Ubiquitin-proteasome Degradation Research Articles

Targeting PPARγ via SIAH1/2-mediated ubiquitin-proteasomal degradation as a new therapeutic approach in luminal-type bladder cancer

Bladder cancer (BC) is the second most prevalent genitourinary malignancy worldwide. Despite recent approvals of immune checkpoint inhibitors and targeted therapy for muscle invasive or recurrent BC, options remain limited for patients with non-muscle invasive BC (NMIBC) refractory to Bacillus Calmette-Guérin (BCG) and chemotherapy. NMIBC is more frequently classified as a luminal subtype, in which increased PPARγ activity is a key feature in promoting tumor growth and evasion of immunosurveillance. Cinobufotalin is one of the major compound of bufadienolides, the primary active components of toad venom that has been utilized in the clinical treatment of cancer. We herein focused on cinobufotalin, examining its anticancer activity and molecular mechanisms in luminal-type NMIBC. Our results newly reveal that cinobufotalin strongly suppresses the viability and proliferation of luminal BC cells with minimal cytotoxic effects on normal uroepithelial cells, and exhibits significant antitumor activity in a RT112 xenograft BC model. Mechanistically, our sub-G1-phase cell accumulation, Annexin V staining, caspase-3/8/9 activation, and PARP activation analyses show that cinobufotalin induces apoptosis in luminal-type BC cells. Cinobufotalin significantly inhibited the levels of PPARγ and its downstream targets, as well as lipid droplet formation and free fatty acid levels in RT112 cells. PPARγ overexpression rescued RT112 cells from cinobufotalin-induced apoptosis and mitigated the downregulation of FASN and PLIN4. Finally, we show seemingly for the first time that cinobufotalin promotes SIAH1/2-mediated proteasomal degradation of PPARγ in luminal BC cells. Together, these findings compellingly support the idea that cinobufotalin could be developed as a promising therapeutic agent for treating luminal-type NMIBC.

Targeting the SMURF2-HIF1α axis: a new frontier in cancer therapy.

The SMAD-specific E3 ubiquitin protein ligase 2 (SMURF2) has emerged as a critical regulator in cancer biology, modulating the stability of Hypoxia-Inducible Factor 1-alpha (HIF1α) and influencing a network of hypoxia-driven pathways within the tumor microenvironment (TME). SMURF2 targets HIF1α for ubiquitination and subsequent proteasomal degradation, disrupting hypoxic responses that promote cancer cell survival, metabolic reprogramming, angiogenesis, and resistance to therapy. Beyond its role in HIF1α regulation, SMURF2 exerts extensive control over cellular processes central to tumor progression, including chromatin remodeling, DNA damage repair, ferroptosis, and cellular stress responses. Notably, SMURF2's ability to promote ferroptotic cell death through GSTP1 degradation offers an alternative pathway to overcome apoptosis resistance, expanding therapeutic options for refractory cancers. This review delves into the multifaceted interactions between SMURF2 and HIF1α, emphasizing how their interplay impacts metabolic adaptations like the Warburg effect, immune evasion, and therapeutic resistance. We discuss SMURF2's dual functionality as both a tumor suppressor and, in certain contexts, an oncogenic factor, underscoring its potential as a highly versatile therapeutic target. Furthermore, modulating the SMURF2-HIF1α axis presents an innovative approach to destabilize hypoxia-dependent pathways, sensitizing tumors to chemotherapy, radiotherapy, and immune-based treatments. However, the complexity of SMURF2's interactions necessitate a thorough assessment of potential off-target effects and challenges in specificity, which must be addressed to optimize its clinical application. This review concludes by proposing future directions for research into the SMURF2-HIF1α pathway, aiming to refine targeted strategies that exploit this axis and address the adaptive mechanisms of aggressive tumors, ultimately advancing the landscape of precision oncology.

Peli1 Deficiency in Macrophages Attenuates Pulmonary Hypertension by Enhancing Foxp1-Mediated Transcriptional Inhibition of IL-6.

