Differential Immune Checkpoint Protein Expression in HNSCC: The Role of HGF/MET Signaling.

Although inhibitors targeting the PD1/PD-L1 immune checkpoint are showing comparably good outcomes, a significant percentage of head and neck squamous cell carcinoma (HNSCC) patients do not respond to treatment. Apart from using different treatment strategies, another possibility would be to target other immune checkpoints operating in these non-responding tumors. To obtain an overview of which checkpoint ligands are expressed on HNSCC tumor cells and if these ligands are affected by HGF/MET signaling, we used mRNA sequencing and antibody-based techniques for identifying checkpoint ligands in six HNSCC tumor cell lines. Furthermore, we compared our results to mRNA sequencing data. From the checkpoint ligands we investigated, VISTA was expressed the highest at the RNA level and was also the most ubiquitously expressed. PD-L2 and B7-H3 were expressed comparably lower and were not present in all cell lines to the same extent. B7-H4, however, was only detectable in the Detroit 562 cell line. Concerning the effect of HGF on the ligand levels, PD-L2 expression was enhanced with HGF stimulation, whereas other checkpoint ligand levels decreased with stimulation. B7-H4 levels in the Detroit 562 cell line drastically decreased with HGF stimulation. This is of interest because both the checkpoint ligand and the growth factor are reported to be connected to epithelial-mesenchymal transition in the literature.

Progestogen-driven B7-H4 contributes to onco-fetal immune tolerance.

Immune tolerance mechanisms are shared in cancer and pregnancy. Through cross-analyzing single-cell RNA-sequencing data from multiple human cancer types and the maternal-fetal interface, we found B7-H4 (VTCN1) is an onco-fetal immune tolerance checkpoint. We showed that genetic deficiency of B7-H4 resulted in immune activation and fetal resorption in allogeneic pregnancy models. Analogously, B7-H4 contributed to MPA/DMBA-induced breast cancer progression, accompanied by CD8+ T cell exhaustion. Female hormone screening revealed that progesterone stimulated B7-H4 expression in placental and breast cancer cells. Mechanistically, progesterone receptor (PR) bound to a newly identified -58 kb enhancer, thereby mediating B7-H4 transcription via the PR-P300-BRD4 axis. PR antagonist or BRD4 degrader potentiated immunotherapy in a murine B7-H4+ breast cancer model. Thus, our work unravels a mechanistic and biological connection of a female sex hormone (progesterone) to onco-fetal immune tolerance via B7-H4 and suggests that the PR-P300-BRD4 axis is targetable for treating B7-H4+ cancer.

Dehydroepiandrosterone attenuated the immune escape of oral squamous cell carcinoma through NF-κB p65/miR-15b-5p/B7-H4 axis.

OBJECTIVES: We aimed to explore the effects and mechanisms of action of dehydroepiandrosterone (DHEA) on immune evasion of oral squamous cell carcinoma (OSCC) to provide evidence for enhancing the effect of immunotherapy. MATERIALS AND METHODS: A xenograft mouse model and immunohistochemistry were used to reveal the patterns of tumor-infiltrating lymphocytes (TILs). The CAL27 and SCC VII cell lines were used for the in vitro study. Western blotting, qPCR, immunofluorescence, and flow cytometry were used to evaluate the expression of B7-H4. Recombinant mouse B7-H4 protein (rmB7-H4) and PG490, an inhibitor of NF-κB p65 were used for the "rescue study." Gain- and loss-of-function, luciferase reporter, and chromatin immunoprecipitation assays were performed to verify this mechanism. RESULTS: DHEA inhibited tumor growth in an OSCC xenograft mouse model, increased CD8 + cells, and decreased FOXP3 + cells in TILs. DHEA reduced the expression of B7-H4 in CAL27 and SCC VII cells RmB7-H4 reverses the effect of DHEA on tumor growth and TIL patterns. DHEA increased the expression of miR-15b-5p and activated its transcriptional factor NF-κB p65. Further experiments demonstrated that miR-15b-5p inhibited B7-H4 expression by binding to its 3'-UTR regions, and NF-κB p65 activated miR-15b transcription. PG490 reversed the effects of DHEA on tumor growth, antitumor immunity in the OSCC xenograft model, and the expression/phosphorylation of NF-κB p65, miR-15b-5p, and B7-H4. CONCLUSIONS: This study indicates that DHEA attenuates the immune escape of OSCC cells by inhibiting B7-H4 expression, providing new insights for cancer immunotherapy.

