Mice were immunized with 106
PFU equivalents of formalin-inactivated RSV/A2 (FI-A2) in the superficial gluteal muscle and then rested a minimum of 4 weeks prior to RSV challenge (14
). Several studies have established that pronounced weight loss, pulmonary eosinophilia, and type 2 cytokine dominance are associated with enhanced FI-RSV vaccine disease in mice (19
). We examined these and other pulmonary inflammatory markers following prophylactic MAb treatment in FI-RSV-vaccinated mice challenged with RSV (Fig. ). No infectious virus or viral RNA was detected in the lungs of mice prophylactically treated with either MAb 131-2G or normal Ig (data not shown). However, treatment with MAb 131-2G was associated with a marked decrease in weight loss (Fig. ) and infiltration by pulmonary cells (Fig. ), including eosinophils, i.e., <11% eosinophilia compared to normal Ig control-treated mice (23 to 42% eosinophilia) (data not shown). FI-A2- or FI-Vero-cell-immunized, mock-challenged mice had a minimal pulmonary inflammatory response that was unaffected by prophylactic administration of MAb 131-2G (data not shown). Prophylactic administration of MAb 131-2G F(ab′)2
was associated with a similar decrease in total BAL fluid cell infiltration (Fig. A) and pulmonary eosinophilia (Fig. ) compared to normal Ig (nIg) treatment. Overall, both intact MAb 131-2G- and MAb 131-2G F(ab′)2
-treated mice had a dramatic reduction in all cell types and at all time points examined compared to nIg-treated mice (Table ). These findings are consistent with previous studies showing that RSV G protein modifies trafficking to the lung of many cells, including CD11b+
, PMN, B220+
, and DX5+
NK cells (33
). Interestingly, at days 3 and 5 p.i. MAb 131-2G F(ab′)2
was associated with a greater percent reduction for most cell types than was intact MAb 131-2G (Table ). The similarity in results with the F(ab′)2
and intact forms of MAb 131-2G demonstrates that the MAb effect is not associated with decreased replication of RSV. MAb 131-2G F(ab′)2
does not increase viral clearance (data not shown). The levels of IL-4 in the cell-free BAL fluid were diminished as early as day 3 postchallenge and significantly reduced by days 5 and 7 postchallenge in mice treated with intact and F(ab′)2
MAb 131-2G (Fig. ).
FIG. 2. Effect of MAb 131-2G prophylaxis on illness in FI-RSV-vaccinated mice. (A) Mean BAL cell numbers (±standard errors) in the lungs of FI-RSV-vaccinated mice treated with either nIg or MAb 131-2G. The data are representative of three independent (more ...)
FIG. 3. Effect of MAb 131-2G F(ab′)2 prophylaxis on illness in FI-RSV-vaccinated mice. (A) Mean BAL fluid cell numbers (±standard errors) in the lungs of nIg-, MAb 131-2G (anti-G MAb)-, or 131-2G F(ab′)2 fragment [anti-G F(ab′) (more ...)
Pulmonary leukocyte infiltration following MAb 131-2G or 131-2G F(ab′)2 antibody treatment 1 day prior to RSV infection in FI-RSV-immunized micea
This study shows that prophylactic treatment with an anti-RSV G protein monoclonal antibody that blocks RSV G binding to CX3CR1 can both decrease the pulmonary inflammatory response to RSV infection and increase viral clearance. These findings are similar to those seen when this antibody was given as treatment after RSV infection of mice (13
). The ability of prophylactic administration of MAb 131-2G to downregulate the pulmonary inflammatory response is broad and involves many cell types, including those associated with inflammation and the innate and adaptive immune responses.
Although prophylactic administration with anti-G MAb (131-2G) resulted in significantly diminished viral load in the lung, the MAb did not totally block viral replication, as indicated by detection of viral RNA by real-time PCR. Studies that have analyzed RSV infection of epithelial cells and macrophages found that addition of anti-F MAbs inhibited productive infection but did not block all viral gene expression (1
), suggesting that intracellular infection is at least in part refractory to these antibodies.
Numerous studies have implicated the G protein in pathogenesis of RSV disease (2
), including several that show that RSV G protein CX3C-CX3CR1 interaction is important for the enhanced-disease phenotype-associated RSV challenge of FI-RSV-immunized mice (33
). In one study, FI-RSV-immunized mice challenged with viruses lacking a functional CX3C motif or pretreated with anti-CX3CR1 antibodies did not develop enhanced disease (14
). Our data are consistent with and support these earlier observations. In the present study, we show that MAb 131-2G given prophylactically as an intact antibody or F(ab′)2
fragments markedly downregulated the Th2-type inflammatory response to RSV challenge in FI-RSV-immunized mice as exemplified by the marked decrease in pulmonary eosinophilia, a hallmark of FI-RSV vaccine-enhanced disease (10
). The marked reduction in weight loss, an indicator of severity of illness in the mouse, is another clear indication of the ability of this MAb to alter the course of enhanced disease after RSV challenge in FI-RSV-immunized mice.
Taken together, the results suggest that MAb 131-2G given as a prophylactic (Fig. ) or for treatment of infection in naïve or FI-RSV-immunized mice can substantially impact the virus-induced inflammatory response and likely decrease RSV disease (13
). The decrease in pulmonary inflammation involves many cell types and is independent of the Fc portion of the antibody and viral clearance (23
). We hypothesize that vaccination to achieve an anti-G protein antibody response with activities similar to that of MAb 131-2G might have a similar impact on the pulmonary inflammation and virus clearance and improve both the safety and efficacy of an RSV vaccine. Our recently reported study showing that vaccination with a polypeptide encompassing the central conserved region of the G protein (the binding site for MAb 131-2G) prevented body weight loss and lung histopathology and decreased lung virus titers after challenge with RSV strain A2 is consistent with this hypothesis (41
). Moreover, serum antibodies from these vaccinated mice demonstrated G protein CX3C-CX3CR1 blocking activities in vitro
). Thus, it is possible that a vaccine that includes an appropriately constructed component that induces antibodies that block G protein binding to CX3CR1 might eliminate the risk of enhanced disease that is otherwise present for a subunit or inactivated RSV vaccine in RSV-naïve infants.