The infiltration of macrophages into the lungs is a common characteristic of perivascular inflammation, contributing to vascular remodeling in pulmonary hypertension (PH). Peli1 (pellino E3 ubiquitin-protein ligase 1) plays a critical role in regulating the production of proinflammatory cytokines and the polarization of macrophages in various diseases. However, the role of Peli1 in PH remains to be investigated. The expression and biological function of Peli1 were investigated in both human and experimental models of PH. Peli1-deficient mice and bone marrow transplant mice were utilized to explore the roles of Peli1 in macrophages in vivo. Proteomic analysis and molecular biology techniques were used to uncover the underlying mechanisms. The upregulation of Peli1 in the lungs and alveolar macrophages was observed in hypoxia-treated mice. Peli1 knockout mice and myeloid Peli1-deficient mice significantly ameliorated hypoxia-induced right ventricular systolic pressure, right ventricular hypertrophy, and pulmonary vascular remodeling. Mechanistically, Peli1 facilitated the ubiquitination and subsequent proteasomal degradation of Foxp1 (forkhead box p1), thereby alleviating its suppression of IL (interleukin)-6 transcription and contributing to macrophage activation. Furthermore, myeloid Foxp1 deficiency partially eliminates the protective effect of myeloid Peli1 deficiency in hypoxia-induced PH mice. Our findings demonstrate that the Peli1-Foxp1-IL-6 pathway plays a crucial role in macrophage activation and recruitment during the development of PH, underscoring the potential of Peli1 as a therapeutic target for PH.

Resurrecting p53 by an Artificial Nano-Protein for Potent Photodynamic Retinoblastoma Therapy.

The treatment of retinoblastoma (RB), a formidable eye cancer that affects infants and children, is not only aimed at saving lives but at preserving ocular function, maintaining optimal visual acuity, and enhancing the overall quality of life. Photodynamic therapy has already been established as a secure and dependable therapeutic modality for the treatment of ocular diseases that effectively preserves ocular function; however, it fails to provide satisfactory outcomes against RB. To address this formidable challenge, groundbreaking advancement is aspired by delving into the genetic characteristics of RB, which initially involves the wild-type p53 pathway but is subsequently suppressed by MDM2 and MDMX. In this study, an artificial nanoprotein (ANP) is developed through the fusion of human serum albumin with a potent MDM2/MDMX degrading peptide and the porphyrin-photosensitizer verteporfin (VP). ANP effectively dismantles MDM2/MDMX proteins within the ubiquitin-proteasome degradation pathway and triggers VP-PDT-mediated cell death through mitochondrial impairment. Remarkably, ANP exhibited remarkable therapeutic efficacy in both subcutaneous and in situ RB mouse models, while maintaining a highly favorable biosafety profile. Collectively, ANP not only provides compelling evidence for sensitizing photodynamic RB therapy by reactivating p53, but also serves as an exceptionally potent photosensitizer with promising potential for clinical translation in RB treatment.

Loss of ERα involved-HER2 induction mediated by the FOXO3a signaling pathway in fulvestrant-resistant breast cancer

Patients with estrogen receptor alpha (ERα)-positive breast cancer are commonly treated with anti-estrogen drugs as an initial treatment strategy. Fulvestrant, an estrogen receptor antagonist, effectively blocks ERα signaling; however, long-term fulvestrant treatment induces drug resistance in the absence of ERα. In this study, we investigated the molecular mechanism underlying the loss of ERα, FOXO3a, and induction of HER2 in fulvestrant-resistant breast cancer. Short-term fulvestrant treatment degraded ERα proteins via the ubiquitin-proteasome degradation pathway in MCF7 cells. MCF7 cells turn into highly proliferative cells (fulvestrant-resistant cells: Ful-R) after long-term fulvestrant treatment. These cells exhibit markedly suppressed estrogen and progesterone receptor levels. The phosphorylation of EGFR, HER2, and ERK was induced in Ful-R, and these phosphorylation inhibitors suppressed cell proliferation in Ful-R. FOXO3a, a transcriptional regulator of ERα was decreased in Ful-R, and a ubiquitin-proteasome inhibitor restored the expression of FOXO3a. These results suggest that the suppression of FOXO3a and ERα led to the increased expression of TGF-α, EGFR, and HER2 and subsequent cell proliferation in Ful-R. This study highlights the potential development of therapeutic drugs targeting FOXO3a for the treatment of HER2-positive, estrogen, and progesterone receptor-negative her2-type proliferative breast cancers.

