BAG3 Is Involved In The Chemotherapy Resistance Of Diffuse Large B-cell Lymphoma To Proteasome Inhibitors

Part ?BAG3 is involved in the chemotherapy resistance of Diffuse Large B-Cell Lymphoma to proteasome inhibitors[Background]Diffuse large B-cell lymphoma is the most common clinical subtype of non-Hodgkin's lymphoma,accounting for about 30%of patients with lymphoma.At present,due to the emergence of rituximab,the treatment of diffuse large B-cell lymphoma has been greatly improved,but there are still 30%-40%of patients with diffuse large B-cell lymphoma recurrence or resistance to chemotherapy in short-term.In order to improve such situation,clinical investigators are working hard to study how to increase the response rate of patients with diffuse large B-cell lymphoma,to reduce their recurrence rate and to avoid resistance.In recent years,many reports have indicated that bortezomib was effective in the treatment of the diffuse large B-cell lymphoma.Bortezomib is a proteasome inhibitor that was initially approved by the US FDA for the treatment of multiple myeloma.Bortezomib could induce apoptosis by inhibiting proteasome inhibitors.Although many studies have shown that bortezomib may have a significant effect on the treatment of patients with the diffuse large B lymphoma,some clinical trials have shown that bortezomib-containing regimens are not effective in improving the treatment of the diffuse large B-cell lymphomas,which may exist some resistance mechanisms that awaiting to be investigated.In the treatment of the diffuse large B-cell lymphoma,in addition to bortezomib,there are many newly founded targeted drugs that are in the study by clinical researchers.Among them,the PI3K/AKT pathway inhibitor is one of the most studied drugs.It has been shown that activation of the PI3K/AKT pathway is a poor prognostic factor for diffuse large B-cell lymphoma.Many studies have shown that the PI3K/AKT pathway inhibitors could enhance the antitumor effect of proteasome inhibitors in many tumors.Therefore,combined treatment with the proteasome inhibitor and the PI3K/AKT pathway inhibitor may become a potential treatment in the treatment of the diffuse large B-cell lymphoma.In order to study the internal mechanisms,we read a large number of literatures and we note that the proteasome inhibitors can induce the expression of an anti-apoptotic protein BAG3(Bcl-2 associated athanogene 3).The BAG3 mediated the resistance of tumor cells to proteasome inhibitors.We hypothesized that proteasome inhibitors can induce the expression of anti-apoptotic protein BAG3 in the diffuse large B-cell lymphoma cell lines,the protease inhibitors and the PI3K/AKT pathway inhibitors have synergistic effects in the diffuse large B-cell lymphoma cell lines,and it is associated with the expression level of BAG3.We designed this experimental study to further confirm our hypothesises.[Objectives]Our study was aimed to detect the expression levle of BAG3 in Lyl and Ly8 cells,the effect of the proteasome inhibitor and the PI3K/AKT inhibitor on the biology activity of Lyl and Ly8 cells and the association with the expression level of BAG3.[Methods]1.Detection of BAG3 mRNA in Lyl and Ly8 cells by PCR.2.The cells were treated with different concentrations of proteasome inhibitors,and then the expression level of BAG3 protein was detected by Western blot.3.The cells were treated with different concentrations of proteasome inhibitors and then the proliferation of cells was detected by CCK8 method.4.The cells were cotreated with proteasome inhibitors and the PI3K/AKT pathway inhibitor,and then the expression of BAG3 was detected by qPCR and Western blot respectively.5.The cells were cotreated with proteasome inhibitors and the PI3K/AKT pathway inhibitor,then the cell proliferation was detected by CCK8 method.6.The cells were cotreated with proteasome inhibitors and the PI3K/AKT pathway inhibitor,and then the cell apoptosis was detected by flow cytometry.[Results]1.The results of RT-PCR showed that the BAG3 mRNA was detected in the cells of diffuse large B cell lymphoma cell lines Lyl and Ly8.2.Proteasome inhibitors could induce the expression of BAG3 in Lyl and Ly8 cells.Western blot showed that proteasome inhibitor could induce the expression of BAG3 in the cells,and its expression level was positively correlated with the concentration of proteasome inhibitor.3.Proteasome inhibitors and the PI3K/AKT pathway inhibitor could inhibit the proliferation of Lyl and Ly8 cells.The effect of cotreatment with proteasome inhibitor and PI3K/AKT pathway inhibitor on the cell proliferation was significantly higher than that of single drug treatment group.4.Western blot analysis showed that the expression level of BAG3 in the cells in the cotreatment group was significantly lower than that in single proteasome inhibitor treatment group.5.The results of flow cytometry showed that the apoptotic level of cells in the cotreatment group was higher than that of single drug treatment group.Each experiments were repeated for 3 times,the results were statistically significant(P<0.05).