The deubiquitinase USP2a promotes tumor immunosuppression by stabilizing immune checkpoint B7-H4 in lung adenocarcinoma harboring EGFR-activating mutants.

B7-H4 is an immune checkpoint crucial for inhibiting CD8+ T-cell activity. A clinical trial is underway to investigate B7-H4 as a potential immunotherapeutic agent. However, the regulatory mechanism of B7-H4 degradation via the ubiquitin-proteasome pathway (UPP) remains poorly understood. In this study, we discovered that proteasome inhibitors effectively increased B7-H4 expression, while EGFR-activating mutants promoted B7-H4 expression through the UPP. We screened B7-H4 binding proteins by co-immunoprecipitation and mass spectrometry and found that USP2a acted as a deubiquitinase of B7-H4 by removing K48- and K63-linked ubiquitin chains from B7-H4, leading to a reduction in B7-H4 degradation. EGFR mutants enhanced B7-H4 stability by upregulating USP2a expression. We further investigated the role of USP2a in tumor growth in vivo. Depletion of USP2a in L858R/LLC cells inhibited tumor cell proliferation, consequently suppressing tumor growth in immune-deficient nude mice by destabilizing downstream molecules such as Cyclin D1. In an immune-competent C57BL/6 mouse tumor model, USP2a abrogation facilitated infiltration of CD95+CD8+ effector T cells and hindered infiltration of Tim-3+CD8+ and LAG-3+CD8+ exhausted T cells by destabilizing B7-H4. Clinical lung adenocarcinoma samples showed a significant correlation between B7-H4 abundance and USP2a expression, indicating the contribution of the EGFR/USP2a/B7-H4 axis to tumor immunosuppression. In summary, this study elucidates the dual effects of USP2a in tumor growth by stabilizing Cyclin D1, promoting tumor cell proliferation, and stabilizing B7-H4, contributing to tumor immunosuppression. Therefore, USP2a represents a potential target for tumor therapy.

Aberrant expression of B7-H4 and B7-H5 contributes to the development of cutaneous squamous cell carcinoma.

Cutaneous squamous cell carcinoma (CSCC) is the second most common malignant tumor of the skin. B7 homolog 4 (B7-H4) and B7-H5 (B7 homolog 5) are associated with a variety of tumors. Investigate the potential role of B7-H4 and B7-H5 in regulating the tumorigenesis and progression of CSCC. B7-H4 and B7-H5 transcriptome data were collected from GEO and TCGA databases and subjected to bioinformatical analysis by protein-protein interaction (PPI) network, functional enrichment analysis, immune analysis, and drug-gene interaction prediction analysis. We characterized the expression of B7-H4 and B7-H5 in carcinoma tissues of CSCC patients by immunohistochemistry. Meanwhile, the clinical correlation of B7-H4 and B7-H5 in CSCC was explored by statistical analysis. B7-H4 and B7-H5 genes were under-expressed in CSCC and correlated with tumor staging. According to GO and KEGG Pathway enrichment analysis, B7-H4, and B7-H5 can regulate the proliferation and activation of T cells, lymphocytes, and monocytes, and the expression of cytokines, such as IL-6 and IL-10, in CSCC. B7-H4 and B7-H5 are also jointly involved in the occurrence and development of CSCC via the JAK-STAT and Notch signaling pathways. We found that B7-H4 and B7-H5 proteins were abnormally highly expressed in CSCC tissue and correlated with tumor size and stage. Our findings offer new insights into the pathogenesis of CSCC and suggest that B7-H4 and B7-H5 are novel tissue biomarkers and promising therapeutic targets for CSCC.

