The ability of human immunodeficiency virus (HIV-1) to persist and cause AIDS is dependent on its avoidance of antibody-mediated neutralization. The virus elicits abundant, envelope-directed antibodies that have little neutralization capacity. This lack of neutralization is paradoxical, given the functional conservation and exposure of receptor-binding sites on the gp120 envelope glycoprotein, which are larger than the typical antibody footprint and should therefore be accessible for antibody binding. Because gp120-receptor interactions involve conformational reorganization, we measured the entropies of binding for 20 gp120-reactive antibodies. Here we show that recognition by receptor-binding-site antibodies induces conformational change. Correlation with neutralization potency and analysis of receptor-antibody thermodynamic cycles suggested a receptor-binding-site 'conformational masking' mechanism of neutralization escape. To understand how such an escape mechanism would be compatible with virus-receptor interactions, we tested a soluble dodecameric receptor molecule and found that it neutralized primary HIV-1 isolates with great potency, showing that simultaneous binding of viral envelope glycoproteins by multiple receptors creates sufficient avidity to compensate for such masking. Because this solution is available for cell-surface receptors but not for most antibodies, conformational masking enables HIV-1 to maintain receptor binding and simultaneously to resist neutralization.

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SUMMARYComplement activation with formation of biologically potent mediators like C5a and the terminal C5b-9 complex (TCC) contributes essentially to development of inflammation and tissue damage in a number of autoimmune and inflammatory conditions. A particular role for complement in the ischaemia/ reperfusion injury of the heart, skeletal muscle, central nervous system, intestine and kidney has been suggested from animal studies. Previous experiments in C3 and C4 knockout mice suggested an important role of the classical or lectin pathway in initiation of complement activation during intestinal ischaemia/reperfusion injury while later use of factor D knockout mice showed the alternative pathway to be critically involved. We hypothesized that alternative pathway amplification might play a more critical role in classical pathway-induced C5 activation than previously recognized and used pathwayselective inhibitory mAbs to further elucidate the role of the alternative pathway. Here we demonstrate that selective blockade of the alternative pathway by neutralizing factor D in human serum diluted 1 : 2 with mAb 166-32 inhibited more than 80% of C5a and TCC formation induced by solid phase IgM and solid-and fluid-phase human aggregated IgG via the classical pathway. The findings emphasize the influence of alternative pathway amplification on the effect of initial classical pathway activation and the therapeutic potential of inhibiting the alternative pathway in clinical conditions with excessive and uncontrolled complement activation.

The terminal complement components play an important role in mediating tissue injury after ischemia and reperfusion (I/R) injury in rats and mice. However, the specific complement pathways involved in I/R injury are unknown. The role of the alternative pathway in I/R injury may be particularly important, as it amplifies complement activation and deposition. In this study, the role of the alternative pathway in I/R injury was evaluated using factor D-deficient (؊/؊) and heterozygote (؉/؊) mice. Gastrointestinal ischemia (GI) was induced by clamping the mesenteric artery for 20 minutes and then reperfused for 3 hours. Sham-operated control mice (؉/؊ versus ؊/؊) had similar baseline intestinal lactate dehydrogenase activity (P ‫؍‬ ns). Intestinal lactate dehydrogenase activity was greater in ؊/؊ mice compared to ؉/؊ mice after GI/R (P ‫؍‬ 0.02) thus demonstrating protection in the ؊/؊ mice. Intestinal myeloperoxidase activity in ؉/؊ mice was significantly greater than ؊/؊ mice after GI/R (P < 0.001). Pulmonary myeloperoxidase activity after GI/R was significantly higher in ؉/؊ than ؊/؊ mice (P ‫؍‬ 0.03). Addition of human factor D to ؊/؊ animals restored GI/R injury and was prevented by a functionally inhibitory antibody against human factor D. These data suggest that the alternative complement pathway plays an important role in local and remote tissue injury after GI/R. Inhibition of factor D may represent an effective therapeutic approach for GI/R injury.

