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Improvement of outcome in patients with pancreatic ductal adenocarcinoma (PDAC) requires exploration of novel therapeutic targets. Thus far, most studies of PDAC therapies, including those inhibiting small ubiquitin-like modifications (SUMOylation), have focused on PDAC epithelial cell biology, yet SUMOylation occurs in a variety of cell types. The mechanisms by which SUMOylation impacts PDAC in the context of its tumor microenvironment are poorly understood. We used clinically relevant orthotopic PDAC mouse models to investigate the effect of SUMOylation inhibition using a specific, clinical-stage compound, TAK-981. In contrast to its inhibition of PDAC cell proliferation in vitro, the survival benefit conferred by TAK-981 in vivo is dependent on the presence of T cells, suggesting that induction of adaptive antitumor immunity is an important antitumor effect of SUMOylation inhibition in vivo. To understand how this adaptive antitumor immunity is promoted, we investigated how SUMOylation inhibition in vivo alters major cell types/subtypes and their communications in the PDAC tumor microenvironment by performing transcriptomic analyses at single-cell resolution, which allowed mapping of cells in our orthotopic mouse model to cells in human PDAC tumors based on gene expression profiles. Findings are further validated by flow cytometry, immunofluorescence, IHC, western blots, and qPCR. The single-cell transcriptome dataset provided here suggests several combination strategies to augment adaptive immune responses that are necessary for durable disease control in patients with PDAC.
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Novel targeted agents used in therapy of lymphoid malignancies are recognized to have complex immune-mediated effects. Sumoylation, a posttranslational modification of target proteins by small ubiquitin-like modifiers (SUMO), regulates a variety of cellular processes indispensable in immune cell activation. Despite this, the role of sumoylation in T-cell biology in context of cancer is not known. TAK-981 (subasumstat) is a small-molecule inhibitor of the SUMO-activating enzyme (SAE) that forms a covalent adduct with an activated SUMO protein. Using T cells derived from patients with chronic lymphocytic leukemia (CLL), we demonstrate that targeting SAE activates type I IFN response. This is accompanied by largely intact T-cell activation in response to T-cell receptor engagement, with increased expression of CD69 and CD38. Furthermore, TAK-981 decreases regulatory T cell (Treg) differentiation and enhances secretion of IFNγ by CD4+ and CD8+ T cells. These findings were recapitulated in mouse models, suggesting an evolutionarily conserved mechanism of T-cell activation regulated by SUMO modification. Relevant to the consideration of TAK-981 as an effective agent for immunotherapy in hematologic malignancies, we demonstrate that the downstream impact of TAK-981 administration is enhancement of the cytotoxic function of CD8+ T cells, thus uncovering immune implications of targeting sumoylation in lymphoid neoplasia.
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Leucemia Linfocítica Crónica de Células B , Ubiquitina , Animales , Ratones , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Procesamiento Proteico-Postraduccional , Inhibidores Enzimáticos , SumoilaciónRESUMEN
PURPOSE: Cancer drug development is currently limited by a paradigm of preclinical evaluation that does not adequately recapitulate the complexity of the intact human tumor microenvironment (TME). To overcome this, we combined trackable intratumor microdosing (CIVO) with spatial biology readouts to directly assess drug effects in patient tumors in situ. EXPERIMENTAL DESIGN: In a first-of-its-kind phase 0 clinical trial, we explored the effects of an investigational stage SUMOylation-activating enzyme (SAE) inhibitor, subasumstat (TAK-981) in 12 patients with head and neck carcinoma (HNC). Patients scheduled for tumor resection received percutaneous intratumor injections of subasumstat and vehicle control 1 to 4 days before surgery, resulting in spatially localized and graded regions of drug exposure (â¼1,000-2,000 µm in diameter). Drug-exposed (n = 214) and unexposed regions (n = 140) were compared by GeoMx Digital Spatial Profiler, with evaluation at single-cell resolution in a subset of these by CosMx Spatial Molecular Imager. RESULTS: Localized regions of subasumstat exposure revealed SUMO pathway inhibition, elevation of type I IFN response, and inhibition of cell cycle across all tumor samples. Single-cell analysis by CosMx demonstrated cell-cycle inhibition specific to the tumor epithelium, and IFN pathway induction commensurate with a TME shift from immune-suppressive to immune-permissive. CONCLUSIONS: Pairing CIVO with spatial profiling enabled detailed investigation of response to subasumstat across a diverse sampling of native and intact TME. We demonstrate that drug mechanism of action can be directly evaluated in a spatially precise manner in the most translationally relevant setting: an in situ human tumor.
