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Circulation Research Aug 2020Increased protein synthesis of profibrotic genes is a common feature in cardiac fibrosis and heart failure. Despite this observation, critical factors and molecular...
RATIONALE
Increased protein synthesis of profibrotic genes is a common feature in cardiac fibrosis and heart failure. Despite this observation, critical factors and molecular mechanisms for translational control of profibrotic genes during cardiac fibrosis remain unclear.
OBJECTIVE
To investigate the role of a bifunctional ARS (aminoacyl-tRNA synthetase), EPRS (glutamyl-prolyl-tRNA synthetase) in translational control of cardiac fibrosis.
METHODS AND RESULTS
Results from reanalyses of multiple publicly available data sets of human and mouse heart failure, demonstrated that EPRS acted as an integrated node among the ARSs in various cardiac pathogenic processes. We confirmed that EPRS was induced at mRNA and protein levels (≈1.5-2.5-fold increase) in failing hearts compared with nonfailing hearts using our cohort of human and mouse heart samples. Genetic knockout of one allele of globally () using CRISPR-Cas9 technology or in a Postn-Cre-dependent manner (; ) strongly reduces cardiac fibrosis (≈50% reduction) in isoproterenol-, transverse aortic constriction-, and myocardial infarction (MI)-induced heart failure mouse models. Inhibition of EPRS using a PRS (prolyl-tRNA synthetase)-specific inhibitor, halofuginone, significantly decreases translation efficiency (TE) of proline-rich collagens in cardiac fibroblasts as well as TGF-β (transforming growth factor-β)-activated myofibroblasts. Overexpression of EPRS increases collagen protein expression in primary cardiac fibroblasts under TGF-β stimulation. Using transcriptome-wide RNA-Seq and polysome profiling-Seq in halofuginone-treated fibroblasts, we identified multiple novel Pro-rich genes in addition to collagens, such as Ltbp2 (latent TGF-β-binding protein 2) and Sulf1 (sulfatase 1), which are translationally regulated by EPRS. SULF1 is highly enriched in human and mouse myofibroblasts. In the primary cardiac fibroblast culture system, siRNA-mediated knockdown of SULF1 attenuates cardiac myofibroblast activation and collagen deposition. Overexpression of SULF1 promotes TGF-β-induced myofibroblast activation and partially antagonizes anti-fibrotic effects of halofuginone treatment.
CONCLUSIONS
Our results indicate that EPRS preferentially controls translational activation of proline codon rich profibrotic genes in cardiac fibroblasts and augments pathological cardiac remodeling. Graphical Abstract: A graphical abstract is available for this article.
Topics: Amino Acyl-tRNA Synthetases; Animals; Case-Control Studies; Collagen; Disease Models, Animal; Enzyme Inhibitors; Female; Fibrosis; Heart Failure; Humans; Latent TGF-beta Binding Proteins; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocytes, Cardiac; Myofibroblasts; NIH 3T3 Cells; Proline-Rich Protein Domains; Protein Biosynthesis; Signal Transduction; Sulfotransferases
PubMed: 32611237
DOI: 10.1161/CIRCRESAHA.119.315999 -
Annals of the Rheumatic Diseases Sep 2016Examine whether osteoarthritis (OA) progression can be delayed by halofuginone in anterior cruciate ligament transection (ACLT) rodent models.
OBJECTIVES
Examine whether osteoarthritis (OA) progression can be delayed by halofuginone in anterior cruciate ligament transection (ACLT) rodent models.
METHODS
3-month-old male C57BL/6J (wild type; WT) mice and Lewis rats were randomised to sham-operated, ACLT-operated, treated with vehicle, or ACLT-operated, treated with halofuginone. Articular cartilage degeneration was graded using the Osteoarthritis Research Society International (OARSI)-modified Mankin criteria. Immunostaining, flow cytometry, RT-PCR and western blot analyses were conducted to detect relative protein and RNA expression. Bone micro CT (μCT) and CT-based microangiography were quantitated to detect alterations of microarchitecture and vasculature in tibial subchondral bone.
