001) for stratifying patients into high- and low-risk levels in terms of overall survival across subgroups with MGMTp methylation statuses, expression subtypes, and chemo/ratio therapies. Immune-related biological processes were significantly different between MRGP risk groups. Compared with the low-risk group, the high-risk group was significantly enriched in humoral immune responses and phagocytosis processes, and had more monocyte infiltration and less activated DC, NK, and γδ T cell infiltration. scRNA-seq dataset analysis identified that the expression levels of 5 MRGs (ABCA1, HMOX1, MTHFD2, PIM1, and PTPRE) in TAMs increased with metabolic risk. With tumor progression, the expression level of ABCA1 in TAMs was positively correlated with the population of TAMs in tumor tissue. Downregulation of ABCA1 levels can promote TAM polarization towards an inflammatory phenotype and control tumor growth. Conclusions: The metabolic signature is expected to be used in the individualized management of primary IDHWT GBM patients.

Distinguishing the pros and cons of metabolic reprogramming in oncolytic virus immunotherapy.

Oncolytic viruses (OVs) represent a class of cancer immunotherapies that rely on hijacking the host cell factory for replicative oncolysis and eliciting immune responses for tumor clearance. An increasing evidence suggests that the metabolic state of tumor cells and immune cells is a putative determinant of the efficacy of cancer immunotherapy. However, how therapeutic intervention with OVs affects metabolic fluxes within the tumor microenvironment (TME) remains poorly understood. Herein, we review the complexities of metabolic reprogramming involving the effects of viruses and their consequences on tumor cells and immune cells. We highlight the inherent drawback of oncolytic virotherapy, namely that treatment with OVs inevitably further exacerbates the depletion of nutrients and the accumulation of metabolic wastes in the TME, leading to a metabolic barrier to antitumor immune responses. We also describe targeted metabolic strategies that can be used to unlock the therapeutic potential of OVs.

Clinical efficacy of probiotics on feeding intolerance in preterm infants: a systematic review and meta-analysis.

Background: The physiological organ system of premature infants is still very immature, so it is easy to result feeding intolerance. Therefore, effective probiotic supplementation plays a very important role and clinical research significance in promoting the growth and development of preterm infants, improving the quality of life and improving the occurrence of feeding intolerance. To explore the clinical effect of probiotics on feeding intolerance (FI) in preterm infants by meta-analysis. Methods: The PubMed, Web of Science, EMBASE and MEDLINE literature databases were searched for relevant literature. The literature related to the clinical effect of probiotics on FI in preterm infants was published from January 2002 to January 2021. RevMan 5.3 was used to calculate the reinforcement mean difference (MD) and evaluate the publication bias. Results: Nine articles were included, involving a total of 1,244 preterm infants with FI. Through the sensitivity analysis of each excluded study, the results showed no significant differences. Compared with patients in the control group, the probiotics group had significant improvements (P<0.1) in the total intestinal feeding time (MD =-2.54, 95% CI: -3.57, -1.52, P<0.00001), weight gain (MD =23.81, 95% CI: 19.75, 27.81, P<0.00001), maximum enteral feeding (MD =6.41, 95% CI: 1.94, 10.88, P=0.005), hospital stay (MD =-5.18, 95% CI: -5.63, -4.74, P<0.00001), incidence of FI [odds rate (OR) =0.38, 95% CI: 0.27, 0.55, P<0.00001] and improvement in the gastrointestinal tract (OR =2.34, 95% CI: 1.07, 5.14, P=0.03). Discussion: Our study shows that the use of probiotics can promote the early growth of preterm infants and effectively improve the occurrence of FI in preterm infants.

An engineered oncolytic vaccinia virus encoding a single-chain variable fragment against TIGIT induces effective antitumor immunity and synergizes with PD-1 or LAG-3 blockade.