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Antineoplásicos , Neoplasias de Cabeza y Cuello , Humanos , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Inhibidores Enzimáticos , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Microambiente TumoralRESUMEN
Neddylation is a sequential enzyme-based process which regulates the function of E3 Cullin-RING ligase (CRL) and thus degradation of substrate proteins. Here we show that CD8+ T cells are a direct target for therapeutically relevant anti-lymphoma activity of pevonedistat, a Nedd8-activating enzyme (NAE) inhibitor. Pevonedistat-treated patient-derived CD8+ T cells upregulated TNFα and IFNγ and exhibited enhanced cytotoxicity. Pevonedistat induced CD8+ T-cell inflamed microenvironment and delayed tumor progression in A20 syngeneic lymphoma model. This anti-tumor effect lessened when CD8+ T cells lost the ability to engage tumors through MHC class I interactions, achieved either through CD8+ T-cell depletion or genetic knockout of B2M. Meanwhile, loss of UBE2M in tumor did not alter efficacy of pevonedistat. Concurrent blockade of NAE and PD-1 led to enhanced tumor immune infiltration, T-cell activation and chemokine expression and synergistically restricted tumor growth. shRNA-mediated knockdown of HIF-1α, a CRL substrate, abrogated the in vitro effects of pevonedistat, suggesting that NAE inhibition modulates T-cell function in HIF-1α-dependent manner. scRNA-Seq-based clinical analyses in lymphoma patients receiving pevonedistat therapy demonstrated upregulation of interferon response signatures in immune cells. Thus, targeting NAE enhances the inflammatory T-cell state, providing rationale for checkpoint blockade-based combination therapy.
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Antineoplásicos , Linfoma , Humanos , Antineoplásicos/uso terapéutico , Linfocitos T CD8-positivos , Línea Celular Tumoral , Linfoma/tratamiento farmacológico , Ciclopentanos/farmacología , Ciclopentanos/uso terapéutico , Proteína NEDD8 , Microambiente Tumoral , Enzimas Ubiquitina-ConjugadorasRESUMEN
Fragility of regulatory T (Treg) cells manifested by the loss of neuropilin-1 (NRP1) and expression of IFNγ undermines the immune suppressive functions of Treg cells and contributes to the success of immune therapies against cancers. Intratumoral Treg cells somehow avoid fragility; however, the mechanisms by which Treg cells are protected from fragility in the tumor microenvironment are not well understood. Here, we demonstrate that the IFNAR1 chain of the type I IFN (IFN1) receptor was downregulated on intratumoral Treg cells. Downregulation of IFNAR1 mediated by p38α kinase protected Treg cells from fragility and maintained NRP1 levels, which were decreased in response to IFN1. Genetic or pharmacologic inactivation of p38α and stabilization of IFNAR1 in Treg cells induced fragility and inhibited their immune suppressive and protumorigenic activities. The inhibitor of sumoylation TAK981 (Subasumstat) upregulated IFNAR1, eliciting Treg fragility and inhibiting tumor growth in an IFNAR1-dependent manner. These findings describe a mechanism by which intratumoral Treg cells retain immunosuppressive activities and suggest therapeutic approaches for inducing Treg fragility and increasing the efficacy of immunotherapies.
