Mystery Rays from Outer Space

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

January 30th, 2008

Herpesvirus immune evasion: An emerging theme?

Herpes Simplex (false color) - Pasteur InstituteCytotoxic T lymphocytes (CTL) are important destroyers of virus, recognizing MHC class I on infected cells and killing the cell before virus replication is complete. Unsurprisingly, many viruses (especially, but not only, the large and complex viruses like herpesviruses and adenoviruses) have evolved mechanisms to block MHC class I antigen processing and, therefore, T cell recognition.

What has been surprising is how ineffective these viral immune evasion mechanisms seem to be. (I’ve talked about this before in a bit more detail.) In cells in the incubator (which is where most of the experiments with the antigen presentation immune evasion molecules have been done), the effects often look pretty dramatic; but when you look at the immune response in humans infected with human cytomegalovirus1 or herpes simplex virus2 (both of which have formidable immune evasion functions in tissue culture3) there are pretty hearty and effective CTL responses, suggesting that the immune evasion doesn’t work all that well in vivo.

Most of the viruses that have had immune evasion genes identified are human viruses; that’s where the clinical interest and the money are. One problem with this is that the large DNA viruses that are likely to have these immune evasion functions tend to be pretty species-specific. Human herpesviruses and adenoviruses don’t infect mice well, if at all, and even if they do infect lab animals it’s not really clear how accurately this infection mimics the natural course in humans. There are a couple of lab animal herpesviruses that may be useful models (though even here the effects of immune evasion molecules are quite small.4) Still, it would be nice to understand what’s going on with immune evasion in, say, herpes simplex infection.

Herpes simplex virus actually will infect mice (the alphaherpesviruses tend to be somewhat less host-restricted than the beta or gammaherpesviruses), and while mouse infection isn’t exactly like that in humans it does seem to be similar enough to offer useful insights. Not into immune evasion, though, because the immune evasion gene of herpes simplex (called “ICP47”) works very poorly in mice.5 Recently, though, Chris Wilson’s group took an ingenious approach to looking at the question, and a theme may be starting to emerge.

Blogging on Peer-Reviewed ResearchWilson reasoned that if ICP47 doesn’t work in mice,6 why not replace it with a functionally-similar gene that does? They made recombinant herpes simplex viruses that have immune evasion genes from mouse or human cytomegalovirus that block mouse MHC class I fairly well. These viruses are more neurovirulent in mice than the control viruses7 and this was because the CTL didn’t work as well in the recombinants. 8,9

Their latest paper10 shows something new and exciting with the recombinant virus: It reactivates better from neurons.

Trigeminal ganglion
Trigeminal ganglion

Remember that HSV establishes a latent infection in neurons and, in humans, periodically reactivates and may shed and infect new hosts this way. HSV latency is still a mysterious and little-understood field in spite of huge amounts of work over the years, but a host of recent findings have started to explain at least bits and pieces. It used to be felt that latent HSV didn’t express any proteins, and so should be invisible to the immune system. As I observed here, it’s now known that the immune system can detect HSV in neurons. That offers a mechanism for reactivation to be coupled with immune suppression. The Orr et al paper here offers more support for this concept, and suggests that one of the (most important?) functions for HSV immune evasion is to tip the scales a little further toward the virus being able to escape recognition in the neuron, and therefore being more easily able to reactivate and infect new hosts — something this ubiquitous virus does with extraordinary efficiency. Orr et al call this an “actively testing the waters” model, in which HSV constantly sticks its toes out into the stream of the immune response to see if it’s safe to reactivate.

What especially interesting about this is that we now have (that I know of) three models of MHC class I immune evasion in more or less authentic infections, and each case the immune evasion seems to be designed to support latency, reactivation, and transmission. Here we have HSV using immune evasion in latency and persistence. Ann Hill’s group has recently shown that mouse cytomegalovirus MHC class I immune evasion proteins help the virus persist in the salivary glands,11 so that (as with the HSV story) it is more able to infect new hosts. And in the third model I know of, mouse herpesvirus 68, the MHC class I immune evasion genes are important for, you guessed it, latency and persistence.12

Three data points aren’t very many, but it does suggest that this may be a common feature of the herpesviruses. Perhaps in acute infection the virus doesn’t really “want” to block immune responses all the effectively, because their lifestyle involves only a brief burst of acute infection followed by a long-term latency; and it’s that latency that allows infection of new hosts. The viruses “let” themselves be driven into apparent submission, but then use their immune evasion functions to mount intermittent stealth campaigns to recruit new hosts.

If this is true for herpesviruses (and of course it’s just a hypothesis) is it also true for other viruses that have MHC class I immune evasion functions? Maybe for some (I wonder about the adenoviruses in particular, since they’re also very capable of long-term persistence in spite of immune responses) but I doubt it’s universal — HIV, for example, doesn’t obviously fit into the same lifestyle pattern as the herpesviruses, and yet if has a protein (nef) that targets MHC class I. Still, if offers a framework for thinking about the problem, which is very valuable.

  1. Manley, T. J., Luy, L., Jones, T., Boeckh, M., Mutimer, H., and Riddell, S. R. (2004). Immune evasion proteins of human cytomegalovirus do not prevent a diverse CD8+ cytotoxic T-cell response in natural infection. Blood 104, 1075-1082.[]
  2. Posavad, C. M., Koelle, D. M., and Corey, L. (1996). High frequency of CD8+ cytotoxic T-lymphocyte precursors specific for herpes simplex viruses in persons with genital herpes. J. Virol. 70, 8165-8168.[]
  3. York, I. A., Roop, C., Andrews, D. W., Riddell, S. R., Graham, F. L., and Johnson, D. C. (1994). A cytosolic herpes simplex virus protein inhibits antigen presentation to CD8+ T lymphocytes. Cell 77, 525-535.
    Jones, T. R., Hanson, L. K., Sun, L., Slater, J. S., Stenberg, R. M., and Campbell, A. E. (1995). Multiple independent loci within the human cytomegalovirus unique short region down-regulate expression of major histocompatibility complex class I heavy chains. J. Virol. 69, 4830-4841.[]
  4. For example, Munks, M. W., Pinto, A. K., Doom, C. M., and Hill, A. B. (2007). Viral interference with antigen presentation does not alter acute or chronic CD8 T cell immunodominance in murine cytomegalovirus infection. J Immunol 178, 7235-7241.[]
  5. Jugovic, P., Hill, A. M., Tomazin, R., Ploegh, H., and Johnson, D. C. (1998). Inhibition of major histocompatibility complex class I antigen presentation in pig and primate cells by herpes simplex virus type 1 and 2 ICP47. Journal of Virology 72, 5076-5084.[]
  6. To be strictly accurate, ICP47 does work in mice, just not very well — 100 times less than in humans, maybe, though what the unit of comparison is or should be isn’t clear[]
  7. I first wrote “than wild-type virus”, but I don’t think we know that — the process of recombination made the virus less virulent and the comparisons were done to this reduced-virulence control virus; I don’t see a direct comparison to wild-type virus.[]
  8. Orr, M. T., Edelmann, K. H., Vieira, J., Corey, L., Raulet, D. H., and Wilson, C. B. (2005). Inhibition of MHC class I is a virulence factor in herpes simplex virus infection of mice. PLoS Pathog 1, e7.[]
  9. This actually parallels and supports previous work showing that ICP47 does in fact have a small effect on neurovirulence in mice.[]
  10. ORR, M., MATHIS, M., LAGUNOFF, M., SACKS, J., WILSON, C. (2007). CD8 T Cell Control of HSV Reactivation from Latency Is Abrogated by Viral Inhibition of MHC Class I. Cell Host & Microbe, 2(3), 172-180. DOI: 10.1016/j.chom.2007.06.013 []
  11. Lu, X., Pinto, A. K., Kelly, A. M., Cho, K. S., and Hill, A. B. (2006). Murine cytomegalovirus interference with antigen presentation contributes to the inability of CD8 T cells to control virus in the salivary gland. J Virol 80, 4200-4202.[]
  12. Bennett, N. J., May, J. S., and Stevenson, P. G. (2005). Gamma-herpesvirus latency requires T cell evasion during episome maintenance. PLoS Biol 3, e120.[]
January 27th, 2008

