Carboxylate isosteres for caspase inhibitors: the acylsulfonamide case revisited.

As part of an ongoing effort to discover inhibitors of caspase-1 with an optimized selectivity and biopharmaceutical profile, acylsulfonamides were explored as carboxylate isosteres for caspase inhibitors. Acylsulfonamide analogues of the clinically investigated caspase-1 inhibitor VRT-043198 and of the pan-caspase inhibitor Z-VAD-CHO were synthesized. The isostere-containing analogues with an aldehyde warhead had inhibitory potencies comparable to the carboxylate references. In addition, the conformational and tautomeric characteristics of these molecules were determined using 1H- and 13C-based NMR. The propensity of acylsulfonamides with an aldehyde warhead to occur in a ring-closed conformation at physiological pH significantly increases the sensitivity to hydrolysis of the acylsulfonamide moiety, yielding the parent carboxylate containing inhibitors. These results indicate that the acylsulfonamide analogues of the aldehyde-based inhibitor VRT-043198 might have potential as a novel type of prodrug for the latter. Finally, inhibition of caspase 1 and 11 mediated inflammation in mouse macrophages was found to correlate with the potencies of the compounds in enzymatic assays.

Ligand-induced conformational changes in prolyl oligopeptidase: a kinetic approach.

Most kinetic studies of prolyl oligopeptidase (PREP) were performed with the porcine enzyme using modified peptide substrates. Yet recent biophysical studies used the human homolog. Therefore, the aim of this study was to compare the kinetic behavior of human and porcine PREP, as well as to find a suitable method to study enzyme kinetics with an unmodified biological substrate. It was found that human PREP behaves identically to the porcine homolog, displaying a double bell-shaped pH profile and a pH-dependent solvent kinetic isotope effect of the kcat/Km, features that set it apart from the related exopeptidase dipeptidyl peptidase IV (DPP IV). However, the empirical temperature coefficient Q10, describing the temperature dependency of the kinetic parameters and the non-linear Arrhenius plot of kcat/Km are common characteristics between PREP and DPP IV. The results also demonstrate the feasibility of microcalorimetry for measuring turn-over of proline containing peptides.

Hearing prevention experience.

Hearing prevention experience. PROBLEMS/OBJECTIVES: Many soldiers suffer from serious NIHL due to the lack of an effective hearing prevention policy. To conduct a study to ameliorate the hearing prevention policy. METHODOLOGY: Literature review Results: Sensitization, innovative hearing protection systems, CEP systems for pilots, using OAEs to identify the actual hearing damage of soldiers and motivating them to better protect themselves, and applying the AHAAH algorithm can determine the number of allowable impulses Conclusions: 1) In line with European and Belgian legislation in this context, tremendous efforts have been made by the Belgian military for information about and sensitization regarding the issue of NIHL. 2) The high variety of positions, functions and environmental factors in a military setting makes the inventory of noise exposures complex and extensive. 3) Efficacious ear protection systems have been rapidly implemented in relation to the most exposed work conditions and, finally, extended to all potential dangerous environments. 4) The use of OAEs was not only a pivotal element in the screening of military staff, but also in their conscientization towards the potentially dramatic consequences of excessive noise exposure. 5) Currently, some "real-life" measurements are being conducted in order to provide military authorities with a complete cadastre on the noise dangers in the Belgian military. 6) The role of military ENT specialists (i.e., ENT and occupational medicine doctors, totally familiar with the working conditions of military personnel), audiologists and specialists in occupational medicine was critical to the success of these initiatives.

Plasma levels of carboxypeptidase U (CPU, CPB2 or TAFIa) are elevated in patients with acute myocardial infarction.

BACKGROUND: Two decades after its discovery, carboxypeptidase U (CPU, CPB2 or TAFIa) has become a compelling drug target in thrombosis research. However, given the difficulty of measuring CPU in the blood circulation and the demanding sample collecton requirements, previous clinical studies focused mainly on measuring its inactive precursor, proCPU (proCPB2 or TAFI). OBJECTIVES: Using a sensitive and specific enzymatic assay, we investigated plasma CPU levels in patients presenting with acute myocardial infarction (AMI) and in controls. METHODS: In this case-control study, peripheral arterial blood samples were collected from 45 patients with AMI (25 with ST segment elevation myocardial infarction [STEMI], 20 with non-ST segment elevation myocardial infarction [NSTEMI]) and 42 controls. Additionally, intracoronary blood samples were collected from 11 STEMI patients during thrombus aspiration. Subsequently, proCPU and CPU plasma concentrations in all samples were measured by means of an activity-based assay, using Bz-o-cyano-Phe-Arg as a selective substrate. RESULTS: CPU activity levels were higher in patients with AMI (median LOD-LOQ, range 0-1277 mU L(-1) ) than in controls (median < LOD, range 0-128 mU L(-1) ). No correlation was found between CPU levels and AMI type (NSTEMI [median between LOD-LOQ, range 0-465 mU L(-1) ] vs. STEMI [median between LOD-LOQ, range 0-1277 mU L(-1) ]). Intracoronary samples (median 109 mU L(-1) , range 0-759 mU L(-1) ) contained higher CPU levels than did peripheral samples (median between LOD-LOQ, range 0-107 mU L(-1) ), indicating increased local CPU generation. With regard to proCPU, we found lower levels in AMI patients (median 910 U L(-1) , range 706-1224 U L(-1) ) than in controls (median 1010 U L(-1) , range 753-1396 U L(-1) ). CONCLUSIONS: AMI patients have higher plasma CPU levels and lower proCPU levels than controls. This finding indicates in vivo generation of functional active CPU in patients with AMI.

