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The proteasome is a proteolytic enzyme complex essential for protein homeostasis in mammalian cells and protozoan parasites like Trichomonas vaginalis (Tv), the cause of the most common, non-viral sexually transmitted disease. Tv and other protozoan 20S proteasomes have been validated as druggable targets for antimicrobials. However, low yields and purity of the native proteasome have hindered studies of the Tv 20S proteasome (Tv20S). We address this challenge by creating a recombinant protozoan proteasome by expressing all seven α and seven β subunits of Tv20S alongside the Ump-1 chaperone in insect cells. The recombinant Tv20S displays biochemical equivalence to its native counterpart, confirmed by various assays. Notably, the marizomib (MZB) inhibits all catalytic subunits of Tv20S, while the peptide inhibitor carmaphycin-17 (CP-17) specifically targets β2 and β5. Cryo-electron microscopy (cryo-EM) unveils the structures of Tv20S bound to MZB and CP-17 at 2.8 Å. These findings explain MZB's low specificity for Tv20S compared to the human proteasome and demonstrate CP-17's higher specificity. Overall, these data provide a structure-based strategy for the development of specific Tv20S inhibitors to treat trichomoniasis.

Protein homeostasis is a tightly conserved process that is regulated through the ubiquitin proteasome system (UPS) in a ubiquitin-independent or ubiquitin-dependent manner. Over the past two decades, the proteasome has become an excellent therapeutic target through inhibition of the catalytic core particle, inhibition of subunits responsible for recognizing and binding ubiquitinated proteins, and more recently, through targeted protein degradation using proteolysis targeting chimeras (PROTACs). The majority of the developed inhibitors of the proteasome's core particle rely on gaining selectivity through binding interactions within the unprimed substrate channel. Although this has allowed for selective inhibitors and chemical probes to be generated for the different proteasome isoforms, much remains unknown about the interactions that could be harnessed within the primed substrate channel to increase potency or selectivity. Herein, we discuss small molecules that interact with the primed substrate pocket and how their differences may give rise to altered activity. Taking advantage of additional interactions with the primed substrate pocket of the proteasome could allow for the generation of improved chemical tools for perturbing or monitoring proteasome activity.

Objectives: Multiple myeloma (MM) is a class of malignant plasma cell diseases. An increasing application of autologous stem cell transplantation (ASCT) and anti-myeloma agents represented by proteasome inhibitors (PIs) has improved the response rates and survival of MM patients. Patients progressing within 12 months were recently categorized with functional high-risk (FHR), which could not be clarified by existing genetic risk factors, with poor outcomes. Our study aimed to investigate clinical indices related to FHR and seek prognostic roles in transplant-eligible MM patients. Methods: Demographic and individual baseline clinical characteristics were compared by using the Pearson's chi-square and Mann-Whitney U test. Progression-free survival (PFS) and overall survival (OS) were described by Kaplan-Meier estimates and compared using the log-rank test. Logistic regression analysis was used to assess the association of baseline characteristics at MM diagnosis with FHR status. Results: From 18th January 2010 to 1st December 2022, 216 patients were included and divided into two groups according to the FHR status. There was no difference in baseline data between the two groups. Renal impairment (RI, Scr > 2 mg/dL) was common in MM patients and made sense in FHR status. AST levels were validated as independent predictors for FHR status (p = 0.019). Discussion: Patients with RI or higher AST levels (AST > 40 U/L) tended to have worse outcomes. However, transplants had apparently improved prognoses. Conclusion: Therefore, in the PIs era, transplantations are still effective therapies for transplant-eligible MM patients. [ABSTRACT FROM AUTHOR]

Objective: In this study, we aimed to obtain real-life data on the use of antimyeloma agents, which significantly increase overall survival (OS) in multiple myeloma (MM) patients, in primary plasma cell leukemia (pPCL) patients with poor prognosis. Materials and Methods: Data from 53 patients who were diagnosed with pPCL between 2011 and 2020 and who used at least one proteasome inhibitor (PI) and/or immunomodulatory (IMID) agent were analyzed retrospectively. Depending on the year of the pPCL diagnosis, 20% leukocytes or ≥2x109/L plasma cells in the peripheral blood was used as a diagnostic criterion. Results: The median age of the patients was 58 years and 23 (43.4%) patients were over 65 years of age. For first-line treatment, PI or IMID alone was used by 31 (58.5%) patients, and PI and IMID were used simultaneously by 15 (28.3%) patients. Additionally, 21 (39.6%) patients received transplantation and 13 (24.5%) patients received maintenance treatment. The median progression-free survival was 4 (range: 1-42) months. When patients whose primary disease was refractory to first-line therapy were excluded, the duration of treatment was 6.5 months. The median OS was 15 months with a median follow-up duration of 15 months. Only 7 (13.2%) of the patients were alive at the last follow-up visit. Those with higher β2- microglobulin levels and International Staging System stage 3 and non-transplant patients receiving first-line treatment had shorter OS (p=0.005, p=0.02, and p=0.008, respectively). The concomitant use of PIs and IMIDs, the addition of chemotherapy to induction therapy, and the response to induction therapy or maintenance therapy did not affect OS. Conclusion: In this study, as in previous similar studies, we could not see an increased survival trend in pPCL, which is observed in MM. New studies are needed for pPCL, which is likely to increase with new diagnostic criteria, based on current agents and information from MM. [ABSTRACT FROM AUTHOR]

Background Carfilzomib, a new proteasome inhibitor indicated for patients with relapsed/refractory myeloma, has been associated with cases of thrombotic microangiopathy (CFZ-TMA). The role of variants in the complement alternative pathway and therapeutic potential of complement blockade with eculizumab remain to be determined. Methods We report 37 cases of CFZ-TMA recorded in the French reference center for TMA with their clinical characteristics, genetic analysis and outcome according to treatments. Results A trigger was identified in more than half of cases, including eight influenza and five severe acute respiratory syndrome coronavirus-2 cases. All patients presented with acute kidney injury (AKI) [KDIGO stage 3 in 31 (84%) patients] while neurological (n  = 13, 36%) and cardiac (n  = 7, 19%) damage were less frequent. ADAMTS13 (a disintegrin and metalloprotease with thrombospondin type I repeats-13) and complement activity were normal (n  = 28 and 18 patients tested) and no pathogenic variant in the alternative complement pathway was found in 7 patients tested. TMA resolved in most (n  = 34, 94%) patients but 12 (44%) still displayed stage 3 AKI at discharge. Nineteen (51%) patients were treated with therapeutic plasma exchange, 14 (38%) patients received corticosteroids and 18 (50%) were treated with eculizumab. However, none of these treatments demonstrated a significant impact on outcomes. Conclusion This study is the largest case series of CFZ-TMA since its approval in 2012. Patients present with severe AKI and experience frequent sequelae. Complement variants and blockade therapy do not seem to play a role in the pathophysiology and prognosis of the disease. [ABSTRACT FROM AUTHOR]

MEDI2228 is an antibody drug conjugate (ADC) comprised of a fully human B-cell maturation antigen (BCMA) antibody conjugated to a pyrrolobenzodiazepine (PBD) dimer. This phase 1 trial evaluated MEDI2228 in patients with relapsed/refractory (R/R) multiple myeloma (MM), who received prior treatment with approved agents from 3 classes of antimyeloma drugs (proteasome inhibitors, immunomodulatory drugs, and monoclonal antibodies). Primary endpoint was safety and tolerability; secondary endpoints included efficacy, pharmacokinetics, and immunogenicity. A total of 107 patients were treated and the maximum tolerated dose (MTD) was 0.14 mg/kg Q3W. Two patients had dose-limiting toxicities (DLTs; thrombocytopenia; 0.20 mg/kg Q3W). The most frequent treatment-related adverse events were photophobia (43.9%), rash (29.0%), and thrombocytopenia (19.6%). In MTD cohort A (n = 41), the objective response rate (ORR) was 56.1%, with 1 stringent complete response, 9 very good partial responses, and 13 partial responses. ORR was 53.3% in triple refractory patients. In cohort B (n=25), ORR was 32%. Although MEDI2228 demonstrated efficacy in R/R MM, ocular toxicity precluded further development of this drug. KEY POINTS: MEDI2228 is an ADC comprised of a fully human anti-BCMA antibody conjugated to the cytotoxic PBD payload, tesirine. MEDI2228 monotherapy demonstrated efficacy across all dose levels; and responses were observed in patients with triple refractory MM. This study further validates BCMA as a target for ADC-based therapy in MM; ocular toxicity precluded further clinical development. [ABSTRACT FROM AUTHOR]

