Ide Inhibitors Research Articles

From Oxetane to Thietane: Extending the Antiviral Spectrum of 2'-Spirocyclic Uridines by Substituting Oxygen with Sulfur.

In continuation of our efforts of finding novel nucleoside inhibitors for the treatment of viral diseases, we initiated a discovery research program aimed at identifying novel nucleos(t)ide inhibitors for emerging diseases like Dengue and Chikungunya. Based on the previously reported 2'-spiro-oxetane uridine derivatives active against Hepatitis C Virus (HCV), we envisaged its sulfur analogue as an interesting congener both from a synthetic as well as biological point of view. Surprisingly, we found the 2'-spirothietane uridine derivatives not only to be active against HCV and Dengue virus (DENV), viruses belonging to the flavivirus family, but also to demonstrate activity against alphaviruses like Chikungunya virus (CHIKV) and Sindbis virus (SINV).

Critical Updates on Chronic Hepatitis B Virus Infection in 2021.

Chronic hepatitis B virus (HBV) infection is a global healthcare burden in the form of chronic liver disease, cirrhosis, liver failure and liver cancer. There is no definite cure for the virus and even though extensive vaccination programs have reduced the burden of liver disease in the future population, treatment options to eradicate the virus from the host are still lacking. In this review, we discuss in detail current updates on the structure and applied biology of the virus in the host, examine updates to current treatment and explore novel and state-of-the-art therapeutics in the pipeline for management of chronic HBV. Furthermore, we also specifically review clinical updates on HBV-related acute on chronic liver failure (ACLF). Current treatments for chronic HBV infection have seen important updates in the form of considerations for treating patients in the immune tolerant phase and some clarity on end points for treatment and decisions on finite therapy with nucleos(t)ide inhibitors. Ongoing cutting-edge research on HBV biology has helped us identify novel target areas in the life cycle of the virus for application of new therapeutics. Due to improvements in the area of genomics, the hope for therapeutic vaccines, vector-based treatments and focused management aimed at targeting host integration of the virus and thereby a total cure could become a reality in the near future. Newer clinical prognostic tools have improved our understanding of timing of specific treatment options for the catastrophic syndrome of ACLF secondary to reactivation of HBV. In this review, we discuss in detail pertinent updates regarding virus biology and novel therapeutic targets with special focus on the appraisal of prognostic scores and treatment options in HBV-related ACLF.

Identification of ebselen as a potent inhibitor of insulin degrading enzyme by a drug repurposing screening

Insulin-degrading enzyme, IDE, is a metalloprotease implicated in the metabolism of key peptides such as insulin, glucagon, β-amyloid peptide. Recent studies have pointed out its broader role in the cell physiology. In order to identify new drug-like inhibitors of IDE with optimal pharmacokinetic properties to probe its multiple roles, we ran a high-throughput drug repurposing screening. Ebselen, cefmetazole and rabeprazole were identified as reversible inhibitors of IDE. Ebselen is the most potent inhibitor (IC50(insulin) = 14 nM). The molecular mode of action of ebselen was investigated by biophysical methods. We show that ebselen induces the disorder of the IDE catalytic cleft, which significantly differs from the previously reported IDE inhibitors. IDE inhibition by ebselen can explain some of its reported activities in metabolism as well as in neuroprotection.

Extracellular Ubiquitin(1-76) and Ubiquitin(1-74) Regulate Cardiac Fibroblast Proliferation.

