The twenty-first century has already recorded more than ten major epidemics or pandemics of viral disease, including the devastating COVID-19. Novel effective antivirals with broad-spectrum coverage are urgently needed. Herein, we reported a novel broad-spectrum antiviral compound PAC5. Oral administration of PAC5 eliminated HBV cccDNA and reduced the large antigen load in distinct mouse models of HBV infection. Strikingly, oral administration of PAC5 in a hamster model of SARS-CoV-2 omicron (BA.1) infection significantly decreases viral loads and attenuates lung inflammation. Mechanistically, PAC5 binds to a pocket near Asp49 in the RNA recognition motif of hnRNPA2B1. PAC5-bound hnRNPA2B1 is extensively activated and translocated to the cytoplasm where it initiates the TBK1-IRF3 pathway, leading to the production of type I IFNs with antiviral activity. Our results indicate that PAC5 is a novel small-molecule agonist of hnRNPA2B1, which may have a role in dealing with emerging infectious diseases now and in the future.
Animals ; Mice ; Antiviral Agents/pharmacology* ; COVID-19 ; Hepatitis B virus ; Interferon Type I/metabolism* ; SARS-CoV-2/drug effects* ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B/antagonists & inhibitors*

OBJECTIVES: Tongue squamous cell carcinoma (TSCC) is a common cancer in the oral and maxillofacial region, which seriously endangers people's life and health.Heterogeneous nuclear ribonucleoprotein A2/B1(hnRNP A2/B1) is an RNA-binding protein that regulates the expression of a variety of genes and participates in the occurrence and development of a variety of cancers. This study aims to investigate the role of hnRNP A2/B1 in TSCC progression. METHODS: The differential expression of hnRNP A2/B1 in oral squamous cell carcinoma (OSCC) and normal oral mucosa cells and tissues was analyzed based on the gene expression profiles of GSE146483 and GSE85195 in the Gene Expression Omnibus (GEO) database. The correlation between hnRNP A2/B1 expression and disease-free survival of TSCC patients was analyzed based on TSCC related chip of GSE4676. TSCC cancer and paracancerous tissue samples of 30 patients were collected in Hunan Cancer Hospital from July to December 2021. Real-time RT-PCR and Western blotting were used to verify the mRNA and protein expression of hnRNP A2/B1 in TSCC patients'samples, respectively. Human TSCC Tca-8113 cells were transfected with hnRNP A2/B1 empty vector (a sh-NC group), knockdown plasmid (a sh-hnRNP A2/B1 group), empty vector overexpression plasmid (an OE-NC group) and overexpression plasmid (an OE-hnRNP A2/B1 group), respectively. The knockdown or overexpression efficiency of hnRNP A2/B1 was detected by Western blotting. The proliferation activity of Tca-8113 cells was detected by cell counting kit-8 (CCK-8), and the apoptosis rate of Tca-8113 cells was detected by flow cytometry. RESULTS: Based on the analysis of OSCC-related chips of GSE146483 and GSE85195 in the GEO database, it was found that hnRNP A2/B1 was differentially expressed in the OSCC and normal oral mucosa cells and tissues (all P<0.01). Meanwhile, the analysis of TSCC related chip GSE4676 confirmed that the expression of hnRNP A2/B1 was negatively correlated with the disease-free survival of TSCC patients (P=0.006). The results of real-time RT-PCR and Western blotting showed that the relative expression levels of hnRNP A2/B1 mRNA and protein in TSCC tissues were significantly up-regulated compared with those in adjacent tissues (all P<0.01). The results of Western blotting showed that the expression level of hnRNP A2/B1 in Tca-8113 cells was significantly inhibited or promoted after knockdown or overexpression of hnRNP A2/B1 (all P<0.01). The results of CCK-8 and flow cytometry showed that inhibition of hnRNP A2/B1 expression in Tca-8113 cells reduced cell proliferation activity (P<0.05) and increased cell apoptic rate (P<0.01). Overexpression of hnRNP A2/B1 in Tca-8113 cells significantly increased cell proliferation (P<0.05) and decreased cell apoptosis (P<0.01). CONCLUSIONS HnRNP A2/B1 is a key factor regulating the proliferation and apoptosis of TSCC cells. Inhibition of hnRNP A2/B1 expression can reduce the proliferation activity of TSCC cells and promote the apoptosis of TSCC cells.
Humans ; Carcinoma, Squamous Cell/genetics* ; Sincalide/metabolism* ; Tongue Neoplasms/genetics* ; Mouth Neoplasms ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B/metabolism* ; RNA, Messenger ; Tongue/metabolism* ; Cell Line, Tumor