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Neoplasias , Linfocitos T Reguladores , Humanos , Microambiente Tumoral , Neuropilina-1 , InmunoterapiaRESUMEN
Effector trogocytosis between malignant cells and tumor-specific cytotoxic T lymphocytes (CTLs) contributes to immune evasion through antigen loss on target cells and fratricide of antigen-experienced CTLs by other CTLs. The mechanisms regulating these events in tumors remain poorly understood. Here, we demonstrate that tumor-derived factors (TDFs) stimulated effector trogocytosis and restricted CTLs' tumoricidal activity and viability in vitro. TDFs robustly altered the CTL's lipid profile, including depletion of 25-hydroxycholesterol (25HC). 25HC inhibited trogocytosis and prevented CTL's inactivation and fratricide. Mechanistically, TDFs induced ATF3 transcription factor that suppressed the expression of 25HC-regulating gene-cholesterol 25-hydroxylase (CH25H). Stimulation of trogocytosis in the intratumoral CTL by the ATF3-CH25H axis attenuated anti-tumor immunity, stimulated tumor growth, and impeded the efficacy of chimeric antigen receptor (CAR) T cell adoptive therapy. Through use of armored CAR constructs or pharmacologic agents restoring CH25H expression, we reversed these phenotypes and increased the efficacy of immunotherapies.
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Linfocitos T Citotóxicos , Trogocitosis , Inmunoterapia , Esteroide Hidroxilasas , Replicación Viral/genéticaRESUMEN
Metabolism and disposition of pevonedistat, an investigational, first-in-class inhibitor of the NEDD8-activating enzyme (NAE), were characterized in patients with advanced solid tumors after intravenous infusion of [14C]pevonedistat at 25 mg/m2 (â¼60-85 µCi radioactive dose). More than 94% of the administered dose was recovered, with â¼41% and â¼53% of drug-related material eliminated in urine and feces, respectively. The metabolite profiles of [14C]pevonedistat were established in plasma using an accelerator mass spectrometer and excreta with traditional radiometric analysis. In plasma, unchanged parent drug accounted for approximately 49% of the total drug-related material. Metabolites M1 and M2 were major (>10% of the total drug-related material) circulating metabolites and accounted for approximately 15% and 22% of the drug-related material, respectively. Unchanged [14C]pevonedistat accounted for approximately 4% and 17% of the dose in urine and feces, respectively. Oxidative metabolites M1, M2, and M3 appeared as the most abundant drug-related components in the excreta and represented approximately 27%, 26%, and 15% of the administered dose, respectively. Based on the unbound plasma exposure in cancer patients and in vitro NAE inhibition, the contribution of metabolites M1 and M2 to overall in vivo pharmacological activity is anticipated to be minimal. The exposure to these metabolites was higher at safe and well tolerated doses in rat and dog (the two preclinical species used in toxicology evaluation) plasma than that observed in human plasma. Reaction phenotyping studies revealed that CYP3A4/5 are primary enzymes responsible for the metabolic clearance of pevonedistat. SIGNIFICANCE STATEMENT: This study details the metabolism and clearance mechanisms of pevonedistat, a first-in-class NEDD8-activating enzyme inhibitor, after intravenous administration to patients with cancer. Pevonedistat is biotransformed to two major circulating metabolites with higher exposure in nonclinical toxicological species than in humans. The pharmacological activity contribution of these metabolites is minimal compared to the overall target pharmacological effect of pevonedistat. Renal clearance was not an important route of excretion of unchanged pevonedistat (â¼4% of the dose).
