Mystery Rays from Outer Space

Meddling with things mankind is not meant to understand. Also, pictures of my kids

May 28th, 2008

A little learning

Kopp et al 1995, proteasomeOne of the problems with genomics research is that the people who interpret it may not know much about the genes they identify. For example:

… single nucleotide polymorphisms (SNPs) in two genes critical for T-cell function are associated with susceptibility to MDD:1 PSMB4 (proteasome 4 subunit), important for antigen processing …2

Kopp et al 1995, HN3/PSMB4
PSMB4 location in proteasome3

OK, first of all, the proteasome is important in antigen processing, but antigen processing is probably about the least important of its myriad tasks. Suggesting that the function of the proteasome is antigen processing is like claiming the the function of my car is to hold coffee, because it happens to have cup-holders.4

Second, PSMB4 per se has never been implicated in antigen processing.5 Not only is it not one of the interferon-inducible trio of proteasome subunits that have been implicated in antigen processing, PSMB4 is not even catalytic.6

This looks like someone had a vague memory of seeing something in a textbook years ago, and didn’t try to look at the literature at all. Or who wanted to squeeze data into a preconceived theory.

  1. MDD: Major depressive disorder[]
  2. Wong ML, Dong C, Maestre-Mesa J, Licinio J.  Polymorphisms in inflammation-related genes are associated with susceptibility to major depression and antidepressant response. Mol Psychiatry. 2008 May 27. [Epub ahead of print] doi:10.1038/mp.2008.59 []
  3. Kopp F, Kristensen P, Hendil KB, Johnsen A, Sobek A, Dahlmann B. The human proteasome subunit HsN3 is located in the inner rings of the complex dimer. J Mol Biol. 1995 Apr 28;248:264-72 doi:10.1016/S0022-2836(95)80049-2 []
  4. Also, in my case, about 2000 little plastic dinosaurs, tucked into every cranny and hidden under the booster seats in the back.  I’m not sure what their function is either[]
  5. As far as I know; which is reasonably far[]
  6. Though it may be involved in proteolysis anyway.  But the point remains — not shown to have a role in antigen processing[]
May 28th, 2008

Alum, take 2: A better answer

Yu et al, Fig 6
Inflammasomes (From Yu et al, 2005)1

We’ve known for quite a while now how adjuvants work. Adjuvants (the components of vaccines that cause an immune response to start up, but that are not themselves the target of the immune response) trigger the parts of the innate immune system that normally identify microbial patterns, so that the immune system becomes aware that there’s a dangerous situation on its hands. (I have a much longer explanation here.)

It was a little annoying, though, that the most important adjuvant — alum, the adjuvant most commonly used for human vaccines — didn’t fit into this explanation. We didn’t know what innate triggers alum tickled to drive immune responses.

A month or two ago, I talked about a paper2 that claimed to answer that long-standing question. Their answer is that alum works through damaging tissue; damaged tissue releases a danger signal, uric acid, and according to Kool and the Gang2 that’s how alum drives immune responses. It’s an interesting suggestion, but I didn’t buy it:

I’m not entirely convinced that this is the whole, or even the main, story. … simple experience says that while vaccines sting, you don’t expect any kind of large-scale necrosis in your injected arm afterward – no more than you’d get from a modest bruise, which isn’t enough to trigger the kind of adjuvant effects we see with alum.

A paper that’s just become available in advance online status3 backs up my skepticism.

This is from Richard Flavell’s group at Yale. They determined that alum activity, at physiological doses, requires an intracellular receptor, Nalp3, that’s a member of the NLR family. NLRs (“Nod-like receptors”) are conceptual parallels to TLRs (“Toll-like receptors”); TLRs and NLRs are receptors for danger signals, and Nalp3 in particular is a receptor for uric acid, among other things. 4

Strictly speaking, uric acid itself does not act as a danger signal. Uric acid is a normal component of extracellular tissues, and if it was inflammatory we’d be in perpetual agony. When dying cells release their own stores of uric acid, though, the surrounding fluid becomes overloaded, and uric acid precipitates out as monosodium urate (MSU) crystals. It’s these MSU crystals that are actually inflammatory. Flavell’s insight was that MSU crystals might be conceptually siimilar to alum — another insoluble, particulate adjuvant — and so the two might act through the same pathway.

Sure enough, knockout mice without Nalp3 (or without other components of the Nalp3 reconition particle) failed to respond to alum (or to uric acid), whereas knockouts for a different NLR member did just fine. The knockouts responded normally when a different adjuvant was used.

Thus, by eliminating signalling through the Nalp3 inflammasome, we have eliminated one critical pathway used by alum to initiate humoral and cellular immunity. In doing so, aluminium hydroxide adjuvants ‘hijack’ an innate immune pathway that is exquisitely sensitive to cellular damage, perhaps as a result of the similarity to MSU in its physical structure.

Eisenbarth et al 2008 Fig 4The effect is pretty striking, actually (for example, the figure to the right).

The previous paper I talked about (Kool et al)2 also pinned alum into the uric acid pathway, but reached the conclusion that alum works through uric acid, rather than parallel to it, by causing tissue damage. Flavell’s group agree that this can happen, but disagree that it’s the normal mode of action:

In vitro, alum induced cell death at very high doses … in WT macrophages and in macrophages deficient in Nalp3 and Caspase-1 (Fig. 3b); however, the induction of IL-1beta by alum did not depend on the presence of MSU because the addition of uricase, which degrades MSU crystals and prevents the induction of IL-1beta (ref. 10), had no effect on IL-1beta production in response to LPS and alum (Fig. 3c).

I’m altogether happier with this explanation.

One interesting question that leaps to my mind — now we know the genes involved in alum recognition — is whether natural variants in these genes exist (I’m sure they do) and whether such variants correlate with vaccine responses in humans. Could we predict whether someone only needs a single dose of vaccine to be protected compared to her cousin who needs three doses? Are there people who lack components of this altogether — like the various TLR3 mutants I talked about earlier — and what happens to their vaccine response?

  1. Yu JW, Wu J, Zhang Z, Datta P, Ibrahimi I, Taniguchi S, Sagara J, Fernandes-Alnemri T, Alnemri ES (2006) Cryopyrin and pyrin activate caspase-1, but not NF-kappaB, via ASC oligomerization. Cell Death Differ 13:236–249. doi:10.1038/sj.cdd.4401734[]
  2. Kool, M., Soullie, T., van Nimwegen, M., Willart, M.A., Muskens, F., Jung, S., Hoogsteden, H.C., Hammad, H., Lambrecht, B.N. (2008). Alum adjuvant boosts adaptive immunity by inducing uric acid and activating inflammatory dendritic cells. Journal of Experimental Medicine DOI: 10.1084/jem.20071087[][][]
  3. Eisenbarth, S.C., Colegio, O.R., O’Connor, W., Sutterwala, F.S., Flavell, R.A. (2008). Crucial role for the Nalp3 inflammasome in the immunostimulatory properties of aluminium adjuvants. Nature DOI: 10.1038/nature06939 []
  4. Martinon F, Petrilli V, Mayor A, Tardivel A, Tschopp J (2006) Gout-associated uric acid crystals activate the NALP3 inflammasome. Nature 440:237-241. doi:10.1038/nature06939[]