TRegs and transplants

Embryonic kidneyLast week I talked about regulatory T cells (TRegs) in cancer. TRegs are often abundant in tumors, and have been linked to poor outcome, presumably because they prevent immune rejection of the tumor. The obvious flip side of this would be in a situation where you want to prevent immune rejection — in organ transplants. TRegs have been frustratingly hard to harness for this, though. (“Tolerance is the future of transplantation, and always will be.” –Norman Shumway)

A paper in the New England Journal of Medicine1 describes an encouraging step forward in this, achieving something that is at least close to the holy grail of transplantation — organ transplants that are maintained indefinitely without immunosuppression. Normally, organ transplants are rejected by the immune system, unless they’re from an identical twin (in which case the donor organ is perceived to be “self” by the immune system). By suppressing immunity, organ transplants can “take” without rejection; usually the immunosuppression is fairly harsh at first, but can be eased up over time, suggesting that a certain degree of tolerance is reached. (Also, the donor organ probably becomes less immunogenic over time, as some of the most immunogenic cells move out of the graft or die off, leaving less immunogenic tissues behind.)

Even though today’s immunosuppression is relatively gentle and focused, it’s only gentle relative to previous brutal treatments; it still leaves the recipient susceptible to infection, so there’s always a juggling act, balancing risk of rejection with risk of infection. The goal, then, has long been to find techniques that will allow the recipient’s immune system to become tolerant of the donor organ, as is seen in tumors.

Embryonic kidneyThe paper describes five kidney transplants that were preceded by bone marrow transfer from the donor. In four of the five cases, they were able to withdraw immunosuppression altogether, and the transplant wasn’t rejected (for at least one to five years, and counting). This is particularly exciting because these transplants weren’t from HLA-matched donors, meaning they were fairly immunogenic. (The same group, and another paper in the same issue of New England Journal, have done the same thing with HLA-matched transplants,2 which is still pretty interesting; but partially-mismatched transplants are much more common these days. )

One particularly interesting observation is that the bone marrow transfer only led to temporary chimerism (i.e. the donor bone marrow didn’t take permanently, and after a while only the original recipient bone marrow cells were present); but the tolerance persisted. They were able to find lots of TRegs infiltrating the donor kidneys, though, and so they believe that the long-term tolerance is probably because of TRegs (peripheral tolerance) although in the early stages thymic effects (central tolerance) may have been more important.

Blogging on Peer-Reviewed ResearchThe same issue of New England Journal describes the case of a young liver transplant recipient who apparently had her bone marrow seeded with stem cells from the donor liver, resulting in a switch of blood type and immune system to the donor’s and, again, a complete take of the graft without immunosuppression.3 That’s the case that’s getting all kinds of press right now, but while it may turn out to be an important guide to future treatment, it was essentially pure luck — the other cases here were the result of deliberate planning and defined conditions, which means that they can be repeated; the flashy case can’t, yet.

  1. Kawai, T. et al., 2008. HLA-Mismatched Renal Transplantation without Maintenance Immunosuppression. N Engl J Med, 358(4), p.353-361. []
  2. Bühler, L.H. et al., 2002. Induction of kidney allograft tolerance after transient lymphohematopoietic chimerism in patients with multiple myeloma and end-stage renal disease. Transplantation, 74(10), p.1405-9.
    Fudaba, Y. et al., 2006. Myeloma responses and tolerance following combined kidney and nonmyeloablative marrow transplantation: in vivo and in vitro analyses. American journal of transplantation, 6(9), p.2121-33.
    Spitzer, T.R. et al., 1999. Combined histocompatibility leukocyte antigen-matched donor bone marrow and renal transplantation for multiple myeloma with end stage renal disease: the induction of allograft tolerance through mixed lymphohematopoietic chimerism. Transplantation, 68(4), p.480-4.
    Scandling, J.D. et al., 2008.
    Tolerance and Chimerism after Renal and Hematopoietic-Cell Transplantation. N Engl J Med, 358(4), p.362-368. []
  3. Alexander, S.I. et al., 2008. Chimerism and Tolerance in a Recipient of a Deceased-Donor Liver Transplant. N Engl J Med, 358(4), p.369-374. []
January 23rd, 2008

Proteasome subunit-dependent signaling in T cells?

Mammalian proteasomeWe all know, now, that the proteasome is the most important factor in generating peptides for MHC class I. (See the antigen processing overview here if you want a quick review.) There are lots of lines of evidence pointing to this conclusion, but one of the early clues was the presence of interferon-inducible proteasome subunits in the MHC region of the genome.

It turns out (to make a long story short) that proteasomes have at least two different options for their catalytic subunits, and one of these options uses three interferon-inducible subunits. Two of the three are MHC-encoded (LMP2 and LMP7); the third is encoded outside the MHC (MECL1). The twin red flags of genomic organization and response to inflammation suggested that they might have a role in MHC class I antigen presentation; biochemical analysis suggested that the proteasome that incorporated these subunits turned out peptides that are better suited for MHC class I antigen presentation.1 (MECL1 identification trailed the other two by a few years2 and by the time it was shown to be interferon-regulated, some of the excitement was over.) The next step was to make knockout mice lacking each of the subunits.

Mammalian proteasomeThe LMP2 and LMP7 knockout mice were duly made3 and sure enough both showed (rather modest) phenotypic changes that could be ascribed to changes in MHC class I antigen presentation. MECL1 knockout mice weren’t made until quite recently (as I say, the excitement had sort of passed on to other things by then); and these knockout mice, too, showed T cell anomalies that could be ascribed to antigen presentation changes.4

Blogging on Peer-Reviewed ResearchA simple straightforward story (in hindsight), right? 5 Not so fast. Double knockout mice, lacking both MECL1 and LMP7, are dysfunctional in a completely unexpected way: the double knockout T cells (but not either of the single knockout LMP7 -/- or MECL1 -/- cells) are hyperproliferative, overresponding to some signals. 6 What’s more, this applies to not only CD8 T cells (which one imagines might be affected by MHC class I antigen oddities) but also to CD4 T cells, which shouldn’t much care about MHC class I antigen presentation.

An even more recent paper, still in press,7 analyzes this further with chimeric mice. It turns out that even when the T cells mature in the same environment (in other words, when MHC class I antigen presentation is held constant) the MECL1-knockout T cells are still different:

On the other hand, we provide evidence that the effects of MECL-1 on CD4 or CD8 T cell expansion are entirely unrelated to its role in antigen processing. Our findings suggest that MECL-1 influences the homeostatic regulatory processes that maintain the relative proportions of both T cell subsets, through a T cell-intrinsic mechanism independent from thymic or lymphoid interaction partners.