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Neoplasias , Pirimidinas , Administración Oral , Animales , Ciclopentanos , Perros , Inhibidores Enzimáticos/uso terapéutico , Heces , Infusiones Intravenosas , Neoplasias/tratamiento farmacológico , Neoplasias/patología , RatasRESUMEN
Small ubiquitin-like modifier (SUMO) is a member of a ubiquitin-like protein superfamily. SUMOylation is a reversible posttranslational modification that has been implicated in the regulation of various cellular processes including inflammatory responses and expression of type 1 interferons (IFN1). In this report, we have explored the activity of the selective small molecule SUMOylation inhibitor subasumstat (TAK-981) in promoting antitumor innate immune responses. We demonstrate that treatment with TAK-981 results in IFN1-dependent macrophage and natural killer (NK) cell activation, promoting macrophage phagocytosis and NK cell cytotoxicity in ex vivo assays. Furthermore, pretreatment with TAK-981 enhanced macrophage phagocytosis or NK cell cytotoxicity against CD20+ target cells in combination with the anti-CD20 antibody rituximab. In vivo studies demonstrated enhanced antitumor activity of TAK-981 and rituximab in CD20+ lymphoma xenograft models. Combination of TAK-981 with anti-CD38 antibody daratumumab also resulted in enhanced antitumor activity. TAK-981 is currently being studied in phase 1 clinical trials (#NCT03648372, #NCT04074330, #NCT04776018, and #NCT04381650; www.clinicaltrials.gov) for the treatment of patients with lymphomas and solid tumors.
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Linfoma , Sumoilación , Citotoxicidad Celular Dependiente de Anticuerpos , Antígenos CD20 , Línea Celular Tumoral , Humanos , Células Asesinas Naturales , Linfoma/tratamiento farmacológico , Linfoma/metabolismo , Macrófagos/metabolismo , Rituximab/metabolismo , Rituximab/farmacología , Rituximab/uso terapéuticoRESUMEN
OBJECTIVES: Young adult cancer survivors often experience altered social relationships which may be a result of social support networks not knowing how to effectively provide the support young adults need. This study aimed to identify and describe themes of young adults' support preferences when engaging in cancer-related conversations and examine whether psychological distress is associated with support-related preferences. METHODS: Young adult survivors (Mage=35.12, N = 59) completed validated self-report measures of depression, cancer-related stress, social isolation, and two open-ended questions on types of preferred support. RESULTS: Listening (81.4%) was most commonly preferred; showing pity/worry (33.9%) was most undesired. Other types of preferred support included empathy, validation, encouragement (42.4%), and honest conversation (23.7%); common types of undesirable support included being uninterested and changing the subject (32.3%), insensitive comments and questions (25.4%), and negative stories/personal comparisons (23.7%). Greater depressive symptoms (OR = 1.21, p = .05) were associated with a preference for honest conversations whereas lower depressive symptoms (OR = 0.83, p = 0.05) and greater cancer-related stress (OR = 1.07, p = .02) were associated with a preference for conversations that did not contain advice. Lastly, lower perceived social isolation (OR = 0.88, p = .05) was associated with a preference for conversations that were not minimizing and that did not contain expressions of pity/worry. CONCLUSIONS: Study findings can inform communication interventions and educate support networks about types of support young adults prefer when discussing cancer-related concerns.
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Supervivientes de Cáncer , Neoplasias , Supervivientes de Cáncer/psicología , Emociones , Empatía , Humanos , Neoplasias/psicología , Neoplasias/terapia , Apoyo Social , Sobrevivientes/psicología , Adulto JovenRESUMEN
Decision-making among adolescents and young adults with cancer (AYA) is often complex, ongoing, and multifaceted, involving caregiver and oncology provider perspectives. Engagement in decision-making against the backdrop of normative developmental processes of acquiring autonomy and gaining independence contributes to the complexity of decision-making. Semi-structured qualitative interviews from 11 AYA and caregiver dyads and eight oncology providers examined decision-making processes with specific attention to the role of shared decision-making, cognitive and emotional processes, and coping with the decision-making experience. Five decision-making patterns were identified, with collaborative decision-making and AYA-driven decisions most commonly described. Utilizing hypothesis coding, AYA and caregivers explained how cognitive (i.e., pros/cons) and emotional (i.e., shock and fear of missing out) processes influenced cancer-related decisions. Coping strategies provided clarity and respite when engaged in decision-making. Our findings illuminate important implications for how to best support decision-making among AYA and caregivers, including the role oncology providers can play during decision-making.
