Microbial proteasomes as drug targets

Proteasomes are compartmentalized, ATP-dependent, N-terminal nucleophile hydrolases that play essentials roles in intracellular protein turnover. They are present in all 3 kingdoms. Pharmacological inhibition of proteasomes is detrimental to cell viability. Proteasome inhibitor rugs revolutionize the treatment of multiple myeloma. Proteasomes in pathogenic microbes such as Mycobacterium tuberculosis (Mtb), Plasmodium falciparum (Pf), and other parasites and worms have been validated as therapeutic targets. Starting with Mtb proteasome, efforts in developing inhibitors selective for microbial proteasomes have made great progress lately. In this review, we describe the strategies and pharmacophores that have been used in developing proteasome inhibitors with potency and selectivity that spare human proteasomes and highlight the development of clinical proteasome inhibitor candidates for treatment of leishmaniasis and Chagas disease. Finally, we discuss the future challenges and therapeutical potentials of the microbial proteasome inhibitors.

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Towards the control of intracellular protein turnover: Mitochondrial Lon protease inhibitors versus proteasome inhibitors

Biochimie ◽

10.1016/j.biochi.2007.10.010 ◽

2008 ◽

Vol 90 (2) ◽

pp. 260-269 ◽

Cited By ~ 37

Author(s):

Aurélien Bayot ◽

Nicolas Basse ◽

Irene Lee ◽

Monique Gareil ◽

Bernard Pirotte ◽

...

Keyword(s):

Protease Inhibitors ◽

Protein Turnover ◽

Proteasome Inhibitors ◽

Lon Protease ◽

Intracellular Protein

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Investigational Agent MLN2238/MLN9708, a Specific, Orally Available, Small Molecule Proteasome Inhibitor, Shows Promising In Vitro Activity Against Multiple Myeloma Cell Lines

Blood ◽

10.1182/blood.v116.21.3014.3014 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3014-3014

Author(s):

Giada Bianchi ◽

Vijay G. Ramakrishnan ◽

Teresa Kimlinger ◽

Jessica Haug ◽

S. Vincent Rajkumar ◽

...

Keyword(s):

