Mystery Rays from Outer Space

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

April 30th, 2009

Swine flu and Mexico

In an interview with Ruben Donis (chief, molecular virology and vaccines branch, CDC) ScienceInsider got a couple of answers that touch on points that I (and lots of others) have noted:

Q: Have you been able to compare isolates from Mexico and the United States?
R.D.: Yes, they are very, very similar. Many genes are identical. In the eight or nine viruses we’ve sequenced, there is nothing different.

Q: Have you compared someone who died with someone who had a mild case?
R.D.: Those data are still slippery. We don’t have good case data. You get age and sex—very limited information. That’s a problem. In the set of samples we know one case was fatal, but we don’t know which one it is.

So, nothing concrete; the circulating Mexican flu probably isn’t different than the sequences we’ve already seen; and reading between the lines at least one fatality wasn’t associated with an unusual viral sequence — though we don’t know how early this case was.

April 30th, 2009

Swine flu, virulence, and jumping viruses

SARS S protein evolution - Zhang et al, 2006
SARS S protein evolution (Zhang et al., 2006)

One of the biggest outstanding questions about the new H1N1 Swine Flu epidemic is the mortality rate.  Most of the cases outside of Mexico have been quite mild, whereas there have been a significant number of deaths linked to this flu infection in Mexico.  A superficial look at the limited data we have suggests that the virus might be similar to routine human flu (0.1% mortality rate) outside Mexico but more like the 1918 flu (2.5% mortality rate) in Mexico (but note that I don’t believe this is the case!) Why the difference?

Vincent at the Virology Blog highlights a number of possible explanations that boil down to three general categories:

  • Different environment – Are people in Mexico City exposed to different opportunistic bacteria? To more pollution? Slower or worse health care?  My opinion — possible, but not really consistent with our knowledge about influenza virus.
  • Different virus – Is the virus circulating in Mexico the same as we’ve seen elsewhere?  My opinion: Although we haven’t yet seen genome sequences from Mexican virus, this is unlikely, because we have seen sequences from patients who presumably caught the virus in Mexico, and these look like all the others.
  • Unknown denominator.  This is my preferred explanation. As I put this in a comment in a previous post, “I still think the most likely reason for the apparent difference in Mexico and the US is the missing denominator, rather than any host or environmental factor. That is, I suspect that the virus is much more widespread in Mexico City than the authorities know. Because they almost entirely tested severely ill hospitalized patients, it’s not surprising that they found high mortality rates; if they were also testing mild cases (as is happening in the US and elsewhere) I think they would likely find a very widespread infection with a low mortality rate.”

However, I want to throw out one more possibility — not because I think it’s the most likely explanation, but because I think it’s the most interesting explanation, at least to me (and it’s my blog, so I get to choose).  (And I’m sure I’m not the first to consider this, by the way — the CDC guys are presumably already thinking about this possibility.)   That is that the virus we are seeing now is not the same virus that caused the mortality in Mexico.  We are now seeing the human-adapted virus, whereas (remember, still speaking hypothetically) the virus that killed the patients was still swine-adapted.  As the virus adapted to humans it became less virulent and better at human-to-human transmission; we are therefore seeing the adapted, lower-virulence virus, and the high-mortality virus has already gone extinct.   To test this, we need not only current samples from Mexico, we need samples from the earliest patients they had.

There are precedents for the pieces of this, though I don’t know offhand of an example where both halves have been seen.  As an example of rapid evolution as a virus shifts species, there’s SARS virus; it jumped from civets to humans; we saw the genomes of the virus evolving at a furious rate as it adapted to humans. (The figure at the top here shows evolution of the SARS S protein1 – click for a larger version.)  Certainly we know that viruses in general change their virulence (in both directions — increased and reduced virulence) as they adapt to a new population.

Some related posts I’ve made:

… and really, a lot more — this has been one of the major themes of my blog.


