The Mechanism Of Enhancing Anti-tumor Immune Response By Blocking TIM3 In Head And Neck Squamous Cell Carcinoma(HNSCC)

Background:Head and neck squamous cell carcinoma(HNSCC)is one of the most common malignancy in the world(Siegel et al.,2018).Local invasion and metastasis are the main features of head and neck squamous cell carcinoma.Chronic exposure to tobacco and alcohol and human papillomavirus(HPV)infection was the main risk factors of HNSCC(Argiris et al.,2008).In recent years,the surgical treatment combined with radiation therapy and chemotherapy effectively improved the prognosis of HNSCC patients,but in terms of global 5-year survival rate is still around 50%over the world(Leemans et al.,2010).More and more studies have shown that the disfunction of the immune system plays an important role in the development of tumor.Head and neck squamous cell carcinoma is an immunosuppressive malignancy.In tumor microenvironment,lymphocyte disorder(Baruah et al.,2012;Whiteside,2005),impaired immune effector cells(Bauernhofer et al.,2003;De Costa et al.,2012;Kloss et al.,2015),poor antigen presenting function(Ferris et al.,2006;Lopez-Albaitero et al.,2006),and existence of a large number of immunosuppressive cells,such as myeloid derived suppressor cells(MDSCs),regulatory T cells(Tregs),tumor associated macrophage(TAMs),induced the suppression of killing tumor cells of the immune system(Ferris,2015).Immune suppression and immune escape induced by disfunction of immune cells are closely related to the development of HNSCC(Ferris,2015).With the discovery of tumor immune checkpoints and its role in the regulation of abnormal immune function,immunotherapy targeting immune checkpoint become a research hotspot in recent years.Immunotherapy targeting immune checkpoint PD-1 and CTLA-4 has achieved effective therapeutic effect and improved the prognosis of tumor patients(Ferris et al.,2016;Hodi et al.,2010;Wolchok et al.,2017).T cell immunoglobulin mucin 3(TIM3),a member of TIM family,is a kind of important immune checkpoint(Sharma and Allison,2015).TIM3 is expressed on T-helper 1 cells,CD8+ T cells,dendritic cells,and other lymphocyte subgroups(Anderson et al.,2007;Dovedi et al.,2014;Freeman et al.,2010;Monney et al.,2002).Binding with galectin-9,TIM3 induced Thl cell death,indicating the negative regulation effect of TIM3 in the Thl immune response(Zhu et al.,2005).At the same time,studies reported TIM3 as an important regulator of CD8+ T cell exhaustion in cancer(Fourcade et al.,2010).While blocking TIM3 signal can promote interferon y(IFN-y)mediated antitumor immune response(Ngiow et al.,2011b).In many other tumors,it has been confirmed that TIM3 is related to the development of tumors(Bu et al.,2016;Wang et al.,2017;Wu et al.,2017),and the blockade of TIM3 can enhance the anti-tumor immunity effect(Hervas-Stubbs et al.,2015;Ngiow et al.,2011b).Therefore,targeted TIM3 signaling pathway is considered as a promising immunotherapy method.Objectives:This study was aimed to investigated TIM3 expression in the established human HNSCC micro-tissue array,analysis the correlation of TIM3 expression with clinicopathological parameters and prognosis of HNSCC patients,detect the different TIM3 expression in recurrence of primary HNSCC,HNSCC with preoperative chemotherapy and postoperative radiotherapy.Additionally,we aime to study the role and mechanism of blocking TIM3 signal in relieving immune suppression and enhancing anti-tumor immune response in the immunocompetent transgenic HNSCC mouse model.Methods:By immunohistochemical staining on human HNSCC micro-tissue array TIM3 expression in human HNSCC expression was detect and its correlation with the clinical classification,TNM stages and prognosis was analyzed.Through the pearson correlation analysis,the correlation of TIM3 expression with Galectin-9,CD8,CD33,CD11b,Foxp3 CD68 and CD163 in HNSCC was performed;The therapy effects of blocking TIM3 T cells by using immunocompetent transgenic mouse model.Tumor growth was detected and the mice tissues were collected for flow cytometry detection.The function of block TIM3 signals on Myeloid derived suppressor cells(MDSCs),regulatory T cells(Tregs),tumor associated macrophage(TAMs)and effect T cells,such as CD4+ T cells and CD8+ T cells was dected by flow cytometry analysis.Results:In this study,we reported that TIM3 expression was significantly up-regulated in HNSCC patients and associated with lymph node metastasis.Additionally,TIM3 expression was increased in patients with recurrent HNSCC and patients with pre-radiotherapy or pre-chemotherapy.However,it is not correlated with clincal classification and prognosis of HNSCC patients.We demonstrated the close positive correlation of TIM3 with Galectin-9,CD33,CD11b,Foxp3,CD68 and CD163,and negative correlation of TIM3 with CD8 in human HNSCC.In the transgenic HNSCC mouse model,blockade of TIM3 by the anti-TIM3 monoclonal antibody induced a reduction of CD11b+Gr-1+ MDSCs and CD4+CD25+Foxp3+ Tregs.The increased IFN-y production on CD8+ T cells in the anti-TIM3 treatment mice suggested that the antitumor immune response was enhanced through relieving the immune suppression.Conclusion:The present study demonstrated that TIM3 is up-regulated in HNSCC and its expression is associated with the immunosuppression in HNSCC.Andtargeting TIM3 can enhance anti-tumor immune response by decreasing MDSCs and Tregs in HNSCC.Our findings suggest that anti-TIM3 immunotherapy maybe a novel therapeutic approach for effective treatment of HNSCC.