RESULTS
Halofuginone attenuated articular cartilage degeneration and subchondral bone deterioration, resulting in substantially lower OARSI scores. Specifically, we found that proteoglycan loss and calcification of articular cartilage were significantly decreased in halofuginone-treated ACLT rodents compared with vehicle-treated ACLT controls. Halofuginone reduced collagen X (Col X), matrix metalloproteinase-13 and A disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS 5) and increased lubricin, collagen II and aggrecan. In parallel, halofuginone-attenuated uncoupled subchondral bone remodelling as defined by reduced subchondral bone tissue volume, lower trabecular pattern factor (Tb.pf) and increased thickness of subchondral bone plate compared with vehicle-treated ACLT controls. We found that halofuginone exerted protective effects in part by suppressing Th17-induced osteoclastic bone resorption, inhibiting Smad2/3-dependent TGF-β signalling to restore coupled bone remodelling and attenuating excessive angiogenesis in subchondral bone.
CONCLUSIONS
Halofuginone attenuates OA progression by inhibition of subchondral bone TGF-β activity and aberrant angiogenesis as a potential preventive therapy for OA.
Topics: Animals; Anterior Cruciate Ligament; Bone Remodeling; Bone Resorption; Bone and Bones; Cartilage, Articular; Disease Models, Animal; Disease Progression; Male; Mice; Mice, Inbred C57BL; Osteoarthritis; Osteoclasts; Piperidines; Quinazolinones; Random Allocation; Rats; Rats, Inbred Lew; Transforming Growth Factor beta
PubMed: 26470720
DOI: 10.1136/annrheumdis-2015-207923 -
Cell Systems May 2016Reporter gene assays are a venerable tool for studying signaling pathways, but they lack the throughput and complexity necessary to contribute to a systems-level...
Reporter gene assays are a venerable tool for studying signaling pathways, but they lack the throughput and complexity necessary to contribute to a systems-level understanding of endogenous signaling networks. We present a parallel reporter assay, transcription factor activity sequencing (TF-seq), built on synthetic DNA enhancer elements, which enables parallel measurements in primary cells of the transcriptome and transcription factor activity from more than 40 signaling pathways. Using TF-seq in Myd88(-/-) macrophages, we captured dynamic pathway activity changes underpinning the global transcriptional changes of the innate immune response. We also applied TF-seq to investigate small molecule mechanisms of action and find a role for NF-κB activation and coordination of the STAT1 response in the macrophage reaction to the anti-inflammatory natural product halofuginone. Simultaneous TF-seq and global gene expression profiling represent an integrative approach for gaining mechanistic insight into pathway activity and transcriptional changes that result from genetic and small molecule perturbations.
Topics: Base Sequence; Gene Expression Profiling; Gene Expression Regulation; High-Throughput Nucleotide Sequencing; NF-kappa B; RNA; Sequence Analysis, RNA; Transcriptome
PubMed: 27211859
DOI: 10.1016/j.cels.2016.04.011 -
BMC Cancer Jun 2022Gastric cancer is one of the deadliest cancers, currently available therapies have limited success. Cancer-associated fibroblasts (CAFs) are pivotal cells in the stroma...
BACKGROUND
Gastric cancer is one of the deadliest cancers, currently available therapies have limited success. Cancer-associated fibroblasts (CAFs) are pivotal cells in the stroma of gastric tumors posing a great risk for progression and chemoresistance. The poor prognostic signature for CAFs is not clear in gastric cancer, and drugs that target CAFs are lacking in the clinic. In this study, we aim to identify a poor prognostic gene signature for CAFs, targeting which may increase the therapeutic success in gastric cancer.