It seems likely that MECL1 somehow affects T cell regulation through affecting some signaling pathways.  This isn’t completely out of the blue — the proteasome has effects on signaling in several systems (in fact the proteasome affects just about every aspect of cell biology, one way or another). And it’s somewhat reassuring that so far there’s no indication that LMP2 or LMP7 might have caused their effects through anything other than antigen presentation. Still, it’s a warning signal not to take the easy explanation every time.

As a side note, it also makes me wonder about the thymus-specific subunit that was recently identified. When that came out I found it hard to imagine that the very large effects associated with the knockout of this subunit could be explained by antigen presentation. Now I wonder even more, because signaling is something that could easily explain the size of the effects they saw, though I’m not quite sure if it could explain the nature of the effects.

  1. There are a bunch of references for this, but I’m going to be lazy and just include a couple that I don’t have to think about: Gaczynska, M., Rock, K. L., and Goldberg, A. L. (1993). Gamma-interferon and expression of MHC genes regulate peptide hydrolysis by proteasomes. Nature 365, 264-267.
    Gaczynska, M., Rock, K. L., Spies, T., and Goldberg, A. L. (1994). Peptidase activities of proteasomes are differentially regulated by the major histocompatibility complex-encoded genes for LMP2 and LMP7. Proc Natl Acad Sci U S A 91, 9213-9217.[]
  2. Groettrup, M., Kraft, R., Kostka, S., Standera, S., Stohwasser, R., and Kloetzel, P. M. (1996). A third interferon-gamma-induced subunit exchange in the 20S proteasome. Eur J Immunol 26, 863-869.
    Hisamatsu, H., Shimbara, N., Saito, Y., Kristensen, P., Hendil, K. B., Fujiwara, T., Takahashi, E., Tanahashi, N., Tamura, T., Ichihara, A., and Tanaka, K. (1996). Newly identified pair of proteasomal subunits regulated reciprocally by interferon gamma. J Exp Med 183, 1807-1816.[]
  3. Van Kaer, L., Ashton-Rickardt, P. G., Eichelberger, M., Gaczynska, M., Nagashima, K., Rock, K. L., Goldberg, A. L., Doherty, P. C., and Tonegawa, S. (1994). Altered peptidase and viral-specific T cell response in LMP2 mutant mice. Immunity 1, 533-541.
    Fehling, H. J., Swat, W., Laplace, C., Kuhn, R., Rajewsky, K., Muller, U., and von Boehmer, H. (1994). MHC class I expression in mice lacking the proteasome subunit LMP-7. Science 265, 1234-1237.[]
  4. Basler, M., Moebius, J., Elenich, L., Groettrup, M., and Monaco, J. J. (2006). An altered T cell repertoire in MECL-1-deficient mice. J Immunol 176, 6665-6672.[]
  5. Especially because I am carefully avoiding mention of some of the blind alleys and red herrings that were puzzling at the time.[]
  6. Caudill, C. M., Jayarapu, K., Elenich, L., Monaco, J. J., Colbert, R. A., and Griffin, T. A. (2006). T cells lacking immunoproteasome subunits MECL-1 and LMP7 hyperproliferate in response to polyclonal mitogens. J Immunol 176, 4075-4082.[]
  7. Zaiss, D.M.W., de Graaf, N., & Sijts, A.J.A.M., 2007. The proteasome immunosubunit MECL-1 is a T cell intrinsic factor influencing homeostatic expansion. Infect Immun. doi:10.1128/IAI.01134-07 []
January 20th, 2008

TRegs and cancer

 Clark & Kupper: Blood, 1 January 2007, Vol. 109, No. 1, pp. 194-202. A while ago, talking about tumor development, I said that “tumors normally develop ways to avoid immunity.” It’s that ability to evade the immune system that allows tumors to escape from the equilibrium phase, when they’re a mere harmless handful of cells held in check by the immune system, and become outright clinically-detectable tumors.

How do tumors avoid immunity?

There are many ways. Remember that tumors are not like viruses; each tumor arises de novo and has only its host’s lifespan in which to evolve. It has no connection to other tumors of the same kind; there is no evolutionary linkage. All human cytomegaloviruses have a common ancestor, but no two colon cancers have the same ancestor. 1 That means that each tumor must find its own solution to every problem rather than relying on its ancestors’ solutions. (And if most colon carcinomas, say, adopt the same solution, we have to look at what factors make that solution particularly accessible to, or appropriate for, that particular type of cancer.)

That said, there are some mechanisms that are very widely used by tumors to evade the immune system, and regulatory T cells (TRegs) are one of them. That’s probably at least in part because TRegs’ natural functions include suppressing immune responses to self antigens and reducing inflammation in chronic exposure to antigen. It’s relatively easy to get the TRegs to do their job a little more enthusiastically.

TReg (J Clin Invest cover)(TRegs are T cells that specifically down-regulate immune responses; without TRegs, immune responses explode and cause massive damage. People without TRegs have terrible, usually fatal, autoimmune and inflammatory disease, so you don’t want to just blithely eliminate them to get a “better” immune response. )

It was suggested nearly 30 years ago that TRegs contributed to tumor growth,2 but the whole I-J fiasco set the field back a long way, and it wasn’t until relatively recently that the questions were revisited. 3 It’s now pretty clear that, in fact, TRegs are often abundant in tumors, and actively shut down immune responses to the tumors. For example, it’s been shown recently that tumors with relatively more TRegs have a worse prognosis:4

In patients with undesirable outcome, the balance is tipped in favor of Tregs (high Tregs and low activated CTLs), whereas in patients with relatively desirable outcome, the balance is tipped toward effector T cells (low Tregs and high activated CTLs).

Eliminating TRegs from mice with experimental tumors caused rejection of the tumors,5 so as you’d expect, there’s a lot of interest in this sort of approach to cancer therapy.

Relevant previous posts
• … and immune escape
• … and TLRs
• … and immunity
• … the Three E’s of
• … and equilibrium
Regulatory T cells:
• … and the I-J story
• … and persistent viruses
A small step forward was described recently in PNAS. 6 “Classic” TRegs are antigen-specific; they recognize antigen just as do T Helper cells, but instead of responding by stimulating immune responses, the TRegs respond to their antigen by suppressing local responses. There are also non-antigen-specific cells that act something like TRegs (though not as effectively). This paper shows that patients with melanoma have circulating (not just tumor-infiltrating) TRegs that are specific for tumor antigens.


This is a nice little paper, but I don’t think it represents a very large or surprising advance.7 It’s already known that tumor-infiltrating TRegs can be tumor-antigen-specific;8 it’s known that patients with melanoma have increased numbers of circulating TRegs;9 and it’s also been shown that tumor-specific T cells are in circulation in patients with melanoma. 10 So this (as far as I can see) extends several prior observations, but in an unsurprising direction.

Also, unless I’m missing something (which wouldn’t be new), I don’t see just how knowing or identifying circulating TReg antigens will help with treatment or prognosis. One possibility that occurs to me is that these circulating TRegs may help protect metastases from rejection. If the TRegs were all infiltrating the tumor, then perhaps the immune system would be able to deal with the small number of metastatic cells that are spreading throughout lymphatics and so on, whereas if TRegs are also circulating then the metastatic cells might be protected. I’m not entirely sure about the actual relevance of this, because even if TRegs were only tumor-infiltrating I think there would be plenty of opportunity to tolerize tumor-specific T cells.

Still, it’s always important to understand the system.

