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The lymphocyte function-associated antigen-1 (LFA-1) antagonist of 'Compound 4' was synthesised by a convenient route using cheap, commercially available starting materials and catalysts under mild reaction conditions and by easily handled reactions. The total yield in the preparation of 'Compound 4' was more than 38% via Sonogashira coupling of an iodide and an alkyne, reduction of the alkyne catalysed by Raney nickel and later steps involving hydrolysis of an ester, condensation of an acid and an amine and a final hydrolysis of an ester. [ABSTRACT FROM AUTHOR]

Aggregatibacter actinomycetemcomitans is an oral pathogen that produces the RTX toxin, leukotoxin (LtxA; Leukothera®). A. actinomycetemcomitans is strongly associated with the development of localized aggressive periodontitis. LtxA acts as a virulence factor for A. actinomycetemcomitans to subvert the host immune response by binding to the β2 integrin lymphocyte function-associated antigen-1 (LFA-1; CD11a/CD18) on white blood cells (WBCs), causing cell death. In this paper, we reviewed the state of knowledge on LtxA interaction with WBCs and the subsequent mechanisms of induced cell death. Finally, we touched on the potential therapeutic applications of LtxA (trade name Leukothera®) toxin therapy for the treatment of hematological malignancies and immune-mediated diseases.

The Gram-negative bacterium, Aggregatibacter actinomycetemcomitans, has been associated with localized aggressive periodontitis (LAP). In particular, highly leukotoxic strains of A. actinomycetemcomitans have been more closely associated with this disease, suggesting that LtxA is a key virulence factor for A. actinomycetemcomitans. LtxA is secreted across both the inner and outer membranes via the Type I secretion system, but has also been found to be enriched within outer membrane vesicles (OMVs), derived from the bacterial outer membrane. We have characterized the association of LtxA with OMVs produced by the highly leukotoxic strain, JP2, and investigated the interaction of these OMVs with host cells to understand how LtxA is delivered to host cells in this OMV-associated form. Our results demonstrated that a significant fraction of the secreted LtxA exists in an OMV-associated form. Furthermore, we have discovered that in this OMV-associated form, the toxin is trafficked to host cells by a cholesterol- and receptor-independent mechanism in contrast to the mechanism by which free LtxA is delivered. Because OMV-associated toxin is trafficked to host cells in an entirely different manner than free toxin, this study highlights the importance of studying both free and OMV-associated forms of LtxA to understand A. actinomycetemcomitans virulence.

Abstract: Lymphocytic-plasmacytic enteritis (LPE) is the most common form of inflammatory bowel disease (IBD) affecting the canine small intestine; however, the molecular basis of the pathogenesis remains unclear. It has recently been hypothesized that the primary defect is impaired innate immune function, as is the case for human IBD. Nuclear factor-kappa B (NFkappaB) plays a central role in innate immunity, and is a major transcriptional regulator of several proinflammatory cytokines, pattern recognition receptors (PRRs) such as toll-like receptors (TLRs), nucleotide-binding oligomerization domain-like receptors and cell adhesion molecules (CAMs). The purpose of this study was to evaluate, in the duodenal mucosa of 21 dogs with LPE and 8 control dogs, the degree of NFkappaB activity and the mRNA expression of two selected cytokines, nucleotide oligomerization domain two (NOD2) receptor and three selected CAMs, all of which are regulated by NFkappaB, using the electrophoretic mobility shift assay and real-time reverse transcription PCR. NFkappaB binding activity was significantly higher in the inflamed duodenal mucosa of the LPE dogs as compared to healthy controls. Furthermore, expression of mRNA for intercellular cell adhesion molecule 1 (ICAM-1) and mucosal addressin cell adhesion molecule 1 (MAdCAM-1) was significantly higher and vascular cell adhesion molecule 1 (VCAM-1) mRNA significantly lower in LPE dogs than in healthy controls. However, there was no significant difference in the mRNA levels for TNFα, IL1β and NOD2 between the two groups. These results suggest that NFkappaB and CAMs may play important roles in the pathogenesis of canine LPE. [Copyright &y& Elsevier]