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Cuidadores , Neoplasias , Adolescente , Toma de Decisiones , Humanos , Neoplasias/terapia , Adulto JovenRESUMEN
Organofluorine compounds are known to be toxic to a broad variety of living beings in different habitats, and chemical fluorination has been historically exploited by mankind for the development of therapeutic drugs or agricultural pesticides. On the other hand, several studies so far have demonstrated that, under appropriate conditions, living systems (in particular bacteria) can tolerate the presence of fluorinated molecules (e.g., amino acids analogues) within their metabolism and even repurpose them as alternative building blocks for the synthesis of cellular macromolecules such as proteins. Understanding the molecular mechanism behind these phenomena would greatly advance approaches to the biotechnological synthesis of recombinant proteins and peptide drugs. However, information about the metabolic effects of long-term exposure of living cells to fluorinated amino acids remains scarce. Hereby, we report the long-term propagation of Escherichia coli (E. coli) in an artificially fluorinated habitat that yielded two strains naturally adapted to live on fluorinated amino acids. In particular, we applied selective pressure to force a tryptophan (Trp)-auxotrophic strain to use either 4- or 5-fluoroindole as essential precursors for the in situ synthesis of Trp analogues, followed by their incorporation in the cellular proteome. We found that full adaptation to both fluorinated Trp analogues requires a low number of genetic mutations but is accompanied by large rearrangements in regulatory networks, membrane integrity, and quality control of protein folding. These findings highlight the cellular mechanisms behind the adaptation to unnatural amino acids and provide the molecular foundation for bioengineering of novel microbial strains for synthetic biology and biotechnology.
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Novel targeted agents used in therapy of lymphoid malignancies, such as inhibitors of B-cell receptor-associated kinases, are recognized to have complex immune-mediated effects. NEDD8-activating enzyme (NAE) has been identified as a tractable target in chronic lymphocytic leukemia (CLL) and non-Hodgkin lymphoma. We and others have shown that pevonedistat (TAK-924), a small-molecule inhibitor of NAE, abrogates NF-κB signaling in malignant B cells. However, NF-κB pathway activity is indispensable in immune response, and T-cell function is altered in patients with CLL. Using T cells derived from patients with CLL, we demonstrate that although targeting NAE results in markedly differential expression of NF-κB-regulated genes and downregulation of interleukin (IL)-2 signaling during T-cell activation, T cells evade apoptosis. Meanwhile, NAE inhibition favorably modulates polarization of T cells in vitro, with decreased Treg differentiation and a shift toward TH1 phenotype, accompanied by increased interferon-γ production. These findings were recapitulated in vivo in immunocompetent mouse models. T cells exposed to pevonedistat in washout experiments, informed by its human pharmacokinetic profile, recover NAE activity, and maintain their response to T-cell receptor stimulation and cytotoxic potential. Our data shed light on the potential immune implications of targeting neddylation in CLL and lymphoid malignancies.
