The marked decline in plasma HIV-1 level that occurs during acute infection strongly suggests that immune responses are capable of partially controlling HIV viremia. Since CTL activity is detectable early in infection, whereas neutralizing antibody is generally absent, immunological control of HIV-1 has been attributed primarily to the CTL response (6
). We have found, however, that in marked contrast to neutralizing activity, antibody directed against infected cells and capable of inhibiting HIV-1 in the presence of NK effector cells is detectable in the majority of patients when viremia is declining and as early as a few days after the onset of symptoms of acute infection. Furthermore, we found that the magnitude of this effector cell-mediated antiviral antibody response is inversely associated with plasma viremia level and that autologous and heterologous primary HIV strains are inhibited. These findings indicate that HIV-1-specific antibody could play an important role in the control of viremia during acute infection.
Our data strongly support ADCC as a mechanism responsible for the antiviral activity in plasma from acutely infected patients. ADCC occurs when antibody forms a bridge between target cells expressing antigens with which the antibody binds and effector cells bearing Fc receptors. We have shown that the antiviral activity is contained within the IgG fraction of antibody and requires NK effector cells (which express Fc receptors [32
]). Furthermore, the antiviral antibody binds to envelope glycoproteins, and activity requires intact antibody rather than Fab fragments. Finally, in cytotoxicity assays directly measuring ADCC, plasma from acutely infected patients, in the presence of effector cells, lysed target cells expressing HIV glycoproteins.
ADCC, like CTL activity, results in the death of infected cells (21
). It is therefore biologically plausible that ADCC plays a role in controlling viremia during HIV-1 infection. Baum et al. have shown that in patients with chronic infection, higher ADCC antibody titer is associated with less rapid progression of disease (4
). We have previously found that ADCC, measured by 51
Cr release assay using target cells transfected with HIV-1 env
and an autologous combination of patient serum and PBMC effector cells, correlates inversely with viral load in chronically infected patients not receiving antiretroviral therapy (14
). Importantly, the effector cells of ADCC—NK cells as well as macrophages—can be found in key sites of HIV replication, such as lymph nodes (M. Lu, N. Kouttab, N. Raja, D. L. Zheng, and G. Skowron, Abstr. 7th Conf. Retroviruses Opportunistic Infect., abstr. 368, 2000; 26). In SIV-infected rhesus macaques with rapidly progressive disease, the passive infusion of plasma from animals with more slowly progressive disease reduces plasma viremia in a manner most consistent with ADCC (5
). Furthermore, during acute SIV infection, a rapid increase in NK activity (measured in a 51
Cr release assay) and in the number of activated NK cells precedes the decline in viremia (15
); since NK cells are a major effector cell for ADCC (38
), it is possible that these activated NK cells are mediating ADCC. In a recent study describing synergism between antibody and immune T cells in protecting mice from herpes simplex virus type 2 genital infection, the authors suggested that T cells responding to the virus produce cytokines that activate NK cells, which in turn mediate ADCC in the presence of antibody (25
). Such a three-way interaction between innate, humoral, and T-cell immunity could also explain the strong correlation between HIV-1-specific CD4+
lymphocyte activity and control of viremia during HIV-1 infection (33
). Finally, the recent demonstration that infusion of an anti-CD8 monoclonal antibody results in a transient increase in plasma SIV viremia is also consistent with a role for ADCC in the control of lentivirus infection, since macaque NK cells generally express CD8 and would likely be depleted along with CTLs (8
). From our results together with those of Connick et al. showing a temporal relationship between antiviral antibodies and the fall in viremia during acute infection (11
), there is mounting evidence supporting an important role for ADCC in controlling viremia during HIV infection.
By definition, ADCC results in the lysis of target cells. Most of our experiments used reduction in viral yield, rather than cell lysis, as an indicator of the biological activity of antibody. It is likely that much of the antiviral activity that we measured in plasma from acutely infected patients was due to the death and removal of virus-producing cells. On the other hand, noncytolytic mechanisms, due to soluble substances secreted as a result of the interaction between target cells, antibody, and effector cells, also reduced viral yield. Some of the noncytolytic antiviral effect was neutralized by antibodies against MIP-1α and RANTES. Furthermore, HIV-specific antibody, in the presence of envelope-expressing target cells, augmented MIP-1α and RANTES release from NK cells. Thus, it is likely that these β-chemokines, released from NK cells via Fc receptor stimulation, were responsible for some or all of the soluble antiviral effect. Their role relative to that of cytotoxicity in inhibiting virus was not ascertained.