BACKGROUND: Circulating cell-free nucleic acids are known to be a noninvasive diagnostic tool for cancer detection. Heterogeneous nuclear ribonucleoprotein (hnRNP) B1, a nuclear core complex, is overexpressed in early stage lung cancer. We intended to evaluate the usefulness of plasma hnRNP B1 mRNA in differentiating non-small cell lung cancer (NSCLC) from other benign lung diseases, especially pulmonary tuberculosis, which is highly prevalent in Korea and often difficult to distinguish from lung cancer. METHODS: Plasma RNA was extracted from 30 patients with NSCLC, 30 patients with benign lung diseases including pulmonary tuberculosis, and 10 healthy controls. Plasma hnRNP B1 mRNA was measured by TaqMan Gene Expression Assay (Applied Biosystems, USA), and pre-developed beta-actin (ACTB) mRNA was used for normalization. We analyzed the relative gene expression data using the delta-delta Ct method. RESULTS: Plasma hnRPN B1 mRNA was measurable in 93.3% (28/30) of NSCLC patients. Normalized 2-DeltaDeltaCt of plasma hnRPN B1 mRNA was 62.2 (95%Cl, 6.4-210.1) in NSCLC patients and 2.7 (95%Cl, 0.5-13.6) in benign lung disease patients (P<0.001, Mann-Whitney U test). CONCLUSIONS: Plasma hnRNP B1 mRNA was significantly increased in patients with lung cancer compared with that in patients with other benign lung diseases. Plasma hnRNP B1 mRNA may be useful as a potential marker for the detection of NSCLC.
Adult ; Aged ; Carcinoma, Non-Small-Cell Lung/*genetics ; Female ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B/*blood/genetics ; Humans ; Lung Diseases/blood/genetics ; Lung Neoplasms/*genetics ; Male ; Middle Aged ; Polymerase Chain Reaction ; RNA, Messenger/*blood/metabolism ; Tumor Markers, Biological

<b>OBJECTIVEb>To screen and identify the proteins related with tumor metastasis of gastric cancer in a nude mouse model bearing orthotopic transplanted tumor.

<b>METHODSb>Zinc finger protein 139 (ZNF139)-specific siRNA was synthesized and transfected into gastric cancer cell line SGC7901, which was then screened by G418. ZNF139-siRNA-transfected cells, negative plasmid-transfected cells and untreated SGC7901 cells were orthotopically transplanted separately on the stomach wall of BALB/c nude mice. The primary tumors and metastatic lymph nodes were harvested to separate the proteins by 2-D fluorescence difference gel electrophoresis (2-D DIGE); after gel digestion, the differential proteins were subjected to liquid chromatography-mass spectrometry (LC-MS) for identification and their functions were analyzed. Western blotting was performed to verify the identified proteins.

<b>RESULTSb>ZNF139 expression was effectively inhibited in siRNA-transfected SGC7901 cells. ZNF139-siRNA-transfected cells showed obviously suppressed tumor growth with a lowered lymph node metastasis rate in nude mice compared with untreated cells and the negative control cells (P<0.05). Proteomic study with 2-D DIGE showed that fascin and hnRNPA2/B1 were down-regulated while ANXA1 was up-regulated in the primary tumors, and ANXA5 was down-regulated in the metastatic lymph nodes in ZNF139-siRNA-transfected group. Western blotting confirmed the results of proteomic analysis.

<b>CONCLUSIONb>ZNF139 gene may promote lymph node metastasis of gastric cancer by regulating fascin, hnRNPA2/B1, ANXA1, and ANXA5.