SDF-1α (stromal cell-derived factor-1α) is a CXCR4-receptor agonist and DPP4 (dipeptidyl peptidase 4) substrate. SDF-1α, particularly when combined with sitagliptin to block the metabolism of SDF-1α by DPP4, stimulates proliferation of cardiac fibroblasts via the CXCR4 receptor; this effect is greater in cells from spontaneously hypertensive rats versus Wistar-Kyoto normotensive rats. Emerging evidence indicates that ubiquitin(1-76) exists in plasma and is a potent CXCR4-receptor agonist. Therefore, we hypothesized that ubiquitin(1-76), similar to SDF-1α, should increase proliferation of cardiac fibroblasts. Contrary to our working hypothesis, ubiquitin(1-76) did not stimulate cardiac fibroblast proliferation, yet unexpectedly antagonized the proproliferative effects of SDF-1α combined with sitagliptin. In this regard, ubiquitin(1-76) was more potent in spontaneously hypertensive versus Wistar-Kyoto cells. In the presence of 6bk (selective inhibitor of insulin-degrading enzyme [IDE]; an enzyme known to convert ubiquitin(1-76) to ubiquitin(1-74)), ubiquitin(1-76) no longer antagonized the proproliferative effects of SDF-1α/sitagliptin. Ubiquitin(1-74) also antagonized the proproliferative effects of SDF-1α/sitagliptin, and this effect of ubiquitin(1-74) was not blocked by 6bk and was >10-fold more potent compared with ubiquitin(1-76). Neither ubiquitin(1-76) nor ubiquitin(1-74) inhibited the proproliferative effects of the non-CXCR4 receptor agonist neuropeptide Y (activates Y1 receptors). Cardiac fibroblasts expressed IDE mRNA, protein, and activity and converted ubiquitin(1-76) to ubiquitin(1-74). Spontaneously hypertensive fibroblasts expressed greater IDE activity. Extracellular ubiquitin(1-76) blocks the proproliferative effects of SDF-1α/sitagliptin via its conversion by IDE to ubiquitin(1-74), a potent CXCR4 antagonist. Thus, IDE inhibitors, particularly when combined with DPP4 inhibitors or hypertension, could increase the risk of cardiac fibrosis.

Hepatitis B and liver transplantation

Chronic Hepatitis B (CHB) infection remains a common cause for liver cirrhosis and hepatocellular carcinoma. Orthotopic liver transplantation (OLT) has been a treatment modality for CHB patients with decompensated cirrhosis and or hepatocellular carcinoma. Previously early outcomes of transplantation for CHB patients were associated with rapid recurrence of the virus and loss of the allograft. Development of hepatitis B immunoglobulin (HBIG) and its use in prophylaxis of CHB patients after liver transplantation resulted in significant improvement in patient and allograft survival. Continued use of HBIG mono-therapy however has been complicated by development of viral resistance. With the development of nucleos(t)ide (NA) inhibitors for treatment of CHB, liver transplant outcomes continued to improve. One of the most important risk factors for viral recurrence in post liver transplant setting is the HBV DNA level. Use of NAs prior to liver transplantation to reduce the viral load has been an important way to prevent recurrence of CHB in post liver transplant patients. Addition of NAs to HBIG was the standard of therapy for liver transplant patients for many years and reduced the development of viral resistance. Long-term use of HBIG however is expensive and inconvenient for patients. Development of more potent NAs, with low viral resistance, has allowed for reduction and in some cases elimination of use of HBIG in post liver transplant prophylaxis of patients with CHB.

The Genotyping Resistance Profile of the Pol Gene Detected in Blood of Newly Diagnosed HIV-Positive Men Is Durably Archived in the Gut Whatever the Time of Initiation of cART

Objective: To evaluate the mutational patterns on the pol gene of the main HIV-1 strain archived in cell genome of 10 chronically infected men according to their clinical and therapeutic history. The genotyping resistance profiles were compared between the first blood plasma available at the time of HIV diagnosis and rectal biopsies and PBMC sampled 1-5 years after the initiation of combined antiretroviral therapy (cART). Methods: HIV-1 RNA and cell-associated HIV-1 DNA were quantified by Abbott Real-Time HIV-1 and Generic HIV® DNA cell (Biocentric) assays. The mutations in protease and reverse transcriptase genes were assessed by the Trugene® assay (Siemens). The C2V3 region was amplified to determine the viral tropism. Results: In 9 patients, slight or no differences were observed between the 3 resistance profiles. Those mostly detected were related to the resistance to nucleos(t)ide (D67N, L210W, T215A, T69D) and nonnucleoside (K103N, V106I, V179I) inhibitors. In 1 rilpivirine-treated patient, the M230I mutation was detected in PBMC. No change of viral tropism was observed between samples. Conclusion: These data suggest that resistance mutations harbored by the main HIV strain in plasma at the time of diagnosis are durably archived in DNA cells whatever the delay between infection and initiation of therapy in patients well controlled by cART.

