JCJC SIMI 7 - JCJC - SIMI 7 - Chimie moléculaire, organique, de coordination, catalyse et chimie biologique

Rational design and synthesis of probes for chemical biology of the crypt-proteases – CHEM-CRYPTIDASES

Explore the role of Insulin degrading enzyme prototype of cryptidases.

Design, synthesize and evaluate small molecules that modulate Insulin Degrading Enzyme, a protein at the crossroads of Alzheimer’s disease and Diabetes.

Untangle the roles of the cryptidase Insulin degrading enzyme.

Alzheimer’s disease is a devastating neurodegenerative disorder concerning about 20 million people. Among key features is the progressive accumulation of amyloid peptide deposits. leading to loss of neurons, cognitive deficit, and finally death. Understanding the mechanism of amyloid pathologically accumulation in the brain is crucial. Recently, pathways of elimination of amyloid peptides by metalloproteases such as the cryptidase insulin degrading-enzyme IDE were discovered. Pathological downregulation of this enzyme is correlated with the development of the pathology. Discovering the exact role of Insulin-Degrading Enzyme may lead to new therapeutic approaches. <br />Diabetes melittus is a life-threatening disorder that targets more than 300 million people. The goal of most anti-diabetic therapies is to improve insulin signaling. Theoretically, it should be possible to reach the same goal by inhibiting Insulin-Degrading Enzyme-mediated insulin catabolism. Despite half a century of research, the development of inhibitors has been an elusive goal. Our objective is to design chemical probes to explore the role of this enzyme in both pathologies, taking into account the peculiar structure of this protein.

In order to obtain probes that will be useful to biologists for untangling the many roles of the cryptidase Insulin Degrading Enzyme in several biological systems, we design and synthesize compounds that modulate the substrate specificity of this enzyme. Importantly for us, IDE cleaves numerous substrates, and stands at the crossroads of several metabolic pathways. A critical point is the specificity of the action of our compounds regarding these substrates. Also, we characterize the selectivity of our probes towards other metalloproteases as well as their physico-chemical, metabolic stability and bioavailability properties.
Also, we design compounds by step-wise optimization of two chemical series from known hits found in house (exosite binders and click-adducts). The binding of these compounds is characterized using X-Ray diffraction and Ligand-based Nuclear Magnetic Resonance.
To identify the best compounds to be used as probes, about 100 must be synthesized and tested. Also, at least one cellular assay is to be developed to assess the activity of the compounds in a more-complex system. Finally, the best compounds have to be tested in mice models of the pathology.

This project has allowed the optimization of the first dual-ligands of IDE . A fluorescence-based assay was developed to measure the hydrolysis of the amyloid peptide and insulin by IDE. Crystallography and NMR provided binding modes of compounds. Assays in pancreatic, muscle or neuronal cells allowed measuring the effect of compounds in a complex environment. At last, we have identified the first pan-, drug-like inhibitor of IDE. Its effect in a in vivo model of glucose tolerance enlightened the role of IDE in diabetes.

This project will deliver innovative chemical biology tools such as the first substrate-dependent enzyme modulators and catalytic-site-driven modulators, to understand ways to modulate a cryptidase like IDE. An integration of all the data obtained (structural, in vitro, on cells) will allow the biologists to chose the best probe to evaluate in vitro and in vivo and possibly to define the profile of an optimal IDE modulator to be used therapeutically in Alzheimer’s disease or diabetes.

We published an original method for the synthesis of benzimidazoles (Tet Lett, 53, 2012, 2440-2443). This project led to two other publications: original binding mode of the first dual-site ligands (EJMC 2014, 79, 184-193); full structure-activity relationships as well as drug-like and pharmacokinetics properties (EJMC 2015,90,547-567). Another paper about the discovery by in situ click-chemistry, in vivo effect of the first catalytic-site inhibitors of IDE is currently in revision.

Cryptidases are Zinc dependent proteases that encapsulate their substrate in a large inner cavity called “crypt”, before hydrolysis. The members of the family differ one from the other by the shape, size and substrate recognition ability of their crypt. Members of this small family of enzymes are found in many living organisms including bacteria, plants and animals.
Insulin-degrading enzyme (IDE) is the prototype of this family of proteases. It hydrolyses many different polypeptides involved in important signaling pathways: amyloid beta, insulin, glucagon, IGF-II,ubiquitin. Although the enzyme is ubiquitous and known for long, and its 3D structure recently solved with several substrates, its role remains poorly understood.
The objective of this chemical biology project held at the Lille Pasteur Institute is to design and synthesize chemical probes that will modulate the substrate specificity of IDE. The probes will be designed using structural data from X-ray, molecular modeling and medicinal chemistry reasoning. The project will build on preliminary structure-activity relationships already obtained by the principal investigator. In particular, her team will exploit series that bind to the catalytic zinc but also to other places in the crypt including a genuine substrate binding exosite. Structural modifications will be made to optimize surface binding (classical) and volume filling (specific of crypt containing enzyme).
The goal is to obtain several nanomolar compounds with various inhibition/activation profiles and physico-chemical properties suitable for a use in complex biological systems (whole cells assays and in vivo experiments). The probes will be useful to biologists for untangling the many roles of IDE in several biological systems. More generally, the concept and the methodology could be used for other cryptidases such as the recently discovered Presequence pepdidase.

Project coordination

Rebecca Deprez-Poulain (INSTITUT PASTEUR DE LILLE) – rebecca.deprez@univ-lille2.fr

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.



Help of the ANR 150,000 euros
Beginning and duration of the scientific project: February 2012 - 24 Months

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