  1. Zhang, C., Wei, J., & He, S. (2006). Adaptive evolution of the spike gene of SARS coronavirus: changes in positively selected sites in different epidemic groups BMC Microbiology, 6 (1) DOI: 10.1186/1471-2180-6-88[]
April 29th, 2009

More Swine Flu genome

Swine flu PB2 gene phylo tree
Swine flu PB2 gene phylogenetic tree

This post will be updated through the day.
Update 1 (near the bottom): I don’t think the Ohio/07 virus is particularly closely related to the current swine flu
Update 2 (at the bottom): A helpful comment from The Virology Blog about the ancestry of the virus

As I noted yesterday, the CDC has released a number of the Swine Flu genome sequences; they’re available here.  (There are  more sequences released yesterday.)  In my quick and primitive glance at them yesterday, I didn’t see anything very remarkable about the sequences — they looked like more or less straightforward swine flu things.

Sandra, at Discovering Biology in a Digital World, has run some more sophisticated analyses on the sequences and suggests that these viruses may be “the same strain that caused an outbreak in 2007 at an Ohio country fair.”  She is refering to the virus A/SW/OH/511445/2007, which is described here:

Vincent, A., Swenson, S., Lager, K., Gauger, P., Loiacono, C., & Zhang, Y. (2009). Characterization of an influenza A virus isolated from pigs during an outbreak of respiratory disease in swine and people during a county fair in the United States Veterinary Microbiology DOI: 10.1016/j.vetmic.2009.01.003

In August 2007, pigs and people became clinically affected by an influenza-like illness during attendance at an Ohio county fair.  … Approximately 26 people in close association with the fair pigs were affected by an influenza-like illness. Viruses from at least two individuals were isolated, sequenced and analyzed at the Centers for Disease Control and determined to be nearly identical to the swine virus studied here (A. Klimov, personal communication).

The accession numbers for the Ohio virus are EU604689-EU604696.

Please see Sandra’s page for her explanation of what she did.  However, I am not yet convinced.  I ran my own analysis, selecting sequences in what I think is a diffferent, and what I think is a more appropriate, way, and I did not see the present swine flu clustering with the Ohio strain.  (See the snippet of a figure at the top left here; click for a larger version.)  I am far from an expert in phylogenetic analysis or on influenza virus, so I’m not saying I’m right by any means — but I would like to see an explanation of why my approach is wrong.

Basically — as far as I can tell; Sandra hasn’t posted her complete methods as I write this, though that will come later today — the difference between our conclusions may be the way we collected sequences for comparison.  She “used H1N1 (and a couple of H1N2) protein sequences from Swine and Humans between Jan 1 2006 and today.”  I didn’t try to restrict the sequences in any way — rather, I ran a BLAST search on the non-redundant nucleotide collection in GenBank and selected the top 100 (or 1000) most similar sequences for comparison.  To me, it makes sense that we should not artificially restrict our sample, but rather should look at all the closest matches we can find.

In my PB2 tree, the nearest neighbours are another California isolate of the new strain, then the closest cluster includes A/duck/NC/91347/01(H1N2), A/mallard duck/South Dakota/Sg-00125/2007(H3N2), A/pintail duck/South Dakota/Sg-00126/2007(H3N2), and A/swine/Korea/CAS05/2004(H3N2).  (The avian PB2 is expected, because current swine flu strains have acquired an avian PB2.)

One way to look more closely at this might be to look at some of the gene sequences more specifically.  Vincent et al. point to some unusual features of the Ohio virus’s HA  gene.  Though it’s not likely to be unique it may help sort out relationships here:

Unique changes at antigenic determinant sites were identified in the OH07 HA at positions 71 and 162 and may play a role in the loss of cross-reactivity. The NA gene was shown to be related to the swine N1 phylogenetic cluster (Fig. 1B). The internal genes (Fig. 2A–F) were shown to be of the triple reassortant SIV lineage and group with those of the cluster IV H3N2 viruses reported by Olsen, et al. (Olsen et al., 2006). The PB2 gene was determined to contain the conserved avian amino acid residue glutamic acid at position 627, reported to be important in avian and human host specifcity (Subbarao et al., 1993).

Right now I need to get my kids ready for school, but I’ll look at those genes later today.