Background: Gilteritinib is a commonly used targeted drug for acute myeloid leukemia (AML), but the emergence of gilteritinib resistance greatly reduces the therapeutic effect. RING finger protein 38 (RNF38), a protein with RING Finger domain and E3 ubiquitin ligase activity, has been implicated in tumorigenesis and drug resistance. However, the role and mechanism of RNF38 in the gilteritinib resistance of AML remains unclear. Methods: Normal AML cells were treated with gilteritinib to construct gilteritinib-resistant cells (MV4-11/Gilteritinib and MOLM-13/Gilteritinib). CCK8 assay, TUNEL staining and EdU assay were used to assess gilteritinib resistance, cell apoptosis and proliferation. The protein levels of autophagy-related markers, RNF38 and LIM homeobox transcription factor 1 alpha (LMX1A) were determined by western blot. Also, RNF38 and LMX1A mRNA levels were tested using qRT-PCR. Autophagic flux was assessed using mRFP-GFP-LC3 labeling, and autophagosome numbers was counted under transmission electron microscopy. Co-IP assay was employed to analyze interaction between RNF38 and LMX1A. The effects of LMX1A and RNF38 on AML tumorigenesis were analyzed by in vivo experiments. Results: In gilteritinib-resistant AML cells, autophagy-related markers, mRFP-GFP-LC3 signals and autophagosome numbers were significantly enhanced. Autophagy inhibitor 3-MA could suppress gilteritinib resistance in AML cells. RNF38 knockdown inhibited gilteritinib resistance and autophagy in AML cells. Mechanistically, RNF38 reduced LMX1A expression by inducing its ubiquitination. RNF38 overexpression reversed the inhibitory effect of LMX1A on gilteritinib resistance and autophagy in AML cells, as well as AML tumor growth in vivo, while these effects could be abolished by proteasome inhibitor MG132. Conclusions: RNF38 induced autophagy to promote gilteritinib resistance in AML by increasing the ubiquitination of LMX1A. 1. Autophagy is activated in gilteritinib-resistant AML cells. 2. Reduction of autophagy suppresses gilteritinib resistance in AML cells. 3. RNF38 knockdown inhibits gilteritinib resistance and autophagy in AML cells. 4. RNF38 reduces LMX1A expression via inducing its ubiquitination. [ABSTRACT FROM AUTHOR]

Human babesiosis is a malaria‐like, tick‐borne infectious disease with a global distribution. Babesiosis is caused by intraerythrocytic, apicomplexan parasites of the genus Babesia. In the United States, human babesiosis is caused by Babesia microti and Babesia duncani. Current treatment for babesiosis includes either the combination of atovaquone and azithromycin or the combination of clindamycin and quinine. However, the side effects of these agents and the resistance posed by these parasites call for alternative approaches for treating human babesiosis. Proteases play several roles in the context of parasitic lifestyle and regulate basic biological processes including cell death, cell progression, and cell migration. Using the SYBR Green‐1 assay, we screened a protease inhibitor library that consisted of 160 compounds against B. duncani in vitro and identified 13 preliminary hits. Dose response assays of hit compounds against B. duncani and B. microti under in vitro conditions identified five effective inhibitors against parasite growth. Of these compounds, we chose ixazomib, a proteasome inhibitor as a potential drug for animal studies based on its lower IC50 and a higher therapeutic index in comparison with other compounds. Our results suggest that Babesia proteasome may be an important drug target and that developing this class of drugs may be important to combat human babesiosis. [ABSTRACT FROM AUTHOR]

Malaria remains a global health challenge, with increasing resistance to frontline antimalarial treatments such as artemisinin (ART) threatening the efficacy of current therapies. In this study, we investigated the potential of FDA-approved drugs to selectively inhibit the malarial proteasome, a novel target for antimalarial drug development. By leveraging pharmacophore modeling, molecular docking, molecular dynamics (MD) simulations, and binding free-energy calculations, we screened a library of compounds to identify inhibitors selective for the Plasmodium proteasome over the human proteasome. Our results highlighted Argatroban, LM-3632, Atazanavir Sulfate, and Pemetrexed Hydrate as promising candidates, with Argatroban and Pemetrexed Hydrate showing the highest binding affinity and selectivity toward the malarial proteasome. MD simulation and gmx_MMPBSA analysis confirmed the compounds' ability to remain within the active site of the malarial proteasome, while some exited or exhibited reduced stability within the human proteasome. This study underscores the potential of proteasome-targeting drugs for overcoming malarial drug resistance and paves the way for the further optimization of these compounds. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica causes amebiasis, a significant global health issue, with millions affected annually, especially in developing countries. EhDUF2419, an important protein involved in E. histolytica's queuine salvage pathway and its interaction network, remains unclear. To explore this, we transfected E. histolytica trophozoites with a plasmid encoding Myc-tagged EhDUF2419 and achieved successful overexpression. Through immunoprecipitation with the Myc antibody followed by mass spectrometry, we identified 335 proteins interacting with Myc-tagged EhDUF2419, including over 100 ribosomal proteins, along with translation initiation and elongation factors, and aminoacyl-tRNA synthetases. Ribosome purification revealed the presence of EhDUF2419 in ribosomal protein-enriched fractions. Treatment with queuosine (Q) significantly reduced the EhDUF2419 protein levels and decreased the Q-modified tRNA in Myc-tagged EhDUF2419 overexpressing trophozoites. This effect, which was Q-dependent, was not observed in strains carrying an empty vector control or overexpressing a truncated form of EhDUF2419 lacking catalytic activity. The reduction in the EhDUF2419 protein levels was regulated by proteasome-mediated degradation, as evidenced by the reduced degradation in the presence of MG132, a proteasome inhibitor. Our study uncovers the novel interaction of EhDUF2419 with ribosomal proteins and its regulation by the proteasome machinery, providing new insights into its role in E. histolytica and potential therapeutic strategies. [ABSTRACT FROM AUTHOR]

Immunomodulatory imide drugs (IMiDs) are central components of therapy for multiple myeloma (MM). IMiDs bind cereblon (CRBN), an adaptor for the CUL4‐DDB1‐RBX1 E3 ligase to change its substrate specificity and induce degradation of 'neosubstrate' transcription factors that are essential to MM cells. Mechanistic studies to date have largely focussed on mediators of therapeutic activity and insight into clinical IMiD toxicities is less developed. We adopted BioID2‐dependent proximity labelling (BioID2‐CRBN) to characterise the CRBN interactome in the presence and absence of various IMiDs and the proteasome inhibitor, bortezomib. We aimed to leverage this technology to further map CRBN interactions beyond what has been achieved by conventional proteomic techniques. In support of this approach, analysis of cells expressing BioID2‐CRBN following IMiD treatment displayed biotinylation of known CRBN interactors and neosubstrates. We observed that bortezomib alone significantly modifies the CRBN interactome. Proximity labelling also suggested that IMiDs augment the interaction between CRBN and proteins that are not degraded, thus designating 'neointeractors' distinct from previously disclosed 'neosubstrates'. Here we identify Non‐Muscle Myosin Heavy Chain IIA (MYH9) as a putative CRBN neointeractor that may contribute to the haematological toxicity of IMiDs. These studies provide proof of concept for proximity labelling technologies in the mechanistic profiling of IMiDs and related E3‐ligase‐modulating drugs. [ABSTRACT FROM AUTHOR]

Proteasome inhibitors (PIs) are crucial in treating multiple myeloma but carry a risk of thrombotic microangiopathy (TMA), especially with carfilzomib use. This systematic review includes 44 studies with 115 cases of PI-induced TMA, where carfilzomib was implicated in 101 cases. Treatment approaches varied: 28 patients received supportive care, 43 underwent therapeutic plasma exchange (TPE), 9 were treated exclusively with eculizumab (ECU), and 13 received both TPE and ECU. Notably, eculizumab significantly improved outcomes for patients unresponsive to initial TPE, achieving complete remission in seven cases. The need for dialysis emerged as a significant predictor of outcomes, often indicating a poor prognosis. For patients suspected of having PI-TMA, it is advisable to discontinue the offending medication promptly, even without definitive laboratory confirmation. In cases where diagnosis is challenging, kidney biopsy may assist if conditions permit. Comprehensive evaluation of the complement system, including genetic mutations, function, and associated complement inhibitory factor antibodies, should be included in the assessment of PI-TMA. Early administration of eculizumab may be beneficial in cases of suspected complement abnormalities or suboptimal response to initial treatments. Highlights: This review systematically addresses for the first time the clinical manifestations, treatments, and prognostic outcomes associated with PI-related TMA. Based on existing research, it is proposed that activation of the complement pathway may be the underlying mechanism of PI-TMA, with further elaboration provided. A comprehensive management model for PI-TMA has been proposed. [ABSTRACT FROM AUTHOR]

