THERAPEUTIC STRATEGIES FOR THE INHIBITION OF INDUCIBLE NITRIC OXIDE SYNTHASE—POTENTIAL FOR A NOVEL CLASS OF ANTI-INFLAMMATORY AGENTS

In recent years, NO, a gas previously considered a potentially toxic chemical, has become established as a diffusible universal messenger mediating cell—cell communication throughout the body. In mammals, NO is a recognized mediator of blood vessel relaxation that helps to maintain blood pressure. In the central nervous system NO acts as a non-conventional neurotransmitter and participates in the establishment of long-term plasticity required for memory formation. In addition, NO is responsible for some parts of the host response to sepsis and inflammation and contributes to certain disease states. A number of strategies have emerged with regard to a pharmacological control of pathological NO overproductions. This review will discuss these novel therapeutic approaches that may provide new means for clinical medicine.     

Surprising negative association between IgG1 allotype disparity and anti-adalimumab formation: a cohort study

Introduction

The human monoclonal antibody adalimumab is known to induce an anti-globulin response in some adalimumab-treated patients. Antibodies against adalimumab (AAA) are associated with non-response to treatment. Immunoglobulins, such as adalimumab, carry allotypes which represent slight differences in the amino acid sequences of the constant chains of an IgG molecule. Immunoglobulins with particular IgG (Gm) allotypes are racially distributed and could be immunogenic for individuals who do not express these allotypes. Therefore, we investigated whether a mismatch in IgG allotypes between adalimumab and IgG in adalimumab-treated patients is associated with the development of AAA.

Methods

This cohort study consisted of 250 adalimumab-treated rheumatoid arthritis (RA) patients. IgG allotypes were determined for adalimumab and for all patients. Anti-idiotype antibodies against adalimumab were measured with a regular radio immunoassay (RIA), and a newly developed bridging enzyme linked immunosorbent assay (ELISA) was used to measure anti-allotype antibodies against adalimumab. The association between AAA and the G1m3 and the G1m17 allotypes was determined. For differences between groups we used the independent or paired samples t-test, Mann-Whitney test or Chi square/Fisher's exact test as appropriate. To investigate the influence of confounders on the presence or absence of AAA a multiple logistic regression-analysis was used.

Results

Adalimumab carries the G1m17 allotype. No anti-allotype antibodies against adalimumab were detected. Thirty-nine out of 249 patients had anti-idiotype antibodies against adalimumab (16%). IgG allotypes of RA patients were associated with the frequency of AAA: patients homozygous for G1m17 had the highest frequency of AAA (41%), patients homozygous for G1m3 the lowest frequency (10%), and heterozygous patients' AAA frequency was 14% (P = 0.0001).

Conclusions

An allotype mismatch between adalimumab and IgG in adalimumab-treated patients did not lead to a higher frequency of AAA. On the contrary, patients who carried the same IgG allotype as present on the adalimumab IgG molecule, had the highest frequency of anti-adalimumab antibodies compared to patients whose IgG allotype differed from adalimumab. This suggests that the allotype of adalimumab may not be highly immunogenic. Furthermore, patients carrying the G1m17-allotype might be more prone to antibody responses.     

Inhibition of L- and P-selectin by a rationally synthesized novel core 2-like branched structure containing GalNAc-Lewisx and Neu5Acalpha2- 3Galbeta1-3GalNAc sequences

The selectins interact in important normal and pathological situations with certain sialylated, fucosylated glycoconjugate ligands containing sialyl Lewisx(Neu5Acalpha2-3Galbeta1-4(Fucalpha1-3)GlcN Ac). Much effort has gone into the synthesis of sialylated and sulfated Lewisxanalogs as competitive ligands for the selectins. Since the natural selectin ligands GlyCAM-1 and PSGL-1 carry sialyl Lewisxas part of a branched Core 2 O-linked structure, we recently synthesized Galbeta1-4(Fucalpha1-3)GlcNAcbeta1-6(SE-3Galbeta1++ +-3)GalNAc1alphaOMe and found it to be a moderately superior ligand for L and P-selectin (Koenig et al. , Glycobiology 7, 79-93, 1997). Other studies have shown that sulfate esters can replace sialic acid in some selectin ligands (Yeun et al. , Biochemistry, 31, 9126-9131, 1992; Imai et al. , Nature, 361, 555, 1993). Based upon these observations, we hypothesized that Neu5Acalpha2-3Galbeta1-3GalNAc might have the capability of interacting with L- and P-selectin. To examine this hypothesis, we synthesized Galbeta1-4(Fucalpha1-3)GlcNAcbeta1-6(Neu5Acalpha2++ +-3Galbeta1-3)- GalNAc alpha1-OB, which was found to be 2- to 3-fold better than sialyl Lexfor P and L selectin, respectively. We also report the synthesis of an unusual structure GalNAcbeta1-4(Fucalpha1- 3)GlcNAcbeta1-OMe (GalNAc- Lewisx-O-methyl glycoside), which also proved to be a better inhibitor of L- and P-selectin than sialyl Lewisx-OMe. Combining this with our knowledge of Core 2 branched structures, we have synthesized a molecule that is 5- to 6-fold better at inhibiting L- and P-selectin than sialyl Lewisx-OMe, By contrast to unbranched structures, substitution of a sulfate ester group for a sialic acid residue in such a molecule resulted in a considerable loss of inhibition ability. Thus, the combination of a sialic acid residue on the primary (beta1-3) arm, and a modified Lexunit on the branched (beta1-6) arm on an O-linked Core 2 structure generated a monovalent synthetic oliogosaccharide inhibitor superior to SLexfor both L- and P-selectin.      