B7-H4 reduces the infiltration of CD8+T cells and induces their anti-tumor dysfunction in gliomas.

B7-H4 is a promising immune checkpoint molecule in tumor immunotherapy. Our previous study showed that high B7-H4 expression was strongly correlated with deficiency in tumor infiltrated lymphocytes (TILs) in glioma patients. On this basis, we investigated the impact of B7-H4 on CD8+TILs in gliomas and the associated molecular mechanism here. B7-H4-positive tumor samples (n=129) from our glioma cohort were used to assess B7-H4 expression and CD8+TIL quantification by immunohistochemistry. CD8+TILs from five glioma patients cultured with B7-H4 protein were used to evaluate anti-tumor dysfunction by flow cytometry and ELISpot. An orthotopic murine glioma model was used to investigate the role of B7-H4 in glioma CD8+TILs by immunohisto- chemistry and flow cytometry. CD8+TILs from glioma patients cultured with B7-H4 protein were used to explore the potential molecular mechanism by RNA sequencing and western blot. Our results showed that glioma CD8+TIL density was negatively correlated with B7-H4 expression both in glioma patient cohort (P < 0.05) and orthotopic glioma murine model (P < 0.01). B7-H4 also lowered the expression of CD137 and CD103 (P < 0.05 for both) in glioma CD8+TILs and reduced their secretion of the anti-tumor cytokines IFN-γ and TNF-α (P < 0.01 for both) in a dose-dependent manner. Furthermore, B7-H4 was found to induce early dysfunction of glioma CD8+TILs by downregulating the phosphorylation of AKT and eNOS (P < 0.05 for both). In conclusion, B7-H4 reduced the infiltration of glioma CD8+TILs and induced an anti-tumor dysfunction phenotype. B7-H4 may also impair the anti-tumor function of glioma CD8+TILs via the AKT-eNOS pathway. These results indicated that B7-H4 may serve as a potential target in future glioma immunotherapy.

Prognostic Value of B7H4 Expression in Patients with Solid Cancers: A Systematic Review and Meta-Analysis.

V-set domain-containing T-cell activation inhibitor 1 (aliases VTCN1, B7H4) participates in tumour immune escape by delivering inhibitory signals to T cells. The purpose of this article was to assess the B7H4 prognostic value in solid cancers. Three databases were searched for relevant articles. The main endpoints were overall survival (OS), disease-specific survival (DSS), progression-free survival (PFS), recurrence-free survival (RFS), and disease-free survival (DFS). Appropriate hazard ratios (HRs) were pooled. The R studio software (version 4.0.3) was used for data analysis. Thirty-one studies met the inclusion criteria. High expression of B7H4 was associated with worse OS (HR = 1.52, 95% CI: 1.37-1.68) but not with DSS (HR = 1.14, 95% CI: 0.49-2.63), RFS (HR = 1.77, 95% CI: 0.75-4.18), DFS (HR = 1.29, 95% CI: 0.8-2.09), or PFS (HR = 1.71, 95% CI: 0.91-3.2) in patients with solid cancers. High expression of B7H4 is associated with a poorer prognosis in patients with solid cancers. B7H4 is a promising prognostic biomarker and immunotherapeutic target for various solid cancers because of its activity in cancer immunity and tumourigenesis.

Interferon-γ in the tumor microenvironment promotes the expression of B7H4 in colorectal cancer cells, thereby inhibiting cytotoxic T cells.

