Current Topics in Medicinal Chemistry - Volume 6, Issue 4, 2006

Protease inhibitors are used to treat and manage a variety of diseases, and have made its mark in drug development. Angiotensin converting enzyme (ACE) inhibitors such as captopril, enalapril, lisinopril, fosinopril, quinapril and ramipril are used to control blood pressure in hypertensive patients. Human immunodeficiency virus (HIV) protease inhibitors, exemplified by amprenavir, indinavir, lopinavir, nelfinavir, ritonavir, and saquinavir are used in treating acquired immunodeficiency syndrome (AIDS). Doxycycline hyclate is the first collagenase inhibitor used for the treatment of periodontal disease. Ximelagatran is a thrombin inhibitor used in venous thrombosis. Broad spectrum inhibitors include camostat mesylate and nafamostat, which help to maintain the pancreatic microcirculation in acute pancreatitis. Clinical applications of protease inhibitors were reviewed in 2005 (Fairlie, D.P.; Abbenante, G. Medicinal Chemistry 2005, 1, 71-104). There is an International Protease Network (www.protease.net) dedicated to protease researchers. This reflects the importance of protease inhibitors as drug candidates for future development. Proteases play an important role in many pathophysiological processes. There are four classes of proteases, categorized by the catalytic residue that effects the hydrolysis of the substrate's amide bond, namely the metallo, aspartic, serine/threonine and cysteine proteases. This special issue gathers reviews on selected protease targets such as matrix metalloprotease, SARSCoronavirus Main protease, cysteine protease, and secretase inhibition. These reviews emphasize the latest developments in synthetic inhibitors, the screening techniques, and the rationale involved in the design of the inhibitors. The first monograph by Amy Sang et al. presents an overview of matrix metalloprotease inhibitors, with particular emphasis on the use of synthetic inhibitors as prospective agents for the prevention and treatment of cardiovascular and neoplastic diseases. Maurizio Pellechia et al. provide an overview of the use of structure- and fragment-based design guided by X-ray crystallography, NMR spectroscopy, computational and/or extended tethering approaches for the identification of potential non-peptidic agents as cysteine and metalloprotease inhibitors. Tanya Schirmeister et al. describe the design of cysteine protease inhibitors with an electrophilic moiety that can covalently bind to the cysteine residue of the active site of the target protease. Examples of such inhibitors are aziridine derivatives and analogues of the diuretic drug, ethacrynic acid. Selective inhibition of cathepsin K has emerged as a potential new therapy for the treatment of osteoporosis. Tae-Seong Kim et al. report the research and development of a series of non-peptidic cathepsin K inhibitors as an alternative to the use of bisphosphonates and anti-RANK ligand therapies. Severe acute respiratory syndrome (SARS) is an emerging infectious disease caused by a novel human coronavirus (CoV). A 3C-like cysteine protease is essential for the life cycle of SARS-CoV. Po-Huang Liang reviews the characterization of SARS-Coronavirus Main protease and various lead inhibitors. This special issue then shifts to the research and development of β- and γ-secretase inhibitors. These two aspartyl proteases cleave amyloid precursor protein to yield amyloid β. Boris Schmidt et al. describe the selective inhibition of both aspartyl proteases, their peptidic and non-peptidic lead inhibitors, and the metabolism of amyloid precursor protein as a potential cause of Alzheimer's disease. Guy Tremblay et al. discuss the importance of tryptase, and chymase in asthma, and the strategies used to inhibit their detrimental action. I would like to thank the authors for their excellent and timely reviews in their respective fields of expertise, and the reviewers for their suggestions and helpful discussions. This special issue is intended to provide the reader with many of the useful concepts and techniques employed in the medicinal chemistry of protease inhibitors. It is not exhaustive, but it provides a concise update for those already familiar with protease research, and an initial reference resource for those wishing to obtain more detailed information on the topic.

Acting on a broad spectrum of extracellular, intracellular, and membrane-associated substrates, the matrix metalloproteinases (MMPs) are critical to the biological processes of organisms; when aberrantly expressed, many pathological conditions may be born or exacerbated. The prospect of MMP inhibition for therapeutic benefit in cancer, cardiovascular disease, and stroke is reviewed here. MMP inhibitor (MMPI) development constitutes an important branch of research in both academic and industrial settings and advances our knowledge on the structure-function relationship of MMPs. Targeting MMPs in disease treatment is complicated by the fact that MMPs are indispensable for normal development and physiology and by their multi-functionality, possible functional redundancy or contradiction, and context-dependent expression and activity. This complexity was revealed by previous efforts to inhibit MMP activity in the treatment of cancer patients that yielded unsatisfactory results. This review focuses on MMPI development since the late 90s, in terms of natural products and their derivatives, and synthetic compounds of low molecular mass incorporating specific zinc-binding groups (ZBGs). A few polyphenols and flavonoids that exhibit MMPI activities may have chemopreventive and neuro- and cardiovascular-protective effects. A new generation of potent and selective MMPIs with novel ZBGs and inhibition mechanisms have been designed, synthesized, and tested. Although only one collagenase inhibitor (Periostat, doxycycline hyclate) has been approved by the Food and Drug Administration as a drug for the treatment of periodontal disease, new hope is emerging in the form of natural and synthetic MMPIs for the prevention and treatment of stroke, cardiovascular disease, cancer, and other medical conditions.

