Ubiquitin-like Modifier-activating Enzyme Research Articles

Efficacy and safety of targeted therapies in VEXAS syndrome: retrospective study from the FRENVEX

ObjectivesVacuoles, E1 enzyme, X-linked, autoinflammatory and somatic (VEXAS) syndrome is an adult-onset autoinflammatory disease associated with somatic ubiquitin-like modifier-activating enzyme 1 (UBA1) mutations. We aimed to evaluate the efficacy and...

UBA6 Inhibition Accelerates Lysosomal TRPML1 Depletion and Exosomal Secretion in Lung Cancer Cells.

Ubiquitin-like modifier-activating enzyme 6 (UBA6) is a member of the E1 enzyme family, which initiates the ubiquitin-proteasome system (UPS). The UPS plays critical roles not only in protein degradation but also in various cellular functions, including neuronal signaling, myocardial remodeling, immune cell differentiation, and cancer development. However, the specific role of UBA6 in cellular functions is not fully elucidated in comparison with the roles of the UPS. It has been known that the E1 enzyme is associated with the motility of cancer cells. In this study, we verified the physiological roles of UBA6 in lung cancer cells through gene-silencing siRNA targeting UBA6 (siUBA6). The siUBA6 treatment attenuated the migration of H1975 cells, along with a decrease in lysosomal Ca2+ release. While autophagosomal proteins remained unchanged, lysosomal proteins, including TRPML1 and TPC2, were decreased in siUBA6-transfected cells. Moreover, siUBA6 induced the production of multivesicular bodies (MVBs), accompanied by an increase in MVB markers in siUBA6-transfected H1975 cells. Additionally, the expression of the exosomal marker CD63 and extracellular vesicles was increased by siUBA6 treatment. Our findings suggest that knock-down of UBA6 induces lysosomal TRPML1 depletion and inhibits endosomal trafficking to lysosome, and subsequently, leads to the accumulation of MVBs and enhanced exosomal secretion in lung cancer cells.

Comprehensive morphologic characterization of bone marrow biopsy findings in a large cohort of patients with VEXAS syndrome: A single-institution longitudinal study of 111 bone marrow samples from 52 patients.

VEXAS syndrome is an adult-onset autoinflammatory disease caused by a somatic pathogenic mutation in the UBA1 (ubiquitin-like modifier activating enzyme 1) gene. Patients present with rheumatologic manifestations and cytopenias and may have an increased predisposition to myelodysplastic syndrome (MDS) and plasma cell neoplasms. Prior studies have reported on the peripheral blood and bone marrow findings in patients with VEXAS syndrome. Due to the protean clinical presentation and lack of specificity of morphologic features (eg, vacuoles in early erythroid and granulocytic precursors), an optimal screening methodology to identify these patients in a timely fashion is desirable. To further evaluate and describe the salient diagnostic morphologic features in VEXAS syndrome, we carried out a comprehensive study of the largest single-institution cohort to date. Diagnostic and follow-up bone marrow biopsy specimens from 52 male patients with molecularly identified VEXAS syndrome underwent central review. Cytopenias were common in all cases, primarily macrocytic anemia, monocytopenia, and thrombocytopenia. Bone marrow aspirate and biopsy were often hypercellular, with an increased myeloid/erythroid ratio, granulocytic hyperplasia with left shift, erythroid left shift, and megakaryocyte hyperplasia, which exhibited a range of striking morphologic findings. Distinctly vacuolated myeloid and erythroid precursors were seen in more than 95% of cases. Our data reveal potential novel diagnostic features, such as a high incidence of monocytopenia and distinct patterns of atypical megakaryopoiesis, that appear different from dysmegakaryopoiesis typically associated with MDS. In our experience, those findings are suggestive of VEXAS, in the appropriate clinical context.

VEXAS syndrome: A review on clinical manifestations

Background: VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome is a recently identified progressive, adult-onset, inflammatory disorder. Somatic mutations in the Ubiquitin-like modifier activating enzyme 1 (UBA1) gene at methionine-41 (p. Met41) were found in the patients. Clinical presentations are not limited to hematological and immunological systems; all organ systems are affected by VEXAS syndrome. The aim of this review is to summarise the scientific evidence accrued from the research studies on VEXAS syndrome, highlighting the clinical features and involvements of different organ systems.
 Conclusion: Although the primary manifestations affect the immunological and haematological systems, they gradually affect different other organ systems. Distinct clinical features necessitate ruling out the other possibilities that will be helpful for early diagnosis. More research should be carried out by the global research community in a collaborative manner for a better understanding and early diagnosis of the syndrome.

