Studies On Effects Of 8-CPT-cAMP And Bortezomib On Biological Behavior Of Lymphoma Cells

As an important cellular signal transducer and major protein degradation regulator,cyclic adenosine monophosphate(cAMP)and proteasome is closely associated with the proliferation and apoptosis of lymphoma cells.This thesis included two parts that aim to study the role of cAMP analogue,8-(4-Chlorophenylthio)adenosine 3’:5’-cyclic monophosphate(8-CPT-cAMP)combined with bortezomib as one of proteasome inhibitors on two different types of lymphoma cells and further explore its mechanisms.Part I was mainly to study the role of 8-CPT-cAMP combined with bortezomib on biological behavior of diffuse large B cell lymphoma(DLBCL)cells.DLBCL cell lines including OCI-Lyl and U2932 cells were used as in vitro model to detect effects of 8-CPT-cAMP on these cells in combination with bortezomib.The proliferation of OCI-Lyl and U2932 cells were evaluated through Cell Counting Kit(CCK-8)assay.Meanwhile,flow cytometry,western blot and small molecule RNA interference assay was applied to explore possible synergistic manner and key interactive module of the two agents.It was shown that 8-CPT-cAMP and bortezomib synergistically enhanced the cell growth inhibition and apoptosis of OCI-Lyl and U2932 cells.However,the same combination exerted little influence on the growth of peripheral blood mononuclear cells isolated from normal individuals.It was also shown that 8-CPT-cAMP synergized with bortezomib to trigger mitochondrial transmembrane potential collapse and induce degradation of apoptosis effectors Caspase-9 and Caspase-3.In addition,intracellular ROS concentrations were also increased.Meanwhile,8-CPT-cAMP could synergize with bortezomib to down-regulate expression of c-myc and p-AKT in these cells.Part II was mainly to study effects of cyclic adenosine monophosphate analogue combined with bortezomib on biological behavior of Burkitt’s lymphoma(BL)cells.Based on our previous experiments,BL cell lines Daudi cells were used as in vitro model to explore possible biological effects of 8-CPT-cAMP combined with bortezomib on BL.It was shown that 8-CPT-cAMP combined with bortezomib could induce cell growth inhibition and apoptosis in Daudi cells.8-CPT-cAMP could synergize with bortezomib to instantly induce G2/M transition arrest in Daudi cells.Meanwhile,8-CPT-cAMP synergized with bortezomib to induce apoptosis in Daudi cells through mitochondrial pathway.Furthermore,it was also shown that 8-CPT-cAMP could significantly inhibit expression of c-Myc in Daudi cells at mRNA and protein levels and bortezomib enhanced these effects.Interestingly,8-CPT-cAMP combined with bortezomib also restrained expression of STAT3,one of major regulators for c-Myc.In summary,8-CPT-cAMP could synergize with bortezomib to induce proliferation inhibition and apoptosis in DLBCL and BL cells,which might be mediated by mitochondrial transmembrane potential collapse,cell cycle G2/M transition arrest,as well as simultaneous suppression of c-Myc and AKT signaling pathways.These results presented in this work may provide experimental and theoretical basis for novel therapeutic regimen based on cAMP-modulating agents combined with bortezomib for DLBCL and BL.