Anti-tumor Action and Clinical Application of Proteasome Inhibitor

Ubiquitin-proteasome pathway mediates the degradation of cell protein, and cell cycle, gene translation and expression, antigen presentation and inflammatory development. Proteasome inhibitor can inhibit growth and proliferation of tumor cell, induce apoptosis and reverse multipledrug resistance of tumor cell, increase the sensitivity of other chemotherapeutic drugs and radiotherapy, and is a novel class of potent anti-tumor agents.

The Apoptosis of Bovine Lens Epithelial Cells Induced by Proteasome Inhibitor MG132

To investigate the effect of proteasome inhibitor MG132 on the apoptosis of bovine lens epithelial cells （BLECs）, the cells were treated with MG132 at different concentrations for12, 24 and 36 h. The cell viability was analyzed by MTT assay and the effect of MG132 on the apoptosis of BLECs was assessed by flow cytometry （FCM）. The results showed that after treatment for the same period, the inhibitory effect of MG132 on BLECs proliferation was enhanced with the increment of the concentration of MG132 （0, 2, 5, 10, μmol/L） （P〈0.05）. The 50% inhibiting concentration （IC50） was 2.03 μmol/L when the BLECs were treated with MG132 for 36 h. MG132 also induced the apop- tosis of BLECs obviously. FCM showed that the apoptosis index of the cells treated by MG132 at 2 μmol/L for 12 h was （20.24±1.51）%, and that of the control was （0.98±0.20）% respectively （P〈0.01, n=3）. It was concluded that MG132 could lead to apoptosis of BLECs. The decrease of proteasome activity may play an important role in the formation and development of cataract.

Prevalence and Clinical Implication of t(11;14)in Light-Chain Amyloidosis with or without Coexistent Multiple Myeloma in the Era of Proteasome Inhibitors

Background Light-chain amyloidosis(AL)and multiple myeloma(MM)may coexist in some patients and,although they share some cytogenetic abnormalities,they usually present with different clinical phenotypes.Translocation(11;14)is the most common cytogenetic abnormality in AL,but the prevalence and clinical implication of t(11;14)in patients with AL,with or without coexistent MM,remains unclear.Methods A total of 119 consecutive newly diagnosed AL patients with available fluorescence in situ hybridization(FISH)data were retrospectively included and classified as primary AL alone(pAL-alone)or AL with coexistent MM(AL-MM).Clinical characteristics,FISH profiles,and hematologic and survival outcomes were analyzed.Results There were 53 patients in the pAL-alone group and 66 in the AL-MM group.The prevalence of t(11;14)was significantly higher in the pAL-alone group than the AL-MM group(49.1%vs.26.2%,P=0.012).A significantly higher proportion of the pAL-alone group achieved hematologic response compared with the AL-MM group(60.4%vs.39.4%,P=0.023).Patients with AL-MM experienced significantly shorter hematologic event-free survival(hemEFS)than those with pAL-alone(median,4.8 months vs.44.3 months,P<0.001),as well as significantly shorter overall survival(OS;median,15.2 months vs.not reached,P<0.001).When stratified by the presence or absence of coexistent MM and t(11;14),AL-MM patients with t(11;14)had the worst hemEFS(median,3.8 months,P<0.001)and OS(median,5.4 months,P=0.001).Conclusions Patients with pAL-alone had a higher prevalence of t(11;14)than those with AL-MM.The AL-MM group had poorer outcomes,despite the availability of proteasome inhibitor treatment,with AL-MM patients with t(11;14)showing the worst outcomes.Better diagnostic and treatment approaches are warranted for this population.