Early activation of inflammatory pathways in UBA1-mutated hematopoietic stem and progenitor cells in VEXAS

VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome is a pleiotropic, severe autoinflammatory disease caused by somatic mutations in the ubiquitin-like modifier activating enzyme 1 (UBA1) gene. To elucidate VEXAS pathophysiology, we performed transcriptome sequencing of single bone marrow mononuclear cells and hematopoietic stem and progenitor cells (HSPCs) from VEXAS patients. HSPCs are biased toward myeloid (granulocytic) differentiation, and against lymphoid differentiation in VEXAS. Activation of multiple inflammatory pathways (interferons and tumor necrosis factor alpha) occurs ontogenically early in primitive hematopoietic cells and particularly in the myeloid lineage in VEXAS, and inflammation is prominent in UBA1-mutated cells. Dysregulation in protein degradation likely leads to higher stress response in VEXAS HSPCs, which positively correlates with inflammation. TCR usage is restricted and there are increased cytotoxicity and IFN-γ signaling in Tcells. In VEXAS syndrome, both aberrant inflammation and myeloid predominance appear intrinsic to hematopoietic stem cells mutated in UBA1.

Recurrent Mutations of the Active Adenylation Domain of UBA1 in Atypical Form of VEXAS Syndrome.

Recurrent Mutations of the Active Adenylation Domain of UBA1 in Atypical Form of VEXAS Syndrome.

UFL1, a UFMylation E3 ligase, plays a crucial role in multiple cellular stress responses.

The UFM1 conjugation system(UFMylation)is a novel type of ubiquitin-like system that plays an indispensable role in maintaining cell homeostasis under various cellular stress. Similar to ubiquitination, UFMylation consists of a three-step enzymatic reaction with E1-like enzymes ubiquitin-like modifier activating enzyme5 (UBA5), E2-like enzymes ubiquitin-fold modifier-conjugating enzyme 1(UFC1), and E3-like ligase UFM1-specific ligase 1 (UFL1). As the only identified E3 ligase, UFL1 is responsible for specific binding and modification of the substrates to mediate numerous hormone signaling pathways and endocrine regulation under different physiological or pathological stress, such as ER stress, genotoxic stress, oncogenic stress, and inflammation. Further elucidation of the UFL1 working mechanism in multiple cellular stress responses is essential for revealing the disease pathogenesis and providing novel potential therapeutic targets. In this short review, we summarize the recent advances in novel UFL1 functions and shed light on the potential challenges ahead, thus hopefully providing a better understanding of UFMylation-mediated cellular stress.