METHODS
We analyzed four GEO datasets with a network-based approach and validated key CAF markers in The Cancer Genome Atlas (TCGA) and The Asian Cancer Research Group (ACRG) cohorts. We implemented stepwise multivariate Cox regression guided by a pan-cancer analysis in TCGA to identify a poor prognostic gene signature for CAF infiltration in gastric cancer. Lastly, we conducted a database search for drugs targeting the signature genes.
RESULTS
Our study revealed the COL1A1, COL1A2, COL3A1, COL5A1, FN1, and SPARC as the key CAF markers in gastric cancer. Analysis of the TCGA and ACRG cohorts validated their upregulation and poor prognostic significance. The stepwise multivariate Cox regression elucidated COL1A1 and COL5A1, together with ITGA4, Emilin1, and TSPAN9 as poor prognostic signature genes for CAF infiltration. The search on drug databases revealed collagenase clostridium histolyticum, ocriplasmin, halofuginone, natalizumab, firategrast, and BIO-1211 as the potential drugs for further investigation.
CONCLUSIONS
Our study demonstrated the central role of extracellular matrix components secreted and remodeled by CAFs in gastric cancer. The gene signature we identified in this study carries high potential as a predictive tool for poor prognosis in gastric cancer patients. Elucidating the mechanisms by which the signature genes contribute to poor patient outcomes can lead to the discovery of more potent molecular-targeted agents and increase the therapeutic success in gastric cancer.
Topics: Cancer-Associated Fibroblasts; Computational Biology; Humans; Prognosis; Stomach Neoplasms
PubMed: 35739492
DOI: 10.1186/s12885-022-09736-5 -
RNA Biology 2018Inhibition of tRNA aminoacylation has proven to be an effective antimicrobial strategy, impeding an essential step of protein synthesis. Mupirocin, the well-known... (Review)
Review
Inhibition of tRNA aminoacylation has proven to be an effective antimicrobial strategy, impeding an essential step of protein synthesis. Mupirocin, the well-known selective inhibitor of bacterial isoleucyl-tRNA synthetase, is one of three aminoacylation inhibitors now approved for human or animal use. However, design of novel aminoacylation inhibitors is complicated by the steadfast requirement to avoid off-target inhibition of protein synthesis in human cells. Here we review available data regarding known aminoacylation inhibitors as well as key amino-acid residues in aminoacyl-tRNA synthetases (aaRSs) and nucleotides in tRNA that determine the specificity and strength of the aaRS-tRNA interaction. Unlike most ligand-protein interactions, the aaRS-tRNA recognition interaction represents coevolution of both the tRNA and aaRS structures to conserve the specificity of aminoacylation. This property means that many determinants of tRNA recognition in pathogens have diverged from those of humans-a phenomenon that provides a valuable source of data for antimicrobial drug development.
Topics: Anti-Bacterial Agents; Escherichia coli; Fatty Alcohols; Humans; Isoleucine-tRNA Ligase; Mupirocin; Piperidines; Protein Synthesis Inhibitors; Quinazolinones; RNA, Transfer, Leu; Species Specificity; Structure-Activity Relationship; Thermus thermophilus; Transfer RNA Aminoacylation
PubMed: 29345185
DOI: 10.1080/15476286.2018.1429879 -
EFSA Journal. European Food Safety... Nov 2020Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific...
Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the coccidiostat STENOROL containing halofuginone hydrobromide (halofuginone HBr) as active substance. The FEEDAP Panel was not able to conclude on the safety of STENOROL for chickens and turkeys for fattening at the highest proposed use level. No incompatibilities or interactions with feedingstuffs, carriers, other approved additives or medicinal drugs are expected. Halofuginone HBr does not have antimicrobial activity at the highest dose proposed; it is not expected to exert adverse effects on chicken gut microbiota or select for resistance and cross-resistance with other antimicrobials. The Panel cannot conclude on the genotoxic potential of halofuginone HBr since an appropriate follow-up to exclude the mutagenic effect of the compound was not available. Therefore, the FEEDAP Panel cannot conclude on the safety of halofuginone HBr for the consumer. The additive is toxic by inhalation, dermal and ocular routes and is very irritant to both the eye and the skin. It is considered also a skin sensitiser. Inhalation exposure is considered a risk to persons handling the additive. Since the lack of genotoxic potential of halofuginone HBr has not been adequately demonstrated, it should be considered as an additional potential concern to users handling the additive. Due to limitations in some of the ecotoxicological studies, no conclusions can be drawn on the safety of the additive for the environment. The FEEDAP Panel is not in the position to conclude on the efficacy of STENOROL in chickens for fattening and in turkeys for fattening.