High-dose melphalan at 200 mg/m2 (MEL-200) is the standard conditioning regimen before autologous stem cell transplantation (ASCT) in patients with multiple myeloma (MM). Busulfan (BU) and cyclophosphamide (CY) can be used as alternatives when MEL is unavailable. However, most studies on BU/CY conditioning regimens were conducted before proteasome inhibitors (PIs) and immunomodulatory drugs (IMIDs) were available. This non-interventional comparative cohort study compared progression-free survival (PFS) between the MEL-200 and BU/CY in patients with MM treated with PIs and/or IMIDs. A total of 137 patients were analyzed (MEL-200,113 patients; BU/CY, 24 patients). The BU/CY group had a higher rate of PI and/or IMID use and very good partial response (VGPR) or complete remission (CR) at ASCT and post-ASCT maintenance. Median PFS was 29.7 and 46.8 months in the MEL-200 and BU/CY groups, respectively. In the multivariate analysis, PFS was significantly better in the BU/CY group. International Staging System Stage I and VGPR or CR at ASCT were significantly associated with longer PFS. No treatment-related mortality was observed in either group by day 100. The BU/CY conditioning regimen may be a viable alternative to the MEL-200 regimen in patients with MM who undergo ASCT after treatment with PIs and/or IMIDs. [ABSTRACT FROM AUTHOR]

Proteasome inhibitors have been applied to anticancer therapy by accumulating toxic misfolded proteins. However, chemical inactivation of proteasome generates aggresome, a Vimentin cage-enclosed subcellular structure quarantining HDAC6-Dynein-transported misfolded proteins before the protein toxicants are degraded by autophagy. Hence, aggresome may attenuate proteasome inhibitor drug-induced cytotoxicity. To solve the problem, it is imperative to characterize how cells assemble aggresome. By examining aggresomes in six cell lines, A549 cells were selectively studied for their bigger cell size and moderate aggresome-forming activity. Aggresome grew in size upon continuous exposure of A549 cells to proteasome inhibitor MG132 and reached a mature size around the 16th to 24th hour of treatment. Mechanistic studies revealed that NF-кB translocated to the nucleus in MG132-treated cells, and chemical activation or knockdown of NF-кB enhanced or prohibited aggresome assembly. Further analyses showed that NF-кB upregulated HDAC6, and HDAC6 maintained the Vimentin cage by interacting with Vimentin p72, a key modification of the intermediate filament contributing to aggresome formation. Remarkably, chemical inactivation of NF-кB synergized MG132-induced cell mortality. All the findings suggest that NF-кB dictates aggresome assembly via upregulating HDAC6, and NF-кB inhibitor may serve as a potential drug potentiating proteasome inhibitor medicine-induced cytotoxicity during the treatment of cancer cells. NEW & NOTEWORTHY: The study reveals a new mechanism guiding MG132-triggered aggresome formation. NF-кB is quickly activated upon exposure to MG132, and NF-кB upregulates the misfolded protein recognizing factor HDCA6. In addition to collecting misfolded proteins, HDAC6 also binds Vimentin and maintains the Vimentin cage, which quarantines toxic misfolded proteins and protects cells from being toxified by those protein toxicants. Therapeutically, chemical inactivation of NF-кB synergizes MG132-induced cytotoxicity, providing a new strategy to defeat cancers. [ABSTRACT FROM AUTHOR]

Minimal residual disease (MRD) is of growing interest in light chain (AL) amyloidosis and is associated with higher rates of cardiac response. A new graded cardiac response criteria has been proposed for better assessment of cardiac improvement. We evaluated MRD status in 63 patients with cardiac AL amyloidosis using next generation flow cytometry (sensitivity ≥ 1*10–5) within four cycles after treatment initiation and cardiac response kinetics. All patients were treated with first-line proteasome inhibitor (100%) and predominantly bortezomib (87.3%). The overall early MRD negative rates were 33.3%. Patients who achieved early MRD negativity were less likely to harbor t(11;14) (21.1% vs 57.5%, P = 0.009). The MRD negative rates amongst patients in hematologic complete response were 66.7% (14/21), and in very good partial response 29.2% (7/24). Early MRD negativity was associated with a higher likelihood of achieving ≥ cardiac partial response (≥ CarPR) (66.7% vs 38.1%, P = 0.032) and ≥ cardiac very good partial response (≥ CarVGPR) (38.1% vs 11.9%, P = 0.023) throughout first-line therapy. The cumulative incidence curve of achieving ≥ CarPR (P = 0.034) and ≥ CarVGPR (P = 0.026) showed significant difference between early MRD negative and positive group. After a median follow-up time of 27.2 months, the median progression free survival was longer in early MRD negative group (not reached vs 31.3 months, P = 0.033). Early MRD eradication in cardiac AL amyloidosis generated deeper and faster cardiac organ response. [ABSTRACT FROM AUTHOR]

Proteasome inhibition emerges as a promising strategy for cancer prevention. PNO1, pivotal for colorectal cancer (CRC) progression, is involved in proteasome assembly in Saccharomyces cerevisiae. Hence, we aimed to explore the role of PNO1 in proteasome assembly and its up- and down-streams in CRC. Here, we demonstrated that PNO1 knockdown suppressed CRC cells growth, proteasome activities and assembly, as well as CDKN1B/p27Kip1 (p27) degradation. Moreover, p27 knockdown partially attenuated the inhibition of HCT116 cells growth by PNO1 knockdown. The up-stream studies of PNO1 identified miR-326 as a candidate miRNA directly targeting to CDS-region of PNO1 and its overexpression significantly down-regulated PNO1 protein expression, resulting in suppression of cell growth, decrease of proteasome activities and assembly, as well as increasing the stability of p27 in CRC cells. These findings indicated that miR-326 overexpression can suppress CRC cell growth, acting as an endogenous proteasome inhibitor by targeting PNO1. [ABSTRACT FROM AUTHOR]

Background: Analysis of existing mutations of Angiotensin-I-Converting Enzyme (ACE) led us to hypothesize that the carriers of damaging ACE mutations (accompanied by low ACE levels) could be at risk for the development of late-onset Alzheimer's disease (AD). Methodology/Principal findings: We quantified blood ACE levels in EDTA-containing plasma from 15 patients with 11 different heterozygous ACE mutations and estimated the effects of these mutations on ACE phenotypes, using a set of mAbs to ACE and two ACE substrates. We confirmed prior observations that the relatively frequent Y215C mutation in the N domain of ACE (present in ~1% of the population) is associated with both Alzheimer's disease (AD) and reduced plasma levels of ACE (~50% of controls), indicating that it likely results in a transport-deficient protein. In addition, we identified another 4 mutations in both ACE domains (M118T, C734Y, V992M and V997M) which are also associated with decreased ACE levels in the blood, and, thus, could be putative risk factors for late-onset AD. One of these mutations, C734Y, is likely transport-deficient, while the other mutations appear to influence ACE catalytic properties. The precipitation of mutant M118T by mAb 2D1 and ACE mutant C734Y by mAb 3F10 increased 2-3-fold compared to native ACE, and therefore, these mAbs could be markers of these mutations. Also, we identified a mutation I989T, which is associated with increased ACE levels in the blood. Conclusions/Significance: Conducting a systematic analysis of blood ACE levels in patients with ACE mutations holds promise for identifying individuals with low blood ACE levels. Such individuals may be at increased risk for late-onset AD. The patients with transport-deficient ACE mutations may benefit from therapeutic treatment with a combination of chemical and pharmacological chaperones and proteasome inhibitors, as was demonstrated previously using a cell model of the transport-deficient ACE mutation, Q1069R [Danilov et al, PLoS One, 2010]. [ABSTRACT FROM AUTHOR]

Inhibitors of the integrated stress response (ISR) have been used to explore the potential beneficial effects of reducing the activation of this pathway in diseases. As the ISR is in essence a protective response, there is, however, a risk that inhibition may compromise the cell's ability to restore protein homeostasis. Here, we show that the experimental compound ISRIB impairs degradation of proteins by the ubiquitin-proteasome system (UPS) during proteotoxic stress in the cytosolic, but not nuclear, compartment. Accumulation of a UPS reporter substrate that is intercepted by ribosome quality control was comparable to the level observed after blocking the UPS with a proteasome inhibitor. Consistent with impairment of the cytosolic UPS, ISRIB treatment caused an accumulation of polyubiquitylated and detergent insoluble defective ribosome products (DRiPs) in the presence of puromycin. Our data suggest that the persistent protein translation during proteotoxic stress in the absence of a functional ISR increases the pool of DRiPs, thereby hindering the efficient clearance of cytosolic substrates by the UPS. Activation of the integrated stress response is important for preserving functionality of the ubiquitin-proteasome system. [ABSTRACT FROM AUTHOR]