Thrombotic Manifestations in Patients with Vexas Syndrome

Introduction: VEXAS syndrome, caused by somatic mutations in ubiquitin-like modifier activating enzyme 1 (UBA1) gene, is a late-onset and progressive autoinflammatory disease. Main hematologic manifestations include macrocytic anemia, thrombocytopenia, and dyspoiesis. Concomitant diagnoses of myelodysplastic syndrome (MDS) and plasma cell dyscrasia (PCD) are common. Venous thromboembolism (VTE) and arterial thrombosis (AT) have been reported. We describe the incidence and characteristics of thrombosis in a large cohort of 86 patients with VEXAS syndrome, along with detailed coagulation profiles in a subgroup of patients. We also explore potential predictors of thrombosis, and effect on overall survival (OS). Methods: Two cohorts of patients with confirmed VEXAS were included. Thirty patients were evaluated at the National Institutes of Health (NIH) Clinical Center and 56 patient records reviewed from referring institutions. Logistic regression models were used to determine predictors of thrombosis. Cox proportional hazards models and Kaplan Meier curves were used for survival analysis. Results: The total population consisted of 86 patients. Eighty-five patients (99%) were male with a median age of 64 years (range, 40-84 years) at disease onset. Hematologic manifestations included macrocytic anemia in 63 (73%), MDS in 25 (29%) and PCD in 17 (20%) patients. Thrombosis occurred in 45 patients (52%); 41 (48%) had VTE and 11 (12%) had AT, with a median time to event from disease onset of 18 and 34 months, respectively; five patients (5%) had both VTE and AT. Among VTE, there were 26 (30%) deep venous thrombosis (DVT), 12 (14%) pulmonary embolism (PE), and 7 (8%) superficial thrombophlebitis. Twenty-two (54%) of VTE cases were unprovoked. Recurrent VTE was seen in 15 (17%) of patients, among which 9 (53%) occurred while receiving anticoagulation. Arterial events included myocardial infarction in 7 (8%) patients, stroke in 2 (2%) patients, and peripheral arterial occlusion in 2 (2%) patients. Recurrent AT was seen in 2 (2%) patients. Patients from the NIH cohort underwent extensive coagulation testing. Fourteen patients (46%) had a positive lupus anticoagulant (LA), which was persistently positive 12 weeks apart in 8 of 9 patients re-tested. Of these 8 patients, 6 had a VTE, 1 had a stroke and 1 did not have thrombotic manifestations. Thirteen patients (43%) with thrombotic events were negative for LA. Median levels of fibrinogen, D-dimer, Factor (F) VII, FIX, and especially von Willebrand factor (VWF) and FVIII, were high in 15 tested VEXAS patients, but there was no significant difference between those with and without thrombosis (Table 1). Of 10 patients with high VWF and 17 with high FVIII activity, 6 (60%) and 9 (53%) had a thrombotic event, respectively; these correlated with high levels of CRP and ESR. No patients were found to have abnormal antithrombin III activity, Factor V Leiden, or prothrombin gene mutations. No difference in age, UBA1 gene mutation, median levels of CRP and ESR, presence of MDS or PCD was found between patients with or without thrombosis; MDS, PCD and UBA1 gene mutation were also not predictive of thrombosis. Among inflammatory disease manifestations, cardiac involvement (myocarditis, pericarditis, or pericardial effusion) correlated with the presence of any thrombosis (p = 0.03). When considering VTE alone, cardiac manifestations strongly predicted PE and DVT (p = 0.004 and p = 0.005 respectively), while the presence of chondritis correlated with PE (p = 0.02). Presence of thrombosis did not impact OS (Figure 1). Conclusions: Patients with VEXAS syndrome are at high risk of thrombosis, mainly venous, which can be recurrent despite anticoagulation therapy. The underlying etiology remains unclear, but many VEXAS patients have positive LA, high VWF and FVIII levels, suggesting inflammation, endothelial activation and/or an immune process as pathophysiologic. Cardiac manifestations, including myocarditis, pericarditis, and pericardial effusion, significantly correlated with thrombosis in general, while chondritis correlated with presence of PE. Presence of thrombosis did not impact OS. Given the high incidence of thrombosis, VEXAS patients should be carefully risk assessed for VTE and considered for prophylactic anticoagulation, and indefinite treatment if thrombosis has occurred. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Genomic screening reveals ubiquitin-like modifier activating enzyme 1 as a potent and druggable target in c-MYC-high triple negative breast cancer models.

Triple negative breast cancer (TNBC) accounts for over 30% of all breast cancer (BC)-related deaths, despite accounting for only 10% to 15% of total BC cases. Targeted therapy development has largely stalled in TNBC, underlined by a lack of traditionally druggable addictions like receptor tyrosine kinases (RTKs). Here, through full genome CRISPR/Cas9 screening of TNBC models, we have uncovered the sensitivity of TNBCs to the depletion of the ubiquitin-like modifier activating enzyme 1 (UBA1). Targeting UBA1 with the first-in-class UBA1 inhibitor TAK-243 induced unresolvable endoplasmic reticulum (ER)-stress and activating transcription factor 4 (ATF4)-mediated upregulation of proapoptotic NOXA, leading to cell death. c-MYC expression correlates with TAK-243 sensitivity and cooperates with TAK-243 to induce a stress response and cell death. Importantly, there was an order of magnitude greater sensitivity of TNBC lines to TAK-243 compared to normal tissue-derived cells. In five patient derived xenograft models (PDXs) of TNBC, TAK-243 therapy led to tumor inhibition or frank tumor regression. Moreover, in an intracardiac metastatic model of TNBC, TAK-243 markedly reduced metastatic burden, indicating UBA1 is a potential new target in TNBC expressing high levels of c-MYC.

Identification of UBA1 as the causative gene of an X-linked non-Kennedy spinal-bulbar muscular atrophy.