The bioactivity of interferon-γ (IFN-γ) in cancer cells in the tumor microenvironment (TME) is not well understood in the current immunotherapy era. We found that IFN-γ has an immunosuppressive effect on colorectal cancer (CRC) cells. The tumor volume in immunocompetent mice was significantly increased after subcutaneous implantation of murine CRC cells followed by IFN-γ stimulation, and RNA sequencing showed high expression of B7 homologous protein 4 (B7H4) in these tumors. B7H4 promotes CRC cell growth by inhibiting the release of granzyme B (GzmB) from CD8+ T cells and accelerating apoptosis in CD8+ T cells. Furthermore, interferon regulatory factor 1 (IRF1), which binds to the B7H4 promoter, is positively associated with IFN-γ stimulation-induced expression of B7H4. The clinical outcome of patients with CRC was negatively related to the high expression of B7H4 in cancer cells or low expression of CD8 in the microenvironment. Therefore, B7H4 is a biomarker of poor prognosis in CRC patients, and interference with the IFN-γ/IRF1/B7H4 axis might be a novel immunotherapeutic method to restore the cytotoxic killing of CRC cells.

Clinical significance of blocking novel immune checkpoint B7-H4 in urothelial carcinoma of bladder as a potential therapeutic target.

Urothelial Carcinoma of Bladder is complex disease with high mortality and recurrence rates. Current standard regimes have exhibited anti-tumor activity but still, a proportion of patients are non-responsive or in-eligible to receive such treatments. Immune checkpoints have emerged as potential class of therapeutics to be tested in UCB patients. Clinical trials targeting PD-1/PD-L1 axis have been tested in UCB but still a proportion of patients are non-responsive to it which stresses upon identifying new targets. New immune checkpoint B7-H4 has been shown to negatively regulate T cell activity in cancer and is a poor prognostic factor in various solid tumors. In this study we assessed the novel immune checkpoint B7-H4 status in UCB patients. We observed elevated expression of B7-H4 and PD-L1 on CD8+ T cells in circulation of UCB patients. Relative mRNA expression and immunohistochemistry displayed upregulation in bladder tumor tissue. Increased expression of B7-H4 along with PD-L1 in periphery and tumor of UCB patients highlights involvement of B7-H4 in disease progression. Combinatorial blocking of B7-H4 and PD-L1 enhanced IFN-γ and granzyme B in CD8+ T cells functional T cell immune response in UCB patients. Also, B7-H4 was significantly associated with clinico-pathological parameters. Our findings highlight B7-H4 as potential therapeutic target for treatment of UCB patients in future after further validation.

An analysis of placental chorionic villous and decidual basalis tissue immunoreactivity in patients after cesarean section due to a placenta accreta spectrum disorder and elective cesarean section followed by the depressed mood.

An analysis of placental chorionic villous and decidual basalis tissue immunoreactivity in patients after cesarean section due to a placenta accreta spectrum disorder and elective cesarean section followed by a depressed mood. RESEARCH BACKGROUND: Over the past few years, interest in investigating immune dysfunction in patients with psychiatric disorders has increased. B7-H4 is a molecule with immunosuppressive properties that seems to play a key role in establishing maternal tolerance against fetal antigens. The aim of this study was to compare the B7-H4 immunoreactivity levels in patients after cesarean section. METHODS: Placental and decidual tissue samples were obtained from 49 women who delivered at Bielanski Hospital in Warsaw between 2009 and 2015. Fifteen of the patients developed postpartum depression and 14 had a diagnosis of placenta accreta spectrum. The control group consisted of 20 healthy patients on whom cesarean section was performed due to breech presentation at term. RESULTS: The highest levels of B7-H4 immunoreactivity were found in the placental chorionic villous and decidual basalis tissue samples of the patients who later developed postpartum depression, while the lowest levels were found in the samples of those patients with a placenta accreta spectrum disorder. The difference between the B7-H4 immunoreactivity levels of these two groups was statistically significant. The B7-H4 expression levels were statistically significantly higher in the women in the postpartum depression group than in the control group. CONCLUSION: Postpartum depression follows a disturbance of the suppressive milieu responsible for rebalancing the maternal immune system after the initial cytotoxic activation during labor.

Analysis of B7-H4 Expression Across Salivary Gland Carcinomas Reveals Adenoid Cystic Carcinoma-Specific Prognostic Relevance.

