RESUMEN
Receptor-interacting serine/threonine-protein kinase 1 (RIPK1) functions as a critical stress sentinel that coordinates cell survival, inflammation, and immunogenic cell death (ICD). Although the catalytic function of RIPK1 is required to trigger cell death, its non-catalytic scaffold function mediates strong pro-survival signaling. Accordingly, cancer cells can hijack RIPK1 to block necroptosis and evade immune detection. We generated a small-molecule proteolysis-targeting chimera (PROTAC) that selectively degraded human and murine RIPK1. PROTAC-mediated depletion of RIPK1 deregulated TNFR1 and TLR3/4 signaling hubs, accentuating the output of NF-κB, MAPK, and IFN signaling. Additionally, RIPK1 degradation simultaneously promoted RIPK3 activation and necroptosis induction. We further demonstrated that RIPK1 degradation enhanced the immunostimulatory effects of radio- and immunotherapy by sensitizing cancer cells to treatment-induced TNF and interferons. This promoted ICD, antitumor immunity, and durable treatment responses. Consequently, targeting RIPK1 by PROTACs emerges as a promising approach to overcome radio- or immunotherapy resistance and enhance anticancer therapies.
Asunto(s)
Muerte Celular Inmunogénica , Proteolisis , Proteína Serina-Treonina Quinasas de Interacción con Receptores , Transducción de Señal , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Humanos , Animales , Ratones , Proteolisis/efectos de los fármacos , Línea Celular Tumoral , Transducción de Señal/efectos de los fármacos , Muerte Celular Inmunogénica/efectos de los fármacos , Necroptosis/efectos de los fármacos , Necroptosis/inmunología , Neoplasias/inmunología , Neoplasias/tratamiento farmacológico , Ratones Endogámicos C57BL , Antineoplásicos/farmacología , Inmunoterapia/métodosRESUMEN
Stress response pathways detect and alleviate adverse conditions to safeguard cell and tissue homeostasis, yet their prolonged activation induces apoptosis and disrupts organismal health1-3. How stress responses are turned off at the right time and place remains poorly understood. Here we report a ubiquitin-dependent mechanism that silences the cellular response to mitochondrial protein import stress. Crucial to this process is the silencing factor of the integrated stress response (SIFI), a large E3 ligase complex mutated in ataxia and in early-onset dementia that degrades both unimported mitochondrial precursors and stress response components. By recognizing bifunctional substrate motifs that equally encode protein localization and stability, the SIFI complex turns off a general stress response after a specific stress event has been resolved. Pharmacological stress response silencing sustains cell survival even if stress resolution failed, which underscores the importance of signal termination and provides a roadmap for treating neurodegenerative diseases caused by mitochondrial import defects.
Asunto(s)
Mitocondrias , Proteínas Mitocondriales , Mutación , Enfermedades Neurodegenerativas , Estrés Fisiológico , Ubiquitina-Proteína Ligasas , Apoptosis/efectos de los fármacos , Ataxia/genética , Supervivencia Celular/efectos de los fármacos , Demencia/genética , Mitocondrias/genética , Mitocondrias/metabolismo , Mitocondrias/patología , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , Complejos Multiproteicos/antagonistas & inhibidores , Complejos Multiproteicos/genética , Complejos Multiproteicos/metabolismo , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/metabolismo , Enfermedades Neurodegenerativas/patología , Estabilidad Proteica/efectos de los fármacos , Transporte de Proteínas/efectos de los fármacos , Proteolisis/efectos de los fármacos , Estrés Fisiológico/efectos de los fármacos , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligasas/antagonistas & inhibidores , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación/efectos de los fármacosRESUMEN
Alkenyl oxindoles have been characterized as autophagosome-tethering compounds (ATTECs), which can target mutant huntingtin protein (mHTT) for lysosomal degradation. In order to expand the application of alkenyl oxindoles for targeted protein degradation, we designed and synthesized a series of heterobifunctional compounds by conjugating different alkenyl oxindoles with bromodomain-containing protein 4 (BRD4) inhibitor JQ1. Through structure-activity relationship study, we successfully developed JQ1-alkenyl oxindole conjugates that potently degrade BRD4. Unexpectedly, we found that these molecules degrade BRD4 through the ubiquitin-proteasome system, rather than the autophagy-lysosomal pathway. Using pooled CRISPR interference (CRISPRi) screening, we revealed that JQ1-alkenyl oxindole conjugates recruit the E3 ubiquitin ligase complex CRL4DCAF11 for substrate degradation. Furthermore, we validated the most potent heterobifunctional molecule HL435 as a promising drug-like lead compound to exert antitumor activity both in vitro and in a mouse xenograft tumor model. Our research provides new employable proteolysis targeting chimera (PROTAC) moieties for targeted protein degradation, providing new possibilities for drug discovery.