[Conclusions]Proteasome inhibitors can induce the expression of BAG3 in diffuse large B-cell lymphoma cell lines Lyl and Ly8.Inhibition of the PI3K/AKT signaling pathway can sensitize diffuse large B cell lymphoma cells to treatment with proteasome Inhibitor by suppression of BAG3.Part ?Weekly versus biweekly bortezomib given in patients with indolent non-Hodgkin lymphoma:A Meta-Analysis[Background]The indolent lymphoma is a group of relatively slow progress of lymphoma,usually including follicular lymphoma,marginal zone lymphoma and small lymphocytic lymphoma.Although the clinical process is slow,but they usually prone to recurrence.In recent years,the emergence of the CD20 monoclonal antibody rituximab improved the prognosis of indolent lymphoma to a large extent,but recurrence and resistance often occur,in order to improve the treatment of the indolent lymphoma,there are many new targeted drugs in study.Bortezomib is receiving increasing attention in the treatment of lymphoma,it is a proteasome inhibitor that has been approved by the US FDA for the treatment of multiple myeloma and recurrent/refractory mantle cell lymphoma,it has been indicated that bortezomib combined with rituximab,in combination with bendamustine,or in combination with other chemotherapeutic agents were effective in the treatment of recurrent and refractory indolent lymphoma.However,there is no uniform guideline for the treatment of bortezomib in indolent lymphoma,many institutions conducted the randomized controlled trials in order to compare the efficacy and safety between once a week and twice a week of the administration schedule of the bortezomib in the treatment of the indolent lymphoma,the results were not consistent.We conducted this meta-analysis to find a better schedule of the bortezomib administration.[Objectives]Though this meta-analysis,we compared efficacy and safety between different administration schedules of bortezomib in the treatment of the indolent lymphoma.[Methods]1.We conducted a comprehensive search of MEDLINE,EMBASE and Cochrane Library database,the keywords included "Follicular lymphoma","Marginal zone lymphoma","Small lymphocytic lymphoma" and "Bortezomib".The literature was written in English,the deadline was July 29,2016.2.Screening the literature.The inclusion criteria were:(1)studies evaluated the efficacy and adverse effects of bortezomib alone or in combination with other drugs in the treatment of indolent lymphoma.(2)randomized,controlled trials,retrospective or prospective cohort studies.(3)The data in each study were separately reported and could be extracted.(4)If more than one article reported the same clinical trial,we selected the most detailed or the most recent literature.(5)The literatures were in English.The exclusion criteria were:(1)the study did not assess the effectiveness and safety of bortezomib in different schedules.(2)There was no report of the original data in the study.(3)case reports,editorials,letters.3.Conduct quality assessment of the literature.The quality of the randomized controlled trials was assessed on the basis of the Jadad scale.For non randomized controlled trials,we used the Newcastle-Ottawa Quality Assessment Scale.4.Extract data.The following information was extracted from the literature:first author's name,publication year,study area,age of patients,sex,study design,study arms,number of total cases and number of patients having response in each arms,number of patients with neuropathy,fatigue,diarrhea,nausea and neutropenia in each arms,number of patients with grade 3 or higher neuropathy,fatigue,diarrhea,nausea and neutropenia in each arms.5.Using Stata 11 software for heterogeneity analysis,drawing the forest map,sensitivity analysis and publication bias analysis.[Results]1.Finally,six studies were included in the study,the average age of patients ranging from 63 to 65 years old.Five of the included studies were phase II clinical trials,and one was phase I clinical trial.Five trials were carried out in the United States,one was practiced in France.The dosing,number of cycles,and synergistic chemotheraphy regimens varied across studies.Four trials reported therapy with bortezomib alone,one trial reported therapy with bortezomib plus rituximab,and one trial reported therapy with bortezomib plus rituximab,cyclophosphamide and prednisone.2.Meta analysis results(1)Response to treatment(6 trials,280 patients)The results showed that there was not statistical heterogeneity(P = 0.291;12 =19.4%).Further analysis using a fixed effect model showed that the efficacy of patients treated with bortezomib twice a week was better than that of patients receiving a weekly schedule,the result was statistically significant(P<0.001)(OR 1.691;95%CI,1.0-2.80).(2)Adverse eventsWe analysed the common adverse events reported in the trials,the main adverse events included neuropathy,fatigue,diarrhea,nausea and neutropenia.The result indicated that there were no differences between the two schedules of bortezomib for the adverse events and no differences were observed between arms for grade 3 or higher toxicities.[Conclusions]This meta-analysis showed that the biweekly schedule of bortezomib was more effective than the weekly schedule in indolent lymphoma.The occurrence of adverse effects was similar.