Update 1. OK, I’ve looked at the sites in the Ohio virus that Vincent et al flagged as being unsual.  The current swine flu viruses do not match all those unusual sites.  They do match some but so do many (dozens of) other swine flus.  I am not really seeing evidence that the A/SW/OH/511445/2007 swine flu isolate is “the same strain that caused an outbreak in 2007 at an Ohio country fair.”

Vincent et al (their Table 3) flagged the following sites as unusual; here are comparisons.  I ran several from the current outbreak — also a couple of HAs from swine flus that a simple BLAST search identified as highly related to the current outbreak (much more similar than is the A/SW/OH/511445/2007).  The figure at bottom shows it graphically (current outbreak in blue — A/SW/OH/511445/2007 in green — unusual residues boxed in red) and the table breaks it down.

71 73 74 142 156 162
A/California/04/2009_H1N1 S A S K N S
A/California/07/2009(H1N1) S A S K N S
A/Texas/05/2009(H1N1) S A S K N S
A/Swine/Ohio/891/01(H1N2) F A S K N S
A/SW/OH/511445/2007 S A S N N N
A/swine/Guangxi/17/2005(H1N2) F A S K N S

Swine flu HA aligned

Update 2. Vincent at The Virology Blog says:

According to ProMED-mail, the NA and MP genes are related to those of influenza viruses from Asian-European swine, and the other genes appear to originate from swine flu viruses from pigs in North America. The data are in accord with the original assertion of the CDC that all genes of the new isolate were derived from swine viruses.

April 28th, 2009

Swine flu genome sequences

Note: I will be updating this post through the day, as I play with the sequences in my spare time.  (Right now I have to get the kids up and get them ready for school.)  Don’t expect to learn anything particularly new from this, since I’m no influenza expert; but since I’m going to be doing this for my own curiosity I may as well save others the time.
Update: Also see my Wednesday post for more info.

For those playing along at home, some of the swine flu genomes are now available.  Only one of them has a complete genome, and these are the California and Texas cases — now we really want to compare them to the Mexico strain to see if the mortality is associated with strain variation or not.

I haven’t looked at the sequences much yet.  I did run some quick alignments of the HA gene; nothing unexpected.  These are all the same strain, but not identical (as you expect from influenza).  It’s significantly different from the HA of the H1N1 strain that was circulating this year (A/Brisbane/59/2007) (I think that was the circulating strain, it’s close anyway).   If you’re interested you can check:
Alignment compared to A/Brisbane/59/2007
Alignment of the 7 swine flu HA proteins
Alignment of the 6 complete swine flu HA proteins

The closest matches to the HA in GenBank are from A/swine/Minnesota/1192/2001(H1N2) and several other swine flu strains, which are ~ 94% identical.  I haven’t compared the other genomic fragments yet to see what other strains got into the mix.

Update 1. I’ve run the segments of the most complete new virus (A/California/04/2009(H1N1)) to see what viruses this looks most like.  The only remarkable thing I see is how unremarkable it is.  It’s pretty much a generic swine flu top to bottom, with close matches only to other strains of swine flu.  Since those other strains didn’t jump into people, that’s just another indicator of how much we have to learn about molecular determinants of virulence in viruses.

Quick results, showing only the first few matches (there are many!):

