Reversible and irreversible inhibitors of coronavirus Nsp15 endoribonuclease.

The emergence of severe acute respiratory syndrome coronavirus 2, the causative agent of coronavirus disease 2019, has resulted in the largest pandemic in recent history. Current therapeutic strategies to mitigate this disease have focused on the development of vaccines and on drugs that inhibit the viral 3CL protease or RNA-dependent RNA polymerase enzymes. A less-explored and potentially complementary drug target is Nsp15, a uracil-specific RNA endonuclease that shields coronaviruses and other nidoviruses from mammalian innate immune defenses. Here, we perform a high-throughput screen of over 100,000 small molecules to identify Nsp15 inhibitors. We characterize the potency, mechanism, selectivity, and predicted binding mode of five lead compounds. We show that one of these, IPA-3, is an irreversible inhibitor that might act via covalent modification of Cys residues within Nsp15. Moreover, we demonstrate that three of these inhibitors (hexachlorophene, IPA-3, and CID5675221) block severe acute respiratory syndrome coronavirus 2 replication in cells at subtoxic doses. This study provides a pipeline for the identification of Nsp15 inhibitors and pinpoints lead compounds for further development against coronavirus disease 2019 and related coronavirus infections.

Syndecan-1 and Free Indoxyl Sulfate Levels Are Associated with miR-126 in Chronic Kidney Disease.

Chronic kidney disease (CKD) is a major cause of death worldwide and is associated with a high risk for cardiovascular and all-cause mortality. In CKD, endothelial dysfunction occurs and uremic toxins accumulate in the blood. miR-126 is a regulator of endothelial dysfunction and its blood level is decreased in CKD patients. In order to obtain a better understanding of the physiopathology of the disease, we correlated the levels of miR-126 with several markers of endothelial dysfunction, as well as the representative uremic toxins, in a large cohort of CKD patients at all stages of the disease. Using a univariate analysis, we found a correlation between eGFR and most markers of endothelial dysfunction markers evaluated in this study. An association of miR-126 with all the evaluated uremic toxins was also found, while uremic toxins were not associated with the internal control, specifically cel-miR-39. The correlation between the expression of endothelial dysfunction biomarker Syndecan-1, free indoxyl sulfate, and total p-cresyl glucuronide on one side, and miR-126 on the other side was confirmed using multivariate analysis. As CKD is associated with reduced endothelial glycocalyx (eGC), our results justify further evaluation of the role of correlated parameters in the pathophysiology of CKD.

TRIM37 is highly expressed during mitosis in CHON-002 chondrocytes cell line and is regulated by miR-223.

Multiple molecular disorders can affect mechanisms regulating proliferation and differentiation of growth plate chondrocytes. Mutations in the TRIM37 gene cause the Mulibrey nanism, a heritable growth disorder. Since chondrocytes are instrumental in long bone growth that is deficient in nanism, we hypothesized that TRIM37 defect could contribute to dysregulation of the chondrocyte cell cycle. Western blotting, confocal microscopy and imaging flow cytometry determined TRIM37 expression in CHON-002 cell lineage. We showed that TRIM37 is expressed during mitosis of chondrocytes and directly impacted their proliferation. During the chondrocyte cell cycle, TRIM37 was present in both nucleus and cytoplasm. During M phase we observed an increase of the TRIM37-Tubulin co-localization in comparison with G1, S and G2 phases. TRIM37 knock down inhibited proliferation, together with cell cycle anomalies and increased autophagy, while overexpression accordingly enhanced cell proliferation. We demonstrated that microRNA-223 directly targets TRIM37, and suggest that miR-223 regulates TRIM37 gene expression during the cell cycle. In summary, our results give clues to explain why TRIM37 deficiency in chondrocytes impacts bone growth. Modulating TRIM37 using miR-223 could be an approach to increase chondrogenesis.

Serum levels of miR-126 and miR-223 and outcomes in chronic kidney disease patients.

Several microRNAs (miRNAs) have been linked to chronic kidney disease (CKD) mortality, cardiovascular (CV) complications and kidney disease progression. However, their association with clinical outcomes remains poorly evaluated. We used real-time qPCR to measure serum levels of miR-126 and miR-223 in a large cohort of 601 CKD patients (CKD stage G1 to G5 patients or on renal replacement therapy - CKD G5D) from Ghent University Hospital and 31 healthy controls. All-cause mortality and cardiovascular and renal events were registered as endpoints over a 6 year follow-up period. miR-126 levels were significantly lower from CKD stage G2 on, compared to controls. The serum levels of miR-223 were significantly lower from CKD stage G3B on. When considering overall mortality, patients with levels of either miR-126 or miR-223 below the median had a lower survival rate. Similar results were observed for CV and renal events. The observed link between the two miRNAs' seric levels and mortality, cardiovascular events or renal events in CKD appears to depend on eGFR. However, this does not preclude their potential role in the pathophysiology of CKD. In conclusion, CKD is associated with a decrease in circulating miR-223 and miR-126 levels.