Spinal-bulbar muscular atrophy (SBMA) (Kennedy's disease) is a motor neuron disease. Kennedy's disease is nearly exclusively caused by mutations in the androgen receptor encoding gene (AR). The results of studies aimed at identification of the genetic cause of a disease that best approximates SBMA in a pedigree (four patients) without mutations in AR are reported. Clinical investigations included thorough neurological and non-neurological examinations and testing. Genetic analysis was performed by exome sequencing using standard protocols. UBA1 mutations were modeled on the crystal structure of UBA1. The clinical features of the patients are described in detail. A missense mutation in UBA1 (c.T1499C; p.Ile500Thr) was identified as the probable cause of the non-Kennedy SBMA in the pedigree. Like AR, UBA1 is positioned on chromosome X. UBA1 is a highly conserved gene. It encodes ubiquitin-like modifier activating enzyme 1 (UBA1) which is the major E1 enzyme of the ubiquitin-proteasome system. Interestingly, UBA1 mutations can also cause infantile-onset X-linked spinal muscular atrophy (XL-SMA). The mutation identified here and the XL-SMA causative mutations were shown to affect amino acids positioned in the vicinity of UBA1's ATP binding site and to cause structural changes. UBA1 was identified as a novel SBMA causative gene. The gene affects protein homeostasis which is one of most important components of the pathology of neurodegeneration. The contribution of this same gene to the etiology of XL-SMA is discussed.

Exome sequencing can misread high variant allele fraction of somatic variants in UBA1 as hemizygous in VEXAS syndrome: a case report

BackgroundVEXAS syndrome (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic syndrome) is a recently described syndrome caused by a somatic missense variant at the methionine-41 (p.(Met41)) position in the ubiquitin-like modifier activating enzyme 1 (UBA1) in Xp11.3. Germline pathogenic variants in UBA1 are associated with a distinct phenotype: a syndrome with severe neurologic features associated with loss of anterior horn cells and infantile death denominated X-Linked Spinal Muscular Atrophy 2 (SMAX2) (OMIM 301,830).Case presentationWe report a male individual with the phenotype of VEXAS syndrome that was initially identified through exome sequencing (ES) as having a hemizygous germline variant in UBA1 due to high variant allele frequency (VAF). Research Sanger sequencing was able to confirm the absence of the p.(Met41Val) variant in a skin biopsy and in gastric mucosa tissue sample confirming the variant happened as a postzygotic event.ConclusionsThe present case exemplifies the diagnostic challenge that was imposed by the high VAF detected by ES that failed to correctly demonstrate that the variant was in a mosaic state. Sequencing of different tissues should be considered when there is conflict between the UBA1 variant status and the clinical findings.

Reduced peripheral blood dendritic cell and monocyte subsets in MDS patients with systemic inflammatory or dysimmune diseases.

Systemic inflammatory and autoimmune diseases (SIADs) occur in 10-20% of patients with myelodysplastic syndrome (MDS). Recently identified VEXAS (Vacuoles, E1 enzyme, X-linked, Autoinflammatory, Somatic) syndrome, associated with somatic mutations in UBA1 (Ubiquitin-like modifier-activating enzyme 1), encompasses a range of severe inflammatory conditions along with hematological abnormalities, including MDS. The pathophysiological mechanisms underlying the association between MDS and SIADs remain largely unknown, especially the roles of different myeloid immune cell subsets. The aim of this study was to quantitatively evaluate peripheral blood myeloid immune cells (dendritic cells (DC) and monocytes) by flow cytometry in MDS patients with associated SIAD (n = 14, most often including relapsing polychondritis or neutrophilic dermatoses) and to compare their distribution in MDS patients without SIAD (n = 23) and healthy controls (n = 7). Most MDS and MDS/SIAD patients had low-risk MDS. Eight of 14 (57%) MDS/SIAD patients carried UBA1 somatic mutations, defining VEXAS syndrome.Compared with MDS patients, most DC and monocyte subsets were significantly decreased in MDS/SIAD patients, especially in MDS patients with VEXAS syndrome. Our study provides the first overview of the peripheral blood immune myeloid cell distribution in MDS patients with associated SIADs and raises several hypotheses: possible redistribution to inflammation sites, increased apoptosis, or impaired development in the bone marrow.

How in vitro maturation changes the proteome of ovine cumulus-oocyte complexes?

