Mystery Rays from Outer Space

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

December 4th, 2008

Controlled TRegs: The future is (almost) now

TRegs (JCI)Our bodies are crammed with millions of tiny time bombs: lymphocytes that could begin to attack our own bodies, causing lethal autoimmune disease. Traditionally, it was said that these self-reactive lymphocytes were rare, because they were eliminated during their development and were never allowed to reach maturity. But it’s been known for quite a few years now that that’s not entirely true. The vast majority of self-reactive T cells may, indeed, be destroyed in the thymus, but by no means all. (Something like a couple million T cells leave a happy, functioning thymus every day. If central tolerance is 99.999% perfect, then 10 self-reactive T cells will enter the system every single day — and it only takes a couple of T cells to initiate a lethal disease.)

Why don’t we all die as infants of autoimmune attack, if circulating self-reactive T cells are so (relatively) common? As with just about everything in our body, there are redundant systems. For autoimmunity, the next line of defense is the regulatory T cell (TReg).

TRegs were identified as a phenomenon long ago, in the 1960s and 1970s; but the concept abruptly fell out of favor in 1984 (for fascinating and rather embarrassing reasons I talked about here), and it wasn’t until the new millennium that immunologists really returned to the field (first firmly changing the name from “suppressor T cells” to “TRegs” to keep their feet out of the muck), and the field really exploded 5 or 6 years ago.

TRegs have proved more important and powerful than just about anybody would have believed ten years ago. Even very powerful immune responses can be controlled by TRegs; strong TReg responses can actually allow a complete “take” of an organ transplant, for example (I mentioned some examples here).

 TRegs infiltrate tumor
Regulatory t cells infiltrate tumor tissue

As well as transplants, being able to turn on TRegs has potential for lots of other diseases. Autoimmunity, obviously, could be controlled this way; but also, less obviously, it’s possible that some virus diseases might benefit from a TReg response. HIV infection, for example, is exacerbated when T cells are activated, and monkeys with SIV are resistant to disease when their T cells are less reactive (see here and here); could a controlled TReg response reduce the harmful activation associated with HIV? It may seem counterintuitive to try to treat a viral disease by reducing immunity, but there is some precedent. Rodents infected with hantaviruses develop a TReg response and don’t have much disease (see here), while humans react with a more conventional immune response and have severe disease. And recently, it was shown that elite suppressors of HIV may have an exceptionally strong TReg response.1

Conversely, there are lots of instances where we’d like to turn off TRegs, in a controlled way. Tumors are often associated with TRegs, which very likely prevent a cleansing immune response to the tumor (discussed here). And the well-known observation that the elderly often have poor immunity against various pathogens is at least partly because TRegs build up over time.

This is a very fast-moving field, and there are a several recent papers that show exciting advances. One is a huge basic step forward, and I’ll talk about that later. The others2 are technical advances, developing new techniques (that are much less cumbersome and finicky than some of the previous approaches) to generate large numbers of TRegs in a controlled way. The obvious use for this is in transplants:

The ex vivo expansion protocol that we describe will very likely increase the success of clinical Treg-based immunotherapy, and will help to induce tolerance to selected antigens, while minimizing general immune suppression. This approach is of particular interest for recipients of HLA mismatched transplants.3

Controlled TRegs have been a holy grail of transplant biology for years, and it’s exciting to see that we may finally be entering an era when TRegs can be produced and used as tools.

  1. Preservation of FoxP3+ regulatory T cells in the peripheral blood of human immunodeficiency virus type 1-infected elite suppressors correlates with low CD4+ T-cell activation.
    Chase AJ, Yang HC, Zhang H, Blankson JN, Siliciano RF
    J Virol 2008 Sep 82(17):8307-15[]
  2. Including, but not limited to:
    W. Tu, Y.-L. Lau, J. Zheng, Y. Liu, P.-L. Chan, H. Mao, K. Dionis, P. Schneider, D. B. Lewis (2008). Efficient generation of human alloantigen-specific CD4+ regulatory T cells from naive precursors by CD40-activated B cells Blood, 112 (6), 2554-2562 DOI: 10.1182/blood-2008-04-152041

    In Vitro Expanded Human CD4+CD25+ Regulatory T Cells are Potent Suppressors of T-Cell-Mediated Xenogeneic Responses. Wu, Jingjing; Yi, Shounan; Ouyang, Li; Jimenez, Elvira; Simond, Denbigh; Wang, Wei; Wang, Yiping; Hawthorne, Wayne J.; O’Connell, Philip J. Transplantation Volume 85(12), 27 June 2008, pp 1841-1848.