TRIMming down to TRIM37: Relevance to Inflammation, Cardiovascular Disorders, and Cancer in MULIBREY Nanism.

TRIpartite motif (TRIM) proteins are part of the largest subfamilies of E3 ligases that mediate the transfer of ubiquitin to substrate target proteins. In this review, we focus on TRIM37 in the normal cell and in pathological conditions, with an emphasis on the MULIBREY (MUscle-LIver-BRain-EYe) genetic disorder caused by TRIM37 mutations. TRIM37 is characterized by the presence of a RING domain, B-box motifs, and a coiled-coil region, and its C-terminal part includes the MATH domain specific to TRIM37. MULIBREY nanism is a rare autosomal recessive caused by TRIM37 mutations and characterized by severe pre- and postnatal growth failure. Constrictive pericarditis is the most serious anomaly of the disease and is present in about 20% of patients. The patients have a deregulation of glucose and lipid metabolism, including type 2 diabetes, fatty liver, and hypertension. Puzzlingly, MULIBREY patients, deficient for TRIM37, are plagued with numerous tumors. Among non-MULIBREY patients affected by cancer, a wide variety of cancers are associated with an overexpression of TRIM37. This suggests that normal cells need an optimal equilibrium in TRIM37 expression. Finding a way to keep that balance could lead to potential innovative drugs for MULIBREY nanism, including heart condition and carcinogenesis treatment.

miR-92a: A Novel Potential Biomarker of Rapid Aortic Valve Calcification.

BACKGROUND AND AIM OF THE STUDY: The study aim was to compare the tissular expression of microRNAs (miRs) in bicuspid and tricuspid valves, and to evaluate their use as potential novel biomarkers of aortic valve calcification in bicuspid valves. METHODS: A prospective single-center observational study was conducted on stenotic bicuspid and tricuspid human aortic valves. According to their potential role in valve vascular and valvular calcification, a decision was taken to include miR- 92a, miR-141, and miR-223 in this analysis. A real-time quantitative polymerase chain reaction was used to measure the expression of each miR, using U6 and Cel-miR-39 as endogenous and exogenous gene controls, respectively. RESULTS: Among a total of 47 human calcified aortic valves collected, 30 (63.8%) were tricuspid valves. The mean preoperative transvalvular gradient was 50.8 mmHg (range: 37-89 mmHg), with no significant difference between bicuspid and tricuspid valves (50 mmHg versus 51.2 mmHg; p = 0.729). The mean aortic valve area was 0.79 cm2 (range: 0.33-1.3 cm2), again with no significant difference between the two groups (p = 0.34). The level of miR-92a expression was twofold higher in bicuspid valves compared to tricuspid valves (0.38 versus 0.17; p = 0.016), but no significant difference in miR-141 and miR-223 expression was observed between the two groups (p = 0.68 and p = 0.35, respectively). A positive correlation was observed between miR-92a expression and mean preoperative transvalvular gradient (r = 0.3257, p = 0.04). CONCLUSIONS: miR-92a is overexpressed in calcified bicuspid aortic valves, and may serve as a potential biomarker of rapid aortic valve calcification. Further studies based on these results may be designed to correlate the relative expression of miR-92a in the serum with its tissular expression in AS.

Serum microRNAs are altered in various stages of chronic kidney disease: a preliminary study.

[This retracts the article DOI: 10.1093/ckj/sfw060.].

Serum microRNAs are altered in various stages of chronic kidney disease: a preliminary study.

BACKGROUND: MicroRNAs (miRNAs) are innovative and informative blood-based biomarkers involved in numerous pathophysiological processes. In this study and based on our previous experimental data, we investigated miR-126, miR-143, miR-145, miR-155 and miR-223 as potential circulating biomarkers for the diagnosis and prognosis of patients with chronic kidney disease (CKD). The primary objective of this study was to assess the levels of miRNA expression at various stages of CKD. METHODS: RNA was extracted from serum, and RT-qPCR was performed for the five miRNAs and cel-miR-39 (internal control). RESULTS: Serum levels of miR-143, -145 and -223 were elevated in patients with CKD compared with healthy controls. They were further increased in chronic haemodialysis patients, but were below control levels in renal transplant recipients. In contrast, circulating levels of miR-126 and miR-155 levels, which were also elevated in CKD patients, were lower in the haemodialysis group and even lower in the transplant group. Four of the five miRNA species were correlated with estimated glomerular filtration rate, and three were correlated with circulating uraemic toxins. CONCLUSIONS: This exploratory study suggests that specific miRNAs could be biomarkers for complications of CKD, justifying further studies to link changes of miRNA levels with outcomes in CKD patients.