Multiple Myeloma ◽

Er Stress ◽

Cell Lines ◽

Small Molecule ◽

Proteasome Inhibitor ◽

Research Funding ◽

Proteasome Inhibitors ◽

Biologically Active ◽

Investigational Agent

Abstract Abstract 3014 Background: Proteasome inhibitors have proven particularly effective in treatment of multiple myeloma, the second most frequent hematologic malignancy in the western world. Bortezomib, the first in class proteasome inhibitor in clinical use, was first approved in 2003 via fast FDA track, given the remarkable activity shown during phase II clinical trials. Nevertheless, more than 50% of multiple myeloma patients did not respond to single agent bortezomib when administered as second line agent. Moreover, bortezomib is only available for intravenous administration, representing a cumbersome therapy for patients, and its use is limited by significant toxicities (especially peripheral neuropathy). MLN9708 (Millennium Pharmaceuticals, Inc.), an investigational orally available, small molecule, is a potent, specific and reversible inhibitor of the 20S proteasome. It is currently under clinical investigation for the treatment of hematologic and non-hematologic malignancies. Upon exposure to aqueous solutions or plasma, MLN9708 rapidly hydrolyzes to MLN2238, the biologically active form, and MLN2238 was used for all of the preclinical studies reported here. In vitro biochemistry studies have shown that MLN2238 has a faster dissociation rate from the proteasome compared to bortezomib, and in vivo studies of MLN2238 have shown antitumor activity in a broader range of tumor xenografts when compared to bortezomib. Given these encouraging preclinical results, we set to investigate the anti-myeloma activity of MLN2238 in vitro. Results: MLN2238 proved to have anti-proliferative and pro-apoptotic activity against a broad range of MM cell lines with EC50 at 24 hours ranging between 10 and 50 nM, even in relatively resistant MM cell lines (OPM2, DOX6, RPMI, etc.). In MM.1S cells, induction of apoptosis was time and dose dependent and related to activation of both caspase 8 and 9. When compared to MM.1S treated for 24 hours with EC50 dose of bortezomib, treatment with EC50 dose of MLN2238 resulted in the same extent of caspases cleavage occurring at an earlier time point (8-12 hours), possibly suggesting more rapid onset and/or irreversibility of apoptosis in cells treated with MLN2238. Treatment with MLN2238 was associated with early, but persistent induction of endoplasmic reticulum (ER) stress with BiP being induced 2–4 hours after treatment with EC50 dose and gradually increasing over time. While bortezomib has been associated with early induction and late decrease in proteins involved in ER stress, MLN2238 appears to induce a persistent rise in these factors, suggesting either more sustained proteasome blockade with stabilization of proteasome substrates or de-novo induction of unfolded protein response (UPR) genes. MLN2238 also proved effective in reducing phosphorylation of ERK1-2 with no overall alteration in the total ERK level, thus accounting for the observed reduction in proliferation upon treatment. Preliminary data indicate potential for additive and synergistic combination with widely used drugs, including doxorubicin and dexamethasone. Conclusion: While further clinical data are needed to establish the effectiveness of MLN2238 in the treatment of multiple myeloma, these preliminary nonclinical data, together with the favorable biochemical and pharmacokinetic properties, including oral bioavailability, make the investigational agent MLN9708 an appealing candidate for treatment of multiple myeloma. Further in vitro data could help establish whether a difference in the apoptotic mechanisms exist between MLN2238 and other proteasome inhibitors, primarily bortezomib, and could also help inform combination treatment approaches aimed at increasing effectiveness, overcoming bortezomib resistance and decreasing toxicity. Disclosures: Kumar: Celgene: Consultancy, Research Funding; Millennium: Research Funding; Merck: Consultancy, Research Funding; Novartis: Research Funding; Genzyme: Consultancy, Research Funding; Cephalon: Research Funding.

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Carfilzomib: A Promising Proteasome Inhibitor for the Treatment of Relapsed and Refractory Multiple Myeloma

Frontiers in Oncology ◽

10.3389/fonc.2021.740796 ◽

2021 ◽

Vol 11 ◽

Author(s):

Shansa Pranami E. Jayaweera ◽

Sacheela Prasadi Wanigasinghe Kanakanamge ◽

Dharshika Rajalingam ◽

Gayathri N. Silva

Keyword(s):

Multiple Myeloma ◽

Proteasome Inhibitor ◽

Proteasome Inhibitors ◽

Biochemical Properties ◽

Regulatory Proteins ◽

Toxicity Profile ◽

The Us ◽

Intracellular Proteins ◽

Ubiquitin Proteasome ◽

Proteasome Pathway

The proteasome is crucial for the degradation of intracellular proteins and plays an important role in mediating a number of cell survival and progression events by controlling the levels of key regulatory proteins such as cyclins and caspases in both normal and tumor cells. However, compared to normal cells, cancer cells are more dependent on the ubiquitin proteasome pathway (UPP) due to the accumulation of proteins in response to uncontrolled gene transcription, allowing proteasome to become a potent therapeutic target for human cancers such as multiple myeloma (MM). Up to date, three proteasome inhibitors namely bortezomib (2003), carfilzomib (2012) and ixazomib (2015) have been approved by the US Food and Drug Administration (FDA) for the treatment of patients with relapsed and/or refractory MM. This review mainly focuses on the biochemical properties, mechanism of action, toxicity profile and pivotal clinical trials related to carfilzomib, a second-generation proteasome inhibitor that binds irreversibly with proteasome to overcome the major toxicities and resistance associated with bortezomib.