PB1 matches
Accession Strain Pct identity
AF342823.1 A/Wisconsin/10/98 (H1N1) 96%
AF250130.1 A/Swine/Indiana/9K035/99 (H1N2) 96%
CY033790.1 A/mallard duck/South Dakota/Sg-00125/2007(H3N2) 96%
PB2 matches
Accession Strain Pct identity
EU301177.2 A/swine/Korea/JNS06/2004(H3N2) 96%
AF455734.1 A/Swine/Minnesota/55551/00 (H1N2) 96%
CY033794.1 A/mallard duck/South Dakota/Sg-00128/2007 96%
PA matches
Accession Strain Pct identity
AF455722.1 A/Swine/Illinois/100084/01 (H1N2) 96%
AF251433.1 A/Swine/Minnesota/593/99 (H3N2) 96%
F251425.1 A/Swine/Iowa/569/99 (H3N2) 96%
HA matches
Accession Strain Pct identity
AF455680.1 A/Swine/Indiana/P12439/00 (H1N2) 95%
AF250124.1 A/Swine/Indiana/9K035/99 (H1N2) 95%
AY038014.1 A/Turkey/MO/24093/99(H1N2) 95%
NP matches
Accession Strain Pct identity
AF251415.2 A/Swine/Iowa/533/99 (H3N2) 96%
EU798854.1 A/swine/Korea/CY05/2007(H3N2) 96%
EU798853.1 A/swine/Korea/CY04/2007(H3N2) 96%
NA matches
Accession Strain Pct identity
AF250366.2 A/Swine/England/195852/92 (H1N1) 94%
CY038009.1 A/swine/England/WVL7/1992(H1N1) 94%
Y038001.1 A/swine/Spain/WVL6/1991(H1N1) 94%
M matches
Accession Strain Pct identity
AY363575.1 A/swine/Hong Kong/5212/99(H3N2) 97%
AY363574.1 A/swine/Hong Kong/5200/99(H3N2) 97%
AY363573.1 A/swine/Hong Kong/5190/99(H3N2) 97%
NS1/NS2  matches
Accession Strain Pct identity
AF153262.1 A/Swine/Minnesota/9088-2/98 (H3N2) 96%
AF153261.1 A/Swine/Texas/4199-2/98 (H3N2) 96%
AF153263.1 A/Swine/Iowa/8548-1/98 96%
April 27th, 2009

On immunology and malaria

Malaria life cycle
Life cycle of Plasmodium falciparum

“In this article we have attempted to make the case that we may not know enough about malaria to make an effective vaccine. If we agree that the development of a malaria vaccine would profit from a better understanding of the basic immunology of the human response to malaria, we then need to ask the following question: have we engaged a sufficient number of immunologists to address the problem? At the moment, probably not. Relative to the magnitude of the global disease burden imposed by malaria, there are only a small number of scientists with the training and expertise in the human immune system who are committed to working at the molecular interface of the parasite and the immune system.”

Pierce, S., & Miller, L. (2009). World Malaria Day 2009: What Malaria Knows about the Immune System That Immunologists Still Do Not The Journal of Immunology, 182 (9), 5171-5177 DOI: 10.4049/jimmunol.0804153

April 22nd, 2009

On HIV and molecular judo

Figure 3, Jern et al
Simulation of hA3G-mediated HIV-1 evolution.

One of the host defenses against HIV1 is “APOBEC3G” and related proteins.  These proteins force HIV to hypermutate, killing its ability to replicate in the next cell it infects.  On the other hand, it looks as if low-level mutation by APOBEC3G over the decades has driven HIV evolution:

We have found that hA3G activity acting on prior generations of virus has left detectable footprints in the HIV-1 genome. 2

HIV can only infect human cells because it has a defense against APOBECs: the viral protein “vif” causes APOBECs to be destroyed, and the virus is able to replicate without being hypermutated. 

So let’s say we develop an antiviral drug that blocks vif.  APOBECs would drive hypermutation of the virus.  This hypermutation would include the gene encoding vif.  Would this mutation, in a kind of molecular judo, drive rapid evolution of vif, so that it becomes resistant to the drug?

According to some experiments 2  both in cells and in silico, perhaps not:

However, since the predicted effect on resistance to standard antiviral drugs is likely to be small, we propose that concerns over increased resistance mutations should not impede development of HIV-1 Vif as a candidate drug target. 2

I’m not quite convinced this paper was actually modeling the phenomenon they say they’re modeling — is vif that’s shut off by deliberate mutations the same as vif that’s blocked by a (hypothetical) drug? But it’s a useful start, anyway.


  1. and other lentiviruses; as well as quite a few other virus types[]
  2. Patric Jern, Rebecca A. Russell, Vinay K. Pathak, & John M. Coffin (2009). Likely Role of APOBEC3G-Mediated G-to-A Mutations in HIV-1 Evolution and Drug Resistance PLoS Pathogens, 5 (4) DOI: 10.1371/journal.ppat.1000367[][][]
April 21st, 2009

1918 flu, dinosaurs, and birds

Update 4-26-09: I see this page is getting a lot of hits from Google in the wake of the Swine Flu outbreak in in California and Mexico. Please note that I am not suggesting this new Swine Flu is likely to be even remotely as dangerous as the 1918 flu! This post was written before the outbreak and was mainly supposed to be pointing out a curiosity.