Chromatin Helicase CHD6 Establishes Pro-inflammatory Enhancers and is a Synthetic Lethal Target in FH-Deficient Renal Cell Carcinoma.

Fumarate hydratase (FH) deficiency causes hereditary leiomyomatosis and renal cell carcinoma (RCC). FH-deficient tumors lack effective therapeutic options. Here, we utilized an epigenetic-focused single-guide RNA library to elucidate potential drug targets in FH-deficient tumors. The screen identified chromodomain helicase DNA binding protein 6 (CHD6) as an essential regulator of the growth of FH-mutated RCC. Mechanically, FH loss induced fumarate-mediated succinylation and inactivation of KEAP1, blocking subsequent ubiquitin-proteasome degradation of CHD6. Stabilized CHD6 formed a complex with p65 to establish pro-inflammatory enhancers and thereby regulate NF-κB-mediated transcription. Moreover, CHD6 recruited mSWI/SNF ATPases to maintain chromatin accessibility at CHD6-bound enhancers. The PROTAC degrader of SMARCA2/4 AU-15330 effectively abolished structures of cis-regulatory elements bound by CHD6 and suppressed the growth of FH-mutated, but not FH-intact, RCC in vivo. Collectively, these data indicate that CHD6 is a molecular bridge between FH deficiency and pro-inflammatory enhancers assembly that endows FH-deficient tumors with epigenetic vulnerabilities.

MicroRNA-145-5p inhibits the tumorigenesis of breast cancer through SENP2-regulated ubiquitination of ERK2

Breast carcinoma exhibits the highest incidence among various cancers and is the foremost cause of mortality in women. Increasing evidence shows that SUMOylation of proteins plays a critical role in the progression of breast cancer; however, the role of SENP2 and its molecular mechanism in breast cancer remain underexplored. Here, we discerned that SENP2 promoted the tumorigenesis of breast cancer both in vitro and in vivo. Furthermore, we identified that ERK2 was SUMOylated and that SENP2 played a role by deconjugating ERK2 SUMOylation in breast cancer. SUMOylation of ERK2 promoted its ubiquitin-proteasomal degradation, thus inhibiting the epithelial-to-mesenchymal transition in breast cancer cells. Furthermore, microRNA-145-5p (miR-145-5p) has emerged as a scarce commodity in breast cancer and binds to the 3’-untranslated region of SENP2 mRNA to govern the regulatory dynamics of SENP2 expression. Finally, miR-145-5p inhibits SENP2 transcription, enhances ERK2 SUMOylation, and ultimately suppresses the progression of breast cancer. These revelations suggest evolving ideas for the miR-145-5p-SENP2 axis in therapeutic intervention, thus heralding transformative prospects for the clinical management of breast cancer.

Mbnl1-mediated alternative splicing of circMlxipl regulates Rbbp6-involved ChREBP turnover to inhibit lipotoxicity-induced β-cell damage