    Jorieke H. Peters, Luuk B. Hilbrands, Hans J. P. M. Koenen, Irma Joosten (2008). Ex Vivo Generation of Human Alloantigen-Specific Regulatory T Cells from CD4posCD25high T Cells for Immunotherapy PLoS ONE, 3 (5) DOI: 10.1371/journal.pone.0002233

    and a review in Piotr Trzonkowski, Magdalena Szary?ska, Jolanta My?liwska, Andrzej My?liwski (2008). Ex vivo expansion of CD4+CD25+ T regulatory cells for immunosuppressive therapy
    Cytometry Part A, 9999A DOI: 10.1002/cyto.a.20659

  3. Jorieke H. Peters, Luuk B. Hilbrands, Hans J. P. M. Koenen, Irma Joosten (2008). Ex Vivo Generation of Human Alloantigen-Specific Regulatory T Cells from CD4posCD25high T Cells for Immunotherapy PLoS ONE, 3 (5) DOI: 10.1371/journal.pone.0002233[]
March 17th, 2008

Controlled TReg production

Saints Cosmas and Damian performing a miraculous cure by transplantation of a leg/The Master of Los Balbases.I’ve previously posted on regulatory T cells (TRegs) and their potential role in transplants. Briefly, TRegs are capable of specifically shutting off immune responses to particular antigens; they’re normal components of an immune system. TRegs can be damaging in some contexts — for example, in cancer, where it seems that TRegs often shut off immune responses to tumors, so that the tumor can escape immune clearance; and they can be beneficial in other context — for example, in some persistent virus infections, where a chronic immune response would be damaging, TRegs apparently modulate the immune response so that the virus persists but doesn’t cause severe damage.

There are a couple of obvious scenarios where it would be nice to be able to control TRegs. There’s a lot of interest in reducing TReg activity in cancer, such as with CTLA4 antagonists. There’s also a lot of interest in increasing TReg activity in organ transplants, and there have actually been a couple of cases where it’s seemed to have worked.

A recent paper in PNAS1 offers steps toward a more general procedure, that could in theory lead to controlled, planned generation of TRegs for any antigen.

A key aspect of TRegs is that they are antigen-specific. They don’t randomly suppress immune responses; they identify particular antigens that should be tolerated, and shut off immunity to those antigens. That allows fine control over the response, but it also makes it harder to catch a TReg; T cells (not just TRegs) that recognize any particular antigen are very rare events, hiding in a blizzard of other specificities. What if you could force T cells for an antigen you choose to enter the TReg pathway?

Regulatory T cells (J Clin Invest cover)This has already been done, in fact, but in a very artificial system — in mice with transgenic T cell receptors. These mice overwhelmingly express a single TcR in all of their T cells — there’s no snowflake in a blizzard problem, because the entire blizzard is made of identical flakes. Harold von Boehmer’s group has shown that you can drive these transgenic T cells into the TReg pathway by offering very, very low levels of antigen, under defined conditions, over a long period. 2 The recent paper1 shows that you can do the same thing in normal, non-transgenic, mice; and by doing this you can force graft tolerance. (They used female mice and drove tolerance to the male antigen H-Y antigen. The tolerized female mice then became tolerant of male grafts, while the control female mice rejected the male grafts.)

The key, at least for this particular protocol, seems to be to use very low dose antigen and “suboptimal” conditions (where “optimal” refers to conditions that drive conventional immune responses. The vocabulary of immune responses is really kind of misleading, because it’s focused on easily-measured responses like protection against viruses or graft rejection. Regulatory T cell responses are just as active, and probably are just about as common and important, but it’s hard to talk about them without giving the impression that they’re somehow passive, or abnormal, or defective).

One problem with moving this into the clinic is that you would need to know what the target antigen is, which in an outbred population like humans you do not know a priori. However, as bioinformatic and experimental techniques for identifying antigen peptides improve, it may become more practical to run this for patients before their transplants. The potential payoff would be very high, because you might be able to remove immunosuppression altogether:

If a procedure as simple as peptide infusion, which permits de novo induction of Tregs from mature T cells, prevents transplant rejection or GVHD, it could offer a realistic opportunity to induce tolerance to a variety of antigens such as allergens, transplantation antigens, and antigens causing autoimmunity while minimizing undesirable side effects often associated with general immunosuppression.