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PROTEASOME INHIBITION IN MULTIPLE MYELOMA: Therapeutic Implication

The Annual Review of Pharmacology and Toxicology ◽

10.1146/annurev.pharmtox.45.120403.100037 ◽

2005 ◽

Vol 45 (1) ◽

pp. 465-476 ◽

Cited By ~ 98

Author(s):

Dharminder Chauhan ◽

Teru Hideshima ◽

Kenneth C. Anderson

Keyword(s):

Multiple Myeloma ◽

Proteasome Inhibitors ◽

Proteasome Inhibition ◽

Protein Translation ◽

26S Proteasome ◽

Intracellular Protein ◽

Cycle Growth ◽

Intracellular Protein Degradation ◽

Ubiquitin Proteasome ◽

Proteasome Pathway

Normal cellular functioning requires processing of proteins regulating cell cycle, growth, and apoptosis. The ubiquitin-proteasome pathway (UBP) modulates intracellular protein degradation. Specifically, the 26S proteasome is a multienzyme protease that degrades misfolded or redundant proteins; conversely, blockade of the proteasomal degradation pathways results in accumulation of unwanted proteins and cell death. Because cancer cells are more highly proliferative than normal cells, their rate of protein translation and degradation is also higher. This notion led to the development of proteasome inhibitors as therapeutics in cancer. The FDA recently approved the first proteasome inhibitor bortezomib (Velcade™), formerly known as PS-341, for the treatment of newly diagnosed and relapsed/refractory multiple myeloma (MM). Ongoing studies are examining other novel proteasome inhibitors, in addition to bortezomib, for the treatment of MM and other cancers.

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Depletion of the Intracellular Akt Kinase, and Its Downstream Signaling, Mediates Synergy between the Proteasome Inhibitor MG-132 and the Heat Shock Protein 90 Inhibitor 17-AAG in the Multiple Myeloma Cell Line U266.

Blood ◽

10.1182/blood.v104.11.3364.3364 ◽

2004 ◽

Vol 104 (11) ◽

pp. 3364-3364

Author(s):

Christopher Maisel ◽

Nizar Bahlis ◽

Yanling Miao ◽

Lili Liu ◽

Stanton Gerson

Keyword(s):

Multiple Myeloma ◽

Heat Shock ◽

Heat Shock Protein ◽

Cell Line ◽

Proteasome Inhibitor ◽

Dose Range ◽

Proteasome Inhibitors ◽

Heat Shock Protein 90 ◽

Hsp 70 ◽

Hsp 90

Abstract While proteasome inhibitors are effective therapy for multiple myeloma (MM), their efficacy could be improved by synergistic targeting of apoptosis. The intracellular serine/threonine kinase Akt has been demonstrated to have a central role in MM cell growth, survival, and drug resistance. Akt is activated by extracellular cytokines such as IGF-1 and IL-6, and contributes to MM resistance by ameliorating the apoptotic effects of proteasome inhibition. Akt requires chaperone proteins for proper stability and function, including the 90 kD molecule Heat Shock Protein 90 (HSP-90). HSP-90 function is abrogated by geldanamycin and its derivative, 17-allylamino-17-demethoxygeldanamycin (17-AAG). In the U266 MM cell line, the IC50 of the proteasome inhibitor MG-132 and 17-AAG was 100 nM and 800 nM, respectively. Following exposure of U266 MM cells to either drug alone, or the combination at a fixed-ratio of their IC50s (1:8), apoptosis was determined by Annexin V staining and FACScan analysis. Synergy analysis was performed using Calcusyn (Biosoft, Cambridge, UK). We found that the combination index (CI) was synergistic (CI<1) throughout the dose range, with a CI = 0.449 ± 0.298 at the combination IC50 (highly significant). For example, the apoptotic effect of 50 nM MG-132 and 400 nM 17-AAG was 6 ±2 % and 23 ±3 %, respectively, whereas the 50:400 nM combination produced apoptosis in 68 ± 2 % of the cells. To analyze effects on Akt and its substrates, we incubated U266 MM cells with MG-132 (50 nM), 17-AAG (400 nM), or the combination. We harvested lysates after zero, two, six, and 24 hours incubation, and Western blot analysis was performed. Co-incubation with MG-132 and 17-AAG, but not either alone, depleted Akt by 24 hours post-therapy. Co-treatment also produced significantly greater upregulation of HSP-90 and HSP-70 than 17-AAG alone, thus demonstrating greater functional inhibition of Akt. The combination also demonstrated the greatest abrogation of Akt-mediated effects on mitochondrial apoptosis: Co-treatment produced the greatest expression of BAD, decreased BCL-XL expression, reduced phosphorylation of GSK-3, and produced the greatest activation of caspase 3. Monotherapy with 17-AAG upregulated HSP-90 and HSP-70, reduced BCL-XL expression, and activated caspase 9. MG-132 monotherapy produced none of these effects. These findings demonstrate that synergy between proteasome inhibitors and 17-AAG is mediated by Akt depletion and abrogation of Akt signaling, predominantly by MG-132 augmentation of 17-AAG-mediated decay of Akt. Down-regulation of Akt-mediated resistance allows dual-apoptotic signaling and synergistic effect of combination therapy. These findings demonstrate a mechanistic rationale for utilizing heat shock protein inhibitors in combination with proteasome inhibitors as therapy for MM.