Just as a final cautionary note: As I said up at the top, tumors are all different. Different classes of tumors may be particularly able to move along certain immune evasion pathways because of their underlying characteristics. For example, inflammatory infiltrate within some tumors is a good sign11 (presumably because it implies that there is an active immune response against the tumor), yet in other classes of tumor the same kind of infiltrate is a bad sign, perhaps because it indicates TReg infiltration and a suppressive environment. Even within classes of tumors there are inevitably variations. It may not be possible to develop universal rules for tumor immune treatment; but it may be possible to find some useful guidelines.

  1. With bizarre exceptions like transmissible canine veneral tumor and Tasmanian Devil tumor. []
  2. Berendt, M.J. & North, R.J., 1980. T-cell-mediated suppression of anti-tumor immunity. An explanation for progressive growth of an immunogenic tumor. The Journal of experimental medicine, 151(1), p.69-80.[]
  3. A nice review: Zou, W., 2006. Regulatory T cells, tumour immunity and immunotherapy. Nat Rev Immunol, 6(4), p.295-307. []
  4. Gao, Q. et al., 2007. Intratumoral Balance of Regulatory and Cytotoxic T Cells Is Associated With Prognosis of Hepatocellular Carcinoma After Resection. J Clin Oncol, 25(18), p.2586-2593. DOI: 10.1200/JCO.2006.09.4565 []
  5. Shimizu, J., Yamazaki, S., & Sakaguchi, S., 1999. Induction of Tumor Immunity by Removing CD25+CD4+ T Cells: A Common Basis Between Tumor Immunity and Autoimmunity. J Immunol, 163(10), p.5211-5218.
    Viehl, C. T., Moore, T. T., Liyanage, U. K., Frey, D. M., Ehlers, J. P., Eberlein, T. J., Goedegebuure, P. S., and Linehan, D. C. (2006). Depletion of CD4+CD25+ regulatory T cells promotes a tumor-specific immune response in pancreas cancer-bearing mice. Ann Surg Oncol 13, 1252-1258.[]
  6. Vence, L. et al., 2007. Circulating tumor antigen-specific regulatory T cells in patients with metastatic melanoma. Proceedings of the National Academy of Sciences, 104(52), p.20884-20889. []
  7. I mention it here mainly because it reminded me that this is a subject I wanted to talk about.[]
  8. see Wang, R., 2006. Functional control of regulatory T cells and cancer immunotherapy. Seminars in Cancer Biology, 16(2), p.106-114. for a review[]
  9. McCarter, M.D. et al., 2007. Immunosuppressive dendritic and regulatory T cells are upregulated in melanoma patients. Annals of surgical oncology, 14(10), p.2854-60.[]
  10. For example, Michalek, J. et al., 2007. Detection and Long-Term In Vivo Monitoring of Individual Tumor-Specific T Cell Clones in Patients with Metastatic Melanoma. J Immunol, 178(11), p.6789-6795.

    and Bioley, G. et al., 2006. Melan-A/MART-1-specific CD4 T cells in melanoma patients: identification of new epitopes and ex vivo visualization of specific T cells by MHC class II tetramers. Journal of immunology 177(10), p.6769-79.[]

  11. For example, Eerola AK, Soini Y, Paakko P. A high number of tumor-infiltrating lymphocytes are associated with a small tumor size, low tumor stage, and a favorable prognosis in operated small cell lung carcinoma. Clin Cancer Res. 2000; 6: 1875-1881[]
January 17th, 2008

Switches and targets: T cell receptor downregulation by viruses

Painting of TcR interacting with artrificial membranes by Raghuveer Parthasarathy
TcR interacting with artificial membrane1

Earlier this week I talked about the phenomenon of viruses that downregulate MHC class II. The “purpose” of this blockade is kind of unclear to me, because the immunity driven by MHC class II is not focused on the cell it’s attached to, but rather spills out broadly over a wide area; it seems that a virus would have to very rapidly infect a very large number of MHC class II-expresing cells to have much effect on the anti-viral immune system.

One possible explanation is that the downregulation may be only peripherally related to MHC class II-based immunity. Instead, the virus could be simply going about its cell-biological business and either targeting MHC class II as an accidental side-effect, or because of some function of MHC class II that isn’t directly related to immunity. Here’s a parallel case that may help think about the problem.

The paper is
Sullivan, B.M., Coscoy, L. (2007). Downregulation of the T-Cell Receptor Complex and Impairment of T-Cell Activation by Human Herpesvirus 6 U24 Protein. Journal of Virology, 82(2), 602-608. DOI: 10.1128/JVI.01571-07

The T-cell receptor (TcR) is what recognizes MHC class I or II. Human herpesvirus 6 (HHV6) infects T helper (CD4) cells and (depending on viral strain) also does a number of things to modulate the immune system.2 Sullivan and Coscoy show here that the virus also down-regulates the TcR on infected T cells. (It’s altogether a more solid paper than the last one I mentioned, with nice experiments that directly show what’s happening to the TcR: “HHV-6 U24 protein inhibits CD3 recycling to the cell surface and, as a consequence, downregulates CD3 cell surface expression and prevents T-cell activation.“)

Sullivan & Coscoy Fig 4
U24 blocks CD3ε access to Rab11 recycling endosomes.3

At first glance this raises exactly the same question as does Vpu’s effect on MHC class II. How does reducing TcR on infected cells benefit the virus? The infected T cell will be less able to recognize its target, but what are the odds that its target is HHV6? Pretty minimal; there are (at least) tens of billions of different TcRs and only a handful of them recognize any particular antigen. The virus might be causing generalized immune suppression if it infects a large fraction of the T cells, but that’s not a particular benefit for the virus. If HHV6 specifically infected cells with TcRs that are specific for the virus then this would be a targeted immune evasion technique, but as far as I know there’s no evidence for this, nor is there an obvious mechanism by which HHV6 could target antigen-specific CD4 T cells.

There is, however, a nice explanation for TcR downregulation that doesn’t involve direct effects on T cell recognition. HHV6 (like all herpesviruses) has two choices when it infects a cell. It can either enter lytic replication — replicating the genome, producing more viruses, and eventually destroying the infected cell — or enter latency — a long-term, perhaps life-long, infection with minimal protein expression and little if any effect on the infected cell. As with any virus, the more prepared a cell is to replicate, the easier it is for a virus to replicate it’s own genome. T cells that receive a signal through their TcR become activated4 and divide very rapidly. In this environment, it’s very easy for HHV6 to replicate — that is, to enter lytic replication and kill the cell, releasing more viruses into the system.

Human herpesvirus 6 (HHV6)
Human herpesvirus 6

Probably HHV6 downregulates the TcR “because” it prevents its host from becoming activated by whatever its random antigen is. That prevents the virus from entering lytic replication and allows it to enter a persistent state, where it can hang about and await the best opportunity to infect a new person.

I know of one other viral protein that downregulates the TcR — the herpesvirus saimiri Tip protein 5 — and there seems to be controversy6 over whether this protein activates T cell signalling (potentially driving the virus into lytic replication) or blocks it (preventing lytic replication and facilitating persistence). 7 The original paper describing the TcR downregulation found that Tip blocked signaling, and proposed the same explanation as Sullivan and Coscoy:

… these associations ultimately block lymphocyte receptor signal transduction. … these interactions likely play an important role in the establishment and maintenance of HVS persistent infection by protecting infected cells from surveillance by the immune system. In fact, animals infected with recombinant HVSΔTip have been shown to have higher levels of cell-associated infectious virus titer compared to other recombinant HVS.