B7-H4 (VTCN1), a member of the B7 family, is overexpressed in several types of cancer. Here we investigated the pattern of expression of B7-H4 in salivary gland carcinomas (SGC) and assessed its potential as a prognostic marker and therapeutic target. Immunohistochemistry (IHC) analyses were performed in a cohort of 340 patient tumors, composed of 124 adenoid cystic carcinomas (ACC), 107 salivary duct carcinomas (SDC), 64 acinic cell carcinomas, 36 mucoepidermoid carcinomas (MEC), 9 secretory carcinomas (SC), as well as 20 normal salivary glands (controls). B7-H4 expression was scored and categorized into negative (<5% expression of any intensity), low (5%-70% expression of any intensity or >70% with weak intensity), or high (>70% moderate or strong diffuse intensity). The associations between B7-H4 expression and clinicopathologic characteristics, as well as overall survival, were assessed. Among all tumors, B7-H4 expression was more prevalent in ACC (94%) compared with those of SC (67%), MEC (44%), SDC (32%), and acinic cell carcinomas (0%). Normal salivary gland tissue did not express B7-H4. High expression of B7-H4 was found exclusively in ACC (27%), SDC (11%), and MEC (8%). In SDC, B7-H4 expression was associated with female gender (P = .002) and lack of androgen receptor expression (P = .012). In ACC, B7-H4 expression was significantly associated with solid histology (P < .0001) and minor salivary gland primary (P = .02). High B7-H4 expression was associated with a poorer prognosis in ACC, regardless of clinical stage and histologic subtype. B7-H4 expression was not prognostic in the non-ACC SGC evaluated. Our comparative study revealed distinct patterns of B7-H4 expression according to SGC histology, which has potential therapeutic implications. B7-H4 expression was particularly high in solid ACC and was an independent prognostic marker in this disease but not in the other SGC assessed.

[Effect of miR-125b on T Cell Activation in Aplastic Anemia by Targetting B7-H4].

OBJECTIVE: To investigate the effect of miR-125b on T cell activation in patients with aplastic anemia (AA) and its molecular mechanism. METHODS: A total of 30 AA patients were enrolled in department of hematology, Binzhou Medical University Hospital from January 2018 to October 2021, as well as 15 healthy individuals as healthy control (HC) group. Peripheral blood mononuclear cells (PBMCs) were isolated, in which the levels of miR-125b and B7-H4 mRNA were detected by RT-qPCR. Immunomagnetic beads were used to separate naive T cells and non-naive T cells from AA patients and healthy people to detect the levels of miR-125b and B7-H4 mRNA. Lentivirus LV-NC inhibitor and LV-miR-125b inhibitor were transfected into cells, and T cell activation was detected by flow cytometry. The dual-luciferase reporter gene assay was used to detect the targetting relationship between miR-125b and B7-H4. RT-qPCR and Western blot were used to detect the levels of miR-125b, CD40L, ICOS, IL-10 mRNA and B7-H4 protein. RESULTS: Compared with HC group, the expression of miR-125b was up-regulated but B7-H4 mRNA was down-regulated in PBMCs of AA patients (P <0.05), and the proportions of CD4+CD69+ T cells and CD8+CD69+ T cells in PBMCs of AA patients were higher (P <0.05). The expression of miR-125b was significantly up-regulated but B7-H4 mRNA was down-regulated in both naive T cells and non-naive T cells of AA patients (P <0.05), and non-naive T cells was more significant than naive T cells (P <0.05). Compared with NC inhibitor group, the expression of miR-125b was significantly decreased, the expression level of CD69 on CD4+ and CD8+ T cells in PBMCs was also significantly decreased, while the luciferase activity was significantly increased after co-transfection of miR-125b inhibitor and B7-H4-3'UTR-WT in the miR-125b inhibitor group (P <0.05). Compared with NC inhibitor group, the mRNA and protein levels of B7-H4 were significantly increased in the miR-125b inhibitor group (P <0.05). Compared with miR-125b inhibitor+shRNA group, the expression levels of CD69 on CD4+ and CD8+ T cells were significantly increased, and the levels of CD40L, ICOS and IL-10 mRNA were also significantly increased in the miR-125b inhibitor+sh-B7-H4 group (P <0.05). CONCLUSION: MiR-125b may promote T cell activation by targetting B7-H4 in AA patients.