BACKGROUND: In addition to directly lysing tumors, oncolytic viruses also induce antitumor immunity by recruiting and activating immune cells in the local tumor microenvironment. However, the activation of the immune cells induced by oncolytic viruses is always accompanied by high-level expression of immune checkpoints in these cells, which may reduce the efficacy of the oncolytic viruses. The aim of this study is to arm the oncolytic vaccinia virus (VV) with immune checkpoint blockade to enhance its antitumor efficacy. METHODS: Through homologous recombination with the parental VV, an engineered VV-scFv-TIGIT was produced, which encodes a single-chain variable fragment (scFv) targeting T-cell immunoglobulin and ITIM domain (TIGIT). The antitumor efficacy of the VV-scFv-TIGIT was explored in several subcutaneous and ascites tumor models. The antitumor efficacy of VV-scFv-TIGIT combined with programmed cell death 1 (PD-1) or lymphocyte-activation gene 3 (LAG-3) blockade was also investigated. RESULTS: The VV-scFv-TIGIT effectively replicated in tumor cells and lysed them, and prompt the infected tumor cells to secret the functional scFv-TIGIT. Compared with control VV, intratumoral injection of VV-scFv-TIGIT in several mouse subcutaneous tumor models showed superior antitumor efficacy, accompanied by more T cell infiltration and a higher degree of CD8+ T cells activation. Intraperitoneal injection of VV-scFv-TIGIT in a mouse model of malignant ascites also significantly improved T cell infiltration and CD8+ T cell activation, resulting in more than 90% of the tumor-bearing mice being cured. Furthermore, the antitumor immune response induced by VV-scFv-TIGIT was dependent on CD8+ T cells which mediated a long-term immunological memory and a systemic antitumor immunity against the same tumor. Finally, the additional combination of PD-1 or LAG-3 blockade further enhanced the antitumor efficacy of VV-scFv-TIGIT, increasing the complete response rate of tumor-bearing mice. CONCLUSIONS: Oncolytic virotherapy using engineered VV-scFv-TIGIT was an effective strategy for cancer immunotherapy. Administration of VV-scFv-TIGIT caused a profound reshaping of the suppressive tumor microenvironment from 'cold' to 'hot' status. VV-scFv-TIGIT also synergized with PD-1 or LAG-3 blockade to achieve a complete response to tumors with poor response to VV or immune checkpoint blockade monotherapy.

Enhanced antitumor efficacy of a novel oncolytic vaccinia virus encoding a fully monoclonal antibody against T-cell immunoglobulin and ITIM domain (TIGIT).

BACKGROUND: Oncolytic virotherapy with vaccinia virus (VV) can lead to effective anti-tumor immunity by turning "cold" tumors into "hot" tumors. However, its therapeutic potential is affected by the tumor's local immunosuppressive tumor microenvironment (TME). Therefore, it is necessary to explore the use of immune checkpoint inhibitors to arm oncolytic VVs to enhance their anti-tumor efficacy. METHODS: A novel recombinant oncolytic VV, VV-α-TIGIT, which encoded a fully monoclonal antibody against T-cell immunoglobulin and ITIM domain (TIGIT) was generated by homologous recombination with a shuttle plasmid. The anti-tumor efficacy of the VV-α-TIGIT was investigated in several subcutaneous and ascites tumor models. FINDINGS: The functional α-TIGIT was sufficiently produced and secreted by tumor cells infected with VV-α-TIGIT, which effectively replicated in tumor cells leading to significant oncolysis. Intratumoral injection of VV-α-TIGIT improved anti-tumor efficacy in several murine subcutaneous tumor models compared to VV-Control (without α-TIGIT insertion). Intraperitoneal injection of VV-α-TIGIT achieved approximately 70% of complete tumor regression in an ascites tumor model. At the same time, treatment with VV-α-TIGIT significantly increased the recruitment and activation of T cells in TME. Moreover, the in vivo anti-tumor activity of VV-α-TIGIT was largely dependent on CD8+ T cell-mediated immunity. Finally, the tumor-bearing mice cured of VV-α-TIGIT treatment resisted rechallenge with the same tumor cells, suggesting a long-term persistence of tumor-specific immunological memory. INTERPRETATION: The recombinant oncolytic virus VV-α-TIGIT successfully combines the advantages of oncolytic virotherapy and intratumorally expression of immune checkpoint inhibitor against TIGIT. This novel strategy can provide information on the optimal design of novel antibody-armed oncolytic viruses for cancer immunotherapy. FUNDING: This work was supported by the National Natural Science Foundation of China (81773255, 81472820, and 81700037), the Science and Technology Innovation Foundation of Nanjing University (14913414), and the Natural Science Foundation of Jiangsu Province of China (BK20171098).

Pan-Cancer Analysis of Immune Cell Infiltration Identifies a Prognostic Immune-Cell Characteristic Score (ICCS) in Lung Adenocarcinoma.

