The Effect Of Proliferation Inhibition Of JQ1 On Ph Positive Acute Lymphoblastic Leukemia Cell SUP-B15 And Its Mechanism

Objective:Ph Positive acute lym Phoblastic leukemia(Ph+ALL) is one of the subtypes of acute lymphoblastic leukemia whose prognosis is poor, chemotherapy and targeted therapy drugs can control the disease but as a result of drug-resistant problem most relapse, looking for a better treatment is necessary. Recent years, foreign study found JQ1(bromine area inhibitors) inhibits the activity of chronic myelogenous leukemia(CML) cell with BCR-ABLT315 I mutation, increase the rate of its apoptosis, and inhibit Brd4( bromine structure domain 4), reduce the myc m RNA level and the combination activity of the myc,overcome the CML resistance;and Ph+ALL has similar gene mutation to CML BCR- ABL,then what on earth Brd4 inhibitors JQ1 for Ph+ALL has the same effect, the study rare reported at home and abroad.This study intends to adopt Ph+ALL cell lines(SUP- B15)with JQ1, through observing cell morphology under light microscopy, determined by MTT method to detect JQ1 role in SUP- B15 cell lines with half inhibitory concentration, flow cytometry instrument to detect cell apoptosis rate,By fluorescence quantitative PCR to detect BCR- ABLm RNA, Brd4 m RNA and its downstream two channels,mycm RNA and P53 m RNA gene’s expression.The purpose is to explore the possible mechanism of JQ1 on Ph+ALL cell lines, providing theoretical basis for clinical application.Methods:1.The cell Proliferation level of SUP-B15 cells were detected by MTT assay;2.The cell apoptosis rate of SUP-B15 cells were determined by flow cytometry(FCM);3.The expressions of BCR-ABLm RNA,Brd4 m RNA,mycm RNA,P53 m RNA were measured by real time-PCR(RT-PCR).Results:1.The results of MTT assay displayed that : when cultured lasted 24 h,48h,72 h,compared with control group, the different concentration of JQ1 could significantly inhibit the proliferation of SUP-B15,cell inhibition rate rises with increased concentration and time expansion,in a time-dose dependent manner. The 48 h half inhibitory concentration(IC50) was 2.576umol/L.2. The results of FCM assay:When1 umol/L, 4 umol/L of JQ1 applied to SUP- B15 cells,after being treated for 48 h,72h, and experiment repeated three times,when cultured lasted 48 h,the average apoptosis rate of control group(3.57 + 0.95) %, 1 umol/L group(19.90 + 1.40)%, 4 umol/L group(35.77 + 1.72)%, the average apoptosis rate of 72 h,respectively, the control group(17.77 + 1.75) %, 1 umol/L group(36.40 + 2.29) %, 4umol/L group(78.83 + 1.40) %, and two experimental apoptosis rate is significantly higher than the control group, difference was statistically significant(F48 = 925.275, F72 =925.275, P < 0.05 and < 0.01).3. Theresults of RT-PCRdisplayed that: when SUP-B15 cells cocultured respectively by1 umol/L and 4 umol/L of JQ1 after 48 h, BCR-ABLm RNA, Brd4 m RNA,mycm RNA expression quantity significantly decreased than the control group,respectively fell by65.67%,8.67%,38.67%;90.00%,67.00%,90.67%;while P53 m RNA expression gradually increased, increased 206.33% and 206.33% respectively, The experimental group compared with control group, the difference had statistical significance(P < 0.01).Conclusions:1.JQ1 can significantly inhibit SUP- B15 cell Proliferation, induce the apoptosis of SUP- B15 cells effectively;2. JQ1 downregulate the expression of Brd4 m RNA then affect the expression of itsdownstream gene,myc m RNA and P53 m RNA, indicate that when JQ1 is applied to SUP-B15 cell,it can obviously affect the expression of the above three genes,proving that Brd4→myc→P53 may be one of the pathways of JQ1 induced SUP- B15 cells apoptosis;3. JQ1 can effectively reduce the transcription level of BCR- ABL m RNA in SUP-B15 cell lines, indicate the drug can affect the expression of BCR- ABL m RNA significantly and it may be another way of apoptosis.