BackgroundDiabetes, a global epidemic, is the leading cause of mortality globally. The aim of this study is to get better understanding of pathophysiology of diabetes.MethodsPalmitic acid (PA)-treated β-cells, db/db mice and high fat diet (HFD)-fed mouse model of type 2 diabetes were established. H&E was used to assess the histological changes of pancreas. IHC, FISH, western blot or qRT-PCR was employed to detect the expression of key molecules in primary islets or lipotoxic β-cells. Cell behaviors were detected by MTT, EdU incorporation assay, TUNEL assay and glucose-induced insulin secretion (GSIS). The associations among circMlxipl, Mbnl1 and Rbbp6 were validated by RIP and RNA pull-down assays, and the direct binding between Hdac3 and Mbnl1 promoter was examined by ChIP and luciferase assays. Co-IP was employed to assess the interaction between ChREBP and Rbbp6, as well as the ubiquitination of ChREBP.ResultsHdac3 and ChREBP were upregulated, but Mbnl1 and circMlxipl were downregulated in islets from diabetic mice and lipotoxic β-cells. Mbnl1 overexpression protected against PA-induced impairments in lipotoxic β-cells through modulating back-splicing of circMlxipl and suppressing ChREBP. Hdac3 served as a transcriptional repressor of Mbnl1, and it was implicated in circMlxipl-mediated protection via regulating ChREBP expression in lipotoxic β-cells. Lack of circMlxipl inhibited Rbbp6-mediated ubiquitin-proteasomal degradation of ChREBP in lipotoxic β-cells. In vivo studies revealed that Hdac3 knockdown or Mbnl1 overexpression alleviated diabetes symptoms through circMlxipl-regulated ChREBP in diabetic mice.ConclusionMbnl1-mediated alternative splicing of circMlxipl regulates Rbbp6-involved ChREBP turnover to inhibit lipotoxicity-induced β-cell damage.

SPOP-mediated RIPK3 destabilization desensitizes LPS/sMAC/zVAD-induced necroptotic cell death

RIPK1/RIPK3-MLKL signaling molecules are fundamental in initiating necroptotic cell death, but their roles in the development of colon cancer are unclear. This study reports that RIPK3 interacted with SPOP, a component of the E3 ligase within the Cul3 complex. This interaction leads to K48-linked ubiquitination and subsequent proteasomal degradation of RIPK3. Two distinct degron motifs, PETST and SPTST, were identified within the linker domain of RIPK3 for SPOP. RIPK3 phosphorylations at Thr403 by PIM2 and at Thr412/Ser413 by ERK2 are essential to facilitate its interaction with SPOP. Computational docking studies and immunoprecipitation analyses showed that these PIM2 and ERK2 phosphorylations bolster the stability of the RIPK3-SPOP interaction. In particular, mutations of RIPK3 at the degron motifs extended the half-life of RIPK3 by preventing its phosphorylation and subsequent ubiquitination. The deletion of SPOP, which led to increased stability of the RIPK3 protein, intensified LPS/sMAC/zVAD-induced necroptotic cell death in colon cancer cells. These findings underscore the critical role of the SPOP-mediated RIPK3 stability regulation pathway in controlling necroptotic cell death.

P53 and the E3 Ubiquitin Ligase MDM2 in Glaucomatous Lamina Cribrosa Cells.

Lamina cribrosa (LC) cells play an integral role in extracellular matrix remodeling and fibrosis in human glaucoma. LC cells bear similarities to myofibroblasts that adopt an apoptotic-resistant, proliferative phenotype, a process linked to dysregulation of tumor suppressor-gene p53 pathways, including ubiquitin-proteasomal degradation via murine-double-minute-2 (MDM2). Here, we investigate p53 and MDM2 in glaucomatous LC cells. Primary human LC cells were isolated from glaucomatous donor eyes (GLC) and age-matched normal controls (NLC) (n = 3 donors/group). LC cells were cultured under standard conditions ± 48-h treatment with p53-MDM2-interaction inhibitor RG-7112. Markers of p53-MDM2, fibrosis, and apoptosis were analyzed by real-time polymerase chain reaction (qRT-PCR), western blotting, and immunofluorescence. Cellular proliferation and viability were assessed using colorimetric methyl-thiazolyl-tetrazolium salt assays (MTS/MTT). In GLC versus NLC cells, protein expression of p53 was significantly decreased (p < 0.05), MDM2 was significantly increased, and immunofluorescence showed reduced p53 and increased MDM2 expression in GLC nuclei. RG-7112 treatment significantly increased p53 and significantly decreased MDM2 gene and protein expression. GLC cells had significantly increased protein expression of αSMA, significantly decreased caspase-3 protein expression, and significantly increased proliferation after 96 h. RG-7112 treatment significantly decreased COL1A1 and αSMA, significantly increased BAX and caspase-3 gene expression, and significantly decreased proliferation in GLC cells. MTT-assay showed equivocal cellular viability in NLC/GLC cells with/without RG-7112 treatment. Our data suggests that proliferation and the ubiquitin-proteasomal pathway are dysregulated in GLC cells, with MDM2-led p53 protein degradation negatively impacting its protective role.