The proteasome is a proteolytically active molecular machine comprising many different protein subunits. It is essential for growth and survival in eukaryotic cells and has long been considered a drug target. Here, we summarize the biology of the proteasome, the early research relating to the development of specific proteasome inhibitors (PIs) for treatment of various cancers, and their translation and eventual evolution as exciting therapies for parasitic diseases. We also highlight the development and adaptation of technologies that have allowed for a deep understanding of the idiosyncrasies of individual parasite proteasomes, as well as the preclinical and clinical advancement of PIs with remarkable therapeutic indices. [ABSTRACT FROM AUTHOR]

Simple Summary: Proteasome inhibitors, such as bortezomib and carfilzomib, are vital for treating relapsed or refractory multiple myeloma, but their potential link to cancer therapy-related cardiovascular dysfunction (CTRCD) raises concerns. Bortezomib, a reversible first-generation inhibitor, and carfilzomib, a second-generation irreversible inhibitor, have been associated with hypertension, heart failure, and arrhythmias. This study explored their impact on cardiac function, focusing on left ventricular ejection fraction (LVEF), and vascular function through arterial stiffness and vascular reactivity assessments. Twelve-week-old male mice were treated with either carfilzomib or bortezomib, with some receiving L-NAME to induce hypertension. Echocardiography was used to evaluate cardiac and vascular parameters at different time points, followed by ex vivo arterial stiffness and reactivity measurements. Results showed no significant changes in arterial stiffness at baseline pressures, but a steeper pressure-stiffness curve was noted in carfilzomib-treated normotensive and hypertensive mice. Carfilzomib also showed a trend toward reduced LVEF in hypertensive mice, with bortezomib showing similar trends. Vascular reactivity remained largely unchanged, but proteasome inhibition appeared to enhance endothelial-independent relaxation. Overall, short-term treatment with both drugs was deemed relatively safe under the tested conditions. Proteasome inhibitors such as bortezomib and carfilzomib induce apoptosis and are a cornerstone in the treatment of relapsed or refractory multiple myeloma. However, concerns have emerged concerning their link to cancer therapy-related cardiovascular dysfunction (CTRCD). Bortezomib, a reversible first-generation inhibitor, and carfilzomib, a second-generation irreversible inhibitor, are associated with hypertension, heart failure, and cardiac arrhythmias. The current study investigated the effects of bortezomib and carfilzomib on cardiac (left ventricular ejection fraction, LVEF) and vascular (arterial stiffness, vascular reactivity) function. Cardiac function assessment aimed to build upon existing evidence of proteasome inhibitors CTRCD, while arterial stiffness served as an early indicator of potential vascular remodeling. Groups of 12-week-old C57BL/6J male mice (n = 8 per group) were randomly assigned to receive vehicle, carfilzomib (8 mg/kg I.P.), or bortezomib (0.5 mg/kg I.P.). Additionally, proteasome inhibition was assessed in mice treated with L-NAME (0.5 mg/kg) to induce hypertension. Cardiac and vascular parameters were evaluated via echocardiography on days 0 and 3. On day 6, mice were sacrificed for ex vivo analysis of arterial stiffness and vascular reactivity. Overall, no changes in arterial stiffness were detected either in vivo or ex vivo at basal pressures. However, a steeper pressure–stiffness curve was observed for carfilzomib in normotensive (p < 0.01) and hypertensive (p < 0.0001) mice ex vivo. Additionally, in hypertensive mice, carfilzomib decreased LVEF (p = 0.06), with bortezomib exhibiting similar trends. Vascular reactivity remained largely unchanged, but proteasome inhibition tended to enhance endothelial-independent relaxations in both control and hypertensive mice. In conclusion, short-term treatment with carfilzomib and bortezomib is considered relatively safe for the protocols assessed in the study. [ABSTRACT FROM AUTHOR]

This case report describes the clinical course of a patient with relapsed IgA kappa multiple myeloma with high-risk cytogenetics. Initially treated with daratumumab–bortezomib–lenalidomide–dexamethasone (Dara-VRD) then transitioned to lenalidomide maintenance. However, he experienced a relapse and was treated with carfilzomib-based therapy (CFZ) but developed drug-induced thrombotic microangiopathy (DI-TMA). Despite receiving eculizumab and supportive care, the patient's condition worsened, leading to encephalopathy and refractory gastrointestinal bleeding in the setting of persistent thrombocytopenia. Ultimately, the decision was made to transition to comfort-focused care. DI-TMA has been documented with various proteasome inhibitors such as ixazomib and bortezomib. Additionally, other medications such as cyclosporine, tacrolimus, clopidogrel, ticlopidine, and interferon have been associated with DI-TMA as well (Pisoni et al. (Drug Saf 24:491–501, 2001) [18]). Here we discuss a case of carfilzomib-induced TMA (CFZ-TMA) refractory to eculizumab as well as a review of the published literature. [ABSTRACT FROM AUTHOR]

In the eukaryotic cells, the ubiquitin–proteasome system (UPS) plays a crucial role in the intracellular protein turnover. It is involved in several cellular functions such as the control of the regular cell cycle progression, the immune surveillance, and the homeostasis. Within the 20S proteasome barrel-like structure, the catalytic subunits, β1, β2 and β5, are responsible for different proteolytic activities: caspase-like (C-L), trypsin-like (T-L) and chymotrypsin-like (ChT-L), respectively. The β5 subunit is particularly targeted for its role in antitumor activity: the synthesis of β5 subunit inhibitors could be a promising strategy for the treatment of solid and hematologic tumors. In the present work, we performed two combination studies of AM12, a recently developed synthetic proteasome inhibitor, with curcumin and quercetin, two nutraceuticals endowed of many pharmacological properties. We measured the combination index (CI), applying the Chou and Talalay method, comparing the two studies, from 50% to 90% of proteasome inhibition. In the case of the combination AM12 + curcumin, an increasing synergism was observed from 50% to 90% of proteasome inhibition, while in the case of the combination AM12 + quercetin an additive effect was observed only from 50% to 70% of β5 subunit inhibition. These results suggest that combining AM12 with curcumin is a more promising strategy than combining it with quercetin for potential therapeutic applications, especially in treating tumors. [ABSTRACT FROM AUTHOR]

Lipocalin-2 (Lcn2), a protein secreted by immune-activated cells, including reactive astrocytes, is detrimental to the brain and induces neurodegeneration. We previously showed that Lcn2 levels are reduced in primary mouse astrocytes after treatment with the proteasome inhibitor bortezomib (BTZ). However, it remains unknown whether a decrease in Lcn2 levels after BTZ treatment can also be observed in vivo and whether it reduces neurotoxicity during lipopolysaccharide (LPS)-induced systemic inflammation in vivo. To answer these questions, we performed LPS challenge experiments by intraperitoneal injection in mice and found that Lcn2 levels were significantly increased in the brain, recapitulating in vitro experiments using astrocytes. Co-administration of LPS and BTZ reduced the Lcn2 levels compared to the levels in LPS-treated controls. Upon LPS challenge, the expression levels of glial marker genes were upregulated in the mouse brain. Of note, this upregulation was hampered by the co-administration of BTZ. Taken together, our results suggested that BTZ can reduce LPS-induced Lcn2 levels and may alleviate LPS-induced neuroinflammation and neurotoxicity in mice. [ABSTRACT FROM AUTHOR]