Natural prevalence of hepatitis C virus (HCV) variants resistant to protease and polymerase inhibitors in patients infected with HCV genotype 1 in Tunisia.

Hepatitis C virus (HCV) protease inhibitors (PIs) and polymerase inhibitors: nucleos(t)ide inhibitors (NS5B-NIs) and non-nucleos(t)ide inhibitors (NS5B-NNIs) have been recently developed to inhibit protease (NS3) or polymerase (NS5B) activities. The drawback of antiviral treatment is the emergence of resistance mutations to the drugs. The prevalence of such mutations conferring resistance to PIs, NS5B-NIs, and NS5B-NNIs before treatment has not been investigated so far in the Tunisian population. The aim of this study was to investigate the prevalence of known substitutions conferring resistance to HCV-PIs, NS5B-NIs, and NS5B-NNIs in 149 untreated patients naïve of any novel or investigational anti-HCV drugs and infected with HCV genotype 1 (genotype 1a = 7; genotype 1b = 142). Twelve sequences (9.2%) of the 131/149 HCV NS3 sequences analyzed showed amino-acid substitutions associated with HCV PIs resistance mutations (T54S, n = 4 (3%); V55A, n = 2 (1.5%); Q80K, n = 4 (3%); R155K, n = 1 (0.7%); A156V, n = 1 (0.7%)). One (1%) of the 95/149 HCV NS5B sequences analyzed showed the substitution V321I conferring resistance to NS5B-NIs, while 34 of 95 (35.8%) showed substitutions conferring resistance to NS5B-NNIs (C316N, n = 2 (2%); M414L, n = 1 (1%); A421V, n = 8 (8.5%); M423A, n = 1 (1%); M423T, n = 2 (2%); I424V, n = 5 (5.2%); C445F, n = 1 (1%); I482T, n = 2 (2%); V494A, n = 1 (1%); P496A, n = 1 (1%); V499A, n = 15 (16%); S556G, n = 5 (5.2%)). Naturally occurring substitutions conferring resistance to NS3 or NS5B inhibitors exist in a substantial proportion of Tunisian treatment-naïve patients infected with HCV genotype 1. Their influence on treatment outcome should be assessed.

New horizons in hepatitis C antiviral therapy with direct-acting antivirals

Most direct-acting antivirals (DAAs) that are being developed as therapy against hepatitis C virus target the NS3/4A protease, the NS5A protein, and the NS5B polymerase. The latter enzyme offers different target sites: the catalytic domain for nucleos(t)ide analogues as well as a number of allosteric sites for nonnucleos(t)ide inhibitors. Two NS3/4A protease inhibitors have been approved recently, and more than 40 new NS3/4A, NS5A, or NS5B inhibitors are in development. These agents can achieve very high cure rates when combined with pegylated interferon-β and ribavirin and show promising clinical results when administered in all-oral combinations. In addition to the more canonical drug targets, new alternative viral targets for small-molecule drug development are emerging, such as p7 or NS4B and viral entry. Future research will need to define well-tolerated and cost-effective DAA combinations that provide the highest rates of viral eradication in all patients (including those with advanced liver disease), the broadest spectrum of action on viral genotypes showing minimal or no clinical resistance, and the shortest treatment duration.

New therapeutic strategies in HCV: polymerase inhibitors

The characterization of the viral life cycle facilitated the development of directly acting antiviral drugs. Among those, several inhibitors of the viral RNA-dependent RNA polymerase have proven effectiveness in clinical trials. The characteristics of different nucleos(t)ide and non-nucleoside polymerase inhibitors, as well as their clinical applications and combinations with other classes of directly acting antiviral drugs are reviewed herein.