The present study evaluated the effects of in vitro maturation (IVM) on the proteome of cumulus-oocyte complexes (COCs) from ewes. Extracted COC proteins were analyzed by LC-MS/MS. Differences in protein abundances (p < 0.05) and functional enrichments in immature versusin vitro-matured COCs were evaluated using bioinformatics tools. There were 2550 proteins identified in the COCs, with 89 and 87 proteins exclusive to immature and mature COCs, respectively. IVM caused downregulation of 84 and upregulation of 34 proteins. Major upregulated proteins in mature COCs were dopey_N domain-containing protein, structural maintenance of chromosomes protein, ubiquitin-like modifier-activating enzyme 2. Main downregulated proteins in mature COCs were immunoglobulin heavy constant mu, inter-alpha-trypsin inhibitor heavy chain 2, alpha-2-macroglobulin. Proteins exclusive to mature COCs and upregulated after IVM related to immune response, complement cascade, vesicle-mediated transport, cell cycle, and extracellular matrix organization. Proteins of immature COCs and downregulated after IVM were linked to metabolic processes, immune response, and complement cascade. KEGG pathways and miRNA-regulated genes attributed to downregulated and mature COC proteins related to complement and coagulation cascades, metabolism, humoral response, and B cell-mediated immunity. Thus, IVM influenced the ovine COC proteome. This knowledge supports the future development of efficient IVM protocols for Ovis aries.

UFMylation System: An Emerging Player in Tumorigenesis.

Simple SummaryThe ubiquitin-fold modifier 1 (UFM1) is a newly identified post-translational modification protein that has been implicated in multiple cellular processes and diseases. Noticeably, an aberrant UFM1 modification system has been closely related to various types of tumorigeneses, implying that the restoration of UFMylation homeostasis may serve as a promising therapeutic strategy. In this review, we summarize the structure, process and biological functions of the UFM1 modification system. In particular, we discuss the relationship between the UFMylation system and tumorigenesis, illustrating the underlying mechanisms and future perspectives. This article aims to improve our understanding of UFM1 modification, as well as provide some new strategies for cancer treatment.Ubiquitin-fold modifier 1 (UFM1), a newly identified ubiquitin-like molecule (UBLs), is evolutionarily expressed in multiple species except yeast. Similarly to ubiquitin, UFM1 is covalently attached to its substrates through a well-orchestrated three-step enzymatic reaction involving E1, the UFM1-activating enzyme (ubiquitin-like modifier-activating enzyme 5, UBA5); E2, the UFM1-conjugating enzyme 1 (UFC1); and E3, the UFM1-specific ligase 1 (UFL1). To date, numerous studies have shown that UFM1 modification is implicated in various cellular processes, including endoplasmic reticulum (ER) stress, DNA damage response and erythroid development. An abnormal UFM1 cascade is closely related to a variety of diseases, especially tumors. Herein, we summarize the process and functions of UFM1 modification, illustrating the relationship and mechanisms between aberrant UFMylation and diversified tumors, aiming to provide novel diagnostic biomarkers or therapeutic targets for cancer treatments.

Immune complexome analysis of serum samples from non-small-cell lung cancer patients identifies predictive biomarkers for nivolumab therapy

BackgroundImmune checkpoint inhibitors (ICIs) have achieved important outcomes in cancer treatment. However, current clinical biomarker tests are not suitable for some patients because they require tumor tissues and have poor predictive value for treatment responses. Therefore, the identification of biomarkers that enable screening tests in all patients is necessary. MethodsWe performed an immune complexome analysis of non-small cell lung cancer patients treated with nivolumab to comprehensively identify and compare antigens incorporated into immune complexes (IC-antigens) in serum samples from the responders (n = 15) and non-responders (n = 20). Additionally, combinations of IC-antigens characteristic to the responder group were evaluated by logistic regression analysis and receiver operating characteristics curves to examine their predictiveness for ICI treatment responses. ResultsThe combination of predictive biomarkers detected before treatment was profilin-1, purine nucleoside phosphorylase, alpha-enolase, and nucleoside diphosphate kinase A [p = 0.0043, odds ratio = 2.26, 95% confidence interval (CI) = 1.19–4.28, area under the curve = 0.76]. The combination of predictive biomarkers detected after treatment was peptidyl-prolyl cis–trans isomerase A, ubiquitin-like modifier-activating enzyme 1, complement component C8 beta chain, and apolipoprotein L1 (p = 0.0039, odds ratio = 2.56, 95% CI = 1.25–5.23, area under the curve = 0.77). ConclusionCombinations of serum IC-antigens may predict the therapeutic effect of nivolumab in non-small cell lung cancer patients.