Trichomonas vaginalis is the causative agent of the common sexually transmitted disease, trichomoniasis, which affects more than a hundred million people worldwide. Metronidazole and tinidazole, agents belonging to the 5-nitroheterocyclic class of antimicrobials, are most often used to treat infection, but increased resistance has been reported and adverse effects of these drugs can be significant. Consequently, an urgent need exists for the development of novel drug entities against trichomoniasis. Critical for antimicrobial drug development is the demonstration of in vivo efficacy. Murine models of vaginal T. vaginalis infection are unreliable for unknown reasons. Meanwhile, murine infections with the related bovine pathogen, Tritrichomonas foetus, tend to be more robust, although susceptibility to different antimicrobials might differ from T. vaginalis. Here, we explored the utility of T. foetus infection as a surrogate model for drug development against T. vaginalis. Four different T. foetus strains caused robust vaginal infection in young mice, while none of four diverse T. vaginalis strains did. Comparison of drug susceptibility profiles revealed that T. foetus and T. vaginalis were similarly susceptible to a range of 5-nitroheterocyclic and gold(I) compounds. By comparison, proteasome inhibitors were 10- to 15-fold less active against T. foetus than T. vaginalis, although one of the proteasome inhibitors, bortezomib, had low micromolar activity or better against multiple strains of both trichomonads. Different strains of T. foetus were used to demonstrate the utility of the murine vaginal infection models for in vivo efficacy testing, including for bortezomib and a gold(I) compound. The differences in susceptibility to proteasome inhibitors may be partially explained by differences in the proteasome subunit sequences between the two trichomonads, although the functional relevance of the proteasome was similar in both organisms. These findings indicate that T. foetus can serve as a reliable surrogate model for T. vaginalis in vitro and in murine infections in vivo, but caution must be exercised for specific drug classes with targets, such as the proteasome, that may display genetic divergence between the trichomonads. [ABSTRACT FROM AUTHOR]

Developing resistance to anticancer drugs complicates the clinical treatment of multiple myeloma patients. Previous studies revealed a link between the unfolded protein response (UPR) and miRNAs with acquired drug resistance. This study aimed to determine the expression profile of XBP1, hsa-miR-34c-5p, hsa-miR-214, and hsa-miR-30c-2* in resistant and sensitive multiple myeloma cell lines to a proteasome inhibitor, bortezomib. After establishing bortezomib-resistant cells, the expression level of XBP1, hsa-miR-214, hsa-miR-34c-5p, and hsa-miR-30c-2* in both cell lines were assessed by qRT-PCR. Hsa-miR-34c-5p was suppressed to study its effect on the expression profile of Bax/Bcl-2. Statistical analysis was done by t-test in two clinically resistant and sensitive cells to bortezomib. MTT assay confirmed the creation of the resistant cell line. The qRT-PCR screening showed a significant difference between XBP1 and miR-34c-5p levels in resistant and sensitive cells. Following hsa-miR-34c-5p blockage, while Bax was overexpressed, Bcl-2 expression was reduced in the resistant cell line, overcoming cells resistant to bortezomib. Our findings demonstrate miR-34c-5p is differentially expressed between bortezomib-sensitive and -resistant MM cells. Inhibiting miR-34c-5p re-sensitized resistant cells to bortezomib by modulating Bax/Bcl-2 expression, suggesting this miRNA regulates apoptosis and drug resistance and may be a promising therapeutic target for overcoming proteasome inhibitor resistance in MM. [ABSTRACT FROM AUTHOR]

Objective: In this study, we aimed to obtain real-life data on the use of antimyeloma agents, which significantly increase overall survival (OS) in multiple myeloma (MM) patients, in primary plasma cell leukemia (pPCL) patients with poor prognosis. Materials and Methods: Data from 53 patients who were diagnosed with pPCL between 2011 and 2020 and who used at least one proteasome inhibitor (PI) and/or immunomodulatory (IMID) agent were analyzed retrospectively. Depending on the year of the pPCL diagnosis, 20% leukocytes or ≥2x109/L plasma cells in the peripheral blood was used as a diagnostic criterion. Results: The median age of the patients was 58 years and 23 (43.4%) patients were over 65 years of age. For first-line treatment, PI or IMID alone was used by 31 (58.5%) patients, and PI and IMID were used simultaneously by 15 (28.3%) patients. Additionally, 21 (39.6%) patients received transplantation and 13 (24.5%) patients received maintenance treatment. The median progression-free survival was 4 (range: 1-42) months. When patients whose primary disease was refractory to first-line therapy were excluded, the duration of treatment was 6.5 months. The median OS was 15 months with a median follow-up duration of 15 months. Only 7 (13.2%) of the patients were alive at the last follow-up visit. Those with higher β2- microglobulin levels and International Staging System stage 3 and non-transplant patients receiving first-line treatment had shorter OS (p=0.005, p=0.02, and p=0.008, respectively). The concomitant use of PIs and IMIDs, the addition of chemotherapy to induction therapy, and the response to induction therapy or maintenance therapy did not affect OS. Conclusion: In this study, as in previous similar studies, we could not see an increased survival trend in pPCL, which is observed in MM. New studies are needed for pPCL, which is likely to increase with new diagnostic criteria, based on current agents and information from MM.

BACKGROUND: Economic differences among currently available proteasome inhibitors (PI)-based lenalidomide-dexamethasone (Rd)-backbone triplet regimens-ixazomib (I), bortezomib (V), and carfilzomib (K) plus Rd-remain poorly understood. OBJECTIVE: To assess health care resource utilization (HCRU) and health care costs of patients with relapsed/refractory multiple myeloma (RRMM) in the United States treated with IRd, VRd, and KRd. METHODS: This retrospective longitudinal cohort study using IQVIA PharMetrics Plus adjudicated claims US data (January 1, 2015, to September 30, 2020) included adult patients with all available data who initiated IRd, VRd, or KRd in second line of therapy or later (LOT2+) on or after September 1, 2015. The index date was the treatment initiation date for each LOT (multiple LOTs per patient were included) and the baseline was 6 months pre-index. MM-related and all-cause HCRU/costs were assessed during follow-up and reported per patient per month (PPPM; 2020 US Dollars). For MM-related costs only, treatment administration costs were excluded from outpatient (OP) costs and instead summed with pharmacy costs. HCRU/costs were compared between treatment groups using generalized linear models (GLMs). Cost variables were compared using 2-part models and GLM with log transformation and γ distribution. Inverse probability of treatment weighting (IPTW) adjusted for imbalance of baseline confounders across treatment groups. RESULTS: The study included 511 patients contributing 542 LOTs (IRd: n = 153; VRd: n = 262; KRd: n = 127). Before IPTW, mean observed time spent on therapy was 8.5, 9.3, and 7.3 months for the IRd, VRd, and KRd cohorts, respectively. During follow-up and after IPTW, IRd and VRd were associated with significantly fewer OP visits vs KRd. Post-IPTW comparisons of MM-related costs for IRd vs KRd yielded lower OP costs for IRd (mean diff. PPPM: -$3,428; P < 0.001), contributing to lower total medical costs (-$3,813; P < 0.001) and total health care cost savings with IRd vs KRd (-$5,813; P = 0.001). MM-related OP costs were lower for VRd (mean diff. PPPM: -$3,543; P < 0.001) than KRd, reducing its total MM-related medical costs (-$3,997; P = 0.002), leading to total MM-related health care cost savings with VRd vs KRd (-$12,357; P < 0.001). All-cause cost comparisons yielded similar results (total health care cost savings for IRd and VRd vs KRd: -$6,371 and -$13,629, respectively; all P < 0.001). CONCLUSIONS: From the US insurance-payer perspective, patients treated with IRd and VRd had significant medical cost savings vs KRd due to lower OP costs when excluding treatment administration costs. The differential economic impacts of PI-Rd regimens in this study may help to inform treatment decisions for patients with MM. DISCLOSURES: This study and article were supported by Takeda Development Center Americas, Inc. Dr Sanchez has no conflicts to declare. Dr Chari has the following relationships: Research Support/Principal Investigator: Amgen, Array Biopharma, Celgene, Glaxo Smith Klein, Janssen, Millenium/Takeda, Novartis Pharmaceuticals, Oncoceutics, Pharmacyclics, Seattle Genetics; Consultant: Amgen, Bristol-Myers Squibb, Celgene, Millenium/Takeda, Janssen, Karyopharm; Scientific Advisory Board: Amgen, Celgene, Millenium/Takeda, Janssen, Karyopharm, Sanofi, Seattle Genetics. Drs Cherepanov, Huang, Dabora, and Noga are current employees of Takeda, while Drs Stull and Young are ex-employees of Takeda; Drs Cherepanov and Huang also own stocks in Takeda. Dr DerSarkissian, Ms Cheng, Ms Zhang, Mr Banatwala, and Dr Duh are employees of Analysis Group, Inc. (AG), a consulting firm that received funding from Takeda to conduct this study. Ms Pi was an employee of AG at the time of the study. Dr Ailawadhi has the following relationships to declare: Research Support and Consulting for BMS, GSK, and Janssen; Research Support from AbbVie, Arch Oncology, Cellectar, Medimmune, Pharmacyclics, and Xencor; Consulting for Beigene, Oncopeptides, Regeneron, Sanofi, and Takeda.