  1. Verginis, P., McLaughlin, K.A., Wucherpfennig, K.W., von Boehmer, H., Apostolou, I. (2008). Induction of antigen-specific regulatory T cells in wild-type mice: Visualization and targets of suppression. Proceedings of the National Academy of Sciences, 105(9), 3479-3484. DOI: 10.1073/pnas.0800149105[][]
  2. Kretschmer, K., Apostolou, I., Hawiger, D., Khazaie, K., Nussenzweig, M. C., and von Boehmer, H. (2005). Inducing and expanding regulatory T cell populations by foreign antigen. Nat Immunol 6, 1219-1227.
    Apostolou, I., and von Boehmer, H. (2004). In vivo instruction of suppressor commitment in naive T cells. J Exp Med 199, 1401-1408.[]
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. []
November 23rd, 2007

Niches and bone marrow transplants

thymocytesStuffed with duck as I am (we don’t do turkey for Thanksgiving in our house) I’m not up to a long post, but I thought a paper in the latest issue of Science was pretty cool. The paper is
Czechowicz, A., Kraft, D., Weissman, I. L., and Bhattacharya, D. (2007). Efficient Transplantation via Antibody-Based Clearance of Hematopoietic Stem Cell Niches. Science 318, 1296-1299. 1

Very briefly, they show that one of the obstacles to bone marrow grafts — even in the absence of host-versus-graft immunity — is that there are a limited number of niches for hematopoietic (bone marrow) stem cells. The native stem cells occupy those niches, so injecting in a donor’s stem cells is very inefficient; only a tiny number can find a home and supply new, desirable progeny. If I’m interpreting the data right, there seem to be only a few hundred open slots, out of maybe 25000 total slots, available for donor stem cells.

Blogging on Peer-Reviewed ResearchThey came up with a protocol that transiently and specifically eliminated host stem cells — opening up niches — before the graft, and the results were pretty dramatic. Without treatment, the chimerism rates (indicating efficiency of donor engraftment) was around 3%; with treatment, it was 90%. If this works in humans as it does in mice, it offers a much gentler alternative to the really brutal and toxic treatments that are usually necessary today.

So what does “niche” mean, in this context? Is it a physical slot into which the tab of a hematopoietic stem cell is tucked? Is it a conceptual niche, a constraint based on available levels of some soluble factor, or on rates of contact with some supporting cell type? I think all three are possible, and2 have parallels in other aspects of the immune system.

For example, growth in the thymus (the figure at top left) probably requires physical niches, cells into which developing thymocytes cuddle up and receive nourishment and advice as they mature. (See, especially, the videos taken by Bousso et al, 3 of thymocytes interacting with thymic stromal cells.) Although I admit I find that the most attractive concept, I don’t really have a good reason for it, and there are probably good examples of non-physical “niches” as well. Survival of naïve lymphocytes outside of the periphery requires intermittent contact with MHC class I molecules — potentially a limiting factor, if they have to compete with others of their kind. There are also several examples of regulation by limiting amounts of certain cytokines, such as IL-7. 4 Still, the various two-photon microscopy videos of in-situ interactions that have been coming out over the past few years have really made me appreciate the importance of physical location and direct interactions in the immune system, which might explain my bias toward physical niches.

  1. As an experiment in aggregation, I am including a second version of the reference here, thus: Czechowicz, A., Kraft, D., Weissman, I.L., Bhattacharya, D. (2007). Efficient Transplantation via Antibody-Based Clearance of Hematopoietic Stem Cell Niches. Science, 318, 1296-1299. DOI: 10.1126/science.1149726[]
  2. According to current understanding, anyway[]
  3. Bousso, P., Bhakta, N. R., Lewis, R. S., and Robey, E. (2002). Dynamics of thymocyte-stromal cell interactions visualized by two-photon microscopy. Science 296, 1876-1880. []
  4. Purton, J. F., Tan, J. T., Rubinstein, M. P., Kim, D. M., Sprent, J., and Surh, C. D. (2007). Antiviral CD4+ memory T cells are IL-15 dependent. J Exp Med 204, 951-961. []