Application of Transcriptome Analysis to Understand the Adverse Effects of Hypotonic Stress on Different Development Stages in the Giant Freshwater Prawn Macrobrachium rosenbergii Post-Larvae

Salinity is one of the important environmental factors affecting survival and growth of aquatic animals. However, the impact of low-salinity stress on M. rosenbergii post-larvae at different development stages remains elusive. Therefore, the aim of this study was to explore the underlying mechanisms of hypotonic stress at different development stages of M. rosenbergii post-larvae through transcriptome analysis and antioxidant parameters detection. The salinity of the control group was 15 psu (S15) and the hypotonic stress group was 6 psu (S6). Samples were collected at 7 days-post-hatch (dph), 14 dph and 21 dph larvae. The results showed that hypotonic stress caused oxidative damage in post-larvae evidenced by decreased glutathione peroxidase (GSH-Px); superoxide dismutase (SOD); anti-superoxide anion free radical (ASAFR); and increased malondialdehyde (MDA); nitric oxide (NO); and inducible nitric oxide synthase (iNOS) levels. Transcriptome analysis showed that there were 1428, 1187, 132 DEGs including 301, 366, 4 up-regulated genes and 1127, 821, 128 down-regulated genes at 7 dph, 14 dph and 21 dph larvae under hypotonic stress, respectively. Furthermore, GO and KEGG enrichment indicated that hypotonic stress led to dysregulation of immune signals including lysosome and autophagy in the 7 dph larvae. The autophagy-related genes including beclin 1-associated autophagy-related key regulator (Barkor); ubiquitin-like modifier-activating enzyme ATG7 (ATG7); Beclin; autophagy-related protein 13 (ATG13); nuclear receptor-binding factor 2 (Nrbf2); ubiquitin-like-conjugating enzyme ATG3 (ATG3); vacuole membrane protein 1 (VMP1); and autophagy-related protein 2 (ATG2) decreased at 7 dph, and 14 dph larvae, and then increased at 21 dph larvae under hypotonic stress. In the 14 dph and 21 dph larvae, the renin-angiotensin system was activated. In conclusion, our data indicated that hypotonic stress reduced the antioxidant capacity and impaired the immune system in post-larvae, but as development progresses, the adaptability of post-larvae to hypotonic stress gradually increased, and might reach a new homeostasis through the RAS signaling pathway.

ABCB1 limits the cytotoxic activity of TAK-243, an inhibitor of the ubiquitin-activating enzyme UBA1.

One of the major concerns of cancer therapy is the emergence of multidrug resistance (MDR). The MDR-associated ATP-binding cassette sub-family B member 1 (ABCB1) transporter is established to mediate resistance against numerous anticancer drugs. In this study, we demonstrated that the Ubiquitin-like modifier activating enzyme 1 (UBA1) inhibitor TAK-243 is transported by the ABCB1. MTT assay was performed to evaluate the cytotoxicity of TAK-243. Western blot was carried out to investigate if TAK-243 affect to ABCB1 protein expression in cancer cells. High Performance Liquid Chromatography (HPLC) and ATPase assay were carried out to confirm TAK-243 as an ABCB1 substrate. [3H]-paclitaxel accumulation assay was used to determine the MDR reversal effect of TAK-243. Computational docking analysis was performed to investigate the drug-transporter binding position. The cytotoxicity profile showed that TAK-243 was less effective in ABCB1-overexpressing cells than in the parental cells, but pharmacological inhibition or knockout the gene of ABCB1 was able to reverse TAK-243 resistance. Furthermore, TAK-243 potently stimulated ABCB1 ATPase activity and the HPLC analysis revealed that TAK-243 accumulation was significantly reduced in ABCB1-overexpressing cells. Finally, the computational docking analysis indicates a high binding affinity between TAK-243 and human ABCB1 transporter. Our in vitro data characterized TAK-243 as a substrate of ABCB1, which may predict limited anticancer effect of this compound in drug resistant tumors.

Perspective on automated in vivo drug screening using the chodl mutant zebrafish line.