Abstract: The innate immune system responds to inflammation, infection and injury by recruiting neutrophils and other leukocytes. These cells are able to leave the intravascular compartment in a process called leukocyte recruitment. This process involves several distinct steps: selectin-mediated rolling, firm adhesion via integrins, postarrest modifications including adhesion strengthening and leukocyte crawling and finally transmigration into tissue. Genetic defects affecting the different steps of the cascade can result in severe impairment in leukocyte recruitment. So far, three leukocyte adhesion deficiencies (LAD I-III) have been described in humans. These LADs are rare autosomal recessive inherited disorders and, although clinically distinct, exhibit several common features including recurrent bacterial infections and leukocytosis. In LAD-I, mutations within the β2-integrin gene result in a severe defect in β2 integrin-mediated firm leukocyte adhesion. Defects in the posttranslational fucosylation of selectin ligands dramatically reduce leukocyte rolling and lead to LAD-II. Finally, LAD-III, also known as LAD-I variant, is caused by impaired integrin activation due to mutations within the kindlin-3 gene. This review provides an overview on the molecular basis of leukocyte adhesion and its deficiencies. [Copyright &y& Elsevier]

Abstract: The S component (LukS-PV) is one of the two components of Panton-Valentine leukocidin (PVL), which is a pore-forming cytotoxin secreted by Staphylococcus aureus, with the ability to lyse leukocytes. In this study, LukS-PV had the ability to induce apoptosis in the human acute myeloid leukemia (AML) cell line THP-1. Therefore, we investigated the mechanisms of LukS-PV-induced apoptosis in THP-1 cells. THP-1 cells treated with LukS-PV, resulted in a significant inhibition of proliferation in a dose- and time-dependent manner, and induced G0/G1 arrest associated with an inhibition of cell cycle arrest regulatory protein (cyclin D1) in a dose- and time-dependent manner, as measured by flow cytometry (FCM). After 12h exposure to LukS-PV (1.00μM), annexin V-EGFP/propidium iodide (PI) FCM revealed that 19.5±3.6% of THP-1 cells were apoptotic, and terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) staining also revealed THP-1 cells were apoptotic. Chip analysis of 84 apoptosis-related genes demonstrated that 9 genes were up-regulated at least 2-fold and that 5 genes were down-regulated at least 2-fold in the treatment group when compared with levels in the control group. Western blotting reveled that the expression of caspase-8 increased significantly (approximately 4-fold). The levels of caspase-9, -3 and Bax increased significantly, and levels of Bcl-2 decreased rapidly with LukS-PV treatment. These data suggest that LukS-PV acts as an anti-leukemia agent and activates AML cell apoptosis via the mitochondrial pathway. Therefore, LukS-PV may be a multi-targeting drug candidate for the prevention and therapy of AML. [Copyright &y& Elsevier]

Six stilbenes were isolated from the methanol extract of Rheum undulatum rhizomes by bioactivity-guided fractionation. The structures of the compounds were determined by spectroscopic analysis (H-, C-NMR and MS), to be desoxyrhapontigenin ( 1), rhapontigenin ( 2), trans-resveratrol ( 3), piceatannol ( 4), piceatannol-3′-O-β-D-glucopyranoside ( 5) and isorhapontin ( 6). Compounds 1-4 inhibited the direct binding between sICAM-1 and LFA-1 of the THP-1 cells in a dose-dependent manner with IC values of 50.1, 25.4, 33.4 and 45.9 μM, respectively. On the other hand, the other compounds 5 and 6 with a glucose moiety in each molecule did not show any inhibitory activity in the cell adhesion assay (IC values of >100.0 μM). Compounds 2, 3 and 4 also had an inhibitory effect on direct binding between sVCAM-1 and VLA-4 of THP-1 cells. This suggests that the stilbenes from Rheum undulatum rhizomes are good candidates for therapeutic strategies towards inflammation. [ABSTRACT FROM AUTHOR]