Proteases are essential enzymes which regulate physiological processes such as inflammation, infection, fertilization, allergic reactions, cell growth and death, blood clotting, tumor growth and bone remodeling. The protease family consists of six major classes of enzymes which are aspartic-, serine-, cysteine-, threonine-, glutamic-, and metalloproteases, all which are implicated in disease propagation. Therefore, protease inhibitors have been of great interest as possible targets for the development of novel therapies. Although, many protease inhibitors have followed a structural design based on either a peptidic or peptidomimetic backbone, other chemical scaffolds have recently emerged. Utilizing structure- and fragment-based design guided by X-ray crystallography, NMR spectroscopy, computational and/or extended tethering approaches, potential non-peptidic therapeutic agents could be identified. In this review, we will report on the recent developments of nonpeptidic cysteine- and metallo- protease inhibitors, focusing on their design by using such strategies.

The roles of cysteine proteases (CP) as protein degrading and protein processing enzymes both in physiological and pathological processes of mammals are well known. Furthermore, the key roles of CP's in the life cycles of infectious agents like protozoa and viruses turn them into new important targets for anti-infective drugs. Thus, the effective inhibition of pathologically relevant cysteine proteases has raised increasing interest in drug development. One strategy to create CP inhibitors is the use of electrophilic moieties, which covalently bind to the cysteine residue of the active site of the target protease. In a previous approach we have selected the aziridine-2,3-dicarboxylic acid as weak electrophilic inhibitor fragment. In order to achieve effective enzyme inhibition this electrophile was incorporated into peptidic or peptidomimetic sequences addressing the substrate binding sites of the protease. High selectivity could be obtained with compounds, which bind into both the primed and non-primed substrate binding pockets. In a second approach the α,β-unsaturated ketone of the well-known diuretic drug ethacrynic acid was found to be another appropriate electrophilic moiety. Derivatives thereof turned out to be new non-peptidic CP inhibitors. Results of inhibition assays obtained with these two inhibitor series on various proteases of human, protozoan, and viral origin, theoretical studies to investigate binding modes and inhibition mechanisms, and structure-activity relationships are presented. Furthermore, the results of in vitro assays on respective pathogens as well as the results of first toxicity studies are summarized.

Cathepsin K is a cysteine protease that plays an important role in the pathological process of bone resorption. Selective cathepsin K inhibitors may thus provide great potential in the treatment of osteoporosis. Pharmaceutical interest in this area is highlighted by the rising number of publications and patent applications. Most recently, the interim results of three clinical trials conducted by Novartis, GlaxoSmithKline, and Merck have strengthened the validation of the target for the therapeutic intervention of osteoporosis. Here we report a series of Cbz-leucyl-(4-piperidinylphenyl)aminoethyl amides based on dipeptidyl anilines for cathepsin K inhibition. These new non-covalent inhibitors exhibit single digit nM inhibition of the cathepsin family. Molecular modeling studies on the interactions responsible for the potency of these inhibitors for cathepsin K will be also discussed.

Severe acute respiratory syndrome (SARS) is an emerging infectious disease caused by a novel human coronavirus (CoV). During the 2003 epidemic, the disease rapidly spread from its origin in southern China to other countries and affected almost 8000 patients, which resulted in about 800 fatalities. A chymotrypsin-like cysteine protease named 3C-like protease (3CLpro) is essential for the life cycle of the SARS-CoV. This main protease is responsible for maturation of functional proteins and represents a key anti-viral target. HPLC and fluorescence-based assays have been used to characterize the protease and to determine the potency of the inhibitors. The fluorogenic method monitoring the increase of fluorescence from the cleavage of a peptide substrate containing an Edans-Dabcyl fluorescence quenching pair at two ends has enabled the use of high throughput screening to speed up the drug discovery process. Several groups of inhibitors have been identified through high throughput screening and rational drug design approaches. Thus, α,β- unsaturated peptidomimetics, anilides, metal-conjugated compounds, boronic acids, quinolinecarboxylate derivatives, thiophenecarboxylates, phthalhydrazide-substituted ketoglutamine analogues, isatin and natural products have been identified as potent inhibitors of the SARS-CoV main protease. The different classes of inhibitors reported in these studies are summarized in this review. Some of these inhibitors could be developed into potential drug candidates, which may provide a solution to combat possible reoccurrence of the SARS and other life-threatening viruses with 3CL proteases.

Most gene mutations associated with Alzheimer's disease point to the metabolism of amyloid precursor protein as potential cause. The β- and γ-secretases are two executioners of amyloid precursor protein processing resulting in amyloid β. Significant progress has been made in the selective inhibition of both proteases, regardless of structural information for γ-secretase. Several peptidic and non-peptidic leads were identified and first drug candidates are in clinical trials. This review focuses on the developments since 2003.

Leukocytes and lung structural cells contribute to the pathophysiology of asthma through the production of numerous mediators including serine proteases. Such proteases include mast cell tryptase and chymase; neutrophil elastase, cathepsin G and myeloblastin (proteinase 3); bronchial epithelial cell-derived transmembrane protease, serine 11D (human airway trypsin-like protease); cytotoxic T lymphocyte- and natural killer cell-derived granzyme B; and, eosinophil serine protease 1 (testisin). Considerable effort to develop potent and selective inhibitors, mostly non-peptidic, especially targeting tryptase and chymase have been made in the last few years. This review presents promising inhibitors, currently in the research and development pipeline. Some endogenous inhibitors and other compounds purified from nonhuman species are also discussed.