The Mechanisms Of IL-17A On Tumor Infiltration Of Antigen-specific CD8+T Cells

Immunotherapy with immune checkpoint blockades(ICBs)which target the programmed death 1(PD1)or PD 1 ligand(PD-L1)and cytotoxic T-lymphocyteassociated protein 4(CTLA4)have revolutionized the management of multiple cancers.To improve the ICB treatment efficacy,combinations of ICBs with radiotherapy,chemotherapeutic agents,or anti-angiogenesis,or tyrosine kinase inhibitors(TKIs),have been developed.Though patients benefit from some combined immunotherapies,adverse event rates increased.The damage can be caused by immunotoxicity,and also by chemotherapeutic regimens,or irradiation.Gastrointestinal organs are involved commonly,with high incidence of gastrointestinal toxicities.It was demonstrated that the inflammatory cytokine IL-17A participate crucially in the pathogenesis of gut inflammation,colitis and some other diseases involved in the barrier surface tissues.Current evidences indicate that multiple types of cells can generate IL-17A that modulates and amplifies signals locally in a context-dependent manner in the pathogenesis of diseases and promoting the disease progression via IL-17A receptor heterodimers,IL-17RA and IL17RC.IL-17A proves to confer suppression of antitumor immunity and to promote tumor angiogenesis.It is uncertain whether ICB-mediated anticancer efficacy is compromised in the patients experiencing acute damage of mucosal barrier,such as colitis.Tumor antigen-specific CD8+ cytotoxic T lymphocytes(CTLs)are critical in antitumor immunity.To mimic gastroenterological toxicity in humans,we induced acute colitis by providing drinking-water containing 2%of dextran sulfate sodium(DSS).The impact of colitis on CTL-mediated anti-tumor effects was examined in several mouse models.Our results indicated that CTL-mediated anti-tumor effect was dampened when colitis occurred.In colitis mice,the top increased in interstitial fluid of colon tissues was IL-17A and colon tissue showed increased infiltration of IL-17A-producing cells infiltration of lamina propria.Tumor growth was significantly inhibited in the anti-IL-17Atreated colitis B16-OVA-bearing mice,similar to that in the normal-mice,which indicated that impairment of CTL-mediated antitumor effects was related to IL-17A elevation in inflamed intestine.We transferred the activated OT-I cells from CD45.2 mice(donor)into CD45.1 mice(recipient)inoculated with B 16-OVA.The infiltration of donorderived CD8+T cells(CD45.2+)reduced more significantly in the colitis mice.Notably,the percentage and the total numbers of donor-derived Ki67+CD8+T cells reduced more profoundly.However,colitis exhibited minor effects on the recipient-derived CD8+T cells.Administration of αIL-17A to the colitis mice restored the infiltration of donor-derived CD8+T cells.These results indicated that colitis-induced IL-17A elevation mainly repressed the CTLs tumor infiltration to dampen the antitumor immunity,with negligibly effect on the antigen-specific T cell generation.Effective ICB-immunotherapy depends on the intra-tumoral accumulation of CTLs that transmigrate across the vasculature into tumor mass.Within the tumors,the CTLs recognize and destroy the tumor cells directly after the inhibitory signals are relieved.Tumor blood vessels are abnormal,both structurally and functionally relative to those of nonmalignant tissues,restraining the effector T cell infiltration.It is required to find out the factors that control the entrance of anti-tumor effector cells into tumors in a specified microenvironment and certain settings such as occurrence of gastrointestinal toxicities.For control of CD8+T-cell extravasation,tumor blood vasculature and certain chemokines that keep the integrins and adhesion molecules on endothelia cells in high-affinity states and retain the effector T cells play crucial roles.The data showed that the transcription levels of several chemokines and ICAM-1 on CD31+endothelial cells(ECs)in the tumors of colitis mice were significantly lower.The density of CD31+blood vessels was significantly higher,but the density of perivascular CD8+T cells was significantly less in the tumors of colitis mice than in that of normal mice.Now,it is clear that ICB-immunotherapy efficacy relies on the presence of intra-tumoral’stem-like exhausted’ CTLs.To develop precisely combined ICB-immunotherapy,it is required to understand how the tumor vasculature interact with CD8+T cells,particularly the ’stem-like exhausted’ CTLs,to suppress the antitumor immunity in the context of intestinal inflammation-related IL-17A generation.In the tumors of colitis mice,we detected mainly the reduction of ’stem-like’ CTLs in the tumors of colitis mice based on the surface Slamf6+Tim3-expression.In addition,‘stem-like exhausted’CTLs might express more IL-17A receptors(IL-17RA and IL-17RC)to transduce the IL-17A signals,becoming more sensitive to the IL-17A stimulation.By interacting with ICAM-1 on vascular ECs,the expression of the leukocyte function-associated antigen-1(LFA-1)integrin on CD8+T cells is required for their efficient extravasation from blood.The data showed that LFA-1 expression on the ’stem-like exhausted’ CTLs was further reduced compared to that of normal mice.However,we observed no difference of LFA-1 expression levels on the ’stem-like exhausted’ CTLs,indicating that the extravasation of‘stem-like exhausted’CTLs were more affected by the colitis-related IL-17A elevation.In vitro experimental model,IL-17A significantly inhibited proliferation of ’stem-like exhausted’ CTLs.Treated the C166 cells with B16-OVA/CM or treated HUVECs HepG2/CM in the presence of IL-17A,ICAM-1 expressions were significantly reduced.C166-based trans-endothelial migration assay showed that he transmigrated numbers of total CD8+T cells,particular the Slamf6+T cells,significantly reduced when C166 cells were treated with B16-OVA/CM in the presence of IL-17A.Production of nitric oxide(NO)by endothelial cells is crucial for vascular function,and the locally generated NO was reported normalizing vascular EC function that improves the ICB-immunotherapy.Addition of IL-17A into the cultured C166 cells significantly reduced the NO generation.The above results indicated that IL-17A exaggerated tumor vascular endothelium dysfunction to suppress the ’stem-like exhausted’ CTLs extravasation.The above results suggest that blocking IL-17A could improve the treatment efficacy of immune checkpoint inhibitors when colitis occurred.Remarkably,the tumor CD31+vessel density was profoundly lower,and CD8+T cell infiltration was markedly increased in the CT26.CL25-bearing colitis mice that received anti-PD1 plus anti-IL-17A.We then extended our observation to a cohort of advanced HCC patients that were treated with antiPD1 plus anti-VEGF.Median progression-free survival of the patients with IL-17A increase post-therapy,prominently shorter than patients with IL-17A no increase.The overall response rate(ORR)of patients with IL-17A increase was 7%,while 44%of the patients with IL-17A no increase.All the patients with progressive disease(PD)tended IL-17 A elevation,while 8 of 9 patients with partial response(PR)tended IL-17A reduction 24h post-therapy.In conclusion,our current study demonstrated that the inhibition of IL-17A on the extravasation of ’stem-like’ CTLs were related to the reduced interaction mediated by LFA-1 on CTLs and ICAM-1 on tumor vascular endothelium.The antigen-specific CTLmediated potent anti-tumor activity was dampened by abnormal IL-17A generation,but restored after neutralizing IL-17A.We provided an option by neutralizing IL-17A to improve ICB-based therapeutic efficacy when abnormal IL-17A generation in the setting,such as colitis,occurred during cancer treatment.