Potent human immunodeficiency virus-neutralizing and complement lysis activities of antibodies are not obligatorily linked

Abstract

To evaluate the contribution of complement-mediated lysis to the in vivo activities of neutralizing antibodies, we analyzed the influence of complement activation on treatment success in a recent passive immunization trial with the neutralizing monoclonal antibodies 2G12, 2F5, and 4E10. Administration of monoclonal antibodies led to an immediate, high activation of the complement system even in the absence of viremia in the 14 participating human immunodeficiency virus-infected individuals. Lysis activity measured in patient plasma increased during passive immunization; however, the increases were modest and only partially attributable to the administration of antibodies. We found that unlike neutralization activity, lysis activity was not associated with treatment success in this trial. Compared to complement lysis mounted by the polyclonal antibody response in vivo, monoclonal antibodies were weak inducers of this activity, suggesting that polyclonal responses are more effective in reaching the required threshold of complement activation. Importantly, strong neutralization activity of the monoclonal antibodies did not predict complement lysis activity against patient and reference viruses, suggesting that these activities are not linked. In summary, our data support the notion that the in vivo activities of 2G12, 2F5, and 4E10 are likely due to direct neutralization or Fc receptor-mediated mechanisms such as phagocytosis and antibody-dependent cellular cytotoxicity.

FIG. 1.

Complement is activated during passive immunization independent of viremia. Plasma levels of complement components were measured before and after the infusion of a mix of MAbs 2G12, 2F5, and 4E10. Levels of C3 before infusion (open circles) and after infusion (closed circles) are shown for patient NAB11 (A) together with the viral load (solid line) for weeks 1 to 11. Shaded areas indicate complement activation. Median levels of C3 (B), C3a (C), and TCC (D) before (open symbols) and after (closed symbols) infusion are shown for all 14 patients. Error bars show 95% confidence intervals.

FIG. 2.

Complement activation by 2G12, 2F5, and 4E10 did not trigger infection enhancement. Activation of complement by MAbs has neither a decreasing nor an enhancing effect on viral set point in noncontrolling patients. Dashed lines represent noncontrollers, and solid lines represent controllers. Means for controllers (closed squares) and noncontrollers (open squares) and 95% confidence intervals (error bars) from a multivariable linear regression model with a robust variance estimator are shown.

FIG. 3.

Passively administered antibodies 2G12, 2F5, and 4E10 can contribute to in vivo complement lysis, but activity is low. Autologous lysis activities in patients with acute (triangles) and chronic (circles) infection (A) as well as in controller (closed squares) and noncontroller patients (open squares) (B). Means and 95% confidence intervals (error bars) from a multivariable linear regression model with robust variance estimator are shown (see Table S3 in the supplemental material). (C) Correlation analysis of MAb concentration in vivo and complement lysis activity. Week 12 plasma levels of MAb 2G12 (dots), 2F5 (triangles), and 4E10 (diamonds) do not correlate with matched plasma complement lysis activity against autologous virus. (D) Neither total lysis activity at week 12 nor the absolute increase in lysis activity from weeks 0 to 12 differs between controller and noncontroller patients. Means and 95% confidence intervals (error bars) are shown. (E and F) MAb-mediated neutralization in patient plasma at week 12 and complement lysis activity from the same time point against autologous (E) and heterologous (F) virus is not correlated. MAb-mediated 70% neutralization (NT₇₀) was determined previously using a TZM-bl cell-based neutralization assay (62).

FIG. 4.

MAbs have weak lysis activity in vitro. (A) Lysis activities of neutralizing MAbs 2G12, 2F5, 4E10, and b12 were tested against the 14 patient virus isolates. Only 4E10, the mix of 2G12/2F5/4E10, and b12 show moderate lysis activity. Error bars show 95% confidence intervals (one-sample t test). (B) Lysis activities of 2G12 (circles), 2F5 (triangles), 4E10 (diamonds), and b12 (squares) against reference viruses. Error bars show standard deviations of triplicate measurements.

FIG. 5.

In vitro neutralization activity does not predict complement lysis activity. Lysis activities (shown as bars) of MAbs 2G12, 2F5, and 4E10 and the mix of these three antibodies against the 14 patient virus isolates do not show a common pattern. Neutralization activities (IC₉₀ values, inverse logarithmic scale) of MAbs 2G12 (circles), 2F5 (triangle), and 4E10 (diamonds) and the mix of the three MAbs (open squares) in a PBMC-based neutralization assay against each virus is depicted above the corresponding bars. The IC₉₀s of the individual MAbs and the equimolar mix of the MAbs were determined using a PBMC-based neutralization assay (61). Error bars show standard deviations of triplicate measurements.