Asunto(s)
Proteínas de Ciclo Celular , Oxindoles , Proteolisis , Ubiquitina-Proteína Ligasas , Humanos , Animales , Proteolisis/efectos de los fármacos , Ratones , Ubiquitina-Proteína Ligasas/metabolismo , Oxindoles/farmacología , Oxindoles/metabolismo , Oxindoles/química , Proteínas de Ciclo Celular/metabolismo , Factores de Transcripción/metabolismo , Línea Celular Tumoral , Ensayos Antitumor por Modelo de Xenoinjerto , Ratones Desnudos , Células HEK293 , Relación Estructura-Actividad , Complejo de la Endopetidasa Proteasomal/metabolismo , Azepinas/farmacología , Azepinas/química , Azepinas/metabolismo , Antineoplásicos/farmacología , Antineoplásicos/química , Femenino , Proteínas que Contienen Bromodominio , Receptores de Interleucina-17RESUMEN
Activation of the Wnt-ß-catenin signaling pathway by CHIR99021, a specific inhibitor of GSK3ß, induces Tcf7l1 protein degradation, which facilitates the maintenance of an undifferentiated state in mouse embryonic stem cells (mESCs); however, the precise mechanism is still unclear. Here, we showed that the overexpression of transducin-ß-like protein 1 (Tbl1, also known as Tbl1x) or its family member Tblr1 (also known as Tbl1xr1) can decrease Tcf7l1 protein levels, whereas knockdown of each gene increases Tcf7l1 levels without affecting Tcf7l1 transcription. Interestingly, only Tbl1, and not Tblr1, interacts with Tcf7l1. Mechanistically, Tbl1 translocates from the cytoplasm into the nucleus in association with ß-catenin (CTNNB1) after the addition of CHIR99021 and functions as an adaptor to promote ubiquitylation of the Tcf7l1 protein. Functional assays further revealed that enforced expression of Tbl1 is capable of delaying mESC differentiation. In contrast, knockdown of Tbl1 attenuates the effect of CHIR99021 on Tcf7l1 protein stability and mESC self-renewal. Our results provide insight into the regulatory network of the Wnt-ß-catenin signaling pathway involved in promoting the maintenance of naïve pluripotency.
Asunto(s)
Células Madre Embrionarias de Ratones , Proteína 1 Similar al Factor de Transcripción 7 , Vía de Señalización Wnt , beta Catenina , Animales , Humanos , Ratones , beta Catenina/metabolismo , Proteínas con Repetición de beta-Transducina/metabolismo , Proteínas con Repetición de beta-Transducina/genética , Diferenciación Celular/efectos de los fármacos , Células Madre Embrionarias de Ratones/metabolismo , Proteolisis/efectos de los fármacos , Piridinas/farmacología , Pirimidinas/farmacología , Proteína 1 Similar al Factor de Transcripción 7/metabolismo , Proteína 1 Similar al Factor de Transcripción 7/genética , UbiquitinaciónRESUMEN
Targeted protein degradation is the selective removal of a protein of interest through hijacking intracellular protein cleanup machinery. This rapidly growing field currently relies heavily on the use of the E3 ligase cereblon (CRBN) to target proteins for degradation, including the immunomodulatory drugs (IMiDs) thalidomide, lenalidomide, and pomalidomide which work through a molecular glue mechanism of action with CRBN. While CRBN recruitment can result in degradation of a specific protein of interest (e.g., efficacy), degradation of other proteins (called CRBN neosubstrates) also occurs. Degradation of one or more of these CRBN neosubstrates is believed to play an important role in thalidomide-related developmental toxicity observed in rabbits and primates. We identified a set of 25 proteins of interest associated with CRBN-related protein homeostasis and/or embryo/fetal development. We developed a targeted assay for these proteins combining peptide immunoaffinity enrichment and high-resolution mass spectrometry and successfully applied this assay to rabbit embryo samples from pregnant rabbits dosed with three IMiDs. We confirmed previously reported in vivo decreases in neosubstrates like SALL4, as well as provided evidence of neosubstrate changes for proteins only examined in vitro previously. While there were many proteins that were similarly decreased by all three IMiDs, no compound had the exact same neosubstrate degradation profile as another. We compared our data to previous literature reports of IMiD-induced degradation and known developmental biology associations. Based on our observations, we recommend monitoring at least a major subset of these neosubstrates in a developmental test system to improve CRBN-binding compound-specific risk assessment. A strength of our assay is that it is configurable, and the target list can be readily adapted to focus on only a subset of proteins of interest or expanded to incorporate new findings as additional information about CRBN biology is discovered.