Hepatitis C therapy in non‐genotype 1 patients: the near future

Worldwide, 50-70 million subjects are infected with an hepatitis C virus (HCV) genotype 2, 3, 4, 5 or 6. In these patients, the combination of PEG-INF-α and ribavirin remains the currently approved standard-of-care treatment. The identification of different potential therapeutic targets in the HCV life cycle has led to the development of both direct antiviral agents (DAAs) and reagents targeting host functions essential for viral replication. DAAs comprise so far first-generation, second-wave and second-generation NS3/4A protease inhibitors (PIs), nucleos(t)ide (NIs) and non-nucleoside inhibitors of the NS5B RNA polymerase and NS5A complex inhibitors. The main host-protein-directed antiviral agents are cyclophilin inhibitors and silibinin. Whereas the launch of first-generation PIs was a major landmark in the management of genotype 1 (GT-1)-infected patients, these drugs are inactive in most non-GT-1-infected patients. Several of these and other drugs have now reached phase II and even phase III clinical stage development. The purpose of this article is to provide an overview of the clinical results recently reported for the treatment for non-GT-1 HCV infection with a focus on the most promising new compounds and combinations.

Insulysin Cleaves the APP Cytoplasmic Fragment at Multiple Sites

The amyloid peptide (Abeta) deposited in Alzheimer's disease (AD) is generated by beta- and gamma-secretase processing of a larger integral membrane protein precursor (APP). Intramembrane processing of APP by gamma-secretase also yields an intracellular fragment, CTFgamma (a.k.a. AICD), which is highly conserved and is believed to regulate the transcription of several genes including KAI-1 and GSK3beta. The intracellular domain of APP is also processed by caspase to a 31 aa fragment that was shown to induce apoptosis by several groups. Although large quantities of CTFgamma are generated continuously by neurons, little if any is normally detected in cell lysates, which suggests that it is very rapidly turned over in vivo. Previous studies demonstrated that insulysin (IDE), an Abeta-degrading enzyme, is responsible for cytosol-mediated CTFgamma degradation in vitro. Consistent with this finding, knockout mice lacking IDE accumulate CTFgamma to detectable levels in the brain, although its levels remain lower than its precursor, suggesting that it continues to be turned over in the brain. Moreover, when we treated cultured cells with IDE inhibitors, we did not observe an increase in CTFgamma in cell lysates, suggesting that pathways other than IDE are also involved in CTFgamma turnover. To understand CTFgamma turnover further, we have mapped the IDE cleavage sites with the intention of mutating them to examine alternative pathways in future studies. Edman degradation revealed that IDE cleaves CTFgamma at multiple sites to small peptides ranging from 5 to 14 aa. The cleavage sites do not reveal the existence of any sequence specificity for IDE cleavage. Understanding the turnover mechanisms of CTFgamma is critical to the understanding of the signaling function of APP mediated by this fragment. The current study presents the interesting specificity of CTFgamma turnover by IDE, which has been previously identified as the major degrading enzyme for Abeta as well as CTFgamma. In addition, the study provides evidence for the presence of alternative CTFgamma-degrading pathways in the cell.

Transepithelial transport of insulin: I. Insulin degradation by insulin-degrading enzyme in small intestinal epithelium.

The purpose of this study was to determine the existence of insulin-degrading enzyme (EC 3.4.22.11) (IDE) in rat intestinal enterocytes. Subcellular fractionation, biochemical characterization, immunoprecipitation, and western blots were employed. Insulin-degrading activity was localized in the cytosol, constituting 92% of total insulin-degrading activity. Cytosolic insulin-degrading activity had a pH optimum of 7.5, was almost completely inhibited by IDE inhibitors (N-ethylmaleimide, 1,10-phenanthroline, EDTA, p-chloromercuribenzoate, bacitracin), but was not or only weakly inhibited by others (aprotinin, chymostatin, leupeptin, and diisopropyl phosphofluoridate.) Further, cytosolic insulin-degrading activity had a Km of 78 nM, sharing a similar Km value with insulin-degrading enzyme in non-purified forms. Approximately, 87 +/- 1.7% of cytosolic insulin-degrading activity was removed by the monoclonal antibody to IDE. On the SDS gel, the molecular weight of cytosolic IDE was 110 KD which is the same as that of human IDE. IDE is the major enzyme which degrades insulin in enterocytes.