Novel splice variants associated with one of the zebrafish <Emphasis Type="Italic">dnmt3</Emphasis>genes

Background  

DNA methylation and the methyltransferases are known to be important in vertebrate development and this may be particularly true for the Dnmt3 family of enzymes because they are thought to be the de novo methyltransferases. Mammals have three Dnmt3 genes; Dnmt3a, Dnmt3b, and Dnmt3L, two of which encode active enzymes and one of which produces an inactive but necessary cofactor. However, due to multiple promoter use and alternative splicing there are actually a number of dnmt3 isoforms present. Six different dnmt3 genes have recently been identified in zebrafish.     

Growth of Variants of Myxococcus virescens

Some observations on variant strains of Myxococcus virescens B2 with special emphasis on characteristics associated with the ability to grow in dispersion are reported. The isolated strains were divided into two major classes according to their mode of growth in shaken and static liquid cultures based on casitone and casamino acids media. Strains growing in dispersion were designated D⁺-strains and those growing in aggregates or as films, D^?-strains. Colony morphology, cell morphology, growth in liquid and on solid medium and morphogenesis were compared. The ability to grow in dispersion shown by D⁺-strains seemed to be associated with a smooth colony on casitone agar, inability to form typical fruiting bodies and a low linear growth rate of colonies on solid medium as compared with the D^?-strains. In contrast D^?-strains produced rough colonies on casitone agar, were able to fruit and evidently formed an adhesive slime in the form of fibrils extending from the cell surface. It is suggested that the observed differences depend on different envelopes of the cells in the two classes.     

Examination of the Cytotoxic and Embryotoxic Potential and Underlying Mechanisms of Next-Generation Synthetic Trioxolane and Tetraoxane Antimalarials

Semisynthetic artemisinin-based therapies are the first-line treatment for P. falciparum malaria, but next-generation synthetic drug candidates are urgently required to improve availability and respond to the emergence of artemisinin-resistant parasites. Artemisinins are embryotoxic in animal models and induce apoptosis in sensitive mammalian cells. Understanding the cytotoxic propensities of antimalarial drug candidates is crucial to their successful development and utilization. Here, we demonstrate that, similarly to the model artemisinin artesunate (ARS), a synthetic tetraoxane drug candidate (RKA182) and a trioxolane equivalent (FBEG100) induce embryotoxicity and depletion of primitive erythroblasts in a rodent model. We also show that RKA182, FBEG100 and ARS are cytotoxic toward a panel of established and primary human cell lines, with caspase-dependent apoptosis and caspase-independent necrosis underlying the induction of cell death. Although the toxic effects of RKA182 and FBEG100 proceed more rapidly and are relatively less cell-selective than that of ARS, all three compounds are shown to be dependent upon heme, iron and oxidative stress for their ability to induce cell death. However, in contrast to previously studied artemisinins, the toxicity of RKA182 and FBEG100 is shown to be independent of general chemical decomposition. Although tetraoxanes and trioxolanes have shown promise as next-generation antimalarials, the data described here indicate that adverse effects associated with artemisinins, including embryotoxicity, cannot be ruled out with these novel compounds, and a full understanding of their toxicological actions will be central to the continuing design and development of safe and effective drug candidates which could prove important in the fight against malaria.     

The relationships of Heberdenia bahamensis and H. penduliflora (Myrsinaceae)

A morphological and anatomical investigation is presented of the two species of Heberdenia, H. bahamensis (Macaronesia) and H. penduliflora (Mexico). A cladistic analysis including 27 taxa of the Myrsinaceae and using Jacquinia (Theophrastaceae) and Manilkara (Sapotaceae) as outgroup was performed to provide a hypothesis of the relationships of the two species of Heberdenia. It was concluded that H. bahamensis and H. penduliflora are not more closely related to each other than either is to many other species of the Myrsinaceae, and that they should not be referred to the same genus. Heberdenia bahamensis appears to be most closely related to the Old World genera Pleiomeris, Embelia , and Grenacheria , whereas H. penduliflora is nested within Ardisia s. L , possibly being most closely related to the segregate genus Gentlea. It is suggested that H. penduliflora is probably best referred to a genus of its own.