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Urologic Oncology Sep 2024Prostate cancer (PCa) is one of the most common tumors affecting men all over the world. PCa has brought a huge health burden to men around the world, especially for... (Review)
Review
Prostate cancer (PCa) is one of the most common tumors affecting men all over the world. PCa has brought a huge health burden to men around the world, especially for elderly men, but its pathogenesis is unclear. In prostate cancer, epigenetic inheritance plays an important role in the development, progression, and metastasis of the disease. An important role in cancer invasion and metastasis is played by matrix metalloproteinases (MMPs), zinc-dependent proteases that break down extracellular matrix. We review two important forms of epigenetic modification and the role of matrix metalloproteinases in tumor regulation, both of which may be of significant value as novel biomarkers for early diagnosis and prognosis monitoring. The author considers that both mechanisms have promising therapeutic applications for therapeutic agent research in prostate cancer, but that efforts should be made to mitigate or eliminate the side effects of drug therapy in order to maximize quality of life of patients. The understanding of epigenetic modification, MMPs, and their inhibitors in the functional regulation of prostate cancer is gradually advancing, it will provide a new technical means for the prevention of prostate cancer, early diagnosis, androgen-independent prostate cancer treatment, and drug research.
Topics: Humans; Male; Prostatic Neoplasms; Matrix Metalloproteinases; Epigenesis, Genetic; Matrix Metalloproteinase Inhibitors; Biomarkers, Tumor
PubMed: 38806387
DOI: 10.1016/j.urolonc.2024.05.002 -
International Journal of Molecular... Jul 2023When attacks its prey, the prey bleeds profusely but feels little pain. We and other research teams have identified several anticoagulant molecules in the saliva of ,...
When attacks its prey, the prey bleeds profusely but feels little pain. We and other research teams have identified several anticoagulant molecules in the saliva of , but the substance that produces the paralyzing effect in is not known. In this study, we successfully isolated, purified, and identified a serine protease inhibitor containing an antistasin-like domain from the salivary secretions of . This peptide (named poeciguamerin) significantly inhibited elastase activity and slightly inhibited FXIIa and kallikrein activity, but had no effect on FXa, trypsin, or thrombin activity. Furthermore, poeciguamerin exhibited analgesic activity in the foot-licking and tail-withdrawal mouse models and anticoagulant activity in the FeCl-induced carotid artery thrombosis mouse model. In this study, poeciguamerin was found to be a promising elastase inhibitor with potent analgesic and antithrombotic activity for the inhibition of pain and thrombosis after surgery or in inflammatory conditions.
Topics: Animals; Mice; Leeches; Serine Proteinase Inhibitors; Serpins; Anticoagulants; Thrombosis; Pancreatic Elastase; Analgesics; Pain
PubMed: 37446275
DOI: 10.3390/ijms241311097 -
Emerging Microbes & Infections Dec 2023The main protease (3-chymotrypsin-like protease, 3CLpro) of SARS-CoV-2 has become a focus of anti-coronavirus research. Despite efforts, drug development targeting...
The main protease (3-chymotrypsin-like protease, 3CLpro) of SARS-CoV-2 has become a focus of anti-coronavirus research. Despite efforts, drug development targeting 3CLpro has been hampered by limitations in the currently available activity assays. Additionally, the emergence of 3CLpro mutations in circulating SARS-CoV-2 variants has raised concerns about potential resistance. Both emphasize the need for a more reliable, sensitive, and facile 3CLpro assay. Here, we report an orthogonal dual reporter-based gain-of-signal assay for measuring 3CLpro activity in living cells. It builds on the finding that 3CLpro induces cytotoxicity and reporter expression suppression, which can be rescued by its inhibitor or mutation. This assay circumvents most limitations in previously reported assays, especially false positives caused by nonspecific compounds and signal interference from test compounds. It is also convenient and robust for high throughput screening of compounds and comparing the drug susceptibilities of mutants. Using this assay, we screened 1789 compounds, including natural products and protease inhibitors, with 45 compounds that have been reported to inhibit SARS-CoV-2 3CLpro among them. Except for the approved drug PF-07321332, only five of these inhibit 3CLpro in our assays: GC376; PF-00835231; S-217622; Boceprevir; and Z-FA-FMK. The susceptibilities of seven 3CLpro mutants prevalent in circulating variants to PF-07321332, S-217622, and GC376 were also assessed. Three mutants were identified as being less susceptible to PF-07321322 (P132H) and S-217622 (G15S, T21I). This assay should greatly facilitate the development of novel 3CLpro-targeted drugs and the monitoring of the susceptibility of emerging SARS-CoV-2 variants to 3CLpro inhibitors.