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The Role of Proteasome Inhibitors and the Trail Apoptotic Pathway in the Treatment of Chronic Lymphocytic Leukemia.

Blood ◽

10.1182/blood.v106.11.5011.5011 ◽

2005 ◽

Vol 106 (11) ◽

pp. 5011-5011

Author(s):

Kristin E. McCrea ◽

Albert Kabore ◽

James B. Johnston ◽

Spencer B. Gibson

Keyword(s):

Multiple Myeloma ◽

Monoclonal Antibodies ◽

B Cells ◽

Chronic Lymphocytic Leukemia ◽

Proteasome Inhibitor ◽

Proteasome Inhibitors ◽

Death Receptors ◽

Lymphocytic Leukemia ◽

Apoptotic Pathway ◽

Novel Approach

Abstract TRAIL (tumour necrosis factor-related apoptosis-inducing ligand) triggers the TRAIL apoptotic pathway selectively in tumour cells by binding to two death receptors, DR4 and DR5, that are present on the surface of many cancer cells. It is effective at inducing apoptosis in a variety of haematological malignancies, such as multiple myeloma. However, chronic lymphocytic leukemia (CLL) cells are relatively resistant to TRAIL-induced apoptosis. We have previously shown that the chemotherapeutic drugs, fludarabine and chlorambucil, increase the cell surface expressions of DR4 and DR5, and give synergistic apoptotic responses when combined with TRAIL. Proteasome inhibitors, that are used in the treatment of multiple myeloma, also upregulate TRAIL and its death receptors in CLL cells but not in normal B cells. Herein we have determined that proteasome inhibitors are effective at inducing apoptosis in CLL cells, and that the activation of the TRAIL apoptotic pathway contributes significantly to proteasome inhibitor cytotoxicity. Combining proteasome inhibitors with TRAIL enhanced apoptosis in CLL cells by approximately 15% over treatment with the proteasome inhibitor alone. Another novel approach to trigger the TRAIL apoptotic pathway is to use activating monoclonal antibodies directed against DR4 and DR5. Similar to TRAIL, we demonstrated that when used alone, the monoclonal antibodies were minimally cytotoxic against CLL cells. Proteasome inhibitors in combination with activating monoclonal antibodies against DR4 or DR5 increased the amount of apoptosis in CLL cells. Cell death was enhanced by 23% and 17% when proteasome inhibitors were combined with monoclonal antibodies against DR4 and DR5, respectively. While proteasome inhibitors may have a potential role in CLL treatment as single therapy, they are also cytotoxic to normal B cells. However, when activating monoclonal antibodies against TRAIL death receptors are given in combination with proteasome inhibitors, that upregulate DR4 and DR5 expression, the anti-tumor specificity and cytotoxicity of these agents may be increased in CLL.

