Many studies have shown that rituximab exerts its effects on B-cell lymphoma through host effector mechanisms, particularly antibodymediated cellular effects in the form of either antibody-dependent, cell-mediated cytotoxicity or antibody-dependent cellular phagocytosis. Although we know that these mechanisms are dependent upon interaction with Fc receptors on the surface of effector cells, we have little information about the exact process that takes place in vivo. Now, Montalvao and colleagues at the Institut Pasteur in Paris report that liver macrophages (i.e., Kupffer cells, KCs) are the key effectors of antibody-mediated B-cell destruction.
Initial studies of B-cell depletion kinetics in different organs following anti-CD20 treatment in mice showed that this process took place most rapidly in the liver and, furthermore, that partial hepatectomy (but not splenectomy) reduced the rate and extent of B-cell depletion. Pre-treatment of B cells with pertussis toxin, which prevents their sequestration in lymph nodes and the splenic white pulp, resulted in even more rapid depletion, supporting the concept that liver-mediated B-cell destruction is the dominant mechanism of action of anti-CD20 therapy and that depletion from other organs is not the result of direct intra-organ cytotoxicity but rather a consequence of redistribution of antibody-targeted B cells to the liver. This interpretation was further supported by the results of experiments that used intravital imaging of B cells labeled with a fluorochrome. Those studies showed rapid accumulation of anti-CD20-treated B cells in the liver sinusoids, followed by local destruction, and this effect was blocked by liposomal clodronate, which is toxic to KCs.
A series of elegant experiments were performed using transgenic mice expressing a fluorescent reporter, which selectively highlights KCs and monocytes/macrophages, allowing direct visualization of phagocytic cells during intravital imaging of the liver. This experimental approach confirmed that normal B cells were rapidly trapped and engulfed by KCs in the liver sinusoids following anti-CD20 treatment, an observation that was replicated when malignant B cells were tested in the same system.
In Brief
We have known for some time that Fc receptor-dependent effects are central to the action of rituximab and other type 1 anti-CD20 antibodies such as ofatumumab, but it has not previously been clear which compartment mediates this action. There was an assumption that lymphoreticular sequestration by macrophages mediated this process, but the studies of the Paris group show that KCs are the primarily agents of antibody-mediated B-cell destruction, at least in the mouse. It also appears that anti-CD20 treated B cells resident at other sites, such as the spleen and lymph nodes, traffic to the liver for destruction rather than being eliminated in situ. These observations have implications for the use of antibodies in the clinic and may provide an explanation for the preferential effect of anti-CD20 therapy on circulating B cells as opposed to those in lymphoid or other organs, where the process of depletion is less rapid. These findings also suggest that type 1 anti-CD20 antibodies may be optimized by targeting Fc receptor engagement on KCs, whereas the type 2 antibodies such as obinutuzumab appear to act through a different mechanism, at least in part by inducing B-cell apoptosis.
For the future, this type of in vivo imaging experiment provides a powerful tool for dissecting the mechanisms of action of antibodies and other complex biotherapeutics, although evidence that the situation in humans is similar to that in the mouse is needed. These experiments have given a strong indication that we should seek this evidence in the liver.
Competing Interests
Dr. Johnson indicated no relevant conflicts of interest.