Topics: Humans; SARS-CoV-2; COVID-19; Mutation; Peptide Hydrolases; Antiviral Agents
PubMed: 37144395
DOI: 10.1080/22221751.2023.2211688 -
Journal of Medicinal Chemistry Aug 2023There is an urgent need for new treatments for Chagas disease, a parasitic infection which mostly impacts South and Central America. We previously reported on the...
There is an urgent need for new treatments for Chagas disease, a parasitic infection which mostly impacts South and Central America. We previously reported on the discovery of GSK3494245/DDD01305143, a preclinical candidate for visceral leishmaniasis which acted through inhibition of the proteasome. A related analogue, active against , showed suboptimal efficacy in an animal model of Chagas disease, so alternative proteasome inhibitors were investigated. Screening a library of phenotypically active analogues against the proteasome identified an active, selective pyridazinone, the development of which is described herein. We obtained a cryo-EM co-structure of proteasome and a key inhibitor and used this to drive optimization of the compounds. Alongside this, optimization of the absorption, distribution, metabolism, and excretion (ADME) properties afforded a suitable compound for mouse efficacy studies. The outcome of these studies is discussed, alongside future plans to further understand the series and its potential to deliver a new treatment for Chagas disease.
Topics: Mice; Animals; Trypanosoma cruzi; Proteasome Inhibitors; Proteasome Endopeptidase Complex; Chagas Disease; Leishmaniasis, Visceral; Trypanocidal Agents
PubMed: 37506194
DOI: 10.1021/acs.jmedchem.3c00582 -
BioRxiv : the Preprint Server For... Oct 2023Nirmatrelvir was the first protease inhibitor (PI) specifically developed against the SARS-CoV-2 main protease (3CL/M) and licensed for clinical use. As SARS-CoV-2...
Nirmatrelvir was the first protease inhibitor (PI) specifically developed against the SARS-CoV-2 main protease (3CL/M) and licensed for clinical use. As SARS-CoV-2 continues to spread, variants resistant to nirmatrelvir and other currently available treatments are likely to arise. This study aimed to identify and characterize mutations that confer resistance to nirmatrelvir. To safely generate M resistance mutations, we passaged a previously developed, chimeric vesicular stomatitis virus (VSV-M) with increasing, yet suboptimal concentrations of nirmatrelvir. Using Wuhan-1 and Omicron M variants, we selected a large set of mutants. Some mutations are frequently present in GISAID, suggesting their relevance in SARS-CoV-2. The resistance phenotype of a subset of mutations was characterized against clinically available PIs (nirmatrelvir and ensitrelvir) with cell-based and biochemical assays. Moreover, we showed the putative molecular mechanism of resistance based on in silico molecular modelling. These findings have implications on the development of future generation M inhibitors, will help to understand SARS-CoV-2 protease-inhibitor-resistance mechanisms and show the relevance of specific mutations in the clinic, thereby informing treatment decisions.
PubMed: 37808638
DOI: 10.1101/2023.09.22.558628 -
Frontiers in Immunology 2023Salivary glands from blood-feeding arthropods secrete several molecules that inhibit mammalian hemostasis and facilitate blood feeding and pathogen transmission. The...
BACKGROUND
Salivary glands from blood-feeding arthropods secrete several molecules that inhibit mammalian hemostasis and facilitate blood feeding and pathogen transmission. The salivary functions from , the main vector of Onchocerciasis in South America, remain largely understudied. Here, we have characterized a salivary protease inhibitor (Guianensin) from the blackfly .
MATERIALS AND METHODS
A combination of bioinformatic and biophysical analyses, recombinant protein production, in vitro and in vivo experiments were utilized to characterize the molecula mechanism of action of Guianensin. Kinetics of Guianensin interaction with proteases involved in vertebrate inflammation and coagulation were carried out by surface plasmon resonance and isothermal titration calorimetry. Plasma recalcification and coagulometry and tail bleeding assays were performed to understand the role of Guianensin in coagulation.