So in this case the downregulation of the TcR (a quintessentially immune molecule) apparently isn’t directly related to immune evasion, but is a way of switching between the lytic and the persistent lifestyles. (It’s also a reminder of the fairly obvious point that we shouldn’t think of viruses as blind replicators, desiring nothing more than maximal replication. At least some viruses have a range of lifestyle options, and can switch between them quite comfortably.)

I still don’t see a direct analogy to the MHC class II downregulation imposed by the HIV Vpu protein. but it’s an example of why we shouldn’t get too focused on single causes and single functions. Life is more complicated than that.

  1. By Raghuveer Parthasarath, then in the Groves lab[]
  2. Lusso, P., 2006. HHV-6 and the immune system: mechanisms of immunomodulation and viral escape. Journal of Clinical Virology, 37(Supplement 1), p.S4-S10. doi:10.1016/S1386-6532(06)70004-X []
  3. Sullivan & Coscoy, Fig 4[]
  4. I am simplifying immensely![]
  5. Park, J. et al., 2002. Herpesviral Protein Targets a Cellular WD Repeat Endosomal Protein to Downregulate T Lymphocyte Receptor Expression. Immunity, 17(2), p.221-233. doi:10.1016/S1074-7613(02)00368-0 []
  6. Brinkmann, M.M. & Schulz, T.F., 2006. Regulation of intracellular signalling by the terminal membrane proteins of members of the Gammaherpesvirinae. J Gen Virol, 87(5), p.1047-1074. DOI 10.1099/vir.0.81598-0[]
  7. I’m more convinced by the argument for blocking signaling, but only because of the very bad reason that I know the people involved. I haven’t looked at the papers pro and con very carefully.[]
January 14th, 2008

HIV, VPU, MHC, and other TLAs

HIV assembling in a macrophage
HIV infecting a macrophage1

It’s never been very clear to me why a virus would want to invest in blocking MHC class II. Since there are viruses that apparently do invest this way2 I may be missing something about virus-host interactions.

MHC class II is certainly critical to the immune response. People without MHC class II, as in some forms of “bare lymphocyte syndrome”, have severe immunodeficiency; unless they receive bone marrow transplants they usually die as children, of overwhelming viral, bacterial, and fungal infections. That’s because MHC class II is recognized by T helper (CD4) T cells, which are the foundation of most adaptive immune responses.  It’s easy to imagine how targeting MHC class II could benefit a pathogen.

So why am I puzzled by viruses that target MHC class II? The thing is that viruses are intracellular parasites, whereas MHC class II mainly presents peptides from extracellular sources. A virus may be able to block MHC class II presentation on the cell it’s infecting, but so what? It’s the neighbouring, uninfected cell that will present the virus peptides to CD4 cells and get the immune response moving.

(By contrast, of course, it’s obvious in principle how blocking MHC class I would benefit a virus. MHC class I molecules present peptide from intracellular molecules, so a virus can block the same molecules that will cause the immune response.)

Blogging on Peer-Reviewed ResearchThe particular paper that reminded me that this bugs me, by the way, is:
Hussain, A., Wesley, C., Khalid, M., Chaudhry, A., Jameel, S. (2007). Human Immunodeficiency Virus Type 1 Vpu Protein Interacts with CD74 and Modulates Major Histocompatibility Complex Class II Presentation. Journal of Virology, 82(2), 893-902. DOI: 10.1128/JVI.01373-07

They show that the HIV Vpu protein binds to invariant chain, a binding partner of MHC class II that (among other things) helps MHC class II traffic to the proper endosomal compartment. I didn’t peer-review this paper: I would have asked for several more experiments. Still, overall the actual association between Vpu and invariant chain looks fairly convincing, and there does seem to be an effect (though not a terribly impressive one) on overall MHC class II surface expression and on antigen presentation.

This is not the first time a viral protein has been shown to affect MHC class II. The human cytomegalovirus proteins US23 and US34 both affect MHC class II as well as MHC class I; herpes simplex virus is supposed to attack several steps in MHC class II antigen processing,5 again as well as MHC class I; and there are a handful of other examples. Aside from the HCMV US2 story, few have been studied in any detail, as far as I know, and in no case is their role in pathogenesis known.

HIV budding from a lymphocyte
HIV budding from a lymphocyte

So how might it make sense for a virus to block MHC class II antigen presentation? There are some ways in which this would make sense, but I don’t think any of them are particularly good answers.

(1) The blocking molecule could be secreted, so it affects neighboring antigen-presenting cells. This would actually be a very simple answer, except that none of the described cases do anything like this; they all affect MHC class II on the infected cell only and are not secreted. (I believe there are bacterial proteins which act somewhat like this.)

(2) The virus infects enough antigen-presenting cells to impact the overall CD4 response. I think this is the most common assumption, but … Eh. Maybe. In the early stages of infection with HIV or HCMV, a significant minority of macrophages and dendritic cells might be infected, but I’m dubious that there would be enough to have much of an effect — even less so for herpes simplex virus, which isn’t known for infecting these kinds of cells much at all.

(3) The virus is protecting against direct recognition by CD4 cells, rather than classical helper-type activity. While MHC class II mainly presents peptides from extracellular sources, the pathway presents a fair amount of intracellular antigen as well,6 and CD4 T cells can be cytotoxic, just like MHC class I-restricted CD8 T cells. This is actually my favorite explanation, but I’m not blown away by it; I’m not sure how common cytotoxic CD4 cells are, for one thing. For another, the rather wimpy effect of Vpu on MHC class II expression that Hussain et al showed here, doesn’t strike me as sufficient to prevent CD4 recognition — though they didn’t test for that, so it’s still a possibility.

(4) The effect on MHC class II is an irrelevant side-effect of the true function of the protein, such as attacking MHC class I. MHC class II molecules are awfully similar to MHC class I (see this post for examples), so maybe the HCMV molecules are binding by accident. Similarity between class I and II doesn’t explain binding the invariant chain, but invariant chain was shown to interact with MHC class I molecules7 as well as class II — a finding no one has known what to do with in the 12-plus years since it was described — so again, maybe Vpu is “really” targeting MHC class I, and the effect on MHC class II is a side-effect. Or there could be some other effect on viral replication or assembly, it doesn’t have to be related to MHC class I.
(5) Similar to (4) — maybe the effect on MHC class II offers a very minor protection for the virus, but it comes cheap because the virus has already developed machinery to attack the closely-related MHC class I pathway. In this argument the payoff of attacking MHC class II is small, but the investment in expanding the function of anti-class I molecules to also interact with class II is small as well, so why not go ahead? (Or, less teleologically, having evolved anti-MHC class I molecules, very minor changes in their sequence allowed an effect on MHC class II as well.)

(6) MHC class II has some effect on viral infection other than through interaction with CD4 T cells, and it’s those (unknown) effects that the viruses are defending themselves against. This is the most intriguing possibility, and the reason I keep going back and studying these papers; but there’s not a lot of actual evidence for it.