Antiprotozoal activity of synthetic amino substituted 1-methyl-1 H-alpha-carbolines.

The antiprotozoal properties of a series of amino substituted 1-methyl-1 H-alpha-carbolines were investigated in a broad panel of parasites. Various substituents were systematically introduced at various positions on the carbocyclic ring of the parent 1-methyl-1 H-alpha-carboline. Most compounds showed a potent antiprotozoal activity, although mostly accompanied by cytotoxicity on MRC-5 cells. One compound, containing the same amino-substitution as chloroquine, showed an IC50 against Plasmodium falciparum of 2.37 microM and was reasonably selective.

A prolyl oligopeptidase inhibitor, KYP-2047, reduces α-synuclein protein levels and aggregates in cellular and animal models of Parkinson's disease.

BACKGROUND AND PURPOSE: The aggregation of α-synuclein is connected to the pathology of Parkinson's disease and prolyl oligopeptidase (PREP) accelerates the aggregation of α-synuclein in vitro. The aim of this study was to investigate the effects of a PREP inhibitor, KYP-2047, on α-synuclein aggregation in cell lines overexpressing wild-type or A30P/A53T mutant human α-syn and in the brains of two A30P α-synuclein transgenic mouse strains. EXPERIMENTAL APPROACH: Cells were exposed to oxidative stress and then incubated with the PREP inhibitor during or after the stress. Wild-type or transgenic mice were treated for 5 days with KYP-2047 (2 × 3 mg·kg(-1) a day). Besides immunohistochemistry and thioflavin S staining, soluble and insoluble α-synuclein protein levels were measured by Western blot. α-synuclein mRNA levels were quantified by PCR. The colocalization of PREP and α-synuclein,and the effect of KYP-2047 on cell viability were also investigated. KEY RESULTS: In cell lines, oxidative stress induced a robust aggregation of α-synuclein,and low concentrations of KYP-2047 significantly reduced the number of cells with α-synuclein inclusions while abolishing the colocalization of α-synuclein and PREP. KYP-2047 significantly reduced the amount of aggregated α-synuclein,and it had beneficial effects on cell viability. In the transgenic mice, a 5-day treatment with the PREP inhibitor reduced the amount of α-synuclein immunoreactivity and soluble α-synuclein protein in the brain. CONCLUSIONS AND IMPLICATIONS: The results suggest that the PREP may play a role in brain accumulation and aggregation of α-synuclein, while KYP-2047 seems to effectively prevent these processes.

Craniofacial embryology and postnatal development of relevant parts of the upper respiratory system.

OBJECTIVES: To compare historical and current knowledge relating to the development of the paranasal sinuses, the nose and face, the Eustachian tube and temporal bones, particularly with respect to chronic inflammation during childhood. METHODOLOGY: Traditional literature data, mainly emanating from text books, were supplemented with information based on a non-structured PubMed search covering the last two decades. RESULTS: Historical knowledge has most often been confirmed, sometimes supplemented and only rarely challenged by present-day studies. Recent studies focus mainly on the clinical application of modern imaging techniques. CONCLUSIONS: Interest in the development of relevant parts of the upper respiratory system remains as lively as ever. Imaging techniques with low or absent radiation exposure may give rise to a novel field of research, especially with respect to paediatric rhinosinusitis.

Clinical aspects of chronic ENT inflammation in children.

In children, all ENT cavities are particularly prone to the development of chronic inflammation. This is due to many predisposing factors, of which the most common are unfavourable anatomy, absence of nasal blowing, day care attendance, allergy, immature immunity, gastro-oesophageal reflux and tobacco smoke exposure. The aim of this paper is to outline the most specific paediatric clinical aspects of chronic pharyngo-tonsillitis, rhinosinusitis, otitis media, adenoiditis and laryngotracheitis and the important influence that some of these pathologies exert on the others.

