Letters in Drug Design & Discovery - Volume 4, Issue 8, 2007

In order to develop novel therapies for the treatment of central nervous system diseases, methods for rapid quantification of different neural cell populations, from healthy or diseased brains, are highly desirable. Today, the method of choice to quantify different cell populations in the brain is immunohistochemistry. This technique is precise and reliable, but is also very time consuming and cost-intensive. In this paper, we describe a rapid procedure where neurons and neural stem/progenitor cells from the adult mouse are isolated, immediately fixed, and quantified with different cell type specific markers by flow cytometry. Our results show that we can reproducibly detect and quantify mature neurons with antibodies towards microtubuli associated protein (MAP2) and neuronal nuclei (NeuN), both in hippocampus and olfactory bulb. Stem/progenitor cell detection and quantification were achieved using antibodies specific for bromodeoxyuridine (BrdU) and doublecortin (DCX). The flow cytometric analysis revealed fractions of positive cells corresponding to the in vivo situation, confirmed and validated by traditional immunohistochemistry and stereological counting procedures. We conclude that the flow cytometric technique can be used as a rapid screening method to accurately quantify neuronal populations and to detect and quantify other brain cell types, such as neural stem/progenitor cells.

We evaluated whether V-5 Immunitor (V5), tableted immunotherapeutic preparation comprising heatinactivated HBV antigens from pooled blood of HBV- and HCV-infected donors, may produce clinical benefit through induction of oral tolerance and reduction of immune-mediated liver injury. Once daily dose of V5 was administered per os to 10 patients with chronic hepatitis B in an open label study that lasted one month. Every patient who entered the study had elevated liver enzyme levels, which at the end of study have decreased in 100% of analyzed patients. The reduction was highly significant, from 112.4 to 44.4 U/L (P=0.00009) and 118.8 to 46.1 U/L (P=0.0032), for ALT and AST respectively. In addition, half of intent-to-treat patients who were HBV surface antigen (HBsAg) positive at study entry, became negative after one month on V5 (P=0.0098). All patients, except one, reported complete recuperation from hepatitisassociated clinical symptoms present at baseline (P=0.0016). No adverse events were observed at any time. Favorable biochemical, virological and clinical responses indicate that V5 is safe and effective means for immunotherapy of chronic hepatitis B. Placebo-controlled, randomized study is required to confirm these findings.

We report the results of the evaluation of a series of benzyl imidazole-based compounds against the enzyme aromatase (AR). The results show that the synthesised compounds were potent inhibitors of AR in comparison to the standard compound aminoglutethimide (AG).

A new series of benzothiazolo-benzothiadiazine hybrids (9a-g) has been synthesized and evaluated for their antibacterial activity against clinical isolates of Gram-positive and Gram-negative bacteria. Some of these hybrids in this series exhibited antibacterial activity comparable to ampicillin. In addition, some of the compounds showed DNA gyrase inhibitory activity at a moderate level compared to ciprofloxacin.

Mitogen-activated protein kinase-activated protein kinase 2 (MK-2) plays an important role in treatment of inflammatory disease such as rheumatoid arthritis. CoMFA was conducted on thirty-one analogs displaying variable inhibition of MK-2 to determine the structural requirements for potency in inhibiting MK-2. The resulting model exhibited good q2 and r2 values up to 0.549 and 0.973 for CoMFA, the standard error of estimation was 0.098. The contributions from the steric and electrostatic fields were 0.586 and 0.414 respectively. The 3D-QSAR model should be very useful for design of novel MK-2 inhibitors.

Two microplate assays for inhibitor testing were established and used to identify new hyaluronidase inhibitors from a series of benzimidazole derivatives. In vitro assays were performed at pH 7 using a “stains-all” based assay and at pH 3.5 using a Morgan-Elson assay. Twelve benzimidazole derivatives were synthesized and tested on hyaluronidase inhibition at a concentration of 100 μM and IC50 values of the compounds with an inhibition of more than 50 % were determined. The most potent compound was 4b (Di(1H-benzo[d]imidazol-2-yl)methane) with an IC50 value of 78 μM at pH 7 and 72 μM at pH 3.5, respectively.

The cysteine hydrolase N ω,N ω-dimethyl-L-arginine dimethylaminohydrolase-1 (DDAH-1) is an important regulator of NO production through the degradation of endogenous Nω-methylated arginines, that are competitive inhibitors for nitric oxide synthase (NOS). Consequently, DDAH-1 is a target for pharmacological drug design to regulate NO production. The appearance of a second isoform DDAH-2, which was assigned through sequence comparison, requires detailed knowledge about the properties of both proteins. This study represents the first attempt for a structural and functional characterization of DDAH-2. However, lack in enzymatic activity together with structural consideration based on a homology model places the designation as a DDAH enzyme into question. On the other hand, it is shown that DDAH-2 is a substrate for two protein-arginine methyltransferases, CARM-1 and PRMT-6, which supports the participation of DDAH-2 in the metabolism of side-chain methylated arginines.

A new method for the synthesis of quinolines has been developed by employing various novel catalysts such as Al(OTf)3, Gd(OTf)3, and TMSCl-NaI. Later, these compounds were condensed with 3-hydrazino benzothiadiazines to obtain quinoline derived [1,2,4]triazolo[1,5-b][1,2,4] benzothiadiazines. Further, these new molecules have been evaluated for their antibacterial activity and some of them have exhibited significant antibacterial activity in comparison to sulfadiazine.

Molecular orbital calculations are used to determine structural and electronic characteristics of a series of sugar mimics, which are correlated with experimentally determined activity toward glycosidases. Differences in activity between α- and β-glycosidase correlate to the position of the HOMO surface of the neutral form of the inhibitor. A correlation is also found between relative activity and energy of the HOMO.

New A-C8 and C-C2-linked 6-chloropurine-pyrrolo[2,1-c][1,4]benzodiazepine hybrids have been prepared and evaluated for their anticancer potential. The molecular modeling studies might be explained in terms of effect of the molecule on binding in the minor groove of DNA and also comparison to both A-C8/C-C2-position of the PBD. These PBD conjugates have shown DNA-binding ability and promising anticancer activity.

To discover new potential sigma ligands, a library of 64 members based on a carbamidic scaffold was designed and a synthetic protocol, based on PASPS and MAOS techniques, was developed and optimized. Applying the developed protocol the library compounds were obtained in high purity, suitable for biological screening.