We have analyzed a panel of eight murine monoclonal antibodies (MAbs) that depend on the V2 domain for binding to human immunodeficiency virus type 1 (HlV-1) gpl20. Each MAb is sensitive to amino acid changes within V2, and some are affected by substitutions elsewhere. With one exception, the MAbs were not reactive with peptides from the V2 region, or only poorly so. Hence their ability to bind recombinant strain IIIB gpl20 depended on the preservation of native structure. Three MAbs cross-reacted with strain RF gpl20, but only one cross-reacted with MN gpl20, and none bound SF-2 gp120. Four MAbs neutralized HIV-1 IIIB with various potencies, and the one able to bind MN gpl20 neutralized that virus. Peptide serology indicated that antibodies cross-reactive with the HxB2 Vi and V2 regions are rarely present in HIV-1-positive sera, but the relatively conserved segment between the Vi and V2 loops was recognized by antibodies in a significant fraction of sera. Antibodies able to block the binding of V2 MAbs to IIIB or MN gpl20 rarely exist in sera from HIV-1-infected humans; more common in these sera are antibodies that enhance the binding of V2 MAbs to gpl20. This enhancement effect of HIV-1-positive sera can be mimicked by several human MAbs to different discontinuous gpl20 epitopes. Soluble CD4 enhanced binding of one V2 MAb to oligomeric gpl20 but not to monomeric gpl20, perhaps by inducing conformational changes in the oligomer.

To define the domains in the envelope glycoprotein important for antibody neutralization of the human immunodeficiency virus type 1 (HIV-1), monoclonal antibodies (mAbs) were generated by immunizing mice with purified glycoprotein gpl20 of the MB isolate. One mAb, G3-4, reacted with the gpl20 of homologous ('fB) and heterologous (RF) isolates. In addition, mAb G3-4 efficiently neutralized both

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The complement system plays an important role in the initial defense against Neisseria meningitidis. In contrast, uncontrolled activation in meningococcal sepsis contributes to the development of tissue damage and shock. In a novel human whole blood model of meningococcal sepsis, we studied the effect of complement inhibition on inflammation and bacterial killing. Monoclonal antibodies (mAbs) blocking lectin and alternative pathways inhibited complement activation by N meningitidis and oxidative burst induced in granulocytes and monocytes. Oxidative burst was critically dependent on CD11b/CD18 (CR3) expression but not on Fc␥-receptors. Specific inhibition of C5a using mAb 137-26 binding the C5a moiety of C5 before cleavage prohibited CR3 up-regulation, phagocytosis, and oxidative burst but had no effect on C5b-9 (TCC) formation, lysis, and bacterial killing. An mAb-blocking cleavage of C5, preventing C5a and TCC formation, showed the same effect on CR3, phagocytosis, and oxidative burst as the anti-C5a mAb but additionally inhibited TCC formation, lysis, and bacterial killing, consistent with a C5b-9-dependent killing mechanism. In conclusion, the anti-C5a mAb 137-26 inhibits the potentially harmful effects of N meningitidis-induced C5a formation while preserving complementmediated bacterial killing. We suggest that this may be an attractive approach for the treatment of meningococcal sepsis. IntroductionThe gram-negative bacterium Neisseria meningitidis is an important human pathogen worldwide. It can cause meningitis or fulminant meningococcal sepsis (FMS); the latter is an overwhelming and often lethal condition that may lead to death within 24 hours. The pathogenic mechanism leading to FMS is a breakdown of homeostasis by a massive activation of diverse inflammatory systems such as the cytokine network and plasma cascades such as the complement, coagulation, fibrinolytic, and kinin/kallikreinin systems. 1,2 The importance of the complement system in meningococcal disease is emphasized by several observations. Complement deficiencies are defined risk factors for meningococcal infections, indicating that complement is crucial in the initial defense against this bacterium. 3-5 On the other hand, during FMS, disease severity, tissue damage, and outcome are closely related to the degree of complement activation. 6,7 Thus, with respect to the pathogenesis of meningococcal disease, the complement system has been aptly named a double-edged sword. 8 Complement is activated on the surfaces of meningococci by one or more of the 3 initial complement-activating pathwaysclassical, lectin, and alternative. After the activation of complement factor-3 (C3) by any of these 3 pathways, a C5 convertase (C4b2a3b through the classical and lectin pathways or C3bBbC3b through the alternative pathway) is formed, and the pivotal C5 molecule is cleaved into C5a and C5b. C5b is the initial molecule in the formation of the terminal C5b-9 complement complex (TCC). Membrane-associated TCC, also designated the C5b-9 membrane attack comple...