Presumably, the β-chemokines act by inhibiting the entry of newly released virus into uninfected cells (13
). β-Chemokine release from NK cells through cross-linking of Fc receptors has been previously documented (28
). However, in that study, Fc receptor cross-linking was accomplished by a specific antibody-antigen interaction directed against the receptor. Our study shows, for the first time, that a physiological interaction between an HIV-1-specific antibody, HIV-1 glycoprotein-expressing cells, and effector cells bearing Fc receptors results in the release of β-chemokines and that a consequent antiviral effect occurs. Thus, we define a new biological activity of antibody, related to ADCC through its component parts but acting through an entirely different mechanism.
In general, antibody alone had little effect on viral yield. Our experimental conditions allowed antibody added to HIV-infected cells to inhibit cell-to-cell spread of virus, either by neutralizing cell-free virus or by interacting with budding virions. In any case, the limited activity of antibody alone is consistent with other studies showing poor neutralizing antibody activity during acute infection (18
). Nonetheless, a recent study reported consistent neutralizing activity early in infection when macrophages, rather than lymphocytes, were used as target cells; however, many of the sera tested were obtained several months after infection and likely well after the steepest declines in viremia (34
). We did not find a strong correlation between the antiviral effect of antibody alone and that of antibody combined with effector cells. Thus, it is unclear whether antibodies that mediate an antiviral effect toward infected cells in the presence of effector cells are the same as those that neutralize cell-free virus. Addressing this question will require further studies focused on determining the antigenic specificity and the antibody-antigen affinity of the early antiviral response.
Plasma antiviral activities were generally similar when measured against a reference strain, other heterologous strains, or autologous strains. Although we did not determine the degree of genetic diversity in the isolates used, these results suggest that the antiviral response during acute infection, when measured in the presence of effector cells, is broadly reactive. Again, this differs from the neutralizing antibody response, which tends to be strain specific, particularly early in infection (23
). If further studies verify the breadth and importance of the effector cell-mediated antiviral response in controlling viremia in vivo, the antigenic determinants of this response may prove to be key components of a protective or therapeutic HIV vaccine.
An unexpected finding of our investigation was the lack of antiviral activity of NK cells in the absence of HIV-specific antibody. NK activity is thought to require a positive signal generated through a specific ligand-receptor interaction or the absence of inhibitory signals mediated by major-histocompatibility complex molecules on target cells and inhibitory receptors on NK cells (3
). HIV infection down regulates the expression of some major histocompatibility complex class I molecules, which should render infected cells targets for NK activity (37
). On the other hand, HLA C and HLA E molecules may not be down regulated and thus remain available to interact with inhibitory receptors (9
). It should be noted that we did not activate NK cells before using them in virus inhibition assays. However, the use of NK effector cells and CD4+
lymphocyte target cells from different donors likely resulted in some activation over the 7-day assay period. Furthermore, it is possible that NK cells could have been effective in reducing viral yield in cells infected for a shorter time. In any case, our results lead us to question any significant role for NK cells—in the absence of Fc receptor cross-linking by antibody—in controlling viremia. If the activation of NK cells, which occurs just prior to the decline in viremia during acute SIV infection (15
), is important in controlling viremia, it may be due to the capacity of NK cells to act as effector cells for an early antiviral antibody response.
In summary, antibody capable of inhibiting autologous and heterologous primary strains of HIV-1, in the presence of NK effector cells, is present early in acute HIV infection, and the magnitude of this antibody response correlates inversely with plasma HIV-1 viremia level. HIV-1-specific antibody may thus be an important contributor to the early control of HIV-1 viremia, and antigens that elicit an effector cell-mediated, antiviral antibody response may be important components of an HIV vaccine.