Multiple myeloma (MM) is a cancer of terminally differentiated plasma cells. MM remains incurable, but overall survival of patients has progressively increased over the past two decades largely due to novel agents such as proteasome inhibitors (PI) and the immunomodulatory agents. While these therapies are highly effective, MM patients can be de novo resistant and acquired resistance with prolonged treatment is inevitable. There is growing interest in early, accurate identification of responsive versus non-responsive patients; however, limited sample availability and need for rapid assays are limiting factors. Here, we test dry mass and volume as label-free biomarkers to monitor early response of MM cells to treatment with bortezomib, doxorubicin, and ultraviolet light. For the dry mass measurement, we use two types of phase-sensitive optical microscopy techniques: digital holographic tomography and computationally enhanced quantitative phase microscopy. We show that human MM cell lines (RPMI8226, MM.1S, KMS20, and AMO1) increase dry mass upon bortezomib treatment. This dry mass increase after bortezomib treatment occurs as early as 1 h for sensitive cells and 4 h for all tested cells. We further confirm this observation using primary multiple myeloma cells derived from patients and show that a correlation exists between increase in dry mass and sensitivity to bortezomib, supporting the use of dry mass as a biomarker. The volume measurement using Coulter counter shows a more complex behavior; RPMI8226 cells increase the volume at an early stage of apoptosis, but MM.1S cells show the volume decrease typically observed with apoptotic cells. Altogether, this cell study presents complex kinetics of dry mass and volume at an early stage of apoptosis, which may serve as a basis for the detection and treatment of MM cells.

Isatuximab, a novel anti‐CD38 monoclonal antibody, is approved in combination with carfilzomib and dexamethasone (Isa‐Kd) in relapsed/refractory multiple myeloma (RRMM) patients. Because of its recent introduction, real‐world efficacy and safety are poorly reported. In this Italian multicenter real‐life observational retrospective study, efficacy and safety of the Isa‐Kd regimen were evaluated in a cohort of 103 RRMM patients. Overall response rate (ORR) was 85%, with stringent (sCR) or complete response (CR) in 18% of cases and very good partial response (VGPR) in 39%. Median PFS and OS were not reached within the study period, while 1‐year PFS and OS were 72% and 77%, respectively. Hematological toxicities were observed in 42% of subjects, and cardiac toxicities occurred in 24% of cases. Moreover, we conducted a subanalysis on patients (N = 69) treated with Isa‐Kd after one prior line of therapy, showing an ORR of 88%, with sCR + CR in 20% of subjects, VGPR in 46%, and PR in 22% of patients. In this group, median PFS and OS were not reached, while 1‐year PFS and OS were 92% and 95%, respectively. In conclusions, our study confirmed Isa‐Kd as an effective treatment option for RRMM with a manageable safety profile even in real‐life settings. [ABSTRACT FROM AUTHOR]

Epstein-Barr virus (EBV) manipulates the ubiquitin-proteasome system and regulators of Bcl-2 family to enable the persistence of the virus and survival of the host cells through the expression of viral proteins in distinct latency patterns. We postulate that the combination of bortezomib (proteasome inhibitor) and venetoclax (Bcl-2 inhibitor) [bort/venetoclax] will cause synergistic killing of post-transplant lymphoproliferative disorder (PTLD) through targeting the pro-survival function of latent viral proteins such as latent membrane protein-1 (LMP-1) and EBV nuclear antigen-3C (EBNA-3C). Bort/venetoclax could synergistically kill spontaneous lymphoblastoid cell lines (sLCLs) derived from patients with PTLD and EBV-associated hemophagocytic lymphohistiocytosis by inducing DNA damage response, apoptosis and G1-S cell cycle arrest in a ROS-dependent manner. Bortezomib potently induced the expression of Noxa, a pro-apoptotic initiator and when combined with venetoclax, inhibited Mcl-1 and Bcl-2 simultaneously. Bortezomib prevented LMP-1 induced proteasomal degradation of IκBα leading to the suppression of the NF-κB signaling pathway. Bortezomib also rescued Bcl-6 from EBNA-3C mediated proteasomal degradation thus maintaining the repression of cyclin D1 and Bcl-2 causing G1-S arrest and apoptosis. Concurrently, venetoclax inhibited Bcl-2 upregulated by either LMP-1 or EBNA-3C. Bort/venetoclax decreased the expression of phosphorylated p65 and Bcl-2 at serine 70 thereby suppressing the NF-κB signaling pathway and promoting apoptosis, respectively. These data corroborated the marked suppression of the growth of xenograft of sLCL in SCID mice (p<0.001). Taken together, the combination of bortezomib and venetoclax targets the pro-survival function of LMP-1 and EBNA-3C of Epstein-Barr virus in spontaneous lymphoblastoid cell lines. Author summary: Epstein-Barr virus (EBV) is an oncogenic virus associated with different cancers and can directly drive the development of a lymphoma type condition in organ transplant patients known as post-transplant lymphoproliferative disorder (PTLD) as a result of weakened immune surveillance of EBV. The malignant spectrum of PTLD will require multi-agent chemotherapy regimen which is not well tolerated by the immunocompromised patients. Here, we combined bortezomib (proteasome inhibitor) with venetoclax (Bcl-2 inhibitor) as a novel strategy to target the pro-survival function of key EBV onco-proteins, namely LMP-1 and EBNA-3C. We demonstrated that this chemotherapy-free regimen could be highly effective in killing patient-derived spontaneous lymphoblastoid cell lines which represent cell models of PTLD. The novel drug regimen can avoid the toxic effects of chemotherapy and may have high efficacy and specificity for the treatment of PTLD. [ABSTRACT FROM AUTHOR]

Background: Relapsed and refractory acute myeloid leukemia (AML) carries a dismal prognosis. CAR T cells have shown limited efficacy in AML, partially due to dysfunctional autologous T cells and the extended time for generation of patient specific CAR T cells. Allogeneic NK cell therapy is a promising alternative, but strategies to enhance efficacy and persistence may be necessary. Proteasome inhibitors (PI) induce changes in the surface proteome which may render malignant cells more vulnerable to NK mediated cytotoxicity. Here, we investigated the potential benefit of combining PIs with CAR-expressing allogeneic NK cells against AML. Methods: We established the IC50 concentrations for Bortezomib and Carfilzomib against several AML cell lines. Surface expression of class-I HLA molecules and stress-associated proteins upon treatment with proteasome inhibitors was determined by multiparameter flow cytometry. Using functional in vitro assays, we explored the therapeutic synergy between pre-treatment with PIs and the anti-leukemic efficacy of NK cells with or without expression of AML-specific CAR constructs against AML cell lines and primary patient samples. Also, we investigated the tolerability and efficacy of a single PI application strategy followed by (CAR-) NK cell infusion in two different murine xenograft models of AML. Results: AML cell lines and primary AML patient samples were susceptible to Bortezomib and Carfilzomib mediated cytotoxicity. Conditioned resistance to Azacitidine/Venetoclax did not confer primary resistance to PIs. Treating AML cells with PIs reduced the surface expression of class-I HLA molecules on AML cells in a time-and-dose dependent manner. Stress-associated proteins were upregulated on the transcriptional level and on the cell surface. NK cell mediated killing of AML cells was enhanced in a synergistic manner. PI pre-treatment increased effector-target cell conjugate formation and Interferon-γ secretion, resulting in enhanced NK cell activity against AML cell lines and primary samples in vitro. Expression of CD33- and CD70-specific CARs further improved the antileukemic efficacy. In vivo, Bortezomib pre-treatment followed by CAR-NK cell infusion reduced AML growth, leading to prolonged overall survival. Conclusions: PIs enhance the anti-leukemic efficacy of CAR-expressing allogeneic NK cells against AML in vitro and in vivo, warranting further exploration of this combinatorial treatment within early phase clinical trials. [ABSTRACT FROM AUTHOR]

Objective: To mine and analyze adverse events (AEs) related to proteasome inhibitors in multiple myeloma based on the FDA Adverse Event Reporting System (FAERS), providing references for rational clinical medication. Methods: AE data related to multiple myeloma proteasome inhibitors were collected from the FAERS from the first quarter of 2010 to the first quarter of 2024. Signal mining of AEs was conducted using the reporting odds ratio method and Bayesian confidence propagation neural network method. Results: A total of 8,805 reports for bortezomib, 5,264 for carfilzomib, and 8,771 for ixazomib were collected, with corresponding AE signals of 474, 279, and 287, respectively, involving 23, 21, and 22 System Organ Classes (SOCs). The report information for the three drugs tended to be consistent: more cases were reported in males than in females; the majority of patients were 65 years and over; AEs mostly occurred within 6 months of medication; the outcomes primarily consisted of hospitalization, prolonged hospital stay, and other serious adverse events; the primary reporting country was the United States. The most affected SOCs were infections and infestations, general disorders and administration site conditions, and blood and lymphatic system disorders. Conclusion: The overall distribution of AEs for the three multiple myeloma proteasome inhibitors was consistent, but there were certain differences in specific AE signal characteristics, which should be noted in clinical applications. [ABSTRACT FROM AUTHOR]