If you want to learn more about the present outbreak I highly recommend the Effect Measure blog.

My own blog posts on influenza are mainly focused on the underlying molecular biology and immunology, rather than the epidemiology that’s of special interest now.

You may also be interested in the Google Flu Tracker; but note that it’s probably not accurately reflecting case number at the moment.


Influenza signThis turns out to be an old hypothesis that I had never before run across, not a brand-new insight; but it’s still a holy crap moment for me.

In 1918, an immensely virulent strain of influenza virus swept across the globe, killing tens if not hundreds of millions of people.   (See here for a little more.)  It came from nowhere, and it disappeared as fast as it came, leaving no descendants; we haven’t seen the 1918 influenza since 1918. 1

Or so I thought.

Turns out the 1918 flu never left.  Its great-great-grandchildren are still out there – mutated, more or less harmless to man, but still recognizable.  In the same way as the dinosaurs still roam the Earth, chirping and clucking, the 1918 influenza virus became swine flu.

Early serological studies linked … the swine H1N1 1930 virus isolate to the 1918 pandemic virus. Laidlaw suggested that the swine influenza virus could be the 1918 pandemic influenza virus which became established in pigs. … Interestingly, the 1930 swine influenza virus may still be circulating in swine. 2

This was proposed — I had no idea — as early as 19353, and several lines of observation have subsequently supported the concept.  The most recent paper,2 from which I learned all this fascinating stuff, directly demonstrated that  “the human 1918 influenza virus can infect and replicate in pigs and cause clinical disease and lesions in the infected animals,” though the disease in swine is fairly mild.  Their reconstruction of the story:

… one could speculate that the initial interspecies transmission of influenza virus during the 1918 pandemic occurred from people to pigs and only later appeared to occasionally transmit back to people, … likely contributing at least regionally to the maintenance and spread of the disease. The virus spread throughout the swine population, adapted to the swine host, and subsequently resulted in the current lineage of the classical H1N1 swine influenza viruses. 2

(Apparently genetic evidence suggests that the circulating H1N1 strains of human influenza are also descendants of the 1918 flu, which I also did not know — clearly I really need to learn a lot more about influenza — making this finding even less surprising to those in the know.  But still.  Holy crap.)


  1. Actually, since early 1919, I guess, but you get my point.[]
  2. Weingartl, H., Albrecht, R., Lager, K., Babiuk, S., Marszal, P., Neufeld, J., Embury-Hyatt, C., Lekcharoensuk, P., Tumpey, T., Garcia-Sastre, A., & Richt, J. (2009). Experimental Infection of Pigs with the Human 1918 Pandemic Influenza Virus Journal of Virology, 83 (9), 4287-4296 DOI: 10.1128/JVI.02399-08[][][]
  3. Laidlaw, P. P. 1935. Epidemic influenza: a virus disease. Lancet 1:1118-1124[]
April 16th, 2009

Is HIV becoming more virulent?

Worldwide HIV/AIDs Epidemic Statistics
Worldwide HIV/AIDs Epidemic Statistics

Viruses (and other pathogens, of course) can become more or less virulent over time.  This is generally a side effect; the virus couldn’t care less whether it kills its host or not, but it does care deeply about how well it is transmitted to new hosts, because if that’s not efficient then the virus will die out.  Sometimes viruses can be better transmitted by reducing virulence — for example, if the host survives longer, it may contact more new victims.  Sometimes viruses can be better transmitted by increasing virulence — if it’s transmitted by the fecal/oral route, then causing more diarrhea and vomitting may improve transmission.

