HIV antibodies. Antigen modification regulates competition of broad and narrow neutralizing HIV antibodies

Abstract

Some HIV-infected individuals develop broadly neutralizing antibodies (bNAbs), whereas most develop antibodies that neutralize only a narrow range of viruses (nNAbs). bNAbs, but not nNAbs, protect animals from experimental infection and are likely a key component of an effective vaccine. nNAbs and bNAbs target the same regions of the viral envelope glycoprotein (Env), but for reasons that remain unclear only nNAbs are elicited by Env immunization. We show that in contrast to germline-reverted (gl) bNAbs, glnNAbs recognized diverse recombinant Envs. Moreover, owing to binding affinity differences, nNAb B cell progenitors had an advantage in becoming activated and internalizing Env compared with bNAb B cell progenitors. We then identified an Env modification strategy that minimized the activation of nNAb B cells targeting epitopes that overlap those of bNAbs.

Fig. 1. Activation by and internalization of Env by glnNAb and glVRC01-class B cells

(A) Calcium flux in B cells expressing the glBCRs of CD4-BS specific nNAbs (1-154, 1-676, 1-695, 1-732, 4-341), CD4-BS specific bNAbs (NIH45-46 and VRC01), or V3-specific nNAbs (1-79, 2-59, 2-1261) challenged with the indicated Env proteins at a 1 μM final concentration. Black arrow indicates time of Env addition. (B) Quantitation of Env internalization in the above B cell lines. Bars represent the mean of two independent experiments (31). Clade A, B, and C rEnvs are colored shades of red, blue and green respectively in (A) and (B).

Fig. 2. Activation by and internalization of Env by glnNAb and glVRC01-class B cells in response to modified Env immunogens

(A) glNIH45-46 or glVRC01 B cells were mixed with the indicated glnNAb B cells. The pooled B cells were then challenged with WT 426c (top row), 426c.NLGS.TM (middle row), or 426c.NLGS.TM.ΔV1-3 (bottom row) at a 1 μM final concentration. Calcium flux was monitored concurrently in both B cell lines with the strategy depicted in fig. S6. Black arrow indicates time of Env addition. (B) Area under the calcium flux curve determined from duplicate experiments in (A). Error bars represent SD from the mean of 10 (glVRC01-class) or 4 replicates (other B cell lines). Green asterisks and blue stars over the bars indicate significant differences (P < 0.05) from glNIH45-46 and glVRC01 B cells, respectively, for each protein using a two-tailed Student’s t test. (C) Quantification of internalization of the indicated Envs by glVRC01 class and glnNAb B cells. Error bars represent SD from the mean of two independent experiments each performed in duplicate (n = 4). Green astrices and blue stars represent significant differences (P < 0.05) from glNIH45-46 and glVRC01, respectively, for each protein using a two-tailed Student’s t test.

Fig. 3. B cell activation and antigen uptake of Env by glVRC01-class B cells in the presence of soluble anti-HIVantibodies

(A) Calcium flux was monitored in glNIH45-46 (solid bars) or glVRC01 (patterned bars) B cells incubated with 426c.NLGS.TM (white bars) or with 426c.NLGS.TM.ΔV1-3 (gray bars) in the presence of equimolar concentrations of the indicated antibodies. Bars represent mean and SD of the area under the calcium flux response curve relative to that of Env in the absence of soluble mAb from three independent experiments. Asterisks represent a significant difference (P < 0.05) in the calcium flux response between 426c.NLGS.TM and 426c.NLGS.TM.ΔV1-3 using a two-tailed Student’s t test. Env internalization by glNIH45-46 (B) or glVRC01 (C) B cells incubated with 426c.NLGS.TM (white bars) or 426c.NLGS.TM.ΔV1-3 (gray bars) in the absence or presence of equimolar concentrations of the indicated nNAbs. Bars represent the mean of two independent experiments. Asterisk denotes significant difference (P < 0.05) in fluorescence compared to cells incubated with Env alone using a two-tailed Student’s t test in (B) and (C). Dotted line corresponds to mean 426c.NLGS.TM internalization by B cells in the absence of Ab in (B) and (C).