Inhibitors of the Purine Salvage Pathway: A Valuable Approach for Antiprotozoal Chemotherapy?

For many years, the purine salvage pathway of parasitic protozoa has been regarded as an attractive chemotherapeutic target. Parasitic protozoa lack de novo synthesis and rely entirely on the purine salvage pathway to meet their purine demands. Because of the great phylogenetic difference between parasite and host, there are often sufficient distinctions that can be exploited to design specific inhibitors for the parasitic enzymes. As a result, this pathway has been thoroughly investigated over the last twenty years. It is only quite recently that the genome studies of Trypanosoma, Leishmania and Plasmodium have been published. Based on these genomic data however, the existence of by-pass mechanisms by other enzymes and transporter systems could be suggested. Taking into account such proposition, the question might arise as to whether inhibition of a single salvage enzyme will be able or not to cause parasite death or growth arrest. In this paper, the key enzymes in the purine salvage pathways of relevant pathogenic species from the genera Trypanosoma, Leishmania and Plasmodium are reviewed. Their potential as drug targets is critically evaluated and where possible, correlated to literature data on antiparasitic activity of their inhibitors. While many studies over the past ten years have yielded contradictory results, this review attempts to clarify these findings by discussing the latest elements of progress in the field. Additionally, as part of a broader discussion on substrate analogue types of inhibitors, special attention is paid to iminoribitol derivatives, serving as transition state analogues of nucleoside-processing enzymes and comprising the most potent inhibitors reported for purine salvage enzymes. More specifically, the development of three generations of immucillins and a newer series of N-(arylmethyl-) substituted iminoribitol derivatives will be discussed. Finally, this review also covers subversive substrates of salvage enzymes: compounds that are transformed by enzymatic activity into cytotoxic agents. Although not by directly intervening in the process of purine recovery, the subversive substrate approach might deliver antiprotozoal compounds that rely on salvage enzymes for their activity.

Synthesis and biochemical evaluation of guanidino-alkyl-ribitol derivatives as nucleoside hydrolase inhibitors.

Nucleoside hydrolase (NH) is a key enzyme in the purine salvage pathway. The purine specificity of the IAG-NH from Trypanosoma vivax is at least in part due to cation-pi-stacking interactions. Guanidinium ions can be involved in cation-pi-stacking interactions, therefore a series of guanidino-alkyl-ribitol derivatives were synthesized in order to examine the binding affinity of these compounds towards the target enzyme. The compounds show moderate to good inhibiting activity towards the IAG-NH from T. vivax.

1,2,3-Triazolylalkylribitol derivatives as nucleoside hydrolase inhibitors.

A range of novel 1,2,3-triazolylalkylribitol derivatives were synthesized and evaluated as nucleoside hydrolase inhibitors. The most active compound (11a) has low micromolar potency and is structurally diverse from previously reported nucleoside hydrolase inhibitors, which, along with the simplicity of the chemistry involved in its synthesis, makes it a good lead for the further development of novel nucleoside hydrolase inhibitors.

The therapeutic potential of inhibitors of dipeptidyl peptidase IV (DPP IV) and related proline-specific dipeptidyl aminopeptidases.

In this review the structural and functional aspects of dipeptidyl peptidase IV (DPP IV) will be described, and the therapeutic potential of DPP IV inhibitors will be highlighted. DPP IV will be situated in clan SC, a group of serine proteases that contains several proline specific peptidases. Structural aspects of DPP IV and its interaction with different types of inhibitors are recently revealed by the publication of several crystal structures. Especially the design and development of new DPP IV inhibitors based on the three-dimensional structure, substrate specificity and catalytic mechanism will be discussed. In the last years there was an important development of new pyrrolidine-2-nitriles with very promising therapeutic properties for the treatment of type 2 diabetes. The role of DPP IV in peptide metabolism of members of the PACAP/glucagon peptide family, neuropeptides and chemokines has been thoroughly investigated during recent years. This is directly related to the promising therapeutic potential of DPP IV inhibitors in the treatment of type 2 diabetes and in the treatment of immunological disorders. Several inhibitors are currently under investigation in clinical trials for the treatment of type 2 diabetes and represent a new class of drugs for the treatment of this disease.

Development of irreversible diphenyl phosphonate inhibitors for urokinase plasminogen activator.

In this letter we report the synthesis and biochemical evaluation of selective, irreversible diphenyl phosphonate inhibitors for urokinase plasminogen activator (uPA). A diphenyl phosphonate group was introduced on the substratelike peptide Z-d-Ser-Ala-Arg, and modification of the guanidine side chain was investigated. A guanylated benzyl group appeared the most promising side chain modification. A k(app) value in the 10(3) M(-1) s(-1) range for uPA was obtained, together with a selectivity index higher than 240 toward other trypsin-like proteases such as tPA, thrombin, plasmin, and FXa.