Plasma cell‐rich acute rejection (PCAR), a relatively rare subtype of acute allograft rejection, is usually associated with a significantly lower treatment response rate and a higher graft failure rate. PCAR is characterized by the presence of more than 10% of plasma cells out of all graft infiltrating cells, with approximately 40%–60% of PCAR resulting in graft failure within a year. Currently, there is no gold standard for the effective treatment of PCAR. This case report demonstrates the potential treatment effect of bortezomib in PCAR. A 37‐year‐old woman with reflux nephropathy received a kidney transplant from a brain‐dead kidney donor. The patient presented with an acute kidney injury with a serum creatinine level over 4 mg/dL 4 months after the surgery. The allograft biopsy showed acute T cell–mediated rejection (TCMR), Grade IIA, plasma cell‐rich variant. There were diffuse polyclonal plasma cells infiltrating the renal parenchyma with marked tubulitis and focal endarteritis. She received a methylprednisolone pulse of 500 mg daily x3, followed by thymoglobulin (rATG) at 4.2 mg/kg. However, a repeated biopsy after 2 months showed persistent plasma cells infiltrate with increased interstitial fibrosis with tubular atrophy. Then, the patient was given one cycle of bortezomib with a total of four subcutaneous injections and continued immunosuppressants of tacrolimus, mycophenolate mofetil, and prednisone. Following the treatment, the patient's serum creatinine level trended down to 2 mg/dL, and a second repeat biopsy after 4 months showed a significant treatment effect with complete resolution of interstitial inflammation and decreased chronicity. Bortezomib is a proteasome inhibitor that prevents cell proliferation by inducing apoptosis in plasma cells and has shown great promise as a therapeutic agent for multiple myeloma. Our case suggests that bortezomib can also be used as a potential therapeutic intervention for patients with PCAR. [ABSTRACT FROM AUTHOR]

The treatment paradigm of multiple myeloma (MM) has shifted in the past years, as continuous therapy is becoming the standard of care for both newly diagnosed and relapsed patients. Although it is indisputable that continuous therapy has added a great benefit on the progression-free as well as overall survival, it is still unclear what the patients' perspective is on this therapeutic approach. Methods: This study included 155 adult MM patients from Fundeni Clinical Institute in Romania, receiving continuous therapy with daratumumab, proteasome inhibitors, immunomodulators, or bi-specific antibodies. The patients had varied economic, social, and educational backgrounds. We developed a questionnaire to interrogate the quantitative and qualitative effect of the therapy on the patients' personal and professional life and to identify the side effects that had the strongest impact on their quality of life. Results: 74.83% of the patients reported that the treatment they received negatively impacted their quality of life. Among them, 40% considered that the most detrimental aspects of the therapy are the financial burden and the negative impact on their professional life. One-third of the patients reported that the therapy negatively impacted their personal life and that it had a deleterious effect on their relationship with their partner and family members. In terms of the side effects experienced, patients considered that tiredness was the main factor causing a decrease in their quality of life, followed by insomnia and bone pain. Despite this, almost none of the patients considered dropping the therapy, and almost half of the patients considered that the frequent visits to the hospital offered them psychological comfort. In addition, more than 70% of the patients declared that they were afraid to stop the therapy if given the choice, with the main concerns being the fear of an early relapse. Conclusions: Although continuous therapy is associated with a high financial burden and a negative impact on both professional and personal life, the frequent visits to the hospital appear to be reassuring. Moreover, the patients would not opt for treatment discontinuation and felt safer when monitored frequently. [ABSTRACT FROM AUTHOR]

The present study provides real-world evidence on the treatment of multiple myeloma (MM) bone disease with various bisphosphonates combined for different myeloma-specific treatments as no validated data regarding the best combination treatment for bone disease associated with MM are available. We examined retrospectively 345 MM patients treated with autologous stem cell transplantation in Finland during 1996–2020. The median age of the patients was 60 years with a median follow-up time of 50 months (1–339). At diagnosis, 72.1% of the patients had myeloma-associated bone disease and 45.8% had fractures. Most patients (58.8%) received proteasome inhibitor (PI)-containing treatment at first line. MM bone disease was treated in 91.6% of the patients; 49.9% received zoledronic acid (ZA) and 29.9% pamidronate. Inferior overall survival was associated with MM bone disease at diagnosis (p = 0.005) or a fracture at diagnosis (p = 0.003). A later fracture was identified in 29% of the patients, and in those patients without MM bone disease at diagnosis later fractures were less common after ZA treatment (p = 0.049). PI-based treatment plus ZA (p = 0.019) seemed to be the best combination to prevent later fractures, even though the same patient subgroup was more likely to experience relapse (p = 0.018), and also when excluding patients with previous induction therapy without novel agents (p = 0.008). To conclude, this study suggests that the best therapy to prevent later fractures in MM might be PI-based treatment combined with ZA. [ABSTRACT FROM AUTHOR]

In this review we explore innovative approaches in the treatment of hematologic cancers by combining various therapeutic modalities. We discuss the synergistic potential of combining inhibitors targeting different cellular pathways with immunotherapies, molecular therapies, and hormonal therapies. Examples include combining PI3K inhibitors with proteasome inhibitors, NF-κB inhibitors with immunotherapy checkpoint inhibitors, and neddylation inhibitors with therapies targeting the tumor microenvironment. Additionally, we discuss the potential use of small molecules and peptide inhibitors in hematologic cancer treatment. These multidimensional therapeutic combinations present promising strategies for enhancing treatment efficacy and overcoming resistance mechanisms. However, further clinical research is required to validate their effectiveness and safety profiles in hematologic cancer patients. [ABSTRACT FROM AUTHOR]

Simple Summary: Multiple myeloma is a heterogenous hematological condition with a worldwide incidence of between 0.5–5 per 100,000. The standard front-line treatment algorithm suggests induction chemotherapy, early stem cell transplant in those eligible, followed by maintenance chemotherapy. Unfortunately, most people subsequently relapse and require salvage treatment. A wide array of options are available in the management of relapse/refractory multiple myeloma. Our review aims to describe the new, approved treatment modalities in this setting while describing data on the efficacy, tolerability, and nuances of each available option. Multiple myeloma is a heterogeneous condition characterized by the proliferation of monoclonal B-cells, for which there is currently no curative treatment available. Relapses are, unfortunately, common after first-line treatment. While the prognosis for relapsed refractory multiple myeloma is generally poor, advances in the treatment of relapsed or refractory multiple myeloma offer hope. However, the expansion of effective options in targeted treatment offers renewed optimism and hope that patients who fail on older therapies may respond to newer modalities, which are often used in combination. We review currently approved and novel investigational agents classified by mechanisms of action, efficacy, approved setting, and adverse events. We delve into future directions of treatment for relapsed/refractory multiple myeloma, reviewing novel agents and therapeutic targets for the future. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is a prevalent yet incurable hematologic malignancy. Despite the proven efficacy of proteasome inhibitors in treating MM, resistance to Bortezomib-based treatments persists in a subset of patients. This case control study explores the potential of circulating endothelial progenitor cells (EPCs) as biomarkers for predicting response to Proteasome Inhibitor based therapy combined with Dexamethasone in MM patients. This study was conducted on 105 MM patients receiving bortezomib plus dexamethasone therapy and 90 healthy individuals as a control group. Utilizing 8-color multi-parameter flow cytometry, we assessed the levels of circulating EPCs, identified through CD34 FITC and CD309 PE markers at diagnosis and after one treatment cycle (4 weeks). Our findings revealed that patients exhibiting poor response to therapy showed significantly higher CD34/CD309 values than those with a good response (p < 0.001). The delineation of response based on CD34/CD309 expression was established with a cutoff ≤ 0.9 for percentage (yielding 100% sensitivity and 94.1% specificity) and ≤ 12.5 for absolute value (also with 100% sensitivity and 94.1% specificity). These results underscore the potential of EPC population levels, as quantified by CD34/CD309, to serve as a predictive biomarker for immunomodulatory treatment in MM patients undergoing Proteasome Inhibitor and Dexamethasone therapy. [ABSTRACT FROM AUTHOR]