Perspective on automated in vivo drug screening using the chodl mutant zebrafish line.

Dermatomyositis: Muscle Pathology According to Antibody Subtypes.

Background and ObjectivesDiscoveries of dermatomyositis-specific antibodies (DMSAs) in patients with dermatomyositis raised awareness of various myopathologic features among antibody subtypes. However, only perifascicular atrophy and perifascicular myxovirus resistant protein A (MxA) overexpression were officially included as definitive pathologic criteria for dermatomyositis classification. We aimed to demonstrate myopathologic features in MxA-positive dermatomyositis to determine characteristic myopathologic features in different DMSA subtypes.MethodsWe performed a retrospective pathology review of muscle biopsies of patients with dermatomyositis diagnosed between January 2009 and December 2020 in a tertiary laboratory for muscle diseases. We included all muscle biopsies with sarcoplasmic expression for MxA and seropositivity for DMSAs. MxA-positive muscle biopsies that tested negative for all DMSAs were included as seronegative dermatomyositis. We evaluated histologic features stratified according to 4 pathology domains (muscle fiber, inflammatory, vascular, and connective tissue) and histologic features of interest by histochemistry, enzyme histochemistry, and immunohistochemical study commonly used in the diagnosis of inflammatory myopathy. We performed ultrastructural studies of 54 available specimens.ResultsA total of 256 patients were included. Of these, 249 patients were positive for 1 of the 5 DMSAs (seropositive patients: 87 anti–transcription intermediary factor 1-γ [TIF1-γ], 40 anti–complex nucleosome remodeling histone deacetylase [Mi-2], 29 anti–melanoma differentiation gene 5 [MDA5], 83 anti–nuclear matrix protein 2 [NXP-2], and 10 anti–small ubiquitin-like modifier-activating enzyme [SAE] dermatomyositis) and 7 patients were negative for all 5 DMSAs (seronegative patients). Characteristic myopathologic features in each DMSA subtype were as follows: anti-TIF1-γ with vacuolated/punched out fibers (64.7%; p < 0.001) and perifascicular enhancement in HLA-ABC stain (75.9%; p < 0.001); anti-Mi-2 with prominent muscle fiber damage (score 4.9 ± 2.1; p < 0.001), inflammatory cell infiltration (score 8.0 ± 3.0; p = 0.002), perifascicular atrophy (67.5%; p = 0.02), perifascicular necrosis (52.5%; p < 0.001), increased perimysial alkaline phosphatase activity (70.0%; p < 0.001), central necrotic peripheral regenerating fibers (45.0%; p = 0.002), and sarcolemmal membrane attack complex deposition (67.5%; p < 0.001); anti-MDA5 with scattered/diffuse staining pattern of MxA (65.5%; p < 0.001) with less muscle pathology and inflammatory features; anti-NXP-2 with microinfarction (26.5%; p < 0.001); and anti-SAE and seronegative dermatomyositis with HLA-DR expression (50.0%; p = 0.02 and 57.1%; p = 0.02, respectively).DiscussionWe describe a comprehensive serologic–pathologic correlation of dermatomyositis primarily using MxA expression as an inclusion criterion. In our study, DMSAs were associated with distinctive myopathologic features suggesting different underlying pathobiologic mechanisms in each subtype.

Knockdown of ubiquitin-like modifier-activating enzyme 2 promotes apoptosis of clear cell renal cell carcinoma cells

Small ubiquitin-related modifier (SUMO) proteins are involved in the development of tumors. Ubiquitin-like modifier-activating enzyme 2 (UBA2) is an important member of the SUMO modification system; however, its role in clear cell renal cell carcinoma (ccRCC) is unclear. Therefore, we investigated the expression and function of UBA2 in ccRCC. Both mRNA and protein expression levels of UBA2 were found to be higher in ccRCC than in normal renal tissues and significantly related to the tumor size, Fuhrman grade, and tumor stage. UBA2 knockdown inhibited ccRCC cell growth, promoted apoptosis in vitro and in vivo, and decreased the abundance of a p53 mutant, c-Myc, and key enzymes of the SUMO modification system. Meanwhile, overexpression of UBA2 had the opposite effects. Overexpression of the p53 mutant or c-Myc alleviated the effects of UBA2 knockdown on ccRCC cell proliferation and apoptosis. In conclusion, targeting UBA2 may have a therapeutic potential against ccRCC.