CT-scan study of the incidence of sinus involvement and nasal anatomic variations in 196 children.

CT-scan was used to examine rhinosinusitis in the developing sinuses; 196 children aged from 3 to 14 years were selected on the base of their chronic rhinorrhea, nasal congestion and cough. The patients were subdivided into six age groups (3-4, 5-6, 7-8, 9-10, 11-12 and 13-14 years). In the youngest age group, the authors noted maxillary involvement in 63%, ethmoidal involvement in 58%, and even sphenoidal sinus involvement in 29% of the children. Involvement decreased gradually with age, with 10% of ethmoidal and 0% of sphenoidal involvement in the 13-14 years age group. Maxillary sinusitis, however, persisted very frequently in the oldest age group (65%). Frontal involvement seems to become significant at the age of 7-8 years (7%) but it never exceeds 15% (11-12 age group). Septal deviations occurred in 16% of the youngest up to 72% in the oldest age group. The prevalence of bullous conchae increased with age too, although less prominently.

Some remarks on nasal polyposis.

After a short historical and literature review the authors present their definition of nasal polyposis. The importance of CT scanning is stressed and the results of a CT-scan study of 350 patients with nasal complaints is presented. The authors discuss the incidence of sinusitis, presence of polypi and nasal anomalies in this population. Furthermore 111 biopsies of nasal polypi were studied and the cellular content, the ducts, glands, veins, aspect of the basal membrane and epithelial layer were described. As an oral ASA provocation test can be hazardous, the authors tested a nasal provocation test with acetylsalicylic acid. Although the test showed some interesting results, the reproducibility was poor. Finally the authors conclude that endoscopy and CT-scanning enable the diagnosis of nasal polyposis in an early stage of the disease.

Incidence and etiology of rhinosinusitis in children.

The authors discuss the incidence of chronic rhinosinusitis in children. From the study of 196 CT-scans, there follows that as well the ethmoidal as the maxillary sinuses are involved in the young child. In older children, however, the incidence of maxillary sinusitis is more frequent. The etiology of chronic rhinosinusitis is discussed stressing the importance of general factors such as environmental factors, congenital and acquired abnormalities and the influence of drugs, as well as of local factors such as again environmental factors, malformations, and upper respiratory infections. Most of these factors are discussed following personal studies.

[CAT-scan study of the prevalence of sinus disorders and anatomical variations in 196 children]. / CAT-scan studie over het voorkomen van sinusaantasting en anatomische variaties bij 196 kinderen.

CT-scan was used to examine rhinosinusitis in the developing sinuses; 196 children aged from 3 to 14 years were selected on the base of their chronic rhinorrhea, nasal congestion and cough. The patient group was subdivided into six age groups (3-4, 5-6, 7-8, 9-10, 11-12 and 13-14 years). In the youngest age group, the authors noted maxillary involvement in 63%, ethmoidal involvement in 58%, and even sphenoidal sinus involvement in 29% of the children. Involvement decreased gradually with age, with 10% of ethmoidal and 0% of sphenoidal involvement in the 13-14 years age group. Maxillary sinusitis, however, persisted very frequently in the oldest age group (65%). Frontal involvement seems to become significant at the age of 7-8 years (7%) but it never exceeds 15% (11-12 age group). Septal deviations occurred in 16% of the youngest up to 72% in the oldest age group. A prevalence of bullous conchae increased with age too, although less prominently.

Recurrent polyposis nasi. Documentation.

After a short historical review a proposal is made for the definition of nasal polyposis. The authors studied 350 CT-scans of patients with nasal complaints. In a high percentage anatomical anomalies were observed. In 57.5% of the CT-scans sinus mucosal disease was visible. In all patients with maxillary sinus disease polyps (rounded structures) could be found; in 31% these polyps were mainly of grade 2. From a retrospective study of 111 biopsies of nasal polyposis (65 patients) it became clear that different polyps from the same patient showed substantial difference in cellular content, i.e. presence of eosinophils, neutrophils, plasma cells, glands, ducti and thickening of the basal membrane. As oral acetylsalicylic acid provocation may be hazardous in ASA-sensitive patients, the authors developed a nasal aspirin provocation test. This nasal ASA test was carried out in 10 normal test subjects, 10 patients with aspecific hyperreactivity, 10 atopic patients and 16 patients with polyposis nasi. The reproducibility of the test, however, was so poor that the nasal ASA challenge test in its present form does not appear to be of any great clinical value. Finally, the authors discuss the physiopathology of nasal polyposis.