PubMed: 33209147
DOI: 10.2903/j.efsa.2020.6169 -
Molecules (Basel, Switzerland) Jan 2015Halofuginone is an analog of febrifugine-an alkaloid originally isolated from the plant Dichroa febrifuga. During recent years, halofuginone has attracted much attention... (Review)
Review
Halofuginone is an analog of febrifugine-an alkaloid originally isolated from the plant Dichroa febrifuga. During recent years, halofuginone has attracted much attention because of its wide range of beneficial biological activities, which encompass malaria, cancer, and fibrosis-related and autoimmune diseases. At present two modes of halofuginone actions have been described: (1) Inhibition of Smad3 phosphorylation downstream of the TGFβ signaling pathway results in inhibition of fibroblasts-to-myofibroblasts transition and fibrosis. (2) Inhibition of prolyl-tRNA synthetase (ProRS) activity in the blood stage of malaria and inhibition of Th17 cell differentiation thereby inhibiting inflammation and the autoimmune reaction by activation of the amino acid starvation and integrated stress responses. This review deals with the history and origin of this natural product, its synthesis, its known modes of action, and it's various biological activities in pre-clinical animal models and in humans.
Topics: Animals; Antimalarials; Antineoplastic Agents; Antiprotozoal Agents; Apoptosis; Clinical Trials as Topic; Humans; Piperidines; Quinazolinones
PubMed: 25569515
DOI: 10.3390/molecules20010573 -
Frontiers in Microbiology 2023The antiparasitic drug halofuginone is important for controlling apicomplexan parasites. However, the occurrence of halofuginone resistance is a major obstacle for it to...
The antiparasitic drug halofuginone is important for controlling apicomplexan parasites. However, the occurrence of halofuginone resistance is a major obstacle for it to the treatment of apicomplexan parasites. Current studies have identified the molecular marker and drug resistance mechanisms of halofuginone in . In this study, we tried to use transcriptomic data to explore resistance mechanisms of halofuginone in apicomplexan parasites of the genus (Apicomplexa: Eimeriidae). After halofuginone treatment of parasites, transcriptome analysis was performed using samples derived from both resistant and sensitive strains. In the sensitive group, DEGs associated with enzymes were significantly downregulated, whereas the DNA damaging process was upregulated after halofuginone treatment, revealing the mechanism of halofuginone-induced parasite death. In addition, 1,325 differentially expressed genes (DEGs) were detected between halofuginone resistant and sensitive strains, and the DEGs related to translation were significantly downregulated after halofuginone induction. Overall, our results provide a gene expression profile for further studies on the mechanism of halofuginone resistance in .
PubMed: 37065111
DOI: 10.3389/fmicb.2023.1141952 -
British Journal of Pharmacology Sep 2021Halofuginone is a febrifugine derivative originally isolated from Chinese traditional herb Chang Shan that exhibits anti-hypertrophic, anti-fibrotic and...
BACKGROUND AND PURPOSE
Halofuginone is a febrifugine derivative originally isolated from Chinese traditional herb Chang Shan that exhibits anti-hypertrophic, anti-fibrotic and anti-proliferative effects. We sought to investigate whether halofuginone induced pulmonary vasodilation and attenuates chronic hypoxia-induced pulmonary hypertension (HPH).