The Methylation in B7-H4 and BTLA Genes are Associated with the Risk of Pulmonary Tuberculosis.

Background: The important roles of B7 homologous body 4 (B7-H4), B and T lymphocyte attenuator (BTLA) in patients with pulmonary tuberculosis (PTB) have been reported. This study aims to evaluate the association among B7-H4 and BTLA genes polymorphism, methylation and PTB susceptibility. Methodology: Here, we assessed the possible relationship of 10 single nucleotide polymorphisms (SNPs) in B7-H4, BTLA genes with PTB susceptibility in a Chinese population (496 PTB patients and 502 controls) by SNPscan technique. Then, the B7-H4, BTLA genes methylation levels among 98 PTB patients and 97 controls were detected using MethylTarget technique. Results: This study found no significant differences in allele and genotype frequencies of B7-H4 gene rs10754339, rs10801935, rs10923223, rs1937956, rs3738414, BTLA gene rs1982809, rs2971205, rs75368388, rs9288953 variants between PTB patients and controls. Haplotype analysis suggested that the lower frequencies of B7-H4 AATTG haplotype, BTLA GATT haplotype and the higher frequency of BTLA AGTC haplotype were found in PTB patients when compared with controls. We also found that the frequency of BTLA gene rs9288953 C allele was significantly increased in PTB patients with drug resistance. Moreover, the methylation levels of B7-H4 and BTLA genes in PTB patients were greater than that in controls, and rs10754339 variant in B7-H4 gene could affect its methylation level in PTB patients. Conclusion: B7-H4, BTLA genes polymorphism might not affect PTB susceptibility, while the abnormal methylation levels of B7-H4, BTLA genes were associated with the genetic background of PTB.

A SOX9-B7x axis safeguards dedifferentiated tumor cells from immune surveillance to drive breast cancer progression.

How dedifferentiated stem-like tumor cells evade immunosurveillance remains poorly understood. We show that the lineage-plasticity regulator SOX9, which is upregulated in dedifferentiated tumor cells, limits the number of infiltrating T lymphocytes in premalignant lesions of mouse basal-like breast cancer. SOX9-mediated immunosuppression is required for the progression of in situ tumors to invasive carcinoma. SOX9 induces the expression of immune checkpoint B7x/B7-H4 through STAT3 activation and direct transcriptional regulation. B7x is upregulated in dedifferentiated tumor cells and protects them from immunosurveillance. B7x also protects mammary gland regeneration in immunocompetent mice. In advanced tumors, B7x targeting inhibits tumor growth and overcomes resistance to anti-PD-L1 immunotherapy. In human breast cancer, SOX9 and B7x expression are correlated and associated with reduced CD8+ T cell infiltration. This study, using mouse models, cell lines, and patient samples, identifies a dedifferentiation-associated immunosuppression mechanism and demonstrates the therapeutic potential of targeting the SOX9-B7x pathway in basal-like breast cancer.

Expression of B7-H4 in endometrial cancer and its impact on patients' prognosis.

OBJECTIVES: The aim of the study was to evaluate the B7-H4 expression in endometrial cancer cells and to investigate its relationship with patient prognosis and clinicopathological features of the disease. MATERIAL AND METHODS: We performed a single-center, retrospective cohort study that included endometrial cancer patients treated between 2012 and 2019. B7-H4 expression in specimens obtained from 63 patients was examined by immunohistochemical staining. The evaluation of B7H4 immunoreactivity was assessed using Immunoreactivity Scoring (IRS) system. RESULTS: B7-H4 reactivity was observed in all, except one, endometrial cancer patients (98%). Staining intensity: no reaction in one case, weak in 16 (24%) patients, moderate in 25 (37%), and strong in 22 (35%). Twenty-nine (46%) patients showed B7-H4 immunoreactivity in more than 60% of cells, while, in 18 (29%) cases and 16 (25%) patients, the percentages were 30-60% and < 30% respectively. Median IRS was 2 (range 0-6). We found a significantly worse overall survival (OS) rate for patients with high versus low B7-H4 IRS (p = 0.03), however, in multivariate analysis, the difference in patient survival was close to the significance level (p = 0.052). B7-H4 expression was not related to histopathological type of the tumor, tumor grade, lymph node metastases, or the FIGO stage of the disease. CONCLUSIONS: Our result suggests that B7-H4 expression might be a useful prognostic factor in endometrial cancer.

