The Research Of Immunotherapy By Blocking B7-H3 In Head Neck Squamous Cell Carcinoma (HNSCC)

Head and neck squamous cell carcinomas(HNSCC)account for 90%of cancer types in head neck,and an increase in the incidence has been noted.During the past 30 years,including operation,reconstruction,infection control,adjuvant radio-and chemotherapy are achieved as well as that of development in basic research including tumorigenesis,pathogenesis,disease development,biomedical and target therapy.However,the overall 5-year survival rate is about 53%to 61.5%.Although many factors contribute to treatment failure in HNSCC,profound immune defects were consistently found in these patients.These defects include generalized T-cell anergy and increased levels of myeloid-derived suppressor cells(MDSCs)and tumor-associated macrophages(TAMs).This problem warrants extensive research into practicable therapeutic targets and corresponding antineoplastic agents in the context of rapidly evolving fundamental biomedical and interdisciplinary research aiming at profound understanding of the mechanisms of tumorigenesis,development,recurrence and metastasis of HNSCC.It is believed such efforts would deepen our understanding of HNSCC and allow doctors to achieve a better prognosis of this lethal disease.B7 homolog 3 protein(B7-H3)is a newly identified member of B7 family in 2001.And B7-H3 is a transmembrane protein with immunoglobulin-like structure,which is known to have immunoregulatory properties with both inhibitory and stimulatory effects on the activation of T cells.Though immunotherapy for cancer has been listed as one of top ten scientific breakthroughs in 2013,the medical community still lacks sufficient research on immunotherapies in the field of HNSCC.MDSCs and TAMs play a pivotal part in the formation and maintenance of immunosuppressive microenvironment.However,the relationship between the B7-H3 expression and clinical features in HNSCC patients remains unclear and the anti-tumor immune response of B7-H3 is still uncertain.In this research,the expression and location of B7-H3 was detected by immunohistochemistry and immunofluorescence.The relation between the expression level and tumor clinical patho-features and patient prognosis were statistically analyzed,and further investigated by in vitro and in vivo function assays.Results show overexpression of B7-H3 in human HNSCC confers poor prognosis of HNSCC patients.Using B7-H3 monoclonal antibody reduced MDSCs and M2 macrophages and improved T cell effector function in Tgfbrl/Pten 2cKO HNSCC mouse model.Part ⅠThe expression level of B7-H3 in HNSCC and its clinical significanceObjective:B7 molecular superfamily in the vast majority of molecules and their ligands in tumor immunotherapy and tumor development process has played a very critical role.Current part aims at studying multi-functional of immunosuppressive molecule B7-H3 and its expression in HNSCC,as well as its subcellular locations and the clinical significance.Methods:Firstly,we analyzed public database Oncomine(cancer microarray database)to study CD276 in available cancer research;then we used tissue microarrays to study B7-H3 expression pattern by immunohistochemistry;expression data were quantified and analyzed by Aperio Scanscope,and differential expression pattern were analyzed by Pearson correlation and hierarchical cluster;association B7-H3 with tumor clinical pathological features and patient prognosis were analyzed by SPSS based statistics.Results:Positive immunostaining of B7-H3 were mainly located in cancer cells,especially in the invasive frontier of cancer.Also,immunoreactivity of B7-H3 can be detected in stromal and infiltrating inflammatory cells.Immunostaining of B7-H3 was significantly increased in human HNSCC tissue as compared with rather lower expressions in oral mucosa(P<0.001)and dysplasia(P<0.001).Kaplan-Meier survival analysis indicated that B7-H3 high expression confers poor overall survival in patient with HNSCC(n = 165,P = 0.0390)with median expression of B7-H3 as a cut-off.Particularly,we found that B7-H3 staining was not significantly altered in different pathology grades,lymph node metastasis status and tumor size.Multivariate analysis by Cox proportional hazard regression model showed B7-H3 was associated with patient poor prognosis(p=0.039),suggesting B7-H3 could be an independent prognostic factor.Conclusions:Overexpression of B7-H3 in human HNSCC confers poor prognosis of HNSCC patients.Part ⅡBlockade of B7-H3 enhances antitumor immune activity in head neck squamous cell carcinomaObjective:Myeloid-derived suppressor cells(MDSCs)and tumor associated macrophages(TAMs)play key roles in the tumor immune suppressive network and tumor progression.As a negative immune checkpoint molecule,B7-H3 also play privital role in anti-tumor immunity.This chapter aims at detecting B7-H3 expression in immature myeloid-derived immunocytes and its functions in anti-tumor immunity in Tgfbr1/Pten 2cKO HNSCC mouse model.Methods:Detecting expression levels of B7-H3 and immature myeloid-derived immunocytes biomarkers(CDllb,Grl,MDSCs;CD68,CD163,TAM)in HNSCC tissue microarray by immunohistochemistry.Results were quantified and analyzed by Aperio Scanscope.Pearson correlation and Cluster analysis were used to analyze differential expression.Double-labeled immunohistofluorescence were used to detect B7-H3 in immature myeloid-derived immunocytes.Using B7-H3 monoclonal antibody in immunocompetent Tgfbrl/Pten conditional double knockout mice and detecting immunosuppressive cells MDSC,TAM;effector cells CD4,CD8 in transgenic mice by flow cytometry,it is possible to assess the function of B7-H3 in disrupting HNSCC immunosuppressive microenvironment.Results:B7-H3 was highly correlated with immature myeloid-derived markers.Double labeled immunofluorescence has shown B7-H3 positive signals in these immunosuppressive cells in the mouse model,Targeting B7-H3 could effectively lower the number of immunosuppressive cells MDSCs and TAM and improved CD4+、CD8+ T cell effector function in Tgfbr1/Pten 2cKO HNSCC mouse model.Conclusion:B7-H3 blockade reduced MDSCs and TAM and improved T cell effector function in Tgfbr1/Pten 2cKO HNSCC mouse model.