The Impact Of The Decline Of EZH2 On The Proliferation Of T-cell Lymphoma And The Sensibility To The Chemotherapy

Objective: T cell lymphoma(TCL), is a malignant T cell clone hyperplastic diseases, accounting for 12%-15% of all lymphoma. With complex pathological patterns, TCL is strongly aggressive, having distinct biologic behavior and clinical characteristics. The other important characteristic is that TCL has obvious racial and area differentiation, with high incidence in HTLV-1 epidemic Asian countries. However, TCL has very poor curative effect to clinical treatment. EZH2 is an important histone methyltransferase, depending on deeply conservative SET domain to methylate H3K27, methylated H3K27 can recruit PRC1 complex to the specific gene site, silenting genes involving cell differentiation and suppressing proliferation, leading to the tumorigenesis. Immunohistochemical staining shows TCL tissues have high level of EZH2. So our study is to observe the impact of EZH2 on cell proliferation and apotosis,by using lentivirus-based EZH2 RNA interference and EZH2 inhibitor UNC1999 to dispose hut78 cells, meanwhile to assess synergistic effect with chemotherapy, aim to provide theoretical basis to bring EZH2 inhibitor to clinical treatment. We also explore the regulation of EZH2 on cell apotosis、drug resistance and adhesive metastasis.Methods: 1.Used lentivirus-delivered EZH2-sh RNA to infect hut78 cells to develop monoclonal cells with low level of EZH2, on the basis of verification by RT-q PCR and Western-blot, to observe the influence on proliferation and apotosis of hut78 cells by MTS、Brdu tests. 2.Used multiple common chemotherapy drugs to dispose hut78、hut78 control and hut78 EZH2-sh RNA, to observe the influence of drugs on cells proliferation by MTS tests. 3.Used EZH2 small-molecule inhibitor UNC1999 to dispose hut78, to observe the influence of inhibitor on cells proliferation by MTS tests. Meanwhile combined chemotherapy drugs to dispose hut78 cells, observed sensibility to the chemotherapy.4.Explored the regulation of EZH2 on cell apotosis、drug resistance and adhesive metastasis molecules by RT-q PCR or Western-blot.Results: 1.After using EZH2-sh RNA to decline the level of EZH2 in hut78 cells, MTS tests show: the proliferation of Hut78 EZH2-sh RNA was obviously reduced., with the inhibition rate of 46.8%. 2.Used various chemotherapy drugs to dispose hut78、hut78 EZH2-sh RNA,the latter cells’ inhibition rate of proliferation was obviously increased, with low level of IC50 value of drugs. There is statistic difference between two cells(P<0.05). 3.Used EZH2 inhibitor to dispose hut78 cells, the proliferation of Hut78 was significantly reduced, having the connection with deposing time and drug dose. 4.Used EZH2 inhibitor and chemotherapy drugs to dispose hut78 cells simultaneously, Q value analysis showed EZH2 can increase the sensibility to the chemotherapy(q value>1.15). 5.Tested the level of main molecules about cell apotosis(P53、Bcl-2)、drug resistance(LRP、MDR-1、MRP-1) and adhesive metastasis(VCAM、ICAM)in hut78 、hut78 EZH2-sh RNA 、 hut78 disposed by EZH2 inhibitor, RQ value(relative quantity) has significant difference(P<0.05).Conclusion: 1.Used lentivirus-delivered EZH2-sh RNA to infect hut78 cells or EZH2 inhibitor to dispose hut78 cells, can decrease the expression of EZH2. 2.Low level of EZH2 can obviously inhibit the proliferation. 3.The decline of EZH2 can increase sensibility of hut78 to the chemotherapy, importantly, EZH2 inhibitor has synergistic effect with chemotherapy drug to hut78 cells. 4.The decline of EZH2 can induce abnormal expression of many apotosis、drug resistance and adhesive metastasis molecules, suggesting EZH2 has extensive regulation in tumorigenesis、drug resistance and metastasis.