The role of proteasomes in tumorigenesis

Protein homeostasis is the basis of normal life activities,and the proteasomefamily plays an extremely important function in this process.The proteasome 2os is a concentric circle structure with twoαrings and twoβrings overlapped.The proteasome 2os can perform both ATP-dependent and non-ATP-dependent ubiquitination proteasome degradation by binding to various subunits(such as 19S,11S,and 200 PA),which is performed by its active subunitβ1,β2,andβ5.The proteasome can degrade misfolded,excess proteins tomaintain homeostasis.At the same time,it can be utilized by tumors to degrade over-proliferate and unwanted proteins to support their growth.Proteasomes can affect the development of tumors from several aspects including tumor signaling pathways such as NF-kB and p53,cell cycle,immune regulation,and drug resistance.Proteasome-encoding genes have been found to be overexpressed in a variety of tumors,providing a potential novel target for cancer therapy.In addition,proteasome inhibitors such as bortezomib,carfilzomib,and ixazomib have been put into clinical application as the first-line treatment of multiple myeloma.More and more studies have shown that it also has different therapeutic effects in other tumors such as hepatocellular carcinoma,non-small cell lung cancer,glioblastoma,and neuroblastoma.However,proteasome inhibitors are not much effective due to their tolerance and singleness in other tumors.Therefore,further studies on their mechanisms of action and drug interactions are needed to investigate their therapeutic potential.

MG132 Induced Apoptosis Pathway in HL-60 Cells and Impact of Allogeneic Mixed Lymphocyte Reaction

Objective： To investigate the proteasome inhibitor MG132-induced apoptosis pathway in HL-60 cells and the role of allogeneic mixed lymphocyte reaction. Methods： Cell apoptosis was analyzed by flow cytometry. The expressions of p21 protein, p27 protein and p53 protein in HL-60 ceils treated with MG132 were measured by Western blot. The proliferation of, peripheral blood mononuclear cells （PBMNCs） after treatment with 75 Gy irradiated HL-60 cells treated with MG132 was measured with CCK-8. Results： High-dose MG132 induced apoptosis in HL-60 cells. No significant change was observed in MG132-induced apoptosis after inhibiting caspase-8 and caspase-9 pathway. The expressions of p21 protein and p27 protein increased in MG132-induced apoptosis. HL-60 cells treated with low-dose MG132 improved the proliferation of PBMNCs from healthy volunteers. Conclusion： High-dose MG132 induced apoptosis and directly killed HL-60 ceils. MG132 induced apoptosis in a caspase-8- and caspase-9-independent pathway, p21 protein and p27 protein were involved in MG132-induced apoptosis in HL-60 cells. HL-60 cells treated with Low-dose MG132 improved the effect of promoting the proliferation of PBMNCs from healthy volunteers.

The dichotomous role of immunoproteasome in cancer:Friend or foe?

Immunoproteasome is a variant of proteasome with structural differences in 20S subunits optimizing them for the production of antigenic peptides with higher binding affinity to major histocompatibility complex(MHC)-I molecules.Apart from this primary function in antigen presentation,immunoproteasome is also responsible for the degradation of proteins,both unfolded proteins for the maintenance of protein homeostasis and tumor suppressor proteins contributing to tumor progression.The altered expression of immunoproteasome is frequently observed in cancers;however,its expression levels and effects vary among different cancer types exhibiting antagonistic roles in tumor development.This review focuses on the dichotomous role of immunoproteasome in different cancer types,as well as summarizes the current progression in immunoproteasome activators and inhibitors.Specifically targeting immunoproteasome may be a beneficial therapeutic intervention in cancer treatment and understanding the role of immunoproteasome in cancers will provide a significant therapeutic insight for the prevention and treatment of cancers.

First line vs delayed transplantation in myeloma:Certainties and controversies

Since the middle of 1990 s autologous stem cell trans-plantation has been the cornerstone for the treatment of young patients with multiple myeloma(MM). In the last decade the introduction of novel agents such as immunomodulatory drugs(IMi Ds) and proteasome inhibitors(PI), has dramatically changed the therapeutic scenario of this yet incurable disease. Due to the impressive results achieved with IMi Ds and PI both in terms of response rates and in terms of progression free and overall survival, and to the toxicity linked to high dose therapy and autologous stem cell transplantation(ASCT), a burning question nowadays is whether all young patients should be offered autotransplanta-tion up front or if this should be reserved for the time of relapse. This article provides a review of the data available regarding ASCT in MM and of the current opinion of the scientific community regarding its optimal timing.