  1. Gross, L., 2006. Reconfirming the Traditional Model of HIV Particle Assembly. PLoS Biology, 4(12), p.e445 EP []
  2. Albeit with rather small investments[]
  3. Tomazin, R. et al., 1999. Cytomegalovirus US2 destroys two components of the MHC class II pathway, preventing recognition by CD4+ T cells. Nat Med, 5(9), p.1039-1043. DOI:10.1038/12478[]
  4. Hegde, N.R. et al., 2002. Inhibition of HLA-DR Assembly, Transport, and Loading by Human Cytomegalovirus Glycoprotein US3: a Novel Mechanism for Evading Major Histocompatibility Complex Class II Antigen Presentation. J. Virol., 76(21), p.10929-10941. DOI: 10.1128/JVI.76.21.10929-10941.2002[]
  5. Neumann, J., Eis-Hubinger, A.M., & Koch, N., 2003. Herpes Simplex Virus Type 1 Targets the MHC Class II Processing Pathway for Immune Evasion . J Immunol, 171(6), p.3075-3083.[]
  6. The phenomenon was known as a curiosity for a long time, but I think it was Ike Eisenlohr who quantified it and showed that it was a fairly common event: Tewari, M.K. et al., 2005. A cytosolic pathway for MHC class II-restricted antigen processing that is proteasome and TAP dependent. Nat Immunol, 6(3), p.287-294. []
  7. Sugita, M. & Brenner, M.B., 1995. Association of the Invariant Chain with Major Histocompatibility Complex Class I Molecules Directs Trafficking to Endocytic Compartments. J. Biol. Chem., 270(3), p.1443-1448. []
January 10th, 2008

Oncolytic viruses and immune clearance

Oncolytic VSV
Oncolytic VSV (gold) infecting lung tumors1

Oncolytic viruses are a concept I’d like to be more excited by than I am.2 It’s an idea that seemed really exciting when I first came across it, but the more I thought about it the more dubious I was. But a recent paper helps me feel better about at least two of my worries.

The concept is a straightforward one. Viruses are good at killing cells.3 Why not have them infect cells that we want to die? That would be, for example, cancer cells. So all you need to do is find or make a virus that only grows in cancer cells, and you’re cured. Simple! Tomorrow we’ll fix global warming!

There’s the obvious problem with this: How can you find (or make) a cancer-specific virus? In principle the answer is the same as with chemotherapy; you use the ways cancer cells are different from normal as targets. This isn’t as hard as you might think. Lots of the things that make cancer cells cancerous are similar to the things viruses like. Viruses often drive infected cells into a cancer-like state that is more hospitable to the virus — friendly to nucleic acid replication, replication, unresponsive to death signals, independent of the signals that normally regulate growth. So lots of viruses are already kind of pre-adapted to replicate well in cells with a cancerous phenotype, and it doesn’t take all that much tweaking to make them adapted to only replicate well in cancer cells.

(After writing this post, it occurred to me that this is actually topical! I don’t usually do the topical blog post thing, but the background in “I Am Legend” has an anti-cancer virus, isn’t it? I haven’t seen it myself, with the grant-writing and the teaching and the two little kids,4 but have I actually tied a current entertainment topic into “Mystery Rays”? Fame and fortune is certain to come my way!)

Oncolysis through the ages

Jennerex Onolytic virusThe first runs at this technique that I knew of5 used mutant herpesviruses,6 but I think that much of the buzz came from work with defective adenoviruxes, especially the ONYX-015 virus.7 The approach here was based on the observation that adenoviruses (like many other viruses) normally inactivate p53 during infection. p53 is a multifunctional growth regulator that is very often also inactivated in cancers, for the same reason as viruses like to inactivate it:it oten triggers death in cells with unchecked growth. Adenoviruses lacking the gene that inactivates p53 (their E1B gene) can only efficiently infect cells lacking p53 — which would usually be, of course, cancer cells.

Blogging on Peer-Reviewed ResearchAs well as herpesviruses and adenoviruses, though, all sorts of other viruses have been used.8 One interesting approach is vesicular stomatitis virus. This is a very, very innocuous virus in normal people, partly because VSV is extremely sensitive to interferon. (VSV is used in bioassays for interferon release, because even tiny amounts of interferon completely block the virus’s replication.) So which kind of cells often aren’t responsive to interferon? Right; cancer cells, as part of their own immune evasion pathway, frequently disable their interferon responses. VSV doesn’t infect normal cells, but does infect, and kill, tumor cells.9

Questions and (maybe) answers

Anyway, the first question, of specificity, is more or less under control.10 Three questions that had made me rather dubious about the concept, though, still remained:

1. Getting the virus to the tumor …
2. Especially in the face of an immune response.
3. Killing all of the cancer cells, not a mere 99% of them (from which the cancer will rapidly recover).

Malignant melanoma cells in lymph node
Malignant melanoma cells in lymph node

A paper in Nature Medicine11 offers encouragement on all of those.

They used VSV as their cancer killer, and their twist here was to deliver it by loading it onto T cells. T cells naturally traffic to lymph nodes, and quite a few tumors metastasize through lymph nodes; the T cell therefore acts as a ferry to deliver its deadly viral cargo to the metastasizing tumor. (The goal here was not to clear the primary tumor, but to prevent metastases, which are often the major problem.  However, they did see some effect on the primary tumor, too, in some cases.) When it reaches the lymphoid tissue, it delivers the passenger virus to the cancer cells, the only ones that the VSV can productively infect (since the cancer cells are the only ones that have mutated their interferon pathway). This is an interesting idea, though limited in this form — I wonder about using antigen-specific T cells instead, to target the virus to a specific site — and it seemed to work quite well.

The two more interesting points to me were kind of peripheral to their main point. First, they find that once the virus killed some cancer cells, there was anti-tumor protection even after the virus was all cleared, and this was probably because of the immune response,12 which was triggered by the cell death initially caused by the virus:

In vivo tumor cell purging resulted both from direct viral oncolysis by virus released from the T cell carriers and from the priming of protective antitumor immunity, which prevents repopulation by further waves of cells metastasizing from the primary tumor.

— just as described in the paper by Apetoh et al13 that I talked about here. The authors suggest that because the cancer metastases are being killed in the lymph nodes, rather than in the bulk of the tumor (which is generally a highly immunosuppressive environment) the immune response was more efficient. That starts to get past my concern #3 above, because it offers multiple attacks on the tumor, not just the virus.

The other point is that the virus could reach the cancer reasonably well even in the face of an anti-viral immune response; the trick was to use just enough virus to kill the tumor cells, without getting enough on the T cells to trigger an immune response:

In virus-immune mice, T cells loaded with large amounts of VSV (MOI 1 or 10) could not keep DLNs or spleens free of tumors. However, T cells loaded with fewer viruses (MOI 0.1) still protected even virus-immune animals from tumor colonization of the DLN and spleen

The data are still very preliminary and inconclusive, but certainly it’s a step in the right direction, and I feel better about this whole approach than I did before reading the paper.