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Antineoplásicos/farmacología , Ciclopentanos/farmacología , Inmunomodulación/efectos de los fármacos , Leucemia Prolinfocítica de Células T/inmunología , Leucemia Prolinfocítica de Células T/metabolismo , Proteína NEDD8/antagonistas & inhibidores , Proteína NEDD8/metabolismo , Pirimidinas/farmacología , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Ciclopentanos/uso terapéutico , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Humanos , Leucemia Prolinfocítica de Células T/tratamiento farmacológico , Leucemia Prolinfocítica de Células T/patología , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/inmunología , Modelos Biológicos , Pirimidinas/uso terapéutico , Receptores de Antígenos de Linfocitos T/metabolismo , Transducción de Señal , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/patología , Linfocitos T Colaboradores-Inductores/efectos de los fármacos , Linfocitos T Colaboradores-Inductores/inmunología , Linfocitos T Colaboradores-Inductores/metabolismoRESUMEN
Meningioma is the most common primary intracranial tumour, and surgical resection is the main therapeutic option. Merlin is a tumour suppressor protein that is frequently mutated in meningioma. The activity of the E3 ubiquitin ligase complex, CRL4-DCAF1, and the Raf/MEK/ERK scaffold protein Kinase suppressor of Ras 1 (KSR1) are upregulated in Merlin-deficient tumours, which drives tumour growth. Identifying small molecules that inhibit these key pathways may provide an effective treatment option for patients with meningioma. We used meningioma tissue and primary cells derived from meningioma tumours to investigate the expression of DDB1 and Cullin 4-associated factor 1 (DCAF1) and KSR1, and confirmed these proteins were overexpressed. We then used primary cells to assess the therapeutic potential of MLN3651, a neddylation inhibitor which impacts the activity of the CRL family of E3 ubiquitin ligases and the MAPK/ERK kinase (MEK1/2) inhibitor selumetinib. MLN3651 treatment reduced proliferation and activated apoptosis, whilst increasing Raf/MEK/ERK pathway activation. The combination of MLN3651 and the MEK1/2 inhibitor selumetinib prevented the increase in Raf/MEK/ERK activity, and had an additive effect compared with either treatment alone. Therefore, the combined targeting of CRL4-DCAF1 and Raf/MEK/ERK activity represents an attractive novel strategy in the treatment of Merlin-deficient meningioma.
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Multicomponent self-assembly of peptides is a powerful strategy to fabricate novel functional materials with synergetic properties that can be used for several nanobiotechnological applications. In the present study, we used a coassembly strategy to generate an injectable ultrashort bioactive peptide hydrogel formed by mixing a dipeptide hydrogelator with a macrophage attracting short chemotactic peptide ligand. Coassembly does not impede hydrogelation as shown by cryo-transmission electron microscopy (cryo-TEM), scanning electron microscopy, and rheology. Biocompatibility was shown by cytotoxicity assays and confocal microscopy. The hydrogels release the entrapped skin antibiotic ciprofloxacin, among others, in a slow and continuous manner. Such bioinspired advanced functional materials can find applications as wound dressing materials to treat chronic wound conditions like diabetic foot ulcer.
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The self-assembly of peptides onto the surface of gold nanoparticles has emerged as a promising strategy towards the creation of artificial enzymes. The resulting high local peptide density surrounding the nanoparticle leads to cooperative and synergistic effects, which result in rate accelerations and distinct catalytic properties compared to the unconjugated peptide. This Minireview summarizes contributions to and progress made in the field of catalytically active peptide-gold nanoparticle conjugates. The origin of distinct properties, as well as potential applications, are also discussed.
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Materiales Biomiméticos/química , Enzimas/metabolismo , Oro/química , Nanopartículas del Metal/química , Péptidos/química , CatálisisRESUMEN
With the advent of proteasome inhibitors (bortezomib) and pleiotropic pathway modulators which target cereblon E3 ligase (lenalidomide), the ubiquitin-proteasome system has emerged as a tractable target in non-Hodgkin lymphoma and multiple myeloma. Here we report that TAK-243, a small molecule inhibitor of the ubiquitin-activating enzyme (UAE), induced ER stress and the unfolded protein response in primary chronic lymphocytic leukemia cells, facilitating cell death. Moreover, targeting UAE was effective in ibrutinib-resistant mantle cell lymphoma cell lines and primary cells in vitro. Thus, UAE is a promising target in lymphoid malignancies, including ibrutinib-resistant lymphomas, an area of unmet medical need.
