Attenuation of inflammation and cytokine production in rat colitis by a novel selective inhibitor of leukotriene A4 hydrolase.

BACKGROUND AND PURPOSE: Leukotriene B(4) (LTB(4)), formed by the sequential actions of the 5-lipoxygenase (5-LO) and leukotriene A(4) hydrolase (LTA(4)H), is a pro-inflammatory mediator implicated in the pathogenesis of inflammatory bowel disease. However, inhibitors of 5-LO have not proved to be consistent in their therapeutic efficacy in colitis. Another approach to inhibiting LTB(4) synthesis is through the use of inhibitors of LTA(4)H, such as the novel, potent and selective compound, JNJ 26993135. EXPERIMENTAL APPROACH: The effect of oral administration of JNJ 26993135 has been evaluated in a rat model of colitis provoked by colonic instillation of trinitrobenzenesulphonic acid (TNBS). The extent and severity of the macroscopic inflammatory response, the colonic levels of myeloperoxidase (MPO) and LTB(4) and of the pro-inflammatory cytokines, tumour necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) were measured. KEY RESULTS: Oral administration of JNJ 26993135 (5, 15 and 30 mg kg(-1), twice a day) dose-dependently reduced both the extent and intensity of the colonic inflammatory damage observed 3 days after TNBS challenge. JNJ 26993135 also dose-dependently reduced the elevated colonic levels of LTB(4), as well as the inflammatory biomarkers, MPO, IL-6 and TNF-alpha. This dosing regimen was supported by the pharmacokinetic profile of JNJ 26993135, along with the demonstration of the inhibition of ex vivo production of LTB(4) in whole blood following oral administration. CONCLUSIONS AND IMPLICATIONS: These results with JNJ 26993135 in the rat TNBS model support the role of LTB(4) in colitis and the potential value of targeting LTA(4)H for the treatment of inflammatory bowel diseases.

Non-specific antiviral activity of antisense molecules targeted to the E1 region of human papillomavirus.

Antisense phosphorothioate oligonucleotides (ODN1 0x5 OMe) directed against the E1 start region of human papillomavirus 11 (HPV11) can inhibit papillomavirus induced growth of implanted human foreskin in a mouse xenograft model. Administration of a mismatch control oligonucleotide (ODN9 0x5 OMe), in which guanine was replaced with adenine in the same model, had no effect on papilloma induced growth. However, the apparent antiviral activity of ODN1 0x5 OMe was also shown in a lethal mouse cytomegalovirus (CMV) model, in which the oligonucleotides are not expected to have antisense activity. To understand the mechanisms of action of these oligonucleotides, a mismatch oligonucleotide (ODN61 0x5 OMe) was prepared which retained the CpG motifs of ODN1 0x5 OMe. This was tested in the mouse xenograft model and shown to have moderate inhibitory activity. As a definitive experiment, a comparison was made between the efficacy of the active oligonucleotide ODN1 0x5 OMe against two papilloma viruses HPV11 and HPV40. Both these viruses cause benign genital warts, but differ by four bases in their E1 sequence that was the target for ODN1 0x5 OMe. Papillomavirus induced growth in the mouse xenograft model was inhibited by ODN1 0x5 OMe in both cases, suggesting that oligonucleotide molecules have a non-specific antiviral activity that is not directly related to their antisense sequence.

Inhibition of virus-encoded thymidine kinase suppresses herpes simplex virus replication in vitro and in vivo.

Both herpes simplex virus type 1 (HSV-1) and HSV-2 encode a thymidine kinase enzyme which differs from cellular thymidine kinase in substrate specificity. Viral thymidine kinase enables the virus to replicate in cells that lack cellular thymidine kinase, namely those of the sensory neurons where the virus establishes, and periodically reactivates from, a latent state. Thymidine kinase-dependent HSV replication following viral reactivation at the site of latency is thought to precede the emergence of virus at mucosal surfaces. The ability to inhibit such an essential viral enzyme would potentially prevent HSV from replicating within neuronal tissue, and thus stop the recurrent disease cycle. Ro 32-2313 was designed as a selective and competitive inhibitor of HSV thymidine kinase and in vitro studies have confirmed this mechanism of action. In vivo evaluation of a soluble prodrug of Ro 32-2313, Ro 32-4397, was undertaken in murine models where pathogenesis was dependent upon viral replication in neuronal tissue. It was shown that in vivo administration of Ro 32-4397 (i) significantly reduced the viral titre detected in isolated dorsal root ganglia; (ii) prevented HSV-2-induced lethality in a systemic infection model; and (iii) reduced zosteriform lesion development in a model of dermal infection. Administration of Ro 32-4397 produced dose-related changes in viral pathogenicity towards those of the phenotype of a thymidine kinase-deficient virus. Overall, the study confirmed that thymidine kinase inhibitors can suppress the replication of HSV in vivo, and suggest that such inhibitors may reduce reactivation of the virus from latency if used prophylactically in recurrent HSV infection.

Detection of toxoplasma-specific immunoglobulin-G: assessment of a slide agglutination test.

The performance of a slide agglutination test for detection of toxoplasma-specific IgG was assessed and compared with that of the dye test and latex agglutination test. The slide agglutination test was as sensitive as, and more specific than, latex agglutination. The predictive value of a negative slide agglutination test was less than the latex agglutination test but produced results within minutes, although quantitative results were not comparable to other assays. Slide agglutination presents a rapid alternative to the latex agglutination test as a screening assay for toxoplasmosis, although patients at risk of life-threatening infection require detailed serological examination using additional methods.

Measurement of bone density by gas displacement.

Small bone volumes, porous and irregular structured objects with volumes 0.05-2 cm3, were measured by differential pressure of displaced air. The differential air pressure was measured by a piezoelectric resistor pressure gauge. The gas-displacement system was used to measure the volumes of 234 bone specimens for calculation of bone density. The operation of the system, calibration, sensitivity, and errors of measurement are described.

Rapid up-regulation of CXC chemokines in the airways after Ag-specific CD4+ T cell activation.

Ag-specific activation of CD4(+) T cells is known to be causative for the cytokine production associated with lung allergy. Chemokine-induced leukocyte recruitment potentially represents a critical early event in Ag-induced lung inflammation. Whether Ag-specific, lung CD4(+) T cell activation is important in lung chemokine production is currently not clear. Using alphabeta-TCR transgenic BALB/c DO11.10 mice, we investigated the ability of Ag-specific CD4(+) T cell activation to induce lung chemokine production and leukocyte recruitment. Within 1 h of exposure of DO11. 10 mice to OVA aerosol, lung mRNA and protein for the neutrophil chemokines KC and macrophage inflammatory protein (MIP)-2 were greatly increased. Accordingly, neutrophils in the airways increased by >50-fold, and KC and MIP-2 proved to be functional because their neutralization significantly reduced airway neutrophilia. CD4(+) T cell activation was critical because CD4(+) but not CD8(+) T cell depletion reduced KC production, which correlated well with the previously observed inhibition of neutrophil influx after CD4(+) T cell depletion. In vitro studies confirmed that OVA-induced KC and MIP-2 production was conditional upon the interaction of CD4(+) T cells with APCs. A likely secondary mediator was TNF-alpha, and a probable source of these chemokines in the lung was alveolar macrophages. Thus, Ag-specific CD4(+) T cell activation in the lung leads to rapid up-regulation of neutrophil chemokines and the recruitment of neutrophils to the site of Ag exposure. This may be a key early event in the pathogenesis of Ag-induced lung inflammation.