The loss of B7-H4 expression in breast cancer cells escaping from T cell cytotoxicity contributes to epithelial-to-mesenchymal transition.

BACKGROUND: B7 homology 4 (B7-H4), a potential target for cancer therapy, has been demonstrated to inhibit T cell cytotoxicity in the early stages of breast cancer. However, B7-H4 manipulating breast tumor immune microenvironment (TIME) in the tumor progression remains unknown. METHODS: We engineered T cells with B7-H4-specific chimeric antigen receptors (CARs) and performed a T cell co-culture assay to characterize B7-H4 expression level in breast cancer cells escaping from T cell cytotoxicity. We generated B7-H4 knockout (KO) and overexpression (OE) breast cancer cells to determine the epithelial-to-mesenchymal transition (EMT) and stemness characteristics in vitro and in vivo, including tumor proliferation, migration, metastasis and chemoresistance. The Cancer Genome Atlas breast cancer database was accessed to investigate the correlation between B7-H4 expression levels and EMT characteristics in patients with breast cancer. RESULTS: Our result found that B7-H4 expression level was significantly reduced in a subset of breast cancer cells that escaped from the cytotoxicity of B7-H4 CAR-T cells. Compared with wild type cells, B7-H4 KO cells prompt EMT and stemness characteristics, including migration, invasion and metastasis, and OE cells vice versa. The increase in H3K27me3 in KO cells confirmed the epigenetic reprogramming of cancer stem cells. The IC50 of doxorubicin or oxaliplatin significantly increased in KO cells, which was in agreement with a decrease in OE cells. Moreover, a trend of downregulated B7-H4 from stage I to stage II breast cancer patients indicates that the low-expressing B7-H4 breast cancer cells escaping from TIME have spread to nearby breast lymph nodes in the cancer progression. CONCLUSIONS: Our study illuminates the novel role of renouncing B7-H4 in breast cancer cells through immune escape, which contributes to EMT processes and provides new insights for breast cancer treatments.

An Investigation of the Effects of B7-H4 Gene rs10754339 and miR-125a Gene rs12976445 on Cancer Susceptibility.

Objective: To investigate the effects of the B7-H4 gene rs10754339 and miR-125a gene rs12976445 on cancer susceptibility through a case-control study and meta-analysis. Methods: A total of 1,490 cancer patients (lung/gastric/liver/: 550/460/480) and 800 controls were recruited in this case-control study. The meta-analysis was performed by pooling the data from previous related studies and the present study. Results: The results of this study showed that in the Hubei Han Chinese population, the rs10754339 gene was significantly associated with the risk of lung and gastric cancer but not liver cancer, and the rs12976445 gene was significantly associated with the risk of lung cancer but not liver or gastric cancer. The meta-analysis results indicated that rs10754339 and rs12976445 contributed to cancer susceptibility in the Chinese population and also revealed a significant association between rs10754339 and breast cancer risk, as well as between rs12976445 and lung cancer risk. Conclusion: The B7-H4 gene rs10754339 and miR-125a gene rs12976445 may be the potential genetic markers for cancer susceptibility in the Chinese population, which should be validated in future studies with larger sample sizes in other ethnic populations.

The immune checkpoint molecule, VTCN1/B7-H4, guides differentiation and suppresses proinflammatory responses and MHC class I expression in an embryonic stem cell-derived model of human trophoblast.