MG132 Inhibits Myocardial Ischemia-reperfusion Injury by Regulating Apoptotic Pathway

Objectives To administrated proteasome inhibitor-MG-132 prior to reperfusion in rat myocardial ischemia-reperfusion model to determine whether MG-132 could reduce myocytic apoptosis. Methods and results MG-132 （0. 75 mg/kg in 2 ml DMSO） injection 5 min prior to reperfusion resulted significant reduction of myocardial reperfusion injury. This effect was accompanied by reduced polymorphonuclear neutrophils （PMN） infiltration in myocardial region surrounding the myocardial infarct, reduced apoptosis in cardiac myocytes, reduced NF-κB activation, as determined by electron microscopy, histology, immunohistochemistry, the terminal deoxynucleotidyl transferase-mediated nick endlabeling （TUNEL） method, reverse transcription-polymerase chain reaction. Functional effects of MG-132 on PMN accumulation, activation of nuclear factor kappa B （p65 mRNA and protein levels ）, and apoptosis were characterized in rat myocardial tissue. MG132 time-dependently inhibited myocardial p65 mRNA expression and reduced myocardial apoptotic index （AI） after reperfusion for 2 h, 6 h and 24 h （ P 〈 0. 01 ）. Moreover, MG-132 time-dependently decreased Bax protein levels, while increased Bcl-2 protein levels in ischemic and reperfused myocardium （ P 〈 0. 05 ）, its effect peaked after reperfusion for 24 h. Conclusions Our results demonstrate that MG-132 reduced myocardial reperfusion injury by inhibiting myosytic apoptotic cell death and blocking activation of NF-κB, down-regulating Bax expression and up-regulating Bcl-2 expression as well as el evating Bcl-2/Bax ratio.

How does conserved dopamine neurotrophic factor protect against and rescue neurodegeneration of PC12 cells？

Conserved dopamine neurotrophic factor protects and rescues dopaminergic neurodegeneration induced by 6-hydroxydopamine in vivo,but its potential value in treating Parkinson＇s disease remains controversial.Here,we used the proteasome inhibitors lactacystin and MG132 to induce neurodegeneration of PC12 cells.Afterwards,conserved dopamine neurotrophic factor was administrated as a therapeutic factor,both pretreatment and posttreatment.Our results showed that（1）conserved dopamine neurotrophic factor enhanced lactacystin/MG132-induced cell viability and morphology,and attenuated alpha-synuclein accumulation in differentiated PC12 cells.（2）Enzyme linked immunosorbent assay showed up-regulated 26S proteasomal activity in MG132-induced PC12 cells after pre-and posttreatment with conserved dopamine neurotrophic factor.Similarly,26S proteasome activity was upregulated in lactacystin-induced PC12 cells pretreated with conserved dopamine neurotrophic factor.（3）With regard proteolytic enzymes（specifically,glutamyl peptide hydrolase,chymotrypsin,and trypsin）,glutamyl peptide hydrolase activity was up-regulated in lactacystin/MG132-administered PC12 cells after pre-and posttreatment with conserved dopamine neurotrophic factor.However,upregulation of chymotrypsin activity was only observed in MG132-administered PC12 cells pretreated with conserved dopamine neurotrophic factor.There was no change in trypsin expression.We conclude that conserved dopamine neurotrophic factor develops its neurotrophic effects by modulating proteasomal activities,and thereby protects and rescues PC12 cells against neurodegeneration.