Canine urothelial carcinoma (cUC) is one of the most malignant tumors affecting dogs; however, its proliferative mechanism is yet to be fully elucidated. The ubiquitin-proteasome system (UPS) is an important metabolic pathway regulating protein degradation, and its dysfunction leads to apoptosis. We investigated the antitumor effect of the proteasome inhibitor bortezomib, which blocks the UPS. Bortezomib inhibited cell growth in cUC cell lines by inducing apoptosis in vitro. These findings suggest the potential of bortezomib as a novel therapeutic drug for dogs with cUC. [ABSTRACT FROM AUTHOR]

Because hepatic stellate cells (HSCs) play a major role in fibrosis, we focused on HSCs as a potential target for the treatment of liver fibrosis. In this study, we attempted to identify drug candidates to inactivate HSCs and found that several proteasome inhibitors (PIs) reduced HSC viability. Our data showed that a second-generation PI, carfilzomib (CZM), suppressed the expression of fibrotic markers in primary murine HSCs at low concentrations of 5 or 10 nM. Since CZM was not toxic to HSCs up to a concentration of 12.5 nM, we examined its antifibrotic effects further. CZM achieved a clear reduction in liver fibrosis in the carbon tetrachloride (CCl4)-induced mouse model of liver fibrosis without worsening of liver injury. Mechanistically, RNA sequence analysis of primary HSCs revealed that CZM inhibits mitosis in HSCs. In the CCl4-injured liver, amphiregulin, which is known to activate mitogenic signaling pathways and fibrogenic activity and is upregulated in murine and human metabolic dysfunction-associated steatohepatitis (MASH), was downregulated by CZM administration, leading to inhibition of mitosis in HSCs. Thus, CZM and next-generation PIs in development could be potential therapeutic agents for the treatment of liver fibrosis via inactivation of HSCs without liver injury. [ABSTRACT FROM AUTHOR]

Proteasome inhibitors (PIs), bortezomib, carfilzomib, and ixazomib, are the first-line treatment for multiple myeloma (MM). They inhibit cytosolic protein degradation in cells, which leads to the accumulation of misfolded and malfunctioned proteins in the cytosol and endoplasmic reticulum, resulting in cell death. Despite being a breakthrough in MM therapy, malignant cells develop resistance to PIs via different mechanisms. Understanding these mechanisms drives research toward new anticancer agents to overcome PI resistance. In this review, we summarize the mechanism of action of PIs and how MM cells adapt to these drugs to develop resistance. Finally, we explore these mechanisms to present strategies to interfere with PI resistance. The strategies include new inhibitors of the ubiquitin–proteasome system, drug efflux inhibitors, autophagy disruption, targeting stress response mechanisms, affecting survival and cell cycle regulators, bone marrow microenvironment modulation, and immunotherapy. We list potential pharmacological targets examined in in vitro, in vivo, and clinical studies. Some of these strategies have already provided clinicians with new anti-MM medications, such as panobinostat and selinexor. We hope that further exploration of the subject will broaden the range of therapeutic options and improve patient outcomes. [ABSTRACT FROM AUTHOR]

Proteasome inhibition interfering with many cell signaling pathways has been extensively explored as a therapeutic strategy for cancers. Proteasome inhibitor YSY01A is a novel agent that has shown remarkable anti-tumor effects; however, its mechanisms of action are not fully understood. Here we report that YSY01A is capable of suppressing cancer cell survival by induction of apoptosis. Paradoxically, we find that YSY01A abrogates constitutive activation of STAT3 via proteasome-independent degradation of gp130 and JAK2, but not transcriptional regulation, in human A549 non-small cell lung cancer cells. The reduction in gp130 and JAK2 can be restored by co-treatment with 3-methyladenine, an early-stage autophagy lysosome and type I/III PI3K inhibitor. YSY01A also effectively inhibits cancer cell migration and lung xenograft tumor growth with little adverse effect on animals. Thus, our findings suggest that YSY01A represents a promising candidate for further development of novel anticancer therapeutics targeting the proteasome. [ABSTRACT FROM AUTHOR]

Computational pharmacogenomics can potentially identify new indications for already approved drugs and pinpoint compounds with similar mechanism-of-action. Here, we used an integrated drug repositioning approach based on transcriptomics data and structure-based virtual screening to identify compounds with gene signatures similar to three known proteasome inhibitors (PIs; bortezomib, MG-132, and MLN-2238). In vitro validation of candidate compounds was then performed to assess proteasomal proteolytic activity, accumulation of ubiquitinated proteins, cell viability, and drug-induced expression in A375 melanoma and MCF7 breast cancer cells. Using this approach, we identified six compounds with PI properties ((-)-kinetin-riboside, manumycin-A, puromycin dihydrochloride, resistomycin, tegaserod maleate, and thapsigargin). Although the docking scores pinpointed their ability to bind to the β5 subunit, our in vitro study revealed that these compounds inhibited the β1, β2, and β5 catalytic sites to some extent. As shown with bortezomib, only manumycin-A, puromycin dihydrochloride, and tegaserod maleate resulted in excessive accumulation of ubiquitinated proteins and elevated HMOX1 expression. Taken together, our integrated drug repositioning approach and subsequent in vitro validation studies identified six compounds demonstrating properties similar to proteasome inhibitors. [ABSTRACT FROM AUTHOR]

Background: Bortezomib (BTZ), a primary treatment for MM, but its effectiveness can be reduced by interactions with vicinal diol moieties (VDMs) in polyphenols. Despite this, it's debated whether BTZ therapy necessitates avoiding polyphenolrich products, given the low bioavailability of polyphenols. Additionally, it remains unclear whether the structure of polyphenols contributes to their BTZ antagonism. Therefore, our study aims to unravel the structure-activity relationship of dietary polyphenols and their BTZ antagonism at daily dietachievable physiological concentrations. Methods: We assessed the antagonistic effects of 25 polyphenols against BTZ using cell viability assays in RPMI 8226 cells. ChemGPS-NP helped analyze the structural similarity. Additionally, long-term cytotoxicity assays evaluated these effects at physiologically relevant concentrations. Results: By cell viability assays, we found a positive correlation between the number of VDMs in gallotannins and their BTZ antagonism. Moreover, the origin and configuration of VDMs, rather than the total VDM concentration, play a pivotal role in the combined antagonistic effects against BTZ in gallotannins. Additionally, ChemGPS-NP analysis indicated that the aromaticity and C-3 hydroxyl group in flavonoids' C-rings enhance their BTZ antagonism. Finally, long-term cytotoxicity assays reveal that gallic acid (GA), epigallocatechin (EGC), and epigallocatechin gallate (EGCG), at their physiological concentrations--attainable through tea consumption--significantly and synergistically antagonize BTZ. Conclusion: Due to the potential for these polyphenols to reduce the effectiveness of BTZ, it is advisable for MM patients undergoing BTZ treatment to reduce their consumption of foods high in VDM-containing polyphenols. [ABSTRACT FROM AUTHOR]

Background and Aims: Poor prognosis in lung cancer is associated with tumor-associated macrophages (TAMs) that exhibit M2clike behaviours that support multiple pathways in the tumour microenvironment. The interplay between epigenetic modifications and the ubiquitin-proteasome pathway involves three key mechanisms: regulation of epigenetic enzymes by ubiquitin, interaction between ubiquitin and epigenetic modifiers, and epigenetic silencing of critical genes involved in cellular processes. Therefore, we investigated the effects of ixazomib, a proteasome inhibitor, on gene expression changes in epigenetic chromatin modification enzymes in a co-culture of M2c macrophages and A549 lung cancer cells. Methods: The IC50 concentration of ixazomib was determined to be 2.19 μM using a real-time cell analyser. THP-1 monocytes were polarised into M0 macrophages with 100 ng/mL phorbol 12-myristate 13-acetate (PMA), rested, and then exposed to 1 mM hydrocortisone to become M2c macrophages. A549 cells were seeded in the lower chamber of a co-culture plate. M2c macrophages were then co-cultured with A549 cells for 24 h with or without 2.19 μM ixazomib. After being isolated, mRNA was converted to cDNA and analysed using a gene panel with RT-PCR. Results: The findings showed that 56 genes had exceptionally high expression levels (up to 1848-fold. Ixazomib downregulated these overexpressed genes. Conclusion: Ixazomib effectively modulates the expression of genes involved in epigenetic chromatin modification in the lung cancer microenvironment, indicating its utility in lung cancer therapy. Further studies are needed to explore the combined use of epigenetic drugs and proteasome inhibitors. [ABSTRACT FROM AUTHOR]