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Apoptosis/efectos de los fármacos , Resistencia a Antineoplásicos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Leucemia Linfocítica Crónica de Células B/metabolismo , Linfoma de Células del Manto/metabolismo , Pirazoles/farmacología , Pirimidinas/farmacología , Enzimas Activadoras de Ubiquitina/metabolismo , Adenina/análogos & derivados , Animales , Biomarcadores , Línea Celular Tumoral , Humanos , Leucemia Linfocítica Crónica de Células B/diagnóstico , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Linfoma de Células del Manto/diagnóstico , Linfoma de Células del Manto/tratamiento farmacológico , Ratones , Piperidinas , Inhibidores de Proteasoma/farmacología , Inhibidores de Proteasoma/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéutico , Pirazoles/uso terapéutico , Pirimidinas/uso terapéutico , Respuesta de Proteína Desplegada/efectos de los fármacosRESUMEN
Three proteasome inhibitors have garnered regulatory approvals in various multiple myeloma settings; but drug resistance is an emerging challenge, prompting interest in blocking upstream components of the ubiquitin-proteasome pathway. One such attractive target is the E1 ubiquitin-activating enzyme (UAE); we therefore evaluated the activity of TAK-243, a novel and specific UAE inhibitor. TAK-243 potently suppressed myeloma cell line growth, induced apoptosis, and activated caspases while decreasing the abundance of ubiquitin-protein conjugates. This was accompanied by stabilization of many short-lived proteins, including p53, myeloid cell leukemia 1 (MCL-1), and c-MYC, and activation of the activating transcription factor 6 (ATF-6), inositol-requiring enzyme 1 (IRE-1), and protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) arms of the ER stress response pathway, as well as oxidative stress. UAE inhibition showed comparable activity against otherwise isogenic cell lines with wild-type (WT) or deleted p53 despite induction of TP53 signaling in WT cells. Notably, TAK-243 overcame resistance to conventional drugs and novel agents in cell-line models, including bortezomib and carfilzomib resistance, and showed activity against primary cells from relapsed/refractory myeloma patients. In addition, TAK-243 showed strong synergy with a number of antimyeloma agents, including doxorubicin, melphalan, and panobinostat as measured by low combination indices. Finally, TAK-243 was active against a number of in vivo myeloma models in association with activation of ER stress. Taken together, the data support the conclusion that UAE inhibition could be an attractive strategy to move forward to the clinic for patients with relapsed and/or refractory multiple myeloma.
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Resistencia a Antineoplásicos/efectos de los fármacos , Mieloma Múltiple/tratamiento farmacológico , Inhibidores de Proteasoma/farmacología , Enzimas Activadoras de Ubiquitina/antagonistas & inhibidores , Respuesta de Proteína Desplegada/efectos de los fármacos , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Sinergismo Farmacológico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Humanos , Estrés Oxidativo/efectos de los fármacos , Terapia Recuperativa/métodos , Células Tumorales Cultivadas , Proteína p53 Supresora de Tumor/efectos de los fármacos , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Alterations in the ubiquitin proteasome system (UPS) leave malignant cells in heightened cellular stress, making them susceptible to proteasome inhibition. However, given the limited efficacy of proteasome inhibitors in non-Hodgkin lymphoma (NHL), novel approaches to target the UPS are needed. Here, we show that TAK-243, the first small-molecule inhibitor of the ubiquitin activating enzyme (UAE) to enter clinical development, disrupts all ubiquitin signaling and global protein ubiquitination in diffuse large B-cell lymphoma (DLBCL) cells, thereby inducing endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). Activation of the ER stress response protein kinase R (PKR)-like ER kinase and phosphorylation of eukaryotic translation initiator factor 2α led to upregulation of the proapoptotic molecule C/EBP homologous protein and cell death across a panel of DLBCL cell lines independent of cell of origin. Concurrently, targeting UAE led to accumulation of Cdt1, a replication licensing factor, leading to DNA rereplication, checkpoint activation, and cell cycle arrest. MYC oncoprotein sensitized DLBCL cells to UAE inhibition; engineered expression of MYC enhanced while genetic MYC knockdown protected from TAK-243-induced apoptosis. UAE inhibition demonstrated enhanced ER stress and UPR and increased potency compared with bortezomib in DLBCL cell lines. In vivo treatment with TAK-243 restricted the growth of xenografted DLBCL tumors, accompanied by reduced cell proliferation and apoptosis. Finally, primary patient-derived DLBCL cells, including those expressing aberrant MYC, demonstrated susceptibility to UAE inhibition. In sum, targeting UAE may hold promise as a novel therapeutic approach in NHL.