  1. Carrier Cell-based Delivery of an Oncolytic Virus Circumvents Antiviral Immunity. Anthony T Power, Jiahu Wang, Theresa J Falls, Jennifer M Paterson, Kelley A Parato, Brian D Lichty, David F Stojdl, Peter A J Forsyth, Harry Atkins and John C Bell. Molecular Therapy (2007) 15, 123-130. []
  2. That sentence needs a road map, but you got here eventually, didn’t you.[]
  3. At least, lytic viruses are.[]
  4. And the running and the screaming and the monkeys in the hair[]
  5. I realize now, though, that the concept arose long before that, apparently in the 1950s. For example: Love R, Sharpless GR. Studies on a transplantable chicken tumor, RPL-12 lymphoma. II. Mechanism of regression following infection with an oncolytic virus. Cancer Res. 1954 Oct;14(9):640-7. though I don’t know much about those studies other than the titles []
  6. Experimental therapy of human glioma by means of a genetically engineered virus mutant. Martuza RL, Malick A, Markert JM, Ruffner KL, Coen DM. Science. 1991 May 10;252(5007):854-6. []
  7. ONYX-015, an E1B gene-attenuated adenovirus, causes tumor-specific cytolysis and antitumoral efficacy that can be augmented by standard chemotherapeutic agents. Heise C, Sampson-Johannes A, Williams A, McCormick F, Von Hoff DD, Kirn DH. Nat Med. 1997 Jun;3(6):639-45.
    An adenovirus mutant that replicates selectively in p53-deficient human tumor cells. Bischoff JR, Kirn DH, Williams A, Heise C, Horn S, Muna M, Ng L, Nye JA, Sampson-Johannes A, Fattaey A, McCormick F. Science. 1996 Oct 18;274(5286):373-6. []
  8. And I have no idea which, if any, is the most promising.[]
  9. Exploiting tumor-specific defects in the interferon pathway with a previously unknown oncolytic virus. David F. Stojdl, Brian Lichty, Shane Knowles, Ricardo Marius, Harold Atkins, Nahum Sonenberg & John C. Bell. Nature Medicine 6, 821 – 825 (2000) doi:10.1038/77558[]
  10. One other point is that you can probably get away with a virus that isn’t completely restricted to tumor cells, because these are usually viruses that cause very mild disease anyway, so even if they can spread to normal cells it’s no more worry than exposure to a standard subway car. Maybe more concern for immunosuppressed cancer patients, of course, but likely not an insurmountable worry.[]
  11. Qiao, J., Kottke, T., Willmon, C., Galivo, F., Wongthida, P., Diaz, R.M., Thompson, J., Ryno, P., Barber, G.N., Chester, J., Selby, P., Harrington, K., Melcher, A., Vile, R.G. (2007). Purging metastases in lymphoid organs using a combination of antigen-nonspecific adoptive T cell therapy, oncolytic virotherapy and immunotherapy. Nature Medicine DOI: 10.1038/nm1681[]
  12. The short-term clearance worked in immune-deficient mice, but the long-term did not[]
  13. Toll-like receptor 4-dependent contribution of the immune system to anticancer chemotherapy and radiotherapy. Apetoh, L., Ghiringhelli, F., Tesniere, A., Obeid, M., Ortiz, C., Criollo, A., Mignot, G., Maiuri, M. C., Ullrich, E., Saulnier, P., Yang, H., Amigorena, S., Ryffel, B., Barrat, F. J., Saftig, P., Levi, F., Lidereau, R., Nogues, C., Mira, J. P., Chompret, A., Joulin, V., Clavel-Chapelon, F., Bourhis, J., Andre, F., Delaloge, S., Tursz, T., Kroemer, G., and Zitvogel, L. (2007). Nat Med 13, 1050 – 1059. []
January 6th, 2008

Clams got herpes!

Oyster reefWhere did herpesviruses come from?

Humans, of course, have 8 different herpesviruses that are remarkably good at infecting us. Humans aren’t exceptional: We know of 200-odd herpesviruses so far, and more are being identified practically daily. It’s likely that virtually every animal species has its own set of unique herpesviruses. This is probably because herpesviruses are very host-restricted (rarely infecting more than a single species) and set up latent, life-long infection. When an animal population speciates, its complement of herpesviruses speciates along with it.

How long has this been happening? What mammal, dinosaur, fish, or trilobite did the first herpesvirus infect?

Herpes simplex structure
Herpes simplex virus structure1

It was certainly before the mammal/reptile split, some 300 million years ago. Mammalian, reptilian, and avian herpesviruses cluster very nicely together by DNA and amino acid sequence, with some 40 genes that are clearly homologous. (Mammalian herpesvirus are grouped into alpha, beta, and gammaherpesvirus based originally on biology and, subsequently, on genome sequence; but bird and reptile herpesviruses so far have all been alphaherpesviruses, so this tripartite split did happen after the synapsid reptiles diverged from the other lineages that led to birds and turtles and so on.)2

How to spot a herpesvirus
• Linear, double-stranded DNA
• 120 – 250,000 base pairs
• Unique short and long regions
• Inverted repeats flank UL and US
• Core containing DNA
• Icosahedral (T=16) capsid
• Amorphous tegument between capsid and envelope
Amphibians and fish herpesviruses are quite different from mammal and reptile groups, and cluster into their own broad family, sharing a number of closely-related genes. Genome similarity is low or non-existent, but the pathognomic physical structure, capsid architecture, and overall DNA organization is entirely consistent. These are clearly herpesviruses that diverged over 400 million years ago, and have evolved away all but the faintest semblance of sequence similarity.

Impressive as it is to be able to reconstruct virus evolution for nearly a half billion years, we’re not done yet. In 1972, a herpesvirus of oysters was identified!3 So far this is the only herpesvirus of invertebrates to be definitively identified (there may be others, such as a herpesvirus of abalones,4 but as far as I know only the Ostreid herpesvirus has had its genome sequenced,5 so it’s quite possible that the abalone virus is the same thing). As with the amphibian and fish herpesviruses, OsHV has the very characteristic herpesvirus structure and organization, but at the sequence level it’s very different from either of the other major branches, making this the only known representative of a third major group of herpesviruses.

…the ancient herpesvirus from which modern herpesviruses are presumed to have descended was in existence around a billion years ago and was recognisably a herpesvirus. This pushes the origins of herpesviruses so far back in time that it is difficult to see how we might discern the nature of even earlier ancestors. 6

ResearchBlogging.orgCan we push ancestry back beyond a billion years? Not easily; DNA sequence and even characteristic capsids and so on can only take us so far. But protein three-dimensional structure is more conserved than is sequence, and a couple of groups have shown common elements in folds of herpesvirus and bacteriophage proteins that suggest a common ancestry past the eukaryote/prokaryote divergence:7

This linkage between the most abundant set of bacteriophages and a major family of eukaryotic viruses highlights the growing evidence of relationships across these major biological divides. In light of this and other recent studies, it is becoming increasingly plausible that extant viruses may have arisen from a relatively small number of primordial progenitors. 8

  1. Three-Dimensional Structure of Herpes Simplex Virus from Cryo-Electron Tomography
    Grünewald et al., Science 302:1396 – 1398 (2003) []
  2. It’s also been suggested that some of the avian and mammalian viruses are too closely related to have diverged that long ago, and may indicate ancient interspecies transmissions some 100 million years ago. See: Toward a Comprehensive Phylogeny for Mammalian and Avian Herpesviruses Duncan J. McGeoch, Aidan Dolan, and Adam C. Ralph Journal of Virology 74:10401-10406 (2000) []
  3. Farley, C.A., Banfield, W.G., Kasnic, G., Foster, W.A. (1972). Oyster herpes-type virus. Science, 178(62), 759-760.[]
  4. Herpes-like virus infection causing mortality of cultured abalone Haliotis diversicolor supertexta in Taiwan. Chang PH, Kuo ST, Lai SH, Yang HS, Ting YY, Hsu CL, Chen HC. Dis Aquat Organ. 2005 Jun 14;65(1):23-7 []
  5. A novel class of herpesvirus with bivalve hosts. Andrew J. Davison, Benes L. Trus, Naiqian Cheng, Alasdair C. Steven, Moira S. Watson, Charles Cunningham, Rose-Marie Le Deuff, and Tristan Renault. J Gen Virol 86 (2005), 41-53 .
    The accession number of the OsHV-1 DNA sequence is AY509253.[]
  6. Davison, A. (2002). Comments on the phylogenetics and evolution of herpesviruses and other large DNA viruses. Virus Res 82, 127-132.[]
  7. Common Ancestry of Herpesviruses and Tailed DNA Bacteriophages. Matthew L. Baker, Wen Jiang, Frazer J. Rixon, and Wah Chiu. Journal of Virology, December 2005, p. 14967-14970, Vol. 79, No. 23.
    Also see Structure of an archaeal virus capsid protein reveals a common ancestry to eukaryotic and bacterial viruses.  Reza Khayat, Liang Tang, Eric T. Larson, C. Martin Lawrence, Mark Young, and John E. Johnson. Proc Natl Acad Sci U S A. 2005 Dec 27;102(52):18944-9
    Structural and functional similarities between the capsid proteins of bacteriophages T4 and HK97 point to a common ancestry.  Fokine A, Leiman PG, Shneider MM, Ahvazi B, Boeshans KM, Steven AC, Black LW, Mesyanzhinov VV, Rossmann MG. Proc Natl Acad Sci U S A. 2005 May 17;102(20):7163-8. []
  8. Common Ancestry of Herpesviruses and Tailed DNA Bacteriophages. Matthew L. Baker, Wen Jiang, Frazer J. Rixon, and Wah Chiu. Journal of Virology, December 2005, p. 14967-14970, Vol. 79, No. 23.[]
January 2nd, 2008