RESULTS
Guianensin was identified in the sialotranscriptome of adult flies and belongs to the Kunitz domain of protease inhibitors. It targets various serine proteases involved in hemostasis and inflammation. Binding to these enzymes is highly specific to the catalytic site and is not detectable for their zymogens, the catalytic site-blocked human coagulation factor Xa (FXa), or thrombin. Accordingly, Guianensin significantly increased both PT (Prothrombin time) and aPTT (Activated partial thromboplastin time) in human plasma and consequently increased blood clotting time . Guianensin also inhibited prothrombinase activity on endothelial cells. We show that Guianensin acts as a potent anti-inflammatory molecule on FXa-induced paw edema formation in mice.
CONCLUSION
The information generated by this work highlights the biological functionality of Guianensin as an antithrombotic and anti-inflammatory protein that may play significant roles in blood feeding and pathogen transmission.
Topics: Mice; Humans; Animals; Simuliidae; Hemostatics; Endothelial Cells; Hemostasis; Anti-Inflammatory Agents; Inflammation; Salivary Proteins and Peptides; Mammals
PubMed: 37469515
DOI: 10.3389/fimmu.2023.1163367 -
Investigative Ophthalmology & Visual... Jul 2023This study investigated the effect of dipeptidyl peptidase-4 inhibitors (DPP-4is) on fibrosis after glaucoma filtering surgery with clinical data and an in vitro model...
PURPOSE
This study investigated the effect of dipeptidyl peptidase-4 inhibitors (DPP-4is) on fibrosis after glaucoma filtering surgery with clinical data and an in vitro model that used transforming growth factor-β (TGF-β) to induce human Tenon's fibroblast (HTF) fibrosis.
METHODS
The medical records of 41 eyes of 35 patients with diabetes with neovascular glaucoma (NVG) who received initial trabeculectomy were retrospectively reviewed. The surgical success rate was compared between cases that received (n = 23) and did not receive (n = 18) DPP-4i treatment for diabetes. The antifibrotic effects of linagliptin (a DPP-4i) were evaluated with quantitative real-time PCR for fibrosis markers (α-smooth muscle actin, collagen Iα, and fibronectin), a scratch assay, and a collagen gel contraction assay of primary cultured HTFs treated with TGF-β1 and linagliptin. Western blotting analysis was performed to evaluate the levels of phosphorylated Smad2 and Smad3 in the presence of linagliptin.
RESULTS
The Kaplan-Meier curve for bleb survival was higher in patients who received DPP-4is (P = 0.017, log-rank test). The in vitro experiments demonstrated that treatment with linagliptin attenuated the elevated levels of fibrosis markers induced by TGF-β1 in HTFs. Linagliptin treatment also prevented the migration and gel contraction of HTFs. Linagliptin inhibited the phosphorylation of Smad2 and Smad3, which is the canonical pathway of TGF-β signaling.
CONCLUSIONS
The current study indicates the potential effect of DPP-4is for maintaining bleb function after glaucoma filtering surgery in patients with diabetes with NVG. Our results demonstrate that linagliptin attenuates fibrotic change in HTFs by inhibiting TGF-β/Smad signaling.
Topics: Humans; Transforming Growth Factor beta1; Dipeptidyl-Peptidase IV Inhibitors; Linagliptin; Retrospective Studies; Glaucoma; Fibrosis; Fibroblasts; Trabeculectomy; Transforming Growth Factor beta; Signal Transduction; Collagen; Cells, Cultured
PubMed: 37405760
DOI: 10.1167/iovs.64.10.2 -
JAMA Network Open Oct 2023Assessing the relative effectiveness and safety of additional treatments when metformin monotherapy is insufficient remains a limiting factor in improving treatment...
IMPORTANCE
Assessing the relative effectiveness and safety of additional treatments when metformin monotherapy is insufficient remains a limiting factor in improving treatment choices in type 2 diabetes.
OBJECTIVE
To determine whether data from electronic health records across the University of California Health system could be used to assess the comparative effectiveness and safety associated with 4 treatments in diabetes when added to metformin monotherapy.