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S-2209, a Novel, Non-Boronic Proteasome Inhibitor, Induces Apoptosis and Cell Growth Arrest in Multiple Myeloma Cells.

Blood ◽

10.1182/blood.v110.11.1513.1513 ◽

2007 ◽

Vol 110 (11) ◽

pp. 1513-1513

Author(s):

Philipp Baumann ◽

Karin Mueller ◽

Sonja Mandl-Weber ◽

Helmut Ostermann ◽

Ralf Schmidmaier ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Growth ◽

Cell Lines ◽

Proteasome Inhibitor ◽

Wistar Rats ◽

Proteasome Inhibitors ◽

Proteasome Inhibition ◽

Inhibition Assay ◽

Induction Of Apoptosis

Abstract Purpose: Multiple Myeloma (MM) is still an incurable disease. Patients become resistant to cytotoxic drugs and die of disease progression. Bortezomib is the first approved member of a new class of antineoplastic agents, the proteasome inhibitors. It has synergistic effects with genotoxic drugs and steroids in vitro and in vivo. However, single agent activity in humans is only moderate and specific toxicity (e.g. neurotoxicity) often limits its clinical use. Further proteasome inhibitors need to be developed to optimize this promising treatment option. Methods: The new proteasome inhibitor S-2209 was characterized by several assays. Inhibition of the chymotryptic activity of the human 20S proteasome was determined with the in-vivo protease inhibition assay. Additionally, proteasome inhibition was determined in isolated PBMCs from S2209-pretreated wistar rats. Inhibition of NFκB activity was determined using a NFκB reporter gene assay. Cell growth rates of MM cells (OPM-2, U266, RPMI-8226 and NCI-H929) were measured with the WST-1 assay. Induction of apoptosis was shown by flow cytometry after staining with annexin-V-FITC and propidium iodide. Intracellular signal modulation was demonstrated by western blotting. Toxicity of the substance was tested in male wistar rats. Results: The proteasome inhibition assay revealed an IC50 at ∼220nM. The NFκB inhibition assay using an A549-NFκB-SEAP transfected cell line showed an EC50 of 0.9μM. Upon incubation with S-2209, cell growth as well as cell proliferation in MM cell lines was significantly inhibited (IC50 100nM – 600nM). Furthermore, the incubation with S-2209 resulted in strong induction of apoptosis in all four MM cell lines even at nanomolar concentrations (IC50 at ∼300nm) as well as primary cells. Western blotting revealed caspase-3 cleavage and upregulon of p53 and increased phosphorylation of IκB. No induction of apoptosis was detected in PBMCs from healthy humans. Despite the administration of 5, 10 or 15mg/kg/day in wistar rats, no toxicity with respect to body weight, hepatic enzymes (ALAT ASAT, ALP), creatinin or hemoglobin was seen. Proteasome inhibition in white blood cells isolated from the treated rats was higher in the S-2209 treated animals than in control animals treated with 0.1mg/kg/d bortezomib (89% vs. 70% respectively). Conclusions: The proteasome inhibitor S-2209 inhibitis MM cell growth and induces apoptosis. This is accompanied by a strong inhibition of proteasome and of the NFκB activity. Because S-2209 shows a favourable toxicity profile in vivo, further clinical development of this promising drug is urgently needed.

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Pre-Clinical Activity of the Novel, First-in-Class p97 Inhibitor, CB-5083, in Multiple Myeloma

Blood ◽

10.1182/blood.v124.21.4701.4701 ◽

2014 ◽

Vol 124 (21) ◽

pp. 4701-4701

Author(s):