Background: The tumor microenvironment (TME) consists of heterogeneous cell populations, including malignant cells and nonmalignant cells that support tumor proliferation, invasion, and metastasis through extensive cross talk. The intra-tumor immune landscape is a critical factor influencing patient survival and response to immunotherapy. Methods: Gene expression data were downloaded from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus databases. Immune cell infiltration was determined by single-sample Gene Set Enrichment Analysis (ssGSEA) depending on the integrated immune gene sets from published studies. Univariate analysis was used to determine the prognostic value of the infiltrated immune cells. Least absolute shrinkage and selection operator (LASSO) regression was performed to screen for the most survival-relevant immune cells. An immune-cell characteristic score (ICCS) model was constructed by using multivariate Cox regression analysis. Results: The immune cell infiltration patterns across 32 cancer types were identified, and patients in the high immune cell infiltration cluster had worse overall survival (OS) but better progression-free interval (PFI) compared to the low immune cell infiltration cluster. However, immune cell infiltration showed inconsistent prognostic value depending on the cancer type. High immune cell infiltration (High CI) indicated a worse prognosis in brain lower grade glioma (LGG), glioblastoma multiforme (GBM), and uveal melanoma (UVM), and favorable prognosis in adrenocortical carcinoma (ACC), cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC), cholangiocarcinoma (CHOL), head and neck squamous cell carcinoma (HNSC), liver hepatocellular carcinoma (LIHC), lung adenocarcinoma (LUAD), sarcoma (SARC), and skin cutaneous melanoma (SKCM). LUAD prognosis was significantly influenced by the infiltration of 13 immune cell types, with high infiltration of all but Type 2 T helper (Th2) cells correlating with a favorable prognosis. The ICCS model based on six most survival-relevant immune cell populations was generated that classified patients into low- and high-ICCS groups with good and poor prognoses, respectively. The multivariate and stratified analyses further revealed that the ICCS was an independent prognostic factor for LUAD. Conclusions: The infiltration of immune cells in 32 cancer types was quantified, and considerable heterogeneity was observed in the prognostic relevance of these cells in different cancer types. An ICCS model was constructed for LUAD with competent prognostic performance, which can further deepen our understanding of the TME of LUAD and can have implications for immunotherapy.

Pd-Catalyzed decarboxylative cross-coupling reactions of epoxides with α,ß-unsaturated carboxylic acids.

A Pd-catalyzed decarboxylative cross-coupling of α,ß-unsaturated carboxylic acids with cyclic and acyclic epoxides has been developed. Both ß-monosubstituted and ß-disubstituted unsaturated carboxylic acids, as well as conjugated diene unsaturated carboxylic acids are suitable reaction substrates. Substituted homoallylic alcohols were obtained in moderate to good yields. The product was obtained as a mixture of diastereomers favoring the anti diastereomer of the cyclic epoxides. This work provides a method for the modification of complex organic molecules containing α,ß-unsaturated carboxylic acids.

Identification of a key candidate gene­phenotype network mediated by glycyrrhizic acid using pharmacogenomic analysis.

Glycyrrhizic acid (GA) is primarily used as an anti­inflammatory agent in cases of chronic hepatitis. However, its underlying mechanisms in diverse biological processes and its reported benefits are yet to be fully elucidated. In the current study, an analytical method based on pharmacogenomics was established to mine disease­modulatory activities mediated by GA. Five primary protein targets and 138 functional partners were identified for GA by querying open­source databases, including Drugbank and STRING. Subsequently, GA­associated primary and secondary protein targets were integrated into Cytoscape to construct a protein­protein interaction network to establish connectivity. GA­associated target genes were then clustered based on Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis. The tumor necrosis factor axis was revealed to be a primary module regulated by GA­associated targets. Furthermore, 12 hub genes were queried to assess their potential anti­cancer effects using cBioPortal. The results indicated that pharmacogenomics­based analysis improved understanding of the underlying drug­target events of GA and provided predictive and definitive leads for future studies.

Cu-catalyzed cross-coupling reactions of vinyl epoxide with organoboron compounds: access to homoallylic alcohols.

Copper-catalyzed cross-coupling reactions of vinyl epoxide with arylboronates to obtain aryl-substituted homoallylic alcohols are described. The reaction selectivity was different to that of previously reported vinyl epoxide ring-opening reactions using aryl nucleophiles. The reaction proceeded under mild conditions, affording aryl-substituted homoallylic alcohols with high selectivity and in good to excellent yields. The reaction provides convenient access to aryl-substituted homoallylic alcohols from vinyl epoxide.

Epichlorohydrin-Cross-linked Hydroxyethyl Cellulose/Soy Protein Isolate Composite Films as Biocompatible and Biodegradable Implants for Tissue Engineering.