EXPERIMENTAL APPROACH
Patch-clamp experiments were conducted to examine the activity of voltage-dependent Ca channels (VDCCs) in pulmonary artery smooth muscle cells (PASMCs). Digital fluorescence microscopy was used to measure intracellular Ca concentration in PASMCs. Isolated perfused and ventilated mouse lungs were used to measure pulmonary artery pressure (PAP). Mice exposed to hypoxia (10% O ) for 4 weeks were used as model of HPH for in vivo experiments.
KEY RESULTS
Halofuginone increased voltage-gated K (K ) currents in PASMCs and K currents through KCNA5 channels in HEK cells transfected with KCNA5 gene. HF (0.03-1 μM) inhibited receptor-operated Ca entry in HEK cells transfected with calcium-sensing receptor gene and attenuated store-operated Ca entry in PASMCs. Acute (3-5 min) intrapulmonary application of halofuginone significantly and reversibly inhibited alveolar hypoxia-induced pulmonary vasoconstriction dose-dependently (0.1-10 μM). Intraperitoneal administration of halofuginone (0.3 mg·kg , for 2 weeks) partly reversed established PH in mice.
CONCLUSION AND IMPLICATIONS
Halofuginone is a potent pulmonary vasodilator by activating K channels and blocking VDCC and receptor-operated and store-operated Ca channels in PASMCs. The therapeutic effect of halofuginone on experimental PH is probably due to combination of its vasodilator effects, via inhibition of excitation-contraction coupling and anti-proliferative effects, via inhibition of the PI3K/Akt/mTOR signalling pathway.
Topics: Animals; Calcium; Hypertension, Pulmonary; Hypoxia; Mice; Myocytes, Smooth Muscle; Pharmaceutical Preparations; Phosphatidylinositol 3-Kinases; Piperidines; Pulmonary Artery; Quinazolinones
PubMed: 33694155
DOI: 10.1111/bph.15442 -
EFSA Journal. European Food Safety... Apr 2023Following a request from the European Commission, EFSA was asked to deliver a new scientific opinion on the coccidiostat halofuginone hydrobromide (STENOROL®) when used...
Safety and efficacy of a feed additive consisting of halofuginone hydrobromide (STENOROL®) for chickens for fattening and turkeys for fattening/reared for breeding (Huvepharma N.V.).
Following a request from the European Commission, EFSA was asked to deliver a new scientific opinion on the coccidiostat halofuginone hydrobromide (STENOROL®) when used as a feed additive for chickens for fattening and turkeys for fattening/reared for breeding. The Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) concludes that the safety for turkeys for fattening established in its previous opinion can be extended to turkeys for breeding up to 12 weeks of age. Based on the new data provided on the safety for consumer, environment and efficacy, the Panel updates its previous conclusions as follows: halofuginone hydrobromide is not genotoxic. Applying an uncertainty factor of 100 to the lowest no observed adverse effect level (NOAEL) of 0.03 mg/kg body weight (bw) per day, an acceptable daily intake (ADI) of 0.3 μg halofuginone/kg bw is established. The chronic exposure of consumers to residues of halofuginone would amount to 6-19% of the ADI after 3 days of withdrawal. Therefore, the Panel considers that the additive is safe for the consumer of tissues obtained from chickens for fattening and turkeys for fattening fed the additive at a maximum level of 3 mg/kg complete feed at a 3-day withdrawal time. For control purposes, the Panel recommends the setting of the following maximum residue limits (MRLs): liver, 50 μg/kg; kidney, 40 μg/kg; muscle, 3 μg/kg; skin/fat, 10 μg/kg wet tissue. Based on an updated environmental risk assessment, no concern for groundwater is expected. Halofuginone is unlikely to bioaccumulate and the risk of secondary poisoning is not likely to occur. No safety concerns are expected for terrestrial and aquatic environments. The additive has the potential to control coccidiosis in chickens for fattening and turkeys for fattening/reared for breeding up to 12 weeks of age at a minimum level of 2 mg/kg complete feed.
PubMed: 37089180
DOI: 10.2903/j.efsa.2023.7978