Fact File

“Minitransplants”

Reduced Intensity Stem Cell Transplants

(RIST) may offer hope for more patients

 Chris Poynton first became interested in transplants and immunology when working in Cambridge with Professor Roy Calne during the early days of liver and heart transplants. He went on to train in haematology in Cambridge and subsequently spent three years at MD Anderson Hospital in Houston, Texas, where he developed a magnetic bone marrow purging system for leukaemia in the monoclonal antibody laboratory with Dr. Chris Reading. Following his return to the UK, he was appointed leukaemia research fellow in London with Professor John Barrett before obtaining his consultancy at the University Hospital of Wales in Cardiff. His main clinical interests are in lymphoma treatment and immunology - both diagnostic and therapeutic.

Introduction

Reduced Intensity Stem cell Transplants (RIST) or minitransplants are a relatively new development designed to make a bone marrow transplant from a donor safer, more tolerable and more efficacious. These goals have not been easy to achieve because of transplant-related problems such as graft-versus-host disease, poor immunity resulting from the ‘conditioning’ high dose treatment regimens, and short survival in older patients.  Intense research is taking place throughout the transplant world to achieve these three objectives, so perhaps it is little surprise that each centre seems to devise a different name for essentially the same procedure.  Thus we have terms such as ‘minigraft’, ‘nonmyeloablative stem cell transplant’ (NMSCT), ‘reduced intensity stem cell transplant’ (RIST) and others, to distinguish the procedure from a conventional stem cell transplant (CST). RIST appears to be the current winner.

Immune system - the major players...

Before we discuss the principles behind reduced intensity stem cell transplants, we need briefly to review the members of the immunological ‘orchestra’ and their roles in fighting cancer and each other.  All these cells are ultimately derived from the bone marrow stem cells, but are schooled in different places around the body to become useful members of the immune system.  There are four major players:

·      T-cells that recognise foreign matter as well as whole cells and produce toxins to kill them 

·      B-cells that, under instruction from T-cells, produce antibodies against foreign material and attract other elements of the immune system to join the fight

·      macrophages, monocytes, dendritic cells and the other types of antigen-presenting cells  that start a specific immune response rolling by recognising what is self and non-self

·      natural killer cells that can recognise tumours directly. 

T-cells go on to develop in the thymus, skin and outer areas of the lymph nodes, whereas B-cells develop in the centre of the lymph nodes and are released to grow up 'on the streets'.  Natural killer cells seem to be released directly into the blood stream from bone marrow and are widely dispersed.

It is important to realise that T-cells will only recognize foreign material if correctly introduced first by the antigen-presenting cells. When this has been done properly they will communicate their information to other cells to recruit help in fighting the intruder.  It is the tissue type molecules or human leucocyte antigens (HLA) in the macrophages, monocytes, dendritic cells and some B-cells