Asunto(s)
Proteolisis , Proteómica , Talidomida , Ubiquitina-Proteína Ligasas , Animales , Conejos , Proteómica/métodos , Ubiquitina-Proteína Ligasas/metabolismo , Talidomida/análogos & derivados , Talidomida/farmacología , Proteolisis/efectos de los fármacos , Femenino , Embrión de Mamíferos/metabolismo , Embrión de Mamíferos/efectos de los fármacos , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Lenalidomida/farmacología , EmbarazoRESUMEN
The ubiquitin (Ub)-proteasome system (UPS) is the major machinery mediating specific protein turnover in eukaryotic cells. By ubiquitylating unwanted, damaged, or harmful proteins and driving their degradation, UPS is involved in many important cellular processes. Several new UPS-based technologies, including molecular glue degraders and PROTACs (proteolysis-targeting chimeras) to promote protein degradation, and DUBTACs (deubiquitinase-targeting chimeras) to increase protein stability, have been developed. By specifically inducing the interactions between different Ub ligases and targeted proteins that are not otherwise related, molecular glue degraders and PROTACs degrade targeted proteins via the UPS; in contrast, by inducing the proximity of targeted proteins to deubiquitinases, DUBTACs are created to clear degradable poly-Ub chains to stabilize targeted proteins. In this review, we summarize the recent research progress in molecular glue degraders, PROTACs, and DUBTACs and their applications. We discuss immunomodulatory drugs, sulfonamides, cyclin-dependent kinase-targeting molecular glue degraders, and new development of PROTACs. We also introduce the principle of DUBTAC and its applications. Finally, we propose a few future directions of these three technologies related to targeted protein homeostasis.
Asunto(s)
Descubrimiento de Drogas , Complejo de la Endopetidasa Proteasomal , Proteolisis , Ubiquitinación , Humanos , Ubiquitinación/efectos de los fármacos , Proteolisis/efectos de los fármacos , Complejo de la Endopetidasa Proteasomal/metabolismo , Enzimas Desubicuitinizantes/metabolismo , Ubiquitina/metabolismo , Animales , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
BACKGROUND & AIMS: Despite the increasing number of treatment options available for liver cancer, only a small proportion of patients achieve long-term clinical benefits. Here, we aim to develop new therapeutic approaches for liver cancer. METHODS: A compound screen was conducted to identify inhibitors that could synergistically induce senescence when combined with cyclin-dependent kinase (CDK) 4/6 inhibitor. The combination effects of CDK4/6 inhibitor and exportin 1 (XPO1) inhibitor on cellular senescence were investigated in a panel of human liver cancer cell lines and multiple liver cancer models. A senolytic drug screen was performed to identify drugs that selectively killed senescent liver cancer cells. RESULTS: The combination of CDK4/6 inhibitor and XPO1 inhibitor synergistically induces senescence of liver cancer cells in vitro and in vivo. The XPO1 inhibitor acts by causing accumulation of RB1 in the nucleus, leading to decreased E2F signaling and promoting senescence induction by the CDK4/6 inhibitor. Through a senolytic drug screen, cereblon (CRBN)-based proteolysis targeting chimera (PROTAC) ARV-825 was identified as an agent that can selectively kill senescent liver cancer cells. Up-regulation of CRBN was a vulnerability of senescent liver cancer cells, making them sensitive to CRBN-based PROTAC drugs. Mechanistically, we find that ubiquitin specific peptidase 2 (USP2) directly interacts with CRBN, leading to the deubiquitination and stabilization of CRBN in senescent liver cancer cells. CONCLUSIONS: Our study demonstrates a striking synergy in senescence induction of liver cancer cells through the combination of CDK4/6 inhibitor and XPO1 inhibitor. These findings also shed light on the molecular processes underlying the vulnerability of senescent liver cancer cells to CRBN-based PROTAC therapy.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Senescencia Celular , Quinasa 4 Dependiente de la Ciclina , Quinasa 6 Dependiente de la Ciclina , Proteína Exportina 1 , Carioferinas , Neoplasias Hepáticas , Inhibidores de Proteínas Quinasas , Receptores Citoplasmáticos y Nucleares , Ubiquitina-Proteína Ligasas , Humanos , Senescencia Celular/efectos de los fármacos , Quinasa 6 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 6 Dependiente de la Ciclina/metabolismo , Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 4 Dependiente de la Ciclina/metabolismo , Carioferinas/antagonistas & inhibidores , Carioferinas/metabolismo , Receptores Citoplasmáticos y Nucleares/antagonistas & inhibidores , Receptores Citoplasmáticos y Nucleares/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/metabolismo , Línea Celular Tumoral , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/antagonistas & inhibidores , Animales , Proteínas de Unión a Retinoblastoma/metabolismo , Proteínas de Unión a Retinoblastoma/genética , Sinergismo Farmacológico , Senoterapéuticos/farmacología , Ensayos Antitumor por Modelo de Xenoinjerto , Transducción de Señal/efectos de los fármacos , Proteolisis/efectos de los fármacos , Hidrazinas/farmacología , Hidrazinas/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Células Hep G2 , Ratones , Piperazinas , Piridinas , TriazolesRESUMEN
