In vitro and in vivo inhibition of cysteine proteinases by EST, a new analog of E-64.

Tamai, Masamitsu; Matsumoto, Kazuko; Omura, Sadafumi; Koyama, Ikuko; Ozawa, Yasuo; Hanada, Kazunori

doi:10.1248/bpb1978.9.672

Cited by 167 publications

(110 citation statements)

References 16 publications

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“…E64d is a membrane permeable inhibitor of cathepsins B, H and L, 25,26 and pepstatin A is an inhibitor of cathepsin D and E. 27,28 As is the case with the other lysosomal marker, Lamp1 (Fig. 1), GFP-LC3 was not colocalized with TMR-Dextran in control cells and treatment with protease inhibitors did not affect the GFP-LC3 distribution pattern ( Fig.…”

Section: Resultsmentioning

confidence: 75%

Dissection of the Autophagosome Maturation Process by a Novel Reporter Protein, Tandem Fluorescent-Tagged LC3

2007

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Section: Resultsmentioning

confidence: 75%

Dissection of the Autophagosome Maturation Process by a Novel Reporter Protein, Tandem Fluorescent-Tagged LC3

2007

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“…In contrast, in cells harboring either the subgenomic or the full-genome-length HCV replicons, Bid expression was restored by pharmacological inhibition of calpains. Although it is notoriously difficult to eliminate the possible cross-specificities in the use of pharmacological enzyme inhibitors, the fact that two chemically unrelated calpain inhibitors 29,30 had a similar activity in our assay argues for these proteases' involvement in Bid degradation.…”

Section: Discussionmentioning

confidence: 97%

Calpain activation by hepatitis C virus proteins inhibits the extrinsic apoptotic signaling pathway

et al. 2009

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An unresolved question regarding the physiopathology of hepatitis C virus (HCV) infection is the remarkable efficiency with which host defenses are neutralized to establish chronic infection. Modulation of an apoptotic response is one strategy used by viruses to escape immune surveillance. We previously showed that HCV proteins down-regulate expression of BH3-only Bcl2 interacting domain (Bid) in hepatocytes of HCV transgenic mice. As a consequence, cells acquire resistance to Fas-mediated apoptosis, which in turn leads to increased persistence of experimental viral infections in vivo. This mechanism might participate in the establishment of chronic infections and the resulting pathologies, including hepatocellular carcinoma. We now report that Bid is also down-regulated in patients in the context of noncirrhotic HCV-linked tumorigenesis and in the HCV RNA replicon system. We show that the nonstructural HCV viral protein NS5A is sufficient to activate a calpain cysteine protease, leading to degradation of Bid. Moreover, pharmacological inhibitors of calpains restore both the physiological levels of Bid and the sensitivity of cells toward a death receptor-mediated apoptotic signal. Finally, human HCV-related tumors and hepatocytes from HCV transgenic mice that display low Bid expression contain activated calpains. Conclusion: Calpains activated by HCV proteins degrade Bid and thus dampen apoptotic signaling. These results suggest that inhibiting calpains could lead to an improved efficiency of immune-mediated elimination of HCV-infected cells. (HEPATOLOGY 2009;50:1370-1379.)P ersistent infection with hepatitis C virus (HCV) is among the most common infectious causes of chronic liver disease. The majority of patients fail to clear the virus and become chronic carriers, with a persistent presence of detectable virus in the serum. 1 Patients with chronic hepatitis C are at risk for hepatic fibrosis, frequently culminating in hepatic cirrhosis and hepatocellular carcinoma (HCC). 2 FL-N/35 transgenic mice, with hepatocyte-targeted expression of the entire open reading frame of the genotype 1b HCV, are at risk for steatosis and hepatocellular adenoma and carcinoma. 3 We previously showed that the FL-N/35 hepatocytes are resistant to apoptosis induced by the Fas/CD95 death receptor stimulation. The lack of sensitivity to apoptotic stimulation was related to decreased expression of BH3-only Bcl2 interacting domain (Bid), a BH3-only member of the Bcl-2 family of apoptosis regulators. 4 Importantly, Bid-deficient hepatocytes are refractory to T lymphocyte-induced cell death, suggesting that apoptosis contributes to HCV persistence and, as a consequence, to liver pathologies characteristic of chronic HCV infection. 4 Here, we report that Bid is also down-regulated in the context of the subgenomic and genome-length HCV replicons and in a subset of HCV-linked human HCC.

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“…This strategy assessed the relative importance of E64d inhibition of the protease of interest, compared with other proteases, in improving TBI. E64d inhibits papain-like cathepsin cysteine proteases and calcium-activated neutral proteases 14 and thus can have multiple protease targets. The similar effects observed in E64d, cathepsin B KO, and E64d-cathepsin B KO mice indicate that E64d likely improves TBI outcomes through inhibiting cathepsin B.…”

Section: E64d Improves Traumatic Brain Injurymentioning

confidence: 99%

The Cysteine Protease Cathepsin B Is a Key Drug Target and Cysteine Protease Inhibitors Are Potential Therapeutics for Traumatic Brain Injury

Hook

Sipes

et al. 2014

Journal of Neurotrauma

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There are currently no effective therapeutic agents for traumatic brain injury (TBI), but drug treatments for TBI can be developed by validation of new drug targets and demonstration that compounds directed to such targets are efficacious in TBI animal models using a clinically relevant route of drug administration. The cysteine protease, cathepsin B, has been implicated in mediating TBI, but it has not been validated by gene knockout (KO) studies. Therefore, this investigation evaluated mice with deletion of the cathepsin B gene receiving controlled cortical impact TBI trauma. Results indicated that KO of the cathepsin B gene resulted in amelioration of TBI, shown by significant improvement in motor dysfunction, reduced brain lesion volume, greater neuronal density in brain, and lack of increased proapoptotic Bax levels. Notably, oral administration of the small-molecule cysteine protease inhibitor, E64d, immediately after TBI resulted in recovery of TBI-mediated motor dysfunction and reduced the increase in cathepsin B activity induced by TBI. E64d outcomes were as effective as cathepsin B gene deletion for improving TBI. E64d treatment was effective even when administered 8 h after injury, indicating a clinically plausible time period for acute therapeutic intervention. These data demonstrate that a cysteine protease inhibitor can be orally efficacious in a TBI animal model when administered at a clinically relevant time point post-trauma, and that E64d-mediated improvement of TBI is primarily the result of inhibition of cathepsin B activity. These results validate cathepsin B as a new TBI therapeutic target.

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In vitro and in vivo inhibition of cysteine proteinases by EST, a new analog of E-64.

Cited by 167 publications

References 16 publications

Dissection of the Autophagosome Maturation Process by a Novel Reporter Protein, Tandem Fluorescent-Tagged LC3

Dissection of the Autophagosome Maturation Process by a Novel Reporter Protein, Tandem Fluorescent-Tagged LC3

Calpain activation by hepatitis C virus proteins inhibits the extrinsic apoptotic signaling pathway

The Cysteine Protease Cathepsin B Is a Key Drug Target and Cysteine Protease Inhibitors Are Potential Therapeutics for Traumatic Brain Injury

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