DESIGN, SETTING, AND PARTICIPANTS
This multicenter, new user, multidimensional propensity score-matched retrospective cohort study with leave-one-medical-center-out (LOMCO) sensitivity analysis used principles of emulating target trial. Participants included patients with diabetes receiving metformin who were then additionally prescribed either a sulfonylurea, dipeptidyl peptidase-4 inhibitor (DPP4I), sodium-glucose cotransporter-2 inhibitor (SGLT2I), or glucagon-like peptide-1 receptor agonist (GLP1RA) for the first time and followed-up over a 5-year monitoring period. Data were analyzed between January 2022 and April 2023.
EXPOSURE
Treatment with sulfonylurea, DPP4I, SGLT2I, or GLP1RA added to metformin monotherapy.
MAIN OUTCOMES AND MEASURES
The main effectiveness outcome was the ability of patients to maintain glycemic control, represented as time to metabolic failure (hemoglobin A1c [HbA1c] ≥7.0%). A secondary effectiveness outcome was assessed by monitoring time to new incidence of any of 28 adverse outcomes, including diabetes-related complications while treated with the assigned drug. Sensitivity analysis included LOMCO.
RESULTS
This cohort study included 31 852 patients (16 635 [52.2%] male; mean [SD] age, 61.4 [12.6] years) who were new users of diabetes treatments added on to metformin monotherapy. Compared with sulfonylurea in random-effect meta-analysis, treatment with SGLT2I (summary hazard ratio [sHR], 0.75 [95% CI, 0.69-0.83]; I2 = 37.5%), DPP4I (sHR, 0.79 [95% CI, 0.75-0.84]; I2 = 0%), GLP1RA (sHR, 0.62 [95% CI, 0.57-0.68]; I2 = 23.6%) were effective in glycemic control; findings from LOMCO sensitivity analysis were similar. Treatment with SGLT2I showed no significant difference in effectiveness compared with GLP1RA (sHR, 1.26 [95% CI, 1.12-1.42]; I2 = 47.3%; no LOMCO) or DPP4I (sHR, 0.97 [95% CI, 0.90-1.04]; I2 = 0%). Patients treated with DPP4I and SGLT2I had fewer cardiovascular events compared with those treated with sulfonylurea (DPP4I: sHR, 0.84 [95% CI, 0.74-0.96]; I2 = 0%; SGLT2I: sHR, 0.78 [95% CI, 0.62-0.98]; I2 = 0%). Patients treated with a GLP1RA or SGLT2I were less likely to develop chronic kidney disease (GLP1RA: sHR, 0.75 [95% CI 0.6-0.94]; I2 = 0%; SGLT2I: sHR, 0.77 [95% CI, 0.61-0.97]; I2 = 0%), kidney failure (GLP1RA: sHR, 0.69 [95% CI, 0.56-0.86]; I2 = 9.1%; SGLT2I: sHR, 0.72 [95% CI, 0.59-0.88]; I2 = 0%), or hypertension (GLP1RA: sHR, 0.82 [95% CI, 0.68-0.97]; I2 = 0%; SGLT2I: sHR, 0.73 [95% CI, 0.58-0.92]; I2 = 38.5%) compared with those treated with a sulfonylurea. Patients treated with an SGLT2I, vs a DPP4I, GLP1RA, or sulfonylurea, were less likely to develop indicators of chronic hepatic dysfunction (sHR vs DPP4I, 0.68 [95% CI, 0.49-0.95]; I2 = 0%; sHR vs GLP1RA, 0.66 [95% CI, 0.48-0.91]; I2 = 0%; sHR vs sulfonylurea, 0.60 [95% CI, 0.44-0.81]; I2 = 0%), and those treated with a DPP4I were less likely to develop new incidence of hypoglycemia (sHR, 0.48 [95% CI, 0.36-0.65]; I2 = 22.7%) compared with those treated with a sulfonylurea.
CONCLUSIONS AND RELEVANCE
These findings highlight familiar medication patterns, including those mirroring randomized clinical trials, as well as providing new insights underscoring the value of robust clinical data analytics in swiftly generating evidence to help guide treatment choices in diabetes.