Ubiquitination of Hemocyanin Mediated by a Mitochondrial E3 Ubiquitin Ligase Regulates Immune Response in Penaeus vannamei.

Ubiquitination is a critical posttranslational modification that regulates host immune responses to pathogens. In this study, we investigated the ubiquitination of hemocyanin (PvHMC [Penaeus vannamei hemocyanin]) mediated by the mitochondrial E3 ubiquitin ligase (PvMulan) in shrimp Penaeus vannamei. We characterized distinct ubiquitination patterns of PvHMC in response to different pathogen challenges, both invitro and invivo. Specifically, we found that Vibrio parahaemolyticus infection led to an increase in PvMulan, which resulted in K48-linked ubiquitination and subsequent proteasomal degradation of PvHMC. In contrast, PvMulan primarily enhanced the SUMOylation of PvHMC, bolstering its immune functions against white spot syndrome virus challenges. Inhibition of PvMulan-mediated PvHMC ubiquitination significantly affected the proliferation of V. parahaemolyticus and the survival rate of infected shrimps. This study sheds light on the role of hemocyanin ubiquitination in immune regulation, illustrating its dual function in response to distinct pathogens.

Spatial and Single Cell Analyses Reveal Heterogeneity of DNAM-1 Receptor-Ligand Interactions that Instructs Intratumoral γδT-Cell Activity.

The dynamic interplay between tumor cells and γδT cells within the tumor microenvironment (TME) significantly influences disease progression and immunotherapy outcome. Here, we delved into the modulation of γδT-cell activation by tumor cell ligands CD112 and CD155, which interact with the activating receptor DNAM-1 on γδT cells. Spatial and single cell RNA sequencing (scRNA-seq), as well as spatial metabolome analysis, from neuroblastoma (NB) revealed that the expression levels and localization of CD112 and CD155 varied across and within tumors, correlating with differentiation status, metabolic pathways, and ultimately disease prognosis and patient survival. Both in vivo tumor xenograft experiments and in vitro co-culture experiments demonstrated that a high CD112/CD155 expression ratio in tumors enhanced γδT-cell-mediated cytotoxicity, while a low-ratio fostered tumor resistance. Mechanistically, CD112 sustained DNAM-1-mediated γδT-cell activation, whereas CD155 downregulated DNAM-1 expression via TRIM21-mediated ubiquitin proteasomal degradation. By interacting with tumor cells differentially expressing CD112 and CD155, intratumoral γδT cells exhibited varying degrees of activation and DNAM-1 expression, representing three major functional subsets. This study underscores the complexity of tumor-immune crosstalk, offering insights into how tumor heterogeneity shapes the immune landscape.

NEDD4 Knockdown Suppresses Human Endometrial Stromal Cell Growth and Invasion by Regulating PTGS2-Mediated Ferroptosis in Endometriosis.

Endometriosis (EM) is a gynecological disease characterized by the benign growth of endometrial tissue outside the uterus. Upregulation of neuronally expressed developmentally downregulated 4 (NEDD4) has been reported to accelerate endometrial cancer progression. We explored whether abnormal expression of NEDD4 is correlated with EM. Endometrial tissue in patients without endometriosis was used to develop the original generation of endometrial stromal cells (ESCs). Different types of endometrial tissue of patients with endometriosis were used to measure the expression of NEDD4 by immunohistochemistry (IHC) and western blotting. Its biological functions in ESCs were investigated using a cell counting kit-8 assay, fluorescein diacetate (FDA) staining, and Transwell invasion assays. Additionally, its involvement in ferroptosis was assessed by measuring Fe2+, malondialdehyde (MDA), glutathione (GSH), and reactive oxygen species (ROS) levels and the expression of ferroptosis markers. Compared with normal controls, NEDD4 levels were significantly elevated in the endometrial tissue of patients with EM. Furthermore, NEDD4 expression was higher in the ectopic endometrium than in the eutopic endometrium. NEDD4 knockdown reduced the viability and invasive capacity of ESCs, increased Fe2+, MDA, and ROS levels, and decreased GSH content. Further analysis revealed that NEDD4 knockdown promoted ferroptosis in ESCs by increasing the expression of prostaglandin-endoperoxide synthase 2 (PTGS2). As an E3 ubiquitin ligase, NEDD4 reduced PTGS2 protein levels by accelerating its ubiquitination and subsequent proteasomal degradation. These findings suggest that inhibiting NEDD4 reduces ESC growth and invasion in EM by regulating PTGS2-dependent ferroptosis.