In females, the pathophysiological mechanism of poor ovarian response (POR) is not fully understood. Considering the expression level of p62 was significantly reduced in the granulosa cells (GCs) of POR patients, this study focused on identifying the role of the selective autophagy receptor p62 in conducting the effect of follicle-stimulating hormone (FSH) on antral follicles (AFs) formation in female mice. The results showed that p62 in GCs was FSH responsive and that its level increased to a peak and then decreased time-dependently either in ovaries or in GCs after gonadotropin induction in vivo. GC-specific deletion of p62 resulted in subfertility, a significantly reduced number of AFs and irregular estrous cycles, which were same as pathophysiological symptom of POR. By conducting mass spectrum analysis, we found the ubiquitination of proteins was decreased, and autophagic flux was blocked in GCs. Specifically, the level of nonubiquitinated Wilms tumor 1 homolog (WT1), a transcription factor and negative controller of GC differentiation, increased steadily. Co-IP results showed that p62 deletion increased the level of ubiquitin-specific peptidase 5 (USP5), which blocked the ubiquitination of WT1. Furthermore, a joint analysis of RNA-seq and the spatial transcriptome sequencing data showed the expression of steroid metabolic genes and FSH receptors pivotal for GCs differentiation decreased unanimously. Accordingly, the accumulation of WT1 in GCs deficient of p62 decreased steroid hormone levels and reduced FSH responsiveness, while the availability of p62 in GCs simultaneously ensured the degradation of WT1 through the ubiquitinâproteasome system and autophagolysosomal system. Therefore, p62 in GCs participates in GC differentiation and AF formation in FSH induction by dynamically controlling the degradation of WT1. The findings of the study contributes to further study the pathology of POR.
Asunto(s)
Hormona Folículo Estimulante , Células de la Granulosa , Folículo Ovárico , Proteína Sequestosoma-1 , Ubiquitinación , Proteínas WT1 , Animales , Hormona Folículo Estimulante/metabolismo , Hormona Folículo Estimulante/farmacología , Femenino , Proteínas WT1/metabolismo , Proteínas WT1/genética , Ratones , Folículo Ovárico/metabolismo , Folículo Ovárico/efectos de los fármacos , Células de la Granulosa/metabolismo , Células de la Granulosa/efectos de los fármacos , Proteína Sequestosoma-1/metabolismo , Proteína Sequestosoma-1/genética , Ratones Endogámicos C57BL , Autofagia/efectos de los fármacos , Proteolisis/efectos de los fármacos , Humanos , Ratones NoqueadosRESUMEN
Multiple myeloma (MM) is the second most common hematological tumor in adults. Immunomodulatory drugs (IMiDs), such as thalidomide and lenalidomide (Len), are effective drugs for the treatment of multiple myeloma. Len can recruit IKZF1 and IKZF3 to cereblon (CRBN), a substrate receptor of the cullin 4-RING E3 ligase (CRL4), promote their ubiquitination and degradation, and finally inhibit the proliferation of myeloma cells. However, MM patients develop resistance to IMiDs over time, leading to disease recurrence and deterioration. To explore the possible approaches that may enhance the sensitivity of IMiDs to MM, in this study, we used the proximity labeling technique TurboID and quantitative proteomics to identify Lys-63-specific deubiquitinase BRCC36 as a CRBN-interacting protein. Biochemical experiments demonstrated that BRCC36 in the BRISC complex protects CRBN from lysosomal degradation by specifically cleaving the K63-linked polyubiquitin chain on CRBN. Further studies found that a small-molecule compound SHIN1, which binds to BRISC complex subunit SHMT2, can upregulate CRBN by elevating BRCC36. The combination of SHIN1 and Len can further increase the sensitivity of MM cells to IMiDs. Therefore, this study provides the basis for the exploration of a possible strategy for the SHIN1 and Len combination treatment for MM.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Lenalidomida , Lisosomas , Mieloma Múltiple , Ubiquitina-Proteína Ligasas , Humanos , Mieloma Múltiple/patología , Mieloma Múltiple/tratamiento farmacológico , Mieloma Múltiple/metabolismo , Lenalidomida/farmacología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Lisosomas/metabolismo , Lisosomas/efectos de los fármacos , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina-Proteína Ligasas/genética , Línea Celular Tumoral , Ubiquitinación/efectos de los fármacos , Proteolisis/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Enzimas Desubicuitinizantes/metabolismo , Enzimas Desubicuitinizantes/antagonistas & inhibidoresRESUMEN
AIMS: Human epidermal growth factor receptor 2 (HER2) is an oncogenic receptor tyrosine kinase amplified in approximately 20% of breast cancer (BC). HER2-targeted therapies are the linchpin of treating HER2-positive BC. However, drug resistance is common, and the main resistance mechanism is unknown. We tested the hypothesis that drug resistance results mainly from inadequate or lack of inhibition of HER2 and its family member epidermal growth factor receptor (EGFR). METHODS: We used clinically relevant cell and tumor models to assess the impact of targeted degradation of HER2 and EGFR on trastuzumab resistance. Trastuzumab is the most common clinically used HER2 inhibitor. Targeted degradation of HER2 and EGFR was achieved using recombinant human protein PEPDG278D, which binds to the extracellular domains of the receptors. siRNA knockdown was used to assess the relative importance of EGFR and HER2 in trastuzumab resistance. RESULTS: Both HER2 and EGFR are overexpressed in all trastuzumab-resistant HER2-positive BC cell and tumor models and that all trastuzumab-resistant models are highly vulnerable to targeted degradation of HER2 and EGFR. Degradation of HER2 and EGFR induced by PEPDG278D causes extensive inhibition of oncogenic signaling in trastuzumab-resistant HER2-positive BC cells. This is accompanied by strong growth inhibition of cultured cells, orthotopic patient-derived xenografts, and metastatic lesions in the brain and lung of trastuzumab-resistant HER2-positive BC. siRNA knockdown indicates that eliminating both HER2 and EGFR is necessary to maximize therapeutic outcome. CONCLUSIONS: This study unravels the therapeutic vulnerability of trastuzumab-resistant HER2-positive BC and shows that an agent that targets the degradation of both HER2 and EGFR is highly effective in overcoming drug resistance in this disease. The findings provide new insights and innovations for advancing treatment of drug-resistant HER2-positive breast cancer that remains an unmet problem.
Asunto(s)
Neoplasias de la Mama , Resistencia a Antineoplásicos , Receptores ErbB , Receptor ErbB-2 , Transducción de Señal , Trastuzumab , Humanos , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Resistencia a Antineoplásicos/efectos de los fármacos , Femenino , Trastuzumab/farmacología , Trastuzumab/uso terapéutico , Receptor ErbB-2/metabolismo , Receptor ErbB-2/antagonistas & inhibidores , Animales , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/metabolismo , Ratones , Transducción de Señal/efectos de los fármacos , Línea Celular Tumoral , Ensayos Antitumor por Modelo de Xenoinjerto , Proteolisis/efectos de los fármacos , Proliferación Celular/efectos de los fármacosRESUMEN
The degradation of oncoproteins mediated by proteolysis-targeting chimera (PROTAC) has emerged as a potent strategy in cancer therapy. However, the clinical application of PROTACs is hampered by challenges such as poor water solubility and off-target adverse effects. Herein, we present an ultrasound (US)-activatable PROTAC prodrug termed NPCe6+PRO for actuating efficient sono-immunotherapy in a spatiotemporally controllable manner. Specifically, US irradiation, which exhibits deep-tissue penetration capability, results in Ce6-mediated generation of ROS, facilitating sonodynamic therapy (SDT) and inducing immunogenic cell death (ICD). Simultaneously, the generated ROS cleaves the thioketal (TK) linker through a ROS-responsive mechanism, realizing the on-demand activation of the PROTAC prodrug in deep tissues. This prodrug activation results in the degradation of the target protein BRD4, while simultaneously reversing the upregulation of PD-L1 expression associated with the SDT process. In the orthotopic mouse model of pancreatic tumors, NPCe6+PRO effectively suppressed tumor growth in conjunction with US stimulation.
Asunto(s)
Inmunoterapia , Neoplasias Pancreáticas , Profármacos , Animales , Profármacos/farmacología , Profármacos/uso terapéutico , Profármacos/química , Neoplasias Pancreáticas/terapia , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/patología , Neoplasias Pancreáticas/inmunología , Ratones , Humanos , Línea Celular Tumoral , Proteolisis/efectos de los fármacos , Terapia por Ultrasonido/métodos , Antígeno B7-H1 , Factores de Transcripción , Proteínas de Ciclo Celular , Especies Reactivas de Oxígeno/metabolismo , Proteínas que Contienen BromodominioRESUMEN
Neuroblastoma (NB), a common solid tumour in young children originating from the sympathetic nervous system during embryonic development, poses challenges despite therapeutic advances like high-dose chemotherapy and immunotherapy. Some survivors still grapple with severe side effects and drug resistance. The role of lncRNA NUTM2A-AS1 has been explored in various cancers, but its function in drug-resistant NB progression is unclear. Our study found that NUTM2A-AS1 expression in cisplatin-resistant NB cells increased in a time- and dose-dependent manner. Knockdown of NUTM2A-AS1 significantly improved NB cell sensitivity to cisplatin and inhibited metastatic abilities. Additionally, we identified B7-H3, an immune checkpoint-related protein, as a NUTM2A-AS1-associated protein in NB cells. NUTM2A-AS1 was shown to inhibit the protein degradation of B7-H3. Moreover, NUTM2A-AS1 modulated immune evasion in cisplatin-resistant NB cells through B7-H3. Furthermore, NUTM2A-AS1 expression in cisplatin-resistant NB cells was transactivated by NR1D1. In summary, our results unveil the molecular or biological relationship within the NR1D1/NUTM2A-AS1/B7-H3 axis in NB cells under cisplatin treatment, providing an intriguing avenue for fundamental research into cisplatin-resistant NB.
Asunto(s)
Antígenos B7 , Cisplatino , Resistencia a Antineoplásicos , Regulación Neoplásica de la Expresión Génica , Neuroblastoma , ARN Largo no Codificante , Humanos , Neuroblastoma/genética , Neuroblastoma/patología , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/metabolismo , Resistencia a Antineoplásicos/genética , Antígenos B7/metabolismo , Antígenos B7/genética , ARN Largo no Codificante/genética , Cisplatino/farmacología , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Evasión Inmune , Animales , Proteolisis/efectos de los fármacos , RatonesRESUMEN
The pathogenesis of trauma-induced heterotopic ossification (HO) in the tendon remains unclear, posing a challenging hurdle in treatment. Recognizing inflammation as the root cause of HO, anti-inflammatory agents hold promise for its management. Malvidin (MA), possessing anti-inflammatory properties, emerges as a potential agent to impede HO progression. This study aimed to investigate the effect of MA in treating trauma-induced HO and unravel its underlying mechanisms. Herein, the effectiveness of MA in preventing HO formation was assessed through local injection in a rat model. The potential mechanism underlying MA's treatment was investigated in the tendon-resident progenitor cells of tendon-derived stem cells (TDSCs), exploring its pathway in HO formation. The findings demonstrated that MA effectively hindered the osteogenic differentiation of TDSCs by inhibiting the mTORC1 signalling pathway, consequently impeding the progression of trauma-induced HO of Achilles tendon in rats. Specifically, MA facilitated the degradation of Rheb through the K48-linked ubiquitination-proteasome pathway by modulating USP4 and intercepted the interaction between Rheb and the mTORC1 complex, thus inhibiting the mTORC1 signalling pathway. Hence, MA presents itself as a promising candidate for treating trauma-induced HO in the Achilles tendon, acting by targeting Rheb for degradation through the ubiquitin-proteasome pathway.
Asunto(s)
Osificación Heterotópica , Complejo de la Endopetidasa Proteasomal , Proteína Homóloga de Ras Enriquecida en el Cerebro , Transducción de Señal , Ubiquitina , Animales , Ratas , Complejo de la Endopetidasa Proteasomal/metabolismo , Osificación Heterotópica/metabolismo , Osificación Heterotópica/etiología , Osificación Heterotópica/patología , Transducción de Señal/efectos de los fármacos , Proteína Homóloga de Ras Enriquecida en el Cerebro/metabolismo , Ubiquitina/metabolismo , Masculino , Osteogénesis/efectos de los fármacos , Tendones/metabolismo , Tendones/patología , Ratas Sprague-Dawley , Traumatismos de los Tendones/metabolismo , Traumatismos de los Tendones/patología , Traumatismos de los Tendones/complicaciones , Proteolisis/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Tendón Calcáneo/metabolismo , Tendón Calcáneo/patología , Tendón Calcáneo/lesiones , Modelos Animales de Enfermedad , Ubiquitinación , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Células Madre/metabolismo , Células Madre/efectos de los fármacosRESUMEN
Proteolysis Targeting Chimeras (PROTACs) represent a significant advancement in therapeutic drug development by leveraging the ubiquitin-proteasome system to enable targeted protein degradation, particularly impacting oncology. This review delves into the various types of PROTACs, such as peptide-based, nucleic acid-based, and small molecule PROTACs, each addressing distinct challenges in protein degradation. It also discusses innovative strategies like bridged PROTACs and conditional switch-activated PROTACs, offering precise targeting of previously "undruggable" proteins. The potential of PROTACs extends beyond oncology, with ongoing research and technological advancements needed to maximize their therapeutic potential. Future progress in this field relies on interdisciplinary collaboration and the integration of advanced computational tools to open new treatment avenues across various diseases.
Asunto(s)
Complejo de la Endopetidasa Proteasomal , Quimera Dirigida a la Proteólisis , Proteolisis , Animales , Humanos , Péptidos/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteolisis/efectos de los fármacos , Ubiquitina/metabolismoRESUMEN
Persistent activation of estrogen receptor alpha (ERα)-mediated estrogen signaling plays a pivotal role in driving the progression of estrogen receptor positive (ER+) breast cancer (BC). In the current study, LINC00173, a long non-coding RNA, was found to bind both ERα and lipopolysaccharide (LPS)-induced tumor necrosis factor alpha (TNFα) factor (LITAF), then cooperatively to inhibit ERα protein degradation by impeding the nuclear export of ERα. Concurrently, LITAF was found to attenuate TNFα transcription after binding to LINC00173, and this attenuating transcriptional effect was quite significant under lipopolysaccharide stimulation. Distinct functional disparities between estrogen subtypes emerge, with estradiol synergistically promoting ER+ BC cell growth with LINC00173, while estrone (E1) facilitated LITAF-transcriptional activation. In terms of therapeutic significance, silencing LINC00173 alongside moderate addition of E1 heightened TNFα and induced apoptosis, effectively inhibiting ER+ BC progression.
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Neoplasias de la Mama , Receptor alfa de Estrógeno , Estrona , ARN Largo no Codificante , Factores de Transcripción , Humanos , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Neoplasias de la Mama/genética , Receptor alfa de Estrógeno/metabolismo , Receptor alfa de Estrógeno/genética , Femenino , Estrona/metabolismo , Estrona/farmacología , Estrona/análogos & derivados , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Factor de Necrosis Tumoral alfa/metabolismo , Células MCF-7 , Línea Celular Tumoral , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Proteolisis/efectos de los fármacos , Animales , Ratones , Silenciador del GenRESUMEN
Mutations resulting in decreased activity of p53 tumor suppressor protein promote tumorigenesis. P53 protein levels are tightly regulated through the Ubiquitin Proteasome System (UPS). Several E3 ligases were shown to regulate p53 stability, including MDM2. Here we report that the ubiquitin E3 ligase XIAP (X-linked Inhibitors of Apoptosis) is a direct ligase for p53 and describe a novel approach for modulating the levels of p53 by targeting the XIAP pathway. Using in vivo (live-cell) and in vitro (cell-free reconstituted system) ubiquitylation assays, we show that the XIAP-antagonist ARTS regulates the levels of p53 by promoting the degradation of XIAP. XIAP directly binds and ubiquitylates p53. In apoptotic cells, ARTS inhibits the ubiquitylation of p53 by antagonizing XIAP. XIAP knockout MEFs express higher p53 protein levels compared to wild-type MEFs. Computational screen for small molecules with high affinity to the ARTS-binding site within XIAP identified a small-molecule ARTS-mimetic, B3. This compound stimulates apoptosis in a wide range of cancer cells but not normal PBMC (Peripheral Blood Mononuclear Cells). Like ARTS, the B3 compound binds to XIAP and promotes its degradation via the UPS. B3 binding to XIAP stabilizes p53 by disrupting its interaction with XIAP. These results reveal a novel mechanism by which ARTS and p53 regulate each other through an amplification loop to promote apoptosis. Finally, these data suggest that targeting the ARTS binding pocket in XIAP can be used to increase p53 levels as a new strategy for developing anti-cancer therapeutics.
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Apoptosis , Proteolisis , Proteína p53 Supresora de Tumor , Ubiquitinación , Proteína Inhibidora de la Apoptosis Ligada a X , Proteína Inhibidora de la Apoptosis Ligada a X/metabolismo , Proteína Inhibidora de la Apoptosis Ligada a X/genética , Humanos , Proteína p53 Supresora de Tumor/metabolismo , Proteína p53 Supresora de Tumor/genética , Apoptosis/efectos de los fármacos , Ubiquitinación/efectos de los fármacos , Proteolisis/efectos de los fármacos , Animales , Ratones , Línea Celular Tumoral , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina-Proteína Ligasas/genética , Regulación hacia Arriba/efectos de los fármacos , Unión ProteicaRESUMEN
Vascular endothelial cells play a critical role in maintaining the health of blood vessels, but dysfunction can lead to cardiovascular diseases. The impact of arsenite exposure on cardiovascular health is a significant concern due to its potential adverse effects. This study aims to explore how NBR1-mediated autophagy in vascular endothelial cells can protect against oxidative stress and apoptosis induced by arsenite. Initially, our observations revealed that arsenite exposure increased oxidative stress and triggered apoptotic cell death in human umbilical vein endothelial cells (HUVECs). However, treatment with the apoptosis inhibitor Z-VAD-FMK notably reduced arsenite-induced apoptosis. Additionally, arsenite activated the autophagy pathway and enhanced autophagic flux in HUVECs. Interestingly, inhibition of autophagy exacerbated arsenite-induced apoptotic cell death. Our findings also demonstrated the importance of autophagy receptor NBR1 in arsenite-induced cytotoxicity, as it facilitated the recruitment of caspase 8 to autophagosomes for degradation. The protective effect of NBR1 against arsenite-induced apoptosis was compromised when autophagy was inhibited using pharmacological inhibitors or through genetic knockdown of essential autophagy genes. Conversely, overexpression of NBR1 facilitated caspase 8 degradation and reduced apoptotic cell death in arsenite-treated HUVECs. In conclusion, our study highlights the vital role of NBR1-mediated autophagic degradation of caspase 8 in safeguarding vascular endothelial cells from arsenite-induced oxidative stress and apoptotic cell death. Targeting this pathway could offer a promising therapeutic approach to mitigate cardiovascular diseases associated with arsenite exposure.
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Apoptosis , Arsenitos , Autofagia , Caspasa 8 , Células Endoteliales de la Vena Umbilical Humana , Estrés Oxidativo , Humanos , Arsenitos/toxicidad , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos , Caspasa 8/metabolismo , Caspasa 8/genética , Estrés Oxidativo/efectos de los fármacos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Proteolisis/efectos de los fármacos , Células CultivadasRESUMEN
Soil salinity pose a significant challenge to global agriculture, threatening crop yields and food security. Understanding the salt tolerance mechanisms of plants is crucial for improving their survival under salt stress. AFP2, a negative regulator of ABA signaling, has been shown to play a crucial role in salt stress tolerance during seed germination. Mutations in AFP2 gene lead to increased sensitivity to salt stress. However, the underline mechanisms by which AFP2 regulates seed germination under salt stress remain elusive. In this study, we identified a protein interaction between AFP2 and SOS2, a Ser/Thr protein kinase known to play a critical role in salt stress response. Using a combination of genetic, biochemical, and physiological approaches, we investigated the role of the SOS2-AFP2 module in regulating seed germination under salt stress. Our findings reveal that SOS2 physically interacts with AFP2 and stabilizes it, leading to the degradation of the ABI5 protein, a negative transcription factor in seed germination under salt stress. This study sheds light on previously unknown connections within salt stress and ABA signaling, paving the way for novel strategies to enhance plant resilience against environmental challenges.
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Proteínas de Arabidopsis , Arabidopsis , Germinación , Estrés Salino , Semillas , Arabidopsis/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Germinación/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Proteolisis/efectos de los fármacos , Tolerancia a la Sal/genética , Semillas/metabolismo , Semillas/efectos de los fármacos , Semillas/crecimiento & desarrollo , Semillas/genética , Transducción de Señal/efectos de los fármacosRESUMEN
Proteolysis-targeting chimera (PROTAC) has become a very important means of protein degradation and a new way of disease treatment. In particular, PROTACs constructed with ligands for E3 ligase cereblon account for more than 90 % of the PROTACs currently in clinical research. Notably, CRBN ligands themselves are a class of molecular glue compounds capable of degrading neo-substrate proteins. Compared to the target proteins degradation, the degradation of neo-substrates, especially IKZF2, has not received enough attention. Therefore, this review summarizes the currently published IKZF2 degraders derived from articles and patents, which are conducive to the design of PROTACs with desired IKZF2 degradation from the perspective of medicinal chemistry.
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Proteolisis , Ubiquitina-Proteína Ligasas , Ubiquitina-Proteína Ligasas/metabolismo , Proteolisis/efectos de los fármacos , Humanos , Diseño de Fármacos , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Ligandos , Factor de Transcripción Ikaros/metabolismo , Quimera Dirigida a la ProteólisisRESUMEN
The proteasome is a multisubunit protease system responsible for the majority of the protein turnover in eukaryotic organisms. Dysregulation of this enzymatic complex leads to protein accumulation, subsequent aggregation, and ultimately diseased states; for that reason, positive modulation of its activity has been recently investigated as a therapeutic strategy for neurodegenerative and age-related diseases. The small molecule AM404 was recently identified as an activator of the 20S isoform of the proteasome and further exploration of the scaffold revealed the importance of the polyunsaturated fatty acid chain to elicit activity. Herein, we report the investigation of the aromatic region of the scaffold and the evaluation of the small molecules in a variety of proteasome activity and protein degradation assays. We found that derivatives A22 and A23, compared to AM404, exhibit enhanced proteasome activity in biochemical and cellular proteasome assays and more favorable cellular viability profiles. Additionally, these compounds demonstrate the ability to degrade intrinsically disordered proteins, regardless of their molecular weight, and the ability to restore the proteasome activity in the presence of toxic oligomeric α-Syn species in a biochemical setting.