Topics: Aged; Female; Humans; Male; Middle Aged; Antiviral Agents; Cohort Studies; Diabetes Mellitus, Type 2; Dipeptidyl-Peptidase IV Inhibitors; Hypoglycemic Agents; Metformin; Protease Inhibitors; Retrospective Studies; Sodium-Glucose Transporter 2 Inhibitors; Sulfonylurea Compounds; Network Meta-Analysis
PubMed: 37782497
DOI: 10.1001/jamanetworkopen.2023.36613 -
Journal of Molecular Biology Dec 2023Deubiquitinases (DUBs) are proteolytic enzymes that catalyze the removal of ubiquitin from protein substrates. The critical role of DUBs in regulating protein...
Deubiquitinases (DUBs) are proteolytic enzymes that catalyze the removal of ubiquitin from protein substrates. The critical role of DUBs in regulating protein ubiquitination makes them attractive drug targets in oncology, neurodegenerative disease, and antiviral development. Biochemical assays for quantifying DUB activity have enabled characterization of substrate preferences and discovery of small molecule inhibitors. However, assessing the efficacy of these inhibitors in cellular contexts to support clinical drug development has been limited by a lack of tractable cell-based assays. To address this gap, we developed a two-color flow cytometry-based assay that allows for sensitive quantification of DUB activity and inhibition in living cells. The utility of this system was demonstrated by quantifying the potency of GRL0617 against the viral DUB SARS-CoV-2 PL, identifying potential GRL0617 resistance mutations, and performing structure-function analysis of the vOTU domain from the recently emerged Yezo virus. In addition, the system was optimized for cellular DUBs by modifying a GFP-targeting nanobody to recruit USP7 and USP28 to benchmark a panel of reported inhibitors and assess inhibition kinetics. Together, these results demonstrate the utility of these assays for studying DUB biology in a cellular context with potential to aid in inhibitor discovery and development.
Topics: Humans; Aniline Compounds; Benzamides; Deubiquitinating Enzymes; Neurodegenerative Diseases; Ubiquitin; Ubiquitin Thiolesterase; Ubiquitin-Specific Peptidase 7; Ubiquitination; Flow Cytometry; Protease Inhibitors; Coronavirus Papain-Like Proteases; Single-Domain Antibodies
PubMed: 37858708
DOI: 10.1016/j.jmb.2023.168316 -
Frontiers in Immunology 2024Protease inhibitors regulate various biological processes and prevent host tissue/organ damage. Specific inhibition/regulation of proteases is clinically valuable for...
Protease inhibitors regulate various biological processes and prevent host tissue/organ damage. Specific inhibition/regulation of proteases is clinically valuable for treating several diseases. Psoriasis affects the skin in the limbs and scalp of the body, and the contribution of cysteine and serine proteases to the development of skin inflammation is well documented. Cysteine protease inhibitors from ticks have high specificity, selectivity, and affinity to their target proteases and are efficient immunomodulators. However, their potential therapeutic effect on psoriasis pathogenesis remains to be determined. Therefore, we tested four tick cystatins (Sialostatin L, Sialostatin L2, Iristatin, and Mialostatin) in the recently developed, innate immunity-dependent mannan-induced psoriasis model. We explored the effects of protease inhibitors on clinical symptoms and histological features. In addition, the number and percentage of immune cells (dendritic cells, neutrophils, macrophages, and γδT cells) by flow cytometry, immunofluorescence/immunohistochemistry and, the expression of pro-inflammatory cytokines (TNF-a, IL-6, IL-22, IL-23, and IL-17 family) by qPCR were analyzed using skin, spleen, and lymph node samples. Tick protease inhibitors have significantly decreased psoriasis symptoms and disease manifestations but had differential effects on inflammatory responses and immune cell populations, suggesting different modes of action of these inhibitors on psoriasis-like inflammation. Thus, our study demonstrates, for the first time, the usefulness of tick-derived protease inhibitors for treating skin inflammation in patients.
Topics: Humans; Cysteine Proteinase Inhibitors; Mannans; Psoriasis; Inflammation; Dermatitis; Protease Inhibitors; Immunity, Innate; Endopeptidases; Peptide Hydrolases
PubMed: 38444844
DOI: 10.3389/fimmu.2024.1344878