PHLDA1 protects intestinal barrier function via restricting intestinal epithelial cells apoptosis in inflammatory bowel disease

The current approach to treating inflammatory bowel disease (IBD) primarily focuses on managing inflammation rather than maintaining the integrity of the intestinal barrier. In our study, we sought to investigate the potential role of PHLDA1 in preserving intestinal barrier function as a promising strategy for treating IBD. We observed a significant decrease in PHLDA1 expression in intestinal epithelial cells (IECs) of both IBD patients and mice with chemically induced colitis. This deficiency of PHLDA1 led to increased apoptosis of IECs, resulting in a compromised epithelial barrier and the invasion of commensal bacteria into the mucosa. Consequently, this microbial invasion substantially exacerbated colonic inflammation in mice with the specific knockout of PHLDA1 in IECs (Phlda1IEC−KO) compared to their control littermates. Mechanistically, we found evidence of PHLDA1 interacting with MCL1 to protect against K48-linked polyubiquitylation at the K40 lysine residue, thus preventing ubiquitin-proteasome degradation through the MCL1 ubiquitin ligase E3 (Mule). We further confirmed that the PHLDA1-MCL1-Mule signaling pathway plays a critical role in the development of IBD. Notably, our study demonstrated that enhancing MCL1 levels or reducing Mule expression using adeno-associated virus (AAV) attenuated experimental colitis in Phlda1IEC−KO mice. Collectively, our findings emphasize the significance of PHLDA1 in the pathogenesis of IBD and propose that targeting the PHLDA1-MCL1-Mule signaling pathway could be a viable approach for combating IBD.

The AKR1C1-CYP1B1-cAMP signaling axis controls tumorigenicity and ferroptosis susceptibility of extrahepatic cholangiocarcinoma.

Extrahepatic cholangiocarcinoma (ECC), a highly malignant type of cancer with increasing incidence, has a poor prognosis due to limited treatment options. Based on genomic analysis of ECC patient samples, here we report that aldo-keto reductase family 1 member C1 (AKR1C1) is highly expressed in human ECC tissues and closely associated with ECC progression and poor prognosis. Intriguingly, we show that inducible AKR1C1 knockdown triggers ECC cells to undergo ferroptosis. Mechanistically, AKR1C1 degrades the protein stability of the cytochrome P450 family member CYP1B1, a newly discovered mediator of ferroptosis, via ubiquitin-proteasomal degradation. Additionally, AKR1C1 decreases CYP1B1 mRNA level through the transcriptional factor aryl-hydrocarbon receptor (AHR). Furthermore, the AKR1C1-CYP1B1 axis modulates ferroptosis in ECC cells via the cAMP-PKA signaling pathway. Finally, in a xenograft mouse model of ECC, AKR1C1 depletion sensitizes cancer cells to ferroptosis and synergizes with ferroptosis inducers to suppress tumor growth. Therefore, the AKR1C1-CYP1B1-cAMP signaling axis is a promising therapeutic target for ECC treatment, especially in combination with ferroptosis inducers.