As viruses enter new populations, such as when the virus jumps from one species to another, they may cast around looking for optimal strategies; their virulence and transmission may work well in one population but not in another.  One such virus is HIV, which entered the human population something like a hundred years ago but didn’t really explode until much later.  There’s been a lot of interest — for obvious reasons — as to whether HIV is becoming more or less virulent, or whether it hasn’t changed at all, since the HIV epidemic started about 30 years ago. It’s hard to come up with a solid theoretical prediction either way, partly because HIV transmission isn’t necessarily linked to virulence in an obvious way.

Unfortunately the answers to the question have been all over the place.  I summarized a dozen or more studies, last time a paper on this came out, by saying that “Experimental evidence has pointed in all directions — some suggests that HIV is becoming less virulent, some that it is becoming more virulent, and some says it’s staying the same.”  The study I was talking about 1 concluded that HIV was not changing in virulence:

Thus, the results of this study do not support the hypothesis that there has been any important change in the virulence of HIV-1 over this time period in this cohort. 1

HIV budding from a lymphocyte
HIV budding from a lymphocyte

A new study2 reaches a different conclusion.  They found that people infected with HIV recently seem to be progressing more rapidly toward AIDS, because they have lower CD4 T cell counts on first presentation. 3

The decrease in the post-seroconversion CD4 cell counts occurred early in the epidemic, with stabilization since the advent of HAART. These data may provide an important clinical correlate to studies suggesting that HIV may have adapted to the host, resulting in a more virulent infection. 2

There’s an excellent accompanying editorial4  that outlines the background of the question, quickly summarizes the theoretical problems in making predictions, and offers a possible different interpretation:

The increased circulation of more-aggressive HIV subtypes could also explain the results of studies that show increased virulence over time. In this case, however, the apparent increased virulence would not be the consequence of a selection process but only an effect of a greater representation of more-virulent subtypes … 4


  1. Herbeck, J.T., Gottlieb, G.S., Li, X., Hu, Z., Detels, R., Phair, J., Rinaldo, C., Jacobson, L.P., Margolick, J.B., Mullins, J.I., Tripathy, S. (2008). Lack of Evidence for Changing Virulence of HIV-1 in North America. PLoS ONE, 3(2), e1525. DOI: 10.1371/journal.pone.0001525[][]
  2. Crum-Cianflone, N., Eberly, L., Zhang, Y., Ganesan, A., Weintrob, A., Marconi, V., Barthel, R., Fraser, S., Agan, B., & Wegner, S. (2009). Is HIV Becoming More Virulent? Initial CD4 Cell Counts among HIV Seroconverters during the Course of the HIV Epidemic: 1985–2007 Clinical Infectious Diseases, 48 (9), 1285-1292 DOI: 10.1086/597777[][]
  3. This is a highly simplified summary! They did do a lot more controls and tests.[]
  4. Has Human Immunodeficiency Virus Become More Virulent? Maria Dorrucci and Andrew Phillips. Clinical Infectious Diseases 2009;48:1293-1295 DOI: 10.1086/597778[][]
April 15th, 2009

Malaria vaccination – a victim of its own (feeble) success

Malaria parasites in mosquito midgut
Malaria parasites in mosquito midgut

A followup study 1 of the most successful malaria vaccine to date is not very impressive:

In cohort 2, adjusted efficacy of the RTS,S/AS02A candidate malaria vaccine against first or only clinical malaria episodes in Mozambican children aged 1 to 4 years was of 35.4% during the first six months of follow up (ATP cohort), decreasing to 9.0% in the subsequent 12 months. 1

(My emphasis)  In  the earlier clinical trial, this vaccine had been moderately effective, 2 something like 35-65% efficacy — pretty feeble, but better than other malaria vaccines.  And in one subset of children, the protection seemed to last reasonably well, remaining roughly constant for 21 months. 3   But in this second group of vaccinated children, the protection dropped like a rock.  Why?

The most likely difference is that children in the second (poor long-term protection) group were treated for malaria.  That means the first group was constantly re-exposed to malaria parasites, which acted as a vaccine booster.  The second group didn’t get that booster effect, and their immunity dropped.  (The first group was also in a higher-risk area, again meaning they were more likely to be re-exposed to the parasite.)