Blake T. Aftab ◽

Daniel J Anderson ◽

Ronan Le Moigne ◽

Stevan Djakovic ◽

Eugen Dhimolea ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Death ◽

Cell Lines ◽

Proteasome Inhibitor ◽

Stromal Cell ◽

Hematological Malignancies ◽

Proteasome Inhibitors ◽

Protein Homeostasis ◽

Ubiquitinated Proteins

Abstract Hematological malignancies such as multiple myeloma (MM) have an increased reliance on the ubiquitin proteasome system (UPS) presumably as a consequence of their high protein synthetic and secretory burden. Chemical agents that target the proteasome, such as bortezomib and carfilzomib, have been successful in treating multiple myeloma; however patients treated with these drugs ultimately relapse. The AAA-ATPase p97/VCP (p97) facilitates ATP-dependent extraction and degradation of ubiquitinated proteins destined for proteasomal elimination. In addition to ubiquitin-dependent protein degradation, p97 is also closely involved in other aspects of protein homeostasis, including endoplasmic reticulum-associated degradation (ERAD) and autophagy. Pharmacologic inhibition of p97 provides a compelling therapeutic approach for hematological malignancies that rely on tight regulation of protein homeostasis as a component of their survival. CB-5083 is a novel small molecule inhibitor of p97 ATPase activity with nanomolar enzymatic and cellular potency. Treatment of cancer cells with CB-5083 causes a dramatic increase in poly-ubiquitinated proteins as well as an accumulation of substrates of the UPS and ERAD. CB-5083 causes a profound induction of the unfolded protein response (UPR) with consequent activation of the DR5 death receptor, caspase 8, caspase 3/7 and ultimately cell death. Induction of the UPR occurs to a greater magnitude with CB-5083 when compared to the proteasome inhibitor, bortezomib, suggesting the potential for increased efficacy in cancers with sensitivity to UPR-mediated cell death. In addition, activation of apoptosis and cell death occur more rapidly with CB-5083 than with bortezomib. Sequencing of cell lines made resistant to CB-5083 reveals missense mutations mapping to the D2 ATPase site in p97, supporting on-target association with cytotoxicity. In an expanded panel of MM cell lines there is no correlation between the cytotoxic sensitivity to CB-5083 and the cytotoxic sensitivity to proteasome inhibitors, suggesting differential mechanisms of cytotoxicity and potential activity of CB-5083 in proteasome inhibitor resistant settings. Compared to myeloma cell lines, CB-5083 has reduced cytotoxic potency in immortalized stromal cell lines and in patient-derived CD138-negative bone marrow mononuclear cells. Furthermore, unlike the reduced potency demonstrated by carfilzomib in the context of MM cell-bone marrow stromal cell (BMSC) interactions, the cyto-reductive potential of CB-5083 is unaffected in co-cultures of MM cells with patient-derived BMSCs or immortalized BMSCs from healthy donors. In vivo, CB-5083 is orally bioavailable, shows a pharmacodynamic effect in tumor tissue (as measured by poly-ubiquitin accumulation) and demonstrates robust anti-tumor activity across several MM models. CB-5083 treatment of mice bearing subcutaneous xenografts leads to tumor stasis and regression in RPMI8226 and AMO1 MM models, respectively. In advanced models of disseminated, ortho-metastatic disease, intermittent oral administration of CB-5083 demonstrates significant inhibition of myeloma burden and improves survival, with an overall efficacy profile that compares favorably to that of clinically approved proteasome inhibitors. Furthermore, in the Vk*Myc genetically engineered mouse model of MM, treatment with CB-5083 results in a significant reduction in M-spike by 55%. Combination treatment of mice bearing the RPMI8226 subcutaneous xenograft model with CB-5083, dexamethasone and lenalidomide results in tumor regression. Taken together, these data demonstrate that CB-5083 is a potent and selective inhibitor of the p97 ATPase with robust activity in vitro and in vivo in numerous MM models and strongly support clinical evaluation. Based on these observations, a phase 1 dose-escalation trial has recently been initiated and is currently underway in patients with relapsed/refractory multiple myeloma. Disclosures Anderson: Cleave Biosciences: Employment. Le Moigne:Cleave Biosciences: Employment. Djakovic:Cleave Biosciences: Employment. Rice:Cleave Biosciences: Employment. Wong:Cleave Biosciences: Employment. Kumar:Cleave Biosciences: Employment. Valle:Cleave Biosciences: Employment. Menon:Cleave Biosciences: Employment. Kiss von Soly:Cleave Biosciences: Employment. Wang:Cleave Biosciences: Employment. Yao:Cleave Biosciences: Employment. Soriano:Cleave Biosciences: Employment. Bergsagel:ONYX: Consultancy; Janssen: Consultancy; BMS: Consultancy; Novartis: Research Funding. Yakes:Cleave Biosciences: Employment. Zhou:Cleave Biosciences: Employment. Wustrow:Cleave Biosciences: Employment. Rolfe:Cleave Biosciences: Employment.