Combined targeting of GPX4 and BCR-ABL tyrosine kinase selectively compromises BCR-ABL+ leukemia stem cells

BackgroundIn the ongoing battle against BCR-ABL+ leukemia, despite significant advances with tyrosine kinase inhibitors (TKIs), the persistent challenges of drug resistance and the enduring presence of leukemic stem cells (LSCs) remain formidable barriers to achieving a cure.MethodsIn this study, we demonstrated that Disulfiram (DSF) induces ferroptosis to synergize with TKIs in inhibiting BCR-ABL+ cells, particularly targeting resistant cells and LSCs, using cell models, mouse models, and primary cells from patients. We elucidated the mechanism by which DSF promotes GPX4 degradation to induce ferroptosis through immunofluorescence, co-immunoprecipitation (CO-IP), RNA sequencing, lipid peroxidation assays, and rescue experiments.ResultsHere, we present compelling evidence elucidating the sensitivity of DSF, an USA FDA-approved drug for alcohol dependence, towards BCR-ABL+ cells. Our findings underscore DSF’s ability to selectively induce a potent cytotoxic effect on BCR-ABL+ cell lines and effectively inhibit primary BCR-ABL+ leukemia cells. Crucially, the combined treatment of DSF with TKIs selectively eradicates TKI-insensitive stem cells and resistant cells. Of particular note is DSF’s capacity to disrupt GPX4 stability, elevate the labile iron pool, and intensify lipid peroxidation, ultimately leading to ferroptotic cell death. Our investigation shows that BCR-ABL expression induces alterations in cellular iron metabolism and increases GPX4 expression. Additionally, we demonstrate the indispensability of GPX4 for LSC development and the initiation/maintenance of BCR-ABL+ leukemia. Mechanical analysis further elucidates DSF’s capacity to overcome resistance by reducing GPX4 levels through the disruption of its binding with HSPA8, thereby promoting STUB1-mediated GPX4 ubiquitination and subsequent proteasomal degradation. Furthermore, the combined treatment of DSF with TKIs effectively targets both BCR-ABL+ blast cells and drug-insensitive LSCs, conferring a significant survival advantage in mouse models.ConclusionIn summary, the dual inhibition of GPX4 and BCR-ABL presents a promising therapeutic strategy to synergistically target blast cells and drug-insensitive LSCs in patients, offering potential avenues for advancing leukemia treatment.

Activation of nuclear receptor pregnane-X-receptor protects against abdominal aortic aneurysm by inhibiting oxidative stress

Abdominal aortic aneurysm (AAA) is a life-threatening condition, but effective medications to prevent its progression and rupture are currently lacking. The nuclear receptor pregnane-X-receptor (PXR) plays a crucial role in vascular homeostasis. However, the role of PXR in AAA development remains unknown. We first detected the PXR expression in human and murine AAA tissues by RT-qPCR and Western blot. To investigate the potential role of PXR in the development of AAA, we used adeno-associated virus-mediated overexpression of PXR and pharmacological activation of PXR by ginkgolide A (GA) in mouse AAA models induced by both angiotensin II (AngII) and calcium phosphate [Ca3(PO4)2]. The underlying mechanism was further explored using RNA-sequencing and molecular biological analyses. We found a significant decrease in both mRNA and protein levels of PXR in both human and murine aortic smooth muscle cells from AAA tissues, accompanied with phenotypic switching of vascular smooth muscle cell and increased oxidative stress. PXR overexpression in abdominal aortas and GA treatment successfully suppressed AAA formation in both mouse AAA models. RNA-sequencing data revealed that PXR activation inhibited gamma-aminobutyric acid type A receptor subunit alpha3 (GABRA3) expression. Additional mechanistic studies identified that PXR suppressed AAA through mitigating GABRA3-induced reactive oxygen species (ROS) generation and subsequent phosphorylation of c-Jun N-terminal kinase (JNK). Interestingly, p-JNK was found to induce ubiquitin-proteasome degradation of PXR. In summary, our data unveiled, for the first time, the protective role of PXR against AAA pathogenesis by inhibiting oxidative stress. These findings suggested PXR as a promising therapeutic target for AAA.

