Another look at how the adjuvant alum works

You’d think scientists would know all there is to know about the biological effects of a substance that has been used for decades to make vaccines. Well, think again. For one thing, they still haven’t figured out how exactly the humble adjuvant alum works—and this concoction of insoluble aluminum salts has been used for more than 80 years to boost the efficacy of childhood immunizations. And for most of that period, it has been the only adjuvant used in human vaccines. 

Researchers at Osaka University in Japan and the University of Liège in Belgium appeared to have shed some light on the mystery a couple of years ago, when they reported in the journal Nature Medicine that the adjuvant kills host cells, and that the DNA released by the dead cells is at least in part responsible for its salutary effects. Their experiments suggested that the DNA plays an important role in driving the migration of antigen presenting cells, such as dendritic cells (DCs), to lymph nodes, where they present antigens to T cells and so fuel the desired immune response (Nat. Med. 17, 996, 2011). 

Trouble was, however, that the researchers injected alum directly into the peritoneal cavity of mice to conduct many of their experiments. This is not how alum is typically delivered to people during immunization: Most human vaccines that contain alum are injected into muscle. 

In a recently published study, Amy McKee, Pippa Marrack, and their colleagues at National Jewish Health, a nonprofit hospital in Denver, looked at the effects of alum in mice after it was injected into the muscles. They found that, following such delivery, the DNA that was released from the host cells did not drive the migration of DCs to lymph nodes. Rather, it appeared to promote the presentation of antigen by DCs to CD4+ T cells in those lymph nodes (Proc. Natl. Acad. Sci. 2013, doi:10.1073/pnas.1300392110). 

McKee, Marrack and colleagues injected alum together with the egg-white protein ovalbumin (Ova) into mouse muscles. To check if host-cell-released DNA was important to boosting the subsequent immune response to the ovalbumin, they co-injected the DNA-degrading enzyme DNAse. Following DNAse co-injection, fewer Ova-presenting DCs showed a stable interaction with Ova-specific CD4+ T cells in the lymph nodes that drain the muscles at the injection site. Further, DCs isolated from mice that had been co-injected with DNAse presented fewer antigens to CD4+ T cells in culture dishes. 

This suggests that the DNA that was released due to alum-induced cell death somehow prompted DCs to present more Ova antigens to CD4+ T cells and interact longer with CD4+ T cells in lymph nodes. 

But where exactly does the released DNA come from? And how does it improve antigen presentation by the DCs? One possible model, Mckee says, is that once the alum-Ova complexes are injected into the muscle, they attract inflammatory cells such as neutrophils. The neutrophils then commit suicide and release their DNA, which binds to the alum particles. Neutrophils are one likely candidate because it’s already known that they sometimes commit suicide to physically trap pathogens such as bacteria with their DNA. Eosinophils and monocytes might also be involved, McKee says, because Michael Munks, another researcher in Marrack’s group, showed three years ago that in mice, alum injection into muscles attracts neutrophils, eosinophils and monocytes that then release their DNA (Blood 116, 5191, 2010). 

After the DNA binds to the alum, DCs then likely ingest the alum-DNA complexes and digest them in their lysosomes, the cellular equivalent of the stomach. The alum particles then probably disrupt the lysosome walls, allowing the neutrophil DNA that’s bound to the alum particles to leak into the cytoplasm of the DCs, where the leaked neutrophil DNA activates DNA receptors and signaling molecules. It is this DNA signal that could potentially enhance the presentation of antigen and the interactions of the DCs with the T cells in the lymph nodes, McKee says.

The challenge now, she adds, is to identify the DNA receptor in DCs and the full range of signaling molecules it activates.