Children in cohort 1 were probably exposed to low-density parasitaemias for a longer time than children in cohort 2, in which the development of this enhanced asexual-stage immune response may have been impaired. We propose this may explain a waning of the vaccine-specific protective response in cohort 2. 1

Does this mean the vaccine is useless?  Not at all.  In high-risk areas this can clearly reduce disease.  But equally clearly, this isn’t likely to be helpful for eradicating malaria, because the more successful this vaccine is at reducing malaria, the less effective it will become.


  1. Guinovart, C., Aponte, J., Sacarlal, J., Aide, P., Leach, A., Bassat, Q., Macete, E., Dobaño, C., Lievens, M., Loucq, C., Ballou, W., Cohen, J., & Alonso, P. (2009). Insights into Long-Lasting Protection Induced by RTS,S/AS02A Malaria Vaccine: Further Results from a Phase IIb Trial in Mozambican Children PLoS ONE, 4 (4) DOI: 10.1371/journal.pone.0005165[][][]
  2. Abdulla, S., Oberholzer, R., Juma, O., Kubhoja, S., Machera, F., Membi, C., Omari, S., Urassa, A., Mshinda, H., Jumanne, A., Salim, N., Shomari, M., Aebi, T., Schellenberg, D. M., Carter, T., Villafana, T., Demoitie, M. A., Dubois, M. C., Leach, A., Lievens, M., Vekemans, J., Cohen, J., Ballou, W. R., and Tanner, M. (2008). Safety and immunogenicity of RTS,S/AS02D malaria vaccine in infants. N. Engl. J. Med. 359, 2533-2544. doi:10.1056/NEJMoa0807773

    Bejon, P., Lusingu, J., Olotu, A., Leach, A., Lievens, M., Vekemans, J., Mshamu, S., Lang, T., Gould, J., Dubois, M. C., Demoitie, M. A., Stallaert, J. F., Vansadia, P., Carter, T., Njuguna, P., Awuondo, K. O., Malabeja, A., Abdul, O., Gesase, S., Mturi, N., Drakeley, C. J., Savarese, B., Villafana, T., Ballou, W. R., Cohen, J., Riley, E. M., Lemnge, M. M., Marsh, K., and von Seidlein, L. (2008). Efficacy of RTS,S/AS01E vaccine against malaria in children 5 to 17 months of age. N. Engl. J. Med. 359, 2521-2532. doi:10.1056/NEJMoa0807381[]

  3. Alonso PL, Sacarlal J, Aponte JJ, Leach A, Macete E, et al. (2005) Duration of protection with RTS,S/AS02A malaria vaccine in prevention of Plasmodium falciparum disease in Mozambican children: single-blind extended follow-up of a randomised controlled trial. Lancet 366: 2012–2018[]
April 14th, 2009

Tumor immunity: The Goldilocks approach

GoldilocksWe know that the immune system can destroy tumors. We also know, unfortunately, that by the time we see a tumor, immunity probably won’t destroy the tumor. There are lots of reasons for that. One is that tumors are essentially part of the normal body, so it’s normal for the immune system to ignore them. It looks as if you need to have immunity that’s just right to get rid of a tumor.

Tumors arise from normal self cells,1 that the immune response has been programmed to ignore. Now, the process of becoming a tumor is not normal, and so tumors are not entirely normal self any more — meaning that there are likely to be some targets in most if not all tumors. But in all but the most reckless tumors the differences between abnormal and normal are relatively small, compared to, say, a virus-infected cell that contains many potential targets.

There’s actually a long list of known tumor antigens; the T-cell tumor peptide database lists many hundreds of them. But most are not truly specific for the tumor.  The’re actually normal self antigens; they’re derived from proteins that are overexpressed in tumors, or that are differentiation antigens or “cancer-germline” antigens that are normally also found in self tissues. What’s more, these normal self antigens are the most interesting tumor antigens, as far as clinical utility is concerned. Mutations can make brand-new, non-self targets for the immune system, but they’re going to be sporadic targets, often unique to individual tumors — not something you can prepare for. The normal antigens, though, are likely to be predictable, common targets; it’s conceivable that tumor vaccines can be prepared in advance.