Animals ; Annexins ; metabolism ; Blotting, Western ; Cell Line, Tumor ; Chromatography, Liquid ; Electrophoresis, Gel, Two-Dimensional ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B ; metabolism ; Humans ; Kruppel-Like Transcription Factors ; metabolism ; Lymphatic Metastasis ; Mice ; Mice, Nude ; Neoplasm Proteins ; metabolism ; Neoplasm Transplantation ; Proteomics ; RNA, Small Interfering ; Stomach Neoplasms ; pathology ; Transfection

<b>OBJECTIVEb>To study the expression of EGFR, survivn and hnRNPA2/B1 proteins in ThinPrep bronchial brushing cytology specimens and their significance in diagnosis of lung cancer.

<b>METHODSb>The protein expression of EGFR, survivn and hnRNPA2/B1 was detected by immunocytochemistry (ICC) in the specimens from 110 cases of non-small cell lung cancer (NSCLC), 32 cases of small cell lung cancer (SCLC) and 37 cases of non-neoplastic lung lesions. The relationship between EGFR, survivn and hnRNPA2/B1 protein expression and clinical characteristics of the patients was analyzed using SPSS 16.0 software.

<b>RESULTSb>(1) Positive expression was observed in 81.1% of NSCLC for EGFR, 66.2% for survivn, 90.9% for hnRNPA2/B1, significantly higher in NSCLC than in the control specimens (P = 0.000, P = 0.000, P = 0.010). The positive expression rate of hnRNPA2/B1 in SCLC was 92.3%, significantly higher than that in the control specimens (P = 0.021). (2) The expression of EGFR was associated with differentiation (P = 0.003), clinical stage (P = 0.023) and lymph node metastasis (P = 0.038), but was not associated with gender, age and histological types. The survivn expression was not related with the above mentioned clinicopathological features (P>0.05). Expression of hnRNPA2/B1 was associated with clinical stage (P = 0.017), but not associated with gender, age, histological type, differentiation and lymph node metastasis. (3) There was a significant difference between the co-expression of EGFR and survivn in NSCLC (98.0%) and benign conditions (2.0%, P = 0.000), also a significant difference between the negative expression of both EGFR and survivn in NSCLC (38.2%) and nonneoplastic lesions (61.8%, P = 0.000).

<b>CONCLUSIONSb>Analysis of EGFR, survivn and hnRNPA2/B1 expression may be an useful adjunct method to stratify controversial cases. The positive expression of EGFR might be associated with invasion, progression and poor prognosis of NSCLC.

Adult ; Aged ; Aged, 80 and over ; Bronchi ; pathology ; Bronchoscopy ; methods ; Carcinoma, Non-Small-Cell Lung ; diagnosis ; metabolism ; pathology ; Cytodiagnosis ; methods ; Female ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B ; metabolism ; Humans ; Inhibitor of Apoptosis Proteins ; metabolism ; Lung Neoplasms ; diagnosis ; metabolism ; pathology ; Lymphatic Metastasis ; Male ; Middle Aged ; Neoplasm Staging ; Receptor, Epidermal Growth Factor ; metabolism ; Small Cell Lung Carcinoma ; diagnosis ; metabolism ; pathology

<b>OBJECTIVEb>To clone the variable region genes of the monoclonal antibody (McAb) against human heterogeneous nuclear ribonucleoprotein A2/B1 (HnRNPA2/B1), ligate them to assemble single chain Fv (ScFv) gene and express in Escherichia coli.

<b>METHODSb>The specificity of the anti-HnRNPA2/B1 McAb 3E8 to synthetic HnRNPA2/B1 peptide, HnRNPA2/B1 protein in lung cancer cells were examined by dot-immunobinding assay, Western blot and immunohistochemistry. The variable region genes of heavy chain (VH) and light chain (VL) were amplified from hybridoma cell by reverse transcription-polymerase chain reaction(RT-PCR), and then were linked by a linker peptide using SOE-PCR (splicing by overlap extension-PCR) to construct recombination ScFv gene. The latter was cloned into the expression vector pET28 (a+) and expressed in E coli BL21. The expressed product was identified by SDS-PAGE and competitive ELISA inhibition test.