Forkhead box M1 transcription factor:a novel target for pulmonary arterial hypertension therapy

Background Forkhead box M1(FoxM1),a member of forkhead family,plays a key role in carcinogenesis,progression,invasion,metastasis and drug resistance.Based on the similarities between cancer and pulmonary arterial hypertension,studies on the roles and mechanisms of FoxM1 in pulmonary arterial hypertension have been increasing.This article aims to review recent advances in the mechanisms of signal transduction associated with FoxM1 in pulmonary arterial hypertension.Data sources Articles were retrieved from PubMed and MEDLINE published after 1990,including-but not limited to FoxM1 and pulmonary arterial hypertension.Results FoxM 1 is overexpressed in pulmonary artery smooth muscle cells in both pulmonary arterial hypertension patients and animal models,and promotes pulmonary artery smooth muscle cell proliferation and inhibits cell apoptosis via regulating cell cycle progression.Multiple signaling molecules and pathways,including hypoxia-inducible factors,transforming growth factor-β/Smad,SET domain-containing 3/vascular endothelial growth factor,survivin,cell cycle regulatory genes and DNA damage response network,are reported to cross talk with FoxM1 in pulmonary arterial hypertension.Proteasome inhibitors are effective in the prevention and treatment of pulmonary arterial hypertension by inhibiting the expression and transcriptional activity of FoxM1.Conclusions FoxM1 has a crucial role in the pathogenesis of pulmonary arterial hypertension and may represent a novel therapeutic target.But more details of interaction between FoxM1 and other signaling pathways need to be clarified in the future.

Overcoming drug resistance by targeting protein homeostasis in multiple myeloma

Multiple myeloma(MM)is a plasma cell disorder typically characterized by abundant synthesis of clonal immunoglobulin or free light chains.Although incurable,a deeper understanding of MM pathobiology has fueled major therapeutical advances over the past two decades,significantly improving patient outcomes.Proteasome inhibitors,immunomodulatory drugs,and monoclonal antibodies are among the most effective anti-MM drugs,targeting not only the cancerous cells,but also the bone marrow microenvironment.However,de novo resistance has been reported,and acquired resistance is inevitable for most patients over time,leading to relapsed/refractory disease and poor outcomes.Sustained protein synthesis coupled with impaired/insufficient proteolytic mechanisms makes MM cells exquisitely sensitive to perturbations in protein homeostasis,offering us the opportunity to target this intrinsic vulnerability for therapeutic purposes.This review highlights the scientific rationale for the clinical use of FDA-approved and investigational agents targeting protein homeostasis in MM.

Therapeutics to harness the immune microenvironment in multiple myeloma

Multiple myeloma(MM)remains an incurable,genetically heterogeneous disease characterized by the uncontrolled proliferation of transformed plasma cells nurtured within a permissive bone marrow(BM)microenvironment.Current therapies leverage the unique biology of MM cells and target the immune microenvironment that drives tumor growth and facilitates immune evasion.Proteasome inhibitors and immunomodulatory drugs were initially introduced to complement and have now supplanted cytotoxic chemotherapy as frontline anti-myeloma agents.Recently,monoclonal antibodies,bispecific antibodies,and chimeric antigen receptor T cells were developed to revamp the immune system to overcome immune suppression and improve patient responses.While current MM therapies have markedly extended patient survival,acquired drug resistance inevitably emerges and drives disease progression.The logical progression for the next generation of MM therapies would be to design and validate agents that prevent and/or overcome acquired resistance to immunotherapies.The complex BM microenvironment promotes resistance to both current anti-myeloma agents and emerging immunotherapies.Myeloma cells are intertwined with a complex BM immune microenvironment that contributes to the development of adaptive drug resistance.Here,we describe recently FDA-approved and investigational anti-myeloma agents that directly or indirectly target the BM microenvironment to prevent or overcome drug resistance.Synergistic effects of anti-myeloma agents may foster the development of rationally-designed drug cocktails that prevent BM-mediated resistance to immunotherapies.