The placenta acts as a protective barrier to pathogens and other harmful substances present in the maternal circulation throughout pregnancy. Disruption of placental development can lead to complications of pregnancy such as preeclampsia, intrauterine growth retardation and preterm birth. In previous work, we have shown that expression of the immune checkpoint regulator, B7-H4/VTCN1, is increased upon differentiation of human embryonic stem cells (hESC) to an in vitro model of primitive trophoblast (TB), that VTCN1/B7-H4 is expressed in first trimester but not term human placenta and that primitive trophoblast may be uniquely susceptible to certain pathogens. Here we report on the role of VTCN1 in trophoblast lineage development and anti-viral responses and the effects of changes in these processes on major histocompatibility complex (MHC) class I expression and peripheral NK cell phenotypes.

Ethacrynic acid suppresses B7-H4 expression involved in epithelial-mesenchymal transition of lung adenocarcinoma cells via inhibiting STAT3 pathway.

Lung cancer is characterized by high incidence and mortality. 90% of cancer deaths are caused by metastases. The epithelial-mesenchymal transition (EMT) process in cancer cells is a prerequisite for the metastatic process. Ethacrynic acid (ECA) is a loop diuretic that inhibits the EMT process in lung cancer cells. EMT has been related to the tumour immunemicroenvironment. However, the effect of ECA on immune checkpoint molecules in the context of cancer has not been fully identified. In the present study, we found that sphingosylphosphorylcholine (SPC) and TGF-ß1, awell-known EMT inducer, induced the expression of B7-H4 in lung cancer cells. We also investigated the involvement of B7-H4 in the SPC-induced EMT process. Knockdown of B7-H4 suppressed SPC-induced EMT, while B7-H4 overexpression enhanced EMT of lung cancer cells. ECA inhibited SPC/TGF-ß1-induced B7-H4 expression via suppression of STAT3 activation. Moreover, ECA inhibits the colonization of mice lung by tail vein-injected LLC1 cells. ECA-treated mice increased the CD4-positive T cells in lung tumour tissues. In summary, these results suggested that ECA inhibits B7-H4 expression via STAT3 inhibition, leading to SPC/TGF-ß1-induced EMT. Therefore, ECA might be an immune oncological drug for B7-H4-positive cancer, especially lung cancer.

Association of B7H3 and B7H4 gene polymorphisms and protein expression with the development and prognosis of autoimmune thyroid diseases.

BACKGROUND: The prognosis of autoimmune thyroid diseases (AITDs), including Graves' disease (GD) and Hashimoto's disease (HD), varies among patients. B7-H3 and B7-H4, members of the B7 family of proteins, regulate immune response. To clarify the association of B7-H3 and B7-H4 with the pathogenesis and prognosis of AITDs, we examined the expression of the soluble and membrane form of B7-H3 and B7-H4 and genotyped single nucleotide polymorphisms (SNPs) in the B7H3 and B7H4 genes. METHODS: We examined the expression of the membrane form of B7-H3 and B7-H4 by flow cytometry and their soluble forms by enzyme-linked immunosorbent assay. We genotyped SNPs in B7H3 and B7H4 in 187 GD patients, 217 HD patients, and 110 healthy volunteers using the PCR-RFLP method. RESULTS: The frequency of the B7H3 rs3816661 CC genotype was higher in patients with severe HD. G carriers of B7H4 rs10754339 A/G and B7H4 rs13505 T/G were more frequent in patients with AITD. A carrier of B7H4 rs10158166 A/G and C carriers of B7H4 rs3806373 C/T were more frequent in patients with intractable GD. The proportion of B7-H3+ monocytes was higher in the CC genotype of B7H3 rs3816661 C/T than in the other genotypes and was lower in patients with GD and HD than in healthy controls. The concentration of soluble B7-H4 was lower in the TG genotype of B7H4 rs13505 T/G than in the TT genotype and was higher in patients with AITD than in healthy controls. CONCLUSION: B7H3 and B7H4 are associated with AITD susceptibility and prognosis.