A series of epichlorohydrin-cross-linked hydroxyethyl cellulose/soy protein isolate composite films (EHSF) was fabricated from hydroxyethyl cellulose (HEC) and soy protein isolate (SPI) using a process involving blending, cross-linking, solution casting, and evaporation. The films were characterized with FTIR, solid-state (13)C NMR, UV-vis spectroscopy, and mechanical testing. The results indicated that cross-linking interactions occurred in the inter- and intramolecules of HEC and SPI during the fabrication process. The EHSF films exhibited homogeneous structure and relative high light transmittance, indicating there was a certain degree of miscibility between HEC and SPI. The EHSF films exhibited a relative high mechanical strength in humid state and an adjustable water uptake ratio and moisture absorption ratio. Cytocompatibility, hemocompatibility and biodegradability were evaluated by a series of in vitro and in vivo experiments. These results showed that the EHSF films had good biocompatibility, hemocompatibility, and anticoagulant effect. Furthermore, EHSF films could be degraded in vitro and in vivo, and the degradation rate could be controlled by adjusting the SPI content. Hence, EHSF films might have a great potential for use in the biomedical field.

Hepatitis C virus core protein induces hypoxia-inducible factor 1α-mediated vascular endothelial growth factor expression in Huh7.5.1 cells.

Hepatitis C virus (HCV) infection is one of the major causes of hepatocellular carcinoma (HCC). It has been demonstrated that the overexpression of angiogenic factors are associated with the maintenance of liver neoplasia. Hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF) are important regulators of angiogenesis and are important in wound healing, the regeneration of new vessels and reproductive functions. The present study investigated the role of the HCV core protein in the induction of HIF-1α and VEGF expression. The HCV core gene and HIF-1α siRNA were transfected into Huh7.5.1 cells. The results demonstrated that the induction of HCV core gene expression in Huh7.5.1 cells leads to the overexpression and stabilization of HIF-1α, and the activation of HIF-1α leads, in turn, to the stimulation of VEGF, which is one of the most important angiogenic factors. These results provide new information to facilitate the understanding of HCC oncogenesis.

Borna disease virus P protein affects neural transmission through interactions with gamma-aminobutyric acid receptor-associated protein.

Borna disease virus (BDV) is one of the infectious agents that causes diseases of the central nervous system in a wide range of vertebrate species and, perhaps, in humans. The phosphoprotein (P) of BDV, an essential cofactor of virus RNA-dependent RNA polymerase, is required for virus replication. In this study, we identified the gamma-aminobutyric acid receptor-associated protein (GABARAP) with functions in neurobiology as one of the viral P protein-interacting cellular factors by using an approach of phage display-based protein-protein interaction analysis. Direct binding between GABARAP and P protein was confirmed by coimmunoprecipitation, protein pull-down, and mammalian two-hybrid analyses. GABARAP originally was identified as a linker between the gamma-aminobutyric acid receptor (GABAR) and the microtubule to regulate receptor trafficking and plays important roles in the regulation of the inhibitory neural transmitter gamma-aminobutyric acid (GABA). We showed that GABARAP colocalizes with P protein in the cells infected with BDV or transfected with the P gene, which resulted in shifting the localization of GABARAP from the cytosol to the nucleus. We further demonstrated that P protein blocks the trafficking of GABAR, a principal GABA-gated ion channel that plays important roles in neural transmission, to the surface of cells infected with BDV or transfected with the P gene. We proposed that during BDV infection, P protein binds to GABARAP, shifts the distribution of GABARAP from the cytoplasm to the nucleus, and disrupts the trafficking of GABARs to the cell membranes, which may result in the inhibition of GABA-induced currents and in the enhancement of hyperactivity and anxiety.

A novel recombinant bacterial vaccine strain expressing dual viral antigens induces multiple immune responses to the Gag and gp120 proteins of HIV-1 in immunized mice.

Recombinant Salmonella enterica serovar Typhi can function as a live vector to deliver foreign antigens to the mammalian immune system and induce both mucosal and systemic immunity. In this study, we generated a recombinant Salmonella Typhi strain pilS-pilT-Gag+ (pVAX1-gp120) harboring the human immunodeficiency virus (HIV) gag gene integrated into the bacterial chromosome and gp120 gene carried by a plasmid. Mice inoculated with this recombinant bacterium through intranasal route produced high titers of IgG to gp120 in sera and IgA to gp120 in fecal washes. In addition, Gag-specific and gp120-specific cytotoxic T lymphocyte (CTL) responses were observed in sorted spleen lymphocytes of immunized mice. These results demonstrated that this recombinant Salmonella Typhi strain elicits multiple immune responses against both Gag and gp120 antigens of HIV, and thus would be a potential vaccine candidate to the prevention of HIV/AIDS.