Intercellular adhesion molecule (ICAM)-1988 is a small molecule lymphocyte function-associated antigen-1 (LFA-1) antagonist being considered for its anti-inflammatory properties. Following intravenous administration of ICAM1988, clearances in mice, rats, dogs, and monkeys were 17.8, 3.31, 15.4, and 6.85 ml min-1 kg-1, respectively. In mass balance studies using [14C]-ICAM1988 in rats dosed intravenously, unchanged ICAM1988 contributed to 25.1% of the dose. In rats, the systemic bioavailability of ICAM1988 was improved to 0.28 when the drug was administered orally as its isobutyl ester, ICAM2660. In rats, this was consistent with the complete in vitro conversion of ICAM2660 to ICAM1988 in plasma, and liver and intestinal S9. In dogs and monkeys, ICAM2660 did not improve the bioavailability of ICAM1988. This is consistent with limited in vitro conversion of ICAM2660 to ICAM1988 in plasma and liver S9. In human in vitro studies, ICAM2660 conversion to ICAM1988 in liver was similar to rats while no conversion in plasma and intestinal S9 fractions were observed. Based on the in vitro metabolism similarities of human and rat, it would be anticipated that in human oral administration of ICAM2660 would improve the systemic exposure of ICAM1988. [ABSTRACT FROM AUTHOR]

Non-infectious uveitis (NIU) is an inflammatory eye disease initiated via CD4+ T-cell activation and transmigration, resulting in focal retinal tissue damage and visual acuity disturbance. Cell adhesion molecules (CAMs) are activated during the inflammatory process to facilitate the leukocyte recruitment cascade. Our review focused on CAM-targeted therapies in experimental autoimmune uveitis (EAU) and NIU. We concluded that CAM-based therapies have demonstrated benefits for controlling EAU severity with decreases in immune cell migration, especially via ICAM-1/LFA-1 and VCAM-1/VLA-4 (integrin) pathways. P-selectin and E-selectin are more involved specifically in uveitis related to vasculitis. These therapies have potential clinical applications for the development of a more personalized and specific treatment. Localized therapies are the future direction to avoid serious systemic side effects. [ABSTRACT FROM AUTHOR]

Aggregatibacter actinomycetemcomitans is an oral pathogen that produces the RTX toxin, leukotoxin (LtxA; Leukothera®). A. actinomycetemcomitans is strongly associated with the development of localized aggressive periodontitis. LtxA acts as a virulence factor for A. actinomycetemcomitans to subvert the host immune response by binding to the β2 integrin lymphocyte function-associated antigen-1 (LFA-1; CD11a/CD18) on white blood cells (WBCs), causing cell death. In this paper, we reviewed the state of knowledge on LtxA interaction with WBCs and the subsequent mechanisms of induced cell death. Finally, we touched on the potential therapeutic applications of LtxA (trade name Leukothera®) toxin therapy for the treatment of hematological malignancies and immune-mediated diseases. [ABSTRACT FROM AUTHOR]

The Gram-negative bacterium, Aggregatibacter actinomycetemcomitans, has been associated with localized aggressive periodontitis (LAP). In particular, highly leukotoxic strains of A. actinomycetemcomitans have been more closely associated with this disease, suggesting that LtxA is a key virulence factor for A. actinomycetemcomitans. LtxA is secreted across both the inner and outer membranes via the Type I secretion system, but has also been found to be enriched within outer membrane vesicles (OMVs), derived from the bacterial outer membrane. We have characterized the association of LtxA with OMVs produced by the highly leukotoxic strain, JP2, and investigated the interaction of these OMVs with host cells to understand how LtxA is delivered to host cells in this OMV-associated form. Our results demonstrated that a significant fraction of the secreted LtxA exists in an OMV-associated form. Furthermore, we have discovered that in this OMV-associated form, the toxin is trafficked to host cells by a cholesterol- and receptor-independent mechanism in contrast to the mechanism by which free LtxA is delivered. Because OMV-associated toxin is trafficked to host cells in an entirely different manner than free toxin, this study highlights the importance of studying both free and OMV-associated forms of LtxA to understand A. actinomycetemcomitans virulence. [ABSTRACT FROM AUTHOR]