<b>RESULTSb>It was shown that the McAb combined specifically with synthetic HnRNPA2/B1 peptide and HnRNPA2/B1 protein in three lung cancer cells. The cloned VH gene and VL gene were 345 bp and 309 bp respectively and were linked successfully to obtain ScFv gene. The ScFv protein was expressed in the form of inclusion body, with molecular weight of 28,000 and immunoreactivity to HnRNPA2/B1.

<b>CONCLUSIONb>VH gene, VL gene and ScFv gene of anti-HnRNPA2/B1 antibody were cloned, constructed and functionally expressed in E coli. These results provide the experimental basis for elucidating the role of HnRNPA2/B1 in lung cancer.

Adenocarcinoma ; metabolism ; pathology ; Antibodies, Monoclonal ; genetics ; immunology ; metabolism ; Carcinoma, Small Cell ; metabolism ; pathology ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Cell Line, Tumor ; Cloning, Molecular ; Escherichia coli ; metabolism ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B ; immunology ; Humans ; Immunoglobulin Fragments ; genetics ; Immunoglobulin Heavy Chains ; genetics ; Immunoglobulin Light Chains ; genetics ; Immunoglobulin Variable Region ; genetics ; Lung Neoplasms ; metabolism ; pathology

<b>OBJECTIVEb>To study the expression of heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) in non-small cell lung cancer (NSCLC), and the interaction between hnRNP A2/B1 protein and mRNA of DNA repair enzymes O(6)-methylguanine DNA-methyltransferase (MGMT), 8-oxoguanine DNA glycosylase (OGG1), redox factor 1(Ref-1), DNA-dependent protein kinase (including DNA-PKcs and ku).

<b>METHODSb>The expression and distribution of hnRNP A2/B1 were detected by immunohistochemistry and Western blot on 50 NSCLC samples from patients who underwent resection in Zhongshan Hospital. The hnRNP A2/B1 mRNA expression was tested by real-time PCR. Co-immunoprecipitation (co-IP) combined RT-PCR was used to investigate whether hnRNP A2/B1 could be bound with the mRNA of the above mentioned 5 DNA repair enzymes in human lung cancer cell line (HTB-182). Then immunohistochemistry and real-time PCR were used to detect the expression of MGMT in the same group of patients.

<b>RESULTSb>HnRNP A2/B1 protein and mRNA expressions were increased in the NSCLC tissues than that in the corresponding normal lung tissues. HnRNP A2/B1 was expressed predominantly in the nuclei of tumor cells. The positive rate and immunohistochemistry score of hnRNP A2/B1 in tumor tissue were significantly higher than that in normal tissue (P < 0.01). In stage III-IV NSCLC, hnRNP A2/B1 expression was higher than that in stage I-II. There was no significant differences of hnRNP A2/B1 expression among patients of different age, sex, histological type, and smoking history. The results of co-IP combined RT-PCR suggested that hnRNP A2/B1 is bound with MGMT mRNA, and MGMT expression is decreased in tumor tissue of NSCLC.

<b>CONCLUSIONSb>The results of this study show that hnRNP A2/B1 protein and mRNA are highly expressed in NSCLC, and hnRNP A2/B1 is bound with MGMT mRNA, which indicate that it might be one of the mechanisms of hnRNP A2/B1 participating in the pathogenesis of NSCLC.

Blotting, Western ; Carcinoma, Non-Small-Cell Lung ; genetics ; DNA-Activated Protein Kinase ; metabolism ; Guanine ; analogs & derivatives ; Heterogeneous-Nuclear Ribonucleoprotein Group A-B ; genetics ; metabolism ; Humans ; Immunohistochemistry ; Immunoprecipitation ; Lung ; chemistry ; Lung Neoplasms ; genetics ; RNA, Messenger ; analysis ; Real-Time Polymerase Chain Reaction ; Reverse Transcriptase Polymerase Chain Reaction