ABSTRACT
Insulin-degrading enzyme (IDE) is a protease that cleaves insulin and other bioactive peptides such as amyloid-ß. Knockout and genetic studies have linked IDE to Alzheimer's disease and type-2 diabetes. As the major insulin-degrading protease, IDE is a candidate drug target in diabetes. Here we have used kinetic target-guided synthesis to design the first catalytic site inhibitor of IDE suitable for in vivo studies (BDM44768). Crystallographic and small angle X-ray scattering analyses show that it locks IDE in a closed conformation. Among a panel of metalloproteases, BDM44768 selectively inhibits IDE. Acute treatment of mice with BDM44768 increases insulin signalling and surprisingly impairs glucose tolerance in an IDE-dependent manner. These results confirm that IDE is involved in pathways that modulate short-term glucose homeostasis, but casts doubt on the general usefulness of the inhibition of IDE catalytic activity to treat diabetes.
Subject(s)
Hydroxamic Acids/chemical synthesis , Insulysin/antagonists & inhibitors , Triazoles/chemical synthesis , Animals , Caco-2 Cells , Catalytic Domain , Diabetes Mellitus/drug therapy , Drug Evaluation, Preclinical , Glucose Tolerance Test , Humans , Hydroxamic Acids/pharmacology , Hydroxamic Acids/therapeutic use , Male , Mice , Mice, Inbred C57BL , Microsomes, Liver , Molecular Targeted Therapy , Random Allocation , Structure-Activity Relationship , Triazoles/pharmacology , Triazoles/therapeutic useABSTRACT
One approach currently being developed in anticancer drug discovery is to search for small compounds capable of occupying and blocking the hydrophobic pocket of anti-apoptotic Bcl-2 family members necessary for interacting with pro-apoptotic proteins. Such an approach led to the discovery of several compounds, such as ABT-737 (which interacts with Bcl-2, Bcl-xl, and Bcl-w) or the latest one, ABT-199, that selectively targets Bcl-2 protein. The efficacy of those compounds is, however, limited by the expression of two other anti-apoptotic Bcl-2 members, Mcl-1 and Bfl-1. Based on the role of Bfl-1 in cancer, especially in chemoresistance associated with its overexpression in B-cell malignancies, we searched for modulators of protein-protein interaction through a high-throughput screening of a designed chemical library with relaxed drug-like properties to identify small molecules targeting Bfl-1 anti-apoptotic protein. We found two compounds that display electrophilic functions, interact with Bfl-1, inhibit Bfl-1 protective activity, and promote cell death of malignant B cells. Of particular interest, we observed a synergistic effect of those compounds with ABT-737 in Bfl-1 overexpressing lymphoma cell lines. Our results provide the basis for the development of Bfl-1 specific antagonists for antitumor therapies.
Subject(s)
Antineoplastic Agents/chemistry , Apoptosis , Biphenyl Compounds/pharmacology , Drug Discovery/methods , Drug Resistance , Lymphoma/drug therapy , Nitrophenols/pharmacology , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Sulfonamides/pharmacology , Apoptosis Regulatory Proteins , Bridged Bicyclo Compounds, Heterocyclic/chemistry , Cell Line, Tumor , Glutathione/chemistry , Glutathione Transferase/metabolism , High-Throughput Screening Assays/methods , Humans , Minor Histocompatibility Antigens , Molecular Conformation , Piperazines/pharmacology , Protein Binding , Protein Interaction Mapping , Spectrometry, Fluorescence , Sulfonamides/chemistryABSTRACT
Malaria is a severe infectious disease that causes between 655,000 and 1.2 million deaths annually. To overcome the resistance to current drugs, new biological targets are needed for drug development. Aminopeptidase M1 (PfAM1), a zinc metalloprotease, has been proposed as a new drug target to fight malaria. Herein, we disclosed the structure-activity relationships of a selective family of hydroxamate PfAM1 inhibitors based on the malonic template. In particular, we performed a "fluoro-scanning" around hit 1 that enlightened the key positions of the halogen for activity. The docking of the best inhibitor 2 is consistent with in vitro results. The stability of 2 was evaluated in microsomes, in plasma, and toward glutathione. The in vivo distribution study performed with the nanomolar hydroxamate inhibitor 2 (BDM14471) revealed that it reaches its site of action. However, it fails to kill the parasite at concentrations relevant to the enzymatic inhibitory potency, suggesting that killing the parasite remains a challenge for potent and druglike catalytic-site binding PfAM1 inhibitors. In all, this study provides important insights for the design of inhibitors of PfAM1 and the validity of this target.
Subject(s)
Aminopeptidases/antagonists & inhibitors , Antimalarials/chemical synthesis , Malonates/chemical synthesis , Metalloproteases/antagonists & inhibitors , Plasmodium falciparum/enzymology , Protozoan Proteins/antagonists & inhibitors , Animals , Antimalarials/blood , Antimalarials/pharmacology , Cell Line , Drug Resistance , Female , Humans , Malaria/drug therapy , Malonates/blood , Malonates/pharmacology , Mice , Molecular Docking Simulation , Plasmodium berghei , Plasmodium falciparum/drug effects , Protein Binding , Rats , Solubility , Stereoisomerism , Structure-Activity Relationship , Tissue Distribution , ZincABSTRACT
Hydroxamates are valuable tools for chemical biology as well as interesting leads for medicinal chemistry. Although many hydroxamates display nanomolar activities against metalloproteases, only three hydroxamates have reached the market, among which is the HDAC inhibitor vorinostat. Failures in development are generally attributed to lack of selectivity, toxicity, or poor stability. To help medicinal chemists with respect to plasma stability, we have performed the first and preliminary study on structure-plasma stability for hydroxamates. We define some structural rules to predict or improve the plasma stability in the preclinical stage.
