The Mechanisms Of T Peptide Inhibiting Tumors Growth In Vivo And In Vitro

Tumor cells survived in the body must be able to escape immune surveillance andkilling by immune cells. When the tumor cells grow to a certain size in local place, tumormicroenvironment is formed. Tumor microenvironment support tumor for proliferation andmetastasis and immune cells out of the tumor microenvironment can't break through thisbarrier to attack cancer cells. So biological response modifiers treatment with patients cannot make effort expectably under this circumstances.Surgery is helpful to reduce tumorburden after resecting most of tumor tissue. On the other hand, tumor microenvironment isdamage quickly by surgery and internal immunosuppression cells and tumor cellsexposured so that the outside activated immune cells have change to identificate and killtumour cells. Based on this principle and combined with clinical cancer therapy principle,we successfully established a novel murine pharmacodynamics model with a postsurgeryresidual tumor. And in this model we find T peptide as a potential immunopotentiator.T peptide is a new derivative of Tuftsin. It not only retains phagocytosis andanti-tumor biological activities, but also greatly improves the enzyme-resistant ability toextend its half-life in vivo. We adopted three tumor model evaluation anti-cancer activity ofT peptide. Compaired with the traditional xenograft tumor mouse model, TP displayedbetter effects on murine model with a pseudosurgery and the inhibition rate reached above80%.T peptide almost no observed toxicity to mice during experiments. And the mice weresignificantly longer life than cyclophosphamide group mice. T peptide also had highinhibition rate of anti-tumor effects on nu/nu and SCID mice operational tumor model. Onmice transfer model, T peptide delayed invasion and transformation by preventingmalignant cells spread to the blood circulation.Combined with the structure characteristics of peptide T drugs and cell biologyfunction, RT-PCR, realtime-PCR, immunohistochemical, ELISA and western blot assayswas tested to study the mechanism of anti-cancer action of TP. Macrophages are one ofspecific target cells for TP. In vitro, T peptide can improve macrophage proliferation, butthis effect was slight and no dose-response relationship. Through detection cell cycle, itconfirmed that T peptide can promote macrophage cell to proliferation by adjusting the G1phase to S phase process. The cell cycle in the regulation of G1/S point is internal andexternal signal transmission through cell, integrate together to the nucleus, so theproliferation of cells can be controlled in the key point. The cell cycle regulation proteinP21WAF1is a negative adjustment factor in G1/S phase. The results showed that T peptidemarkedly decreased P21WAF1expression with CDK2phosphorylation level increased. Tpeptide impacted macrophage proliferation via ERK/JNK/MAPK signal transductionpathways.On the other hand, T peptide activated macrophage through NF-κB signaling pathwaysin a short time. Macrophage phagocytosis was activated and its cytotoxic function obviously enhanced after TP treated. Activated macrophage by TP had been polarized toM1phenotype. M1macrophage secretes Th1proinflammatory cytokines and chemokinessuch as TNF-α,IL-12, NO, IFN α/β,CXCL9and CXCL11. In vitro, T peptide can alsotrigger M2macrophages polarized by IL-4to release Th1cytokines expression of cytokines.Marker gene of M2macrophages like Arg-1was obviously down-regulated and theexpression of M1macrophages marker gene rised, such as iNOS and IL-12.The experimentresults suggested that T peptide could be reverse phenotype of M2macrophage. Tumorassociated macrophage in tumor microenvironment appears M2phenotype. In TAM frompostoperative residual tumor tissue of T peptide group and Tuftsin group, the transcriptionsof Th1and Th2cytikines were decreased except in IL-1β. T peptide activated macrophagethrough NF-κB signaling pathways. NF-κB signaling pathways can not work in TAM resultthat TP could not swith TAM phenotype from M1to M2.The suppression of Th2cytokinesalso prompted that T peptide influenced TAM through other pathways.It is reported in2006that NRP-1is a receptor of Tuftsin. Tuftsin can block VEGFR2phosphorylation by competition with VEGF165via NRP-1and inhibited tumorangiogenesis. As a new Tuftsin polymerization analogues, T peptide inhibited VEGF andMMP-9in tumor and M2macopahe. To NRP-1receptor, T peptide significantly improvedmacrophages to express NRP-1.But the expression of NRP-1in HUVEC had not changed.This activity of T peptide may be involved in macrophage proliferation and activation Tpeptideas as well as anticancer activity. T peptides block PD-1/PD-L1signaling pathwaysin tumor and macrophage.4-1BB, PD-L2and OX40molecular expression were higher thancontrol and PD-L1as well as PD-1protein decreased. The results confirmed that T peptidemodulated immunity of macrophage and T cells.In conclusion, we successfully established a novel murine model with a postsurgeryresidual tumors for anticancer pharmacodynamics evaluation. And we researched theanticancer effects of TP on this novel murine model in vivo and in vitro. On the basis of thenew model, we studied the mechanisms of TP on macrophages, TAMs.The results and theknown low toxicity are clues that TP might be a promising candidate drug for the treatmentof clinical patients with postsurgery residual tumors or micrometastasis.In short, T peptide induced macrophage to M1phenotype polarization. Activemacrophage by TP released Th1cytokines, strengthen antibody-dependent cell-mediatedcytotoxicity function. IL-12and IFN-γ can also activate T lymphocytes and NK cells. Theall results are clues that TP might be a promising candidate drug for the treatment ofclinical patients with postsurgery residual tumors or micrometastasis.