Antibody-based vaccines

Broadly neutralizing anti-HIV antibody Viruses replicate inside cells, which shields them from some components of the immune system. In particular, antibodies can’t penetrate inside a cell1 to bind to a virus there, so antibodies are not much use for eliminating a viral infection.2 For some viruses that have to exit the cell to spread to a new target cell, antibody may help limit spread, but many viruses can spread directly from one cell to its neighbor without ever being exposed to antibodies. So once a virus has entered a host’s cells, you probably want mostly cell-mediated immune responses, such as T helper cells and cytotoxic T lymphocytes, to get rid of the virus.

That’s for eliminating viral infections. What antibodies are often extremely good at is blocking infections–stopping the virus from ever getting a foothold. A virus that enters your body has to be exposed to extracellular components at least briefly before it can burrow into its protective cell. During that phase the virus is vulnerable to antibody-mediated inhibition. Antibodies therefore may be relatively unhelpful for getting rid of an ongoing infection, but they can be very good at protecting against new infections.

Not surprisingly, then, most3 antiviral vaccines depend on inducing a strong and specific antibody response. That also means you can often get away with killed virus vaccines like the Salk polio vaccine, or even subunit vaccines like Hepatitis B vaccine; these are very poor at inducing cytotoxic T lymphocyte (CTL) responses, but they don’t have to. Killed vaccines are, in principle, intrinsically safer than the attenuated viruses, or even vector-based recombinant vaccines.

Why are researchers looking for alternatives?

So why is there so much interest in developing vaccines that stimulate CTL? Why are so many groups working on vector-based vaccines or attenuating viruses? One reason is that these vaccines are (again, in principle) intrinsically more immunogenic than killed vaccines. If you can give one dose of vaccine, and then have your antigens stick around for a couple of weeks, or even amplify themselves as they replicate in situ, then you may not need to give a second (booster) dose of the vaccine. That’s a moderate advantage in the first world, and potentially a huge advantage in the third world, where you may only have one chance to visit your patients.

Another reason is that to a large extent we’ve already nailed the simple problems. If a killed vaccine can protect against a major pathogen, we probably already have that vaccine up and running. We’re left with those virus diseases that, for one reason or another, are not easily prevented by antibody-type responses, and so cellular CTL-type responses are the most promising next step.

HIVWhat keeps a virus from being blocked by antibodies? There are a number of reasons, but the most obvious is that the virus offers a moving target to antibodies. HIV is probably the most famous example of this approach. The HIV surface is dominated by glycoproteins that are enormously variable; an antibody that blocks one particular HIV strain does nothing against a different strain. Hepatitis C virus (HCV) is another virus with highly variable surface proteins. Malaria, a parasite rather than a virus, has enough room in its genome to take this strategy even further. As well as using strain variation, individual parasites can dynamically change their surface proteins, stepping methodically through some 60 variants.4

Surface antigens in these pathogens have probably evolved to be variable; there’s been selective pressure for a pathogen to be different from the majority, since that way they’re less likely to infect an immune host.5 Internal antigens–those that are not exposed to antibodies-tend to be more highly conserved. Internal antigens aren’t exposed to antibodies, but they’re perfectly good targets for CTL. This is one reason for the interest in developing vaccines that induce good CTL responses.

Back to the future: Workarounds for antibody-based vaccines

There’s another approach, though. We have a lot of experience with antibody-based vaccines. It would be nice if there was a way to use them against HIV and HCV. Are there sections of the virus surface that are not variable? If so, then designing a vaccine that raises antibodies against these regions might be effective against many different strains. That’s been a hot topic for quite a while, and in fact there have been some steps forward on this front for HIV.6 More recently, in the latest issue of Nature Medicine7 there’s an article suggesting that some antibodies may be able to neutralize many hepatitis C strains.

The results provide evidence that broadly neutralizing antibodies to HCV protect against heterologous viral infection and suggest that a prophylactic vaccine against HCV may be achievable.

  1. Yes, I know that some forms of antibody routinely penetrate cells as they’re pumped into the gut, for example, but let’s stay relevant.[]
  2. Perhaps antibodies are important in some cases for triggering antibody-dependent cell-mediated cytotoxicity (ADCC) by NK cells, but again let’s not get sidetracked.[]
  3. If not all. I can’t think of a counterexample offhand[]
  4. Developmental selection of var gene expression in Plasmodium falciparum. Qijun Chen, Victor Fernandez, Annika Sundstram, Martha Schlichtherle, Santanu Datta, Per Hagblom & Mats Wahlgren. Nature 394, 392-395 (23 July 1998) []
  5. That being said, I don’t know that this has been formally shown for any of these agents; and in fact the only study I know of off the top of my head specifically did not find evidence for frequency-dependent selection in malaria surface protein alleles: Sequence Variation in the T-Cell Epitopes of the Plasmodium falciparum Circumsporozoite Protein among Field Isolates Is Temporally Stable: a 5-Year Longitudinal Study in Southern Vietnam. Amadu Jalloh, Huynh van Thien, Marcelo U. Ferreira, Jun Ohashi, Hiroyuki Matsuoka, Toshio Kanbe, Akihiko Kikuchi, and Fumihiko Kawamoto. Journal of Clinical Microbiology, April 2006, p. 1229-1235, Vol. 44, No. 4  []
  6. For example, Structural definition of a conserved neutralization epitope on HIV-1 gp120. Tongqing Zhou, Ling Xu, Barna Dey, Ann J. Hessell, Donald Van Ryk, Shi-Hua Xiang, Xinzhen Yang, Mei-Yun Zhang, Michael B. Zwick, James Arthos, Dennis R. Burton, Dimiter S. Dimitrov, Joseph Sodroski, Richard Wyatt, Gary J. Nabel & Peter D. Kwong. Nature 445, 732-737 (15 February 2007)–the source of the image at top here[]
  7. Broadly neutralizing antibodies protect against hepatitis C virus quasispecies challenge. Mansun Law, Toshiaki Maruyama, Jamie Lewis, Erick Giang, Alexander W Tarr, Zania Stamataki, Pablo Gastaminza, Francis V Chisari, Ian M Jones, Robert I Fox, Jonathan K Ball, Jane A McKeating, Norman M Kneteman & Dennis R Burton. Nature Medicine Published online: 6 December 2007  []