Melanoma cell (Eva-Maria Schnäker, University of Münster)
Human melanoma cell

If these antigens were common (which they are, in some tumor types — like melanoma), and they were good targets for the immune system, then we wouldn’t see much cancer. We do see melanomas quite often, and part of the reason may be that the immune system generally responds quite weakly to these antigens.  Why is that? And, more to the point, how can we make the immune system respond more strongly? A recent paper in the Journal of Experimental Medicine2 offers answers for both of these questions.

From work in the past couple of years, we now have decent estimates of how many T cells there are that can react with any particular target. (See here and here for my discussion of the earlier papers.) A reasonably strong immune response to a non-self epitope might originate from maybe 100 or so precursor T cells. There’s a rather wide range of frequency for these precursor cells, say from 20 to 1000; and to some extent, the fewer T cells there are the weaker (the less immunodominant) the immune response.

We expect T cells against normal self targets to be less common, because they should be eliminated as they mature in the thymus. Some may survive, though, and we would count on these survivors to attack the normal (albeit overexpressed, or abnormally present) target in the cancer cells. But just how rare are they?

Rizzuto et al say they’re really rare (this was in mice, by the way); at least ten times less abundant than T cells against non-self antigens.  If you look at the range I gave for “normal” precursors, that could mean there are fewer than 5 or 10 precursors.  If the average is “fewer than five”, then quite possibly some mice have only two, or one, or no precursors.  You can’t have much of a response with no precursors.

So there’s a weak anti-tumor response because there aren’t many T cells in the body that can respond to the normal self targets in the tumor. That’s not really a surprise, but it does raise the question, What if there were more of the T cells? To ask that question, Rizzuto et al. tried transferring more of these precursor T cells into tumor-bearing mice — starting at around the normal level for a precursor to non-self antigen, and going up from there — and then vaccinating with the appropriate target.

The effects were pretty dramatic. With no supplemental T cells (that is, with the natural, very low, level of T cell precursors) the mice all died of the tumor quickly. At the middle of the range, almost all of the mice rejected the tumor. And at the highest levels of transfers? The mice all died again. Having enough T cells to respond was protective, but putting in too many made them useless.

These results identify vaccine-specific CD8+ precursor frequency as a remarkably significant predictor of treatment and side-effect outcome. Paradoxically, above a certain threshold there is an inverse relationship between pmel-1 clonal frequency and vaccine-induced tumor rejection.2

Melanoma cell
Mouse melanoma cell

(My emphasis) This paradoxical effect is probably because the numerous T cells started to compete with each other so that none of them were properly activated; they only saw effective-looking polyfunctional T cells at the lower transfer levels.

In other words, if you’re going to transfer T cells to try to eliminate a tumor, more is not necessarily better. Quality and quantity are both important factors, and quantity helps determine quality.

One question I have is how this relates to tumor immune evasion. Many tumor types  acquire mutations, as they develop, that block presentation of antigen to T cells. Are these mutations perhaps only partially effective — giving the tumors sufficient protection against the tiny handful of natural precursors they “expect” to deal with, but not against a larger attack after, say, vaccination — or are they more complete, and protective even if the optimal number of T cells are transfered? I’d guess that it would depend on the tumor, but it looks as if it might be a relevant question and it would be nice to have more than a guess.

Our results show that combining lymphodepletion with physiologically relevant numbers of naive tumor-specific CD8+ cells and in vivo administration of an effective vaccine generates a high-quality, antitumor response in mice. This approach requires strikingly low numbers of naive tumor-specific cells, making it a new and truly potent treatment strategy.   2


  1. I’m ignoring here crazy things like the contagious tumors of Tasmanian Devils and dogs[]
  2. Rizzuto, G., Merghoub, T., Hirschhorn-Cymerman, D., Liu, C., Lesokhin, A., Sahawneh, D., Zhong, H., Panageas, K., Perales, M., Altan-Bonnet, G., Wolchok, J., & Houghton, A. (2009). Self-antigen-specific CD8+ T cell precursor frequency determines the quality of the antitumor immune response Journal of Experimental Medicine, 206 (4), 849-866 DOI: 10.1084/jem.20081382[][][]