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Proteasome inhibitors bortezomib and carfilzomib used for the treatment of multiple myeloma do not inhibit the serine protease HtrA2/Omi

Toxicology Research ◽

10.1039/c6tx00220j ◽

2016 ◽

Vol 5 (6) ◽

pp. 1619-1628 ◽

Cited By ~ 2

Author(s):

Vilmos Csizmadia ◽

Paul Hales ◽

Christopher Tsu ◽

Jingya Ma ◽

Jiejin Chen ◽

...

Keyword(s):

Multiple Myeloma ◽

Peripheral Neuropathy ◽

Serine Protease ◽

Proteasome Inhibitor ◽

Proteasome Inhibitors

The proteasome inhibitor bortezomib is associated with the development of peripheral neuropathy in patients, but the mechanism is not fully understood.

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Inhibition of p-glycoprotein does not increase the efficacy of proteasome inhibitors in multiple myeloma cells

10.1101/2020.07.16.206102 ◽

2020 ◽

Author(s):

Rachel L. Mynott ◽

Craig T. Wallington-Beddoe

Keyword(s):

Gene Expression ◽

Multiple Myeloma ◽

Proteasome Inhibitor ◽

Proteasome Inhibitors ◽

Specific Inhibitor ◽

Myeloma Cell ◽

Drug Transporter ◽

Myeloma Cells ◽

Multiple Myeloma Cell ◽

P Glycoprotein

AbstractThe aim of this study is to determine whether manipulation of the drug transporter P-glycoprotein improves the efficacy of proteasome inhibitors in multiple myeloma cells. P-glycoprotein is a well-known drug transporter that is associated with chemotherapy resistance in a number of cancers but its role in modulating the efficacy of proteasome inhibitors in multiple myeloma is not well understood. Research has shown that the second generation proteasome inhibitor carfilzomib is a substrate of P-glycoprotein and as such its efficacy may correlate with P-glycoprotein activity. In contrast to carfilzomib, research concerning the first-in-class proteasome inhibitor bortezomib is inconsistent with some reports suggesting that inhibition of P-glycoprotein increases bortezomib cytotoxicity in multiple myeloma cells whereas others have shown no effect. Through the mining of publicly available gene expression microarrays of patient bone marrow, we show that P-glycoprotein gene expression increases with the disease stages leading to multiple myeloma. However, RNA-seq on LP-1 cells treated with bortezomib or carfilzomib demonstrated minimal basal P-glycoprotein expression which did not increase with treatment. Moreover, only one (KMS-18) of nine multiple myeloma cell lines expressed P-glycoprotein, including RPMI-8226 cells that are resistant to bortezomib or carfilzomib. We hypothesised that by inhibiting P-glycoprotein, multiple myeloma cell sensitivity to proteasome inhibitors would increase, thus providing a potential approach to improving responses and reversing resistance to these agents. However, the sensitivity of multiple myeloma cells lines to proteasome inhibition was not enhanced by inhibition of P-glycoprotein with the specific inhibitor tariquidar. In addition, targeting glucosylceramide synthase with eliglustat did not inhibit P-glycoprotein activity and also did not improve proteasome inhibitor efficacy except at a high concentration. We conclude that P-glycoprotein is poorly expressed in multiple myeloma cells, its inhibition does not enhance the efficacy of proteasome inhibitors, and it is unlikely to be a useful avenue for further translational research.

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