USP10 promotes cell proliferation, migration, and invasion in NSCLC through deubiquitination and stabilization of EIF4G1

Lung cancer is one of the most common types of malignant cancer worldwide, causing a serious social and economic burden. It is classified into non-small cell lung cancer (NSCLC) and small cell lung cancer, with NSCLC accounting for 80–85% of cases. Eukaryotic translation initiation factor 4 gamma 1 (EIF4G1) is highly expressed in NSCLC, playing an important role in regulating tumor growth, angiogenesis, malignant transformation, and phagocytosis. Ubiquitin-specific protease 10 (USP10) functions as a deubiquitinating enzyme to regulate substrate protein deubiquitination and reverse the ubiquitin proteasome degradation pathway. Our previous study identified an interaction between EIF4G1 and USP10; however, their regulatory mechanism remains unclear. Herein, we found that USP10 positively regulates EIF4G1 in NSCLC cells. An in vivo ubiquitination assay demonstrated deubiquitination of EIF4G1 by USP10, which reversed the ubiquitin proteasomal degradation of EIF4G1, thereby increasing its stability. Upregulation of EIF4G1 promoted cell proliferation, migration, and invasion in NSCLC cells. The current study not only reveals a novel mechanism through which USP10 positively regulates EIF4G1 in NSCLC, but also demonstrates the potential of USP10 as a therapeutic target to treat NSCLC.

Multi-omics analyses were combined to construct ubiquitination-related features in colon adenocarcinoma and identify ASNS as a novel biomarker.

As one of the malignant tumors with the highest incidence and fatality in the world, colon adenocarcinoma (COAD) has a very complex pathogenic mechanism, which has not yet been fully elucidated. Ubiquitin can regulate cell proliferation, cell cycle, apoptosis, DNA damage repair, and other processes by changing the activity of substrate proteins or causing ubiquitin-proteasome degradation. These are the key links in the pathogenesis of COAD, and ubiquitin plays an important role in the occurrence and development of COAD. We integrated transcriptomics, single-cell and clinical omics, and TCGA and GEO databases of COAD patient data. Cox and Lasso regression was employed to assess ubiquitination genes in COAD for generating ubiquitination-related features. The aim was to evaluate the prognostic value of these features for tumors and their impact on the immune microenvironment. At the same time, the expression level of model genes was further analyzed using single-cell data. Finally, the expression and function of ASNS, a key gene for this trait, were detected in vitro. In our study, based on identifiable changes in the expression of marker genes, this feature can be used to classify patients with COAD. Kaplan-Meier survival analysis indicated that those with elevated risk scores in each cohort experienced inferior outcomes. There is good validation in both the training queue and the validation queue. The results of the immune infiltration analysis showed that the immune infiltration rate was significantly increased in the high-risk group. After the knockdown of ASNS, an important gene in the signature, the activity and migration capacity of SW620 and RKO cell lines and colony formation capacity were dramatically reduced in cell tests. We screened ubiquitination-related genes and constructed ubiquitination-related features, which can be used as reliable prognostic indicators of COAD. ASNS was identified as a possible biomarker for COAD.

Penfluridol inhibits melanoma growth and metastasis through enhancing von Hippel‒Lindau tumor suppressor-mediated cancerous inhibitor of protein phosphatase 2A (CIP2A) degradation.

Melanoma's high metastatic potential, especially to the brain, poses significant challenges to patient survival. The blood‒brain barrier (BBB) is a major obstacle to the effective treatment of melanoma brain metastases. We screened antipsychotic drugs capable of crossing the BBB and identified penfluridol (PF) as the most active candidate. PF reduced melanoma cell viability and induced apoptosis. In animal models, PF effectively inhibited melanoma growth and metastasis to the lung and brain. Using immunoprecipitation combined with high-resolution mass spectrometry, and other techniques such as drug affinity responsive target stability, we identified CIP2A as a direct binding protein of PF. CIP2A is highly expressed in melanoma and its metastases, and is linked to poor prognosis. PF can restore Protein Phosphatase 2A activity by promoting CIP2A degradation, thereby inhibiting several key oncogenic pathways, including AKT and c-Myc. Additionally, von Hippel‒Lindau (VHL) is the endogenous E3 ligase for CIP2A, and PF enhances the interaction between VHL and CIP2A, promoting the ubiquitin‒proteasome degradation of CIP2A, thereby inhibiting melanoma growth and metastasis. Overall, this study not only suggests PF's potential in treating melanoma and its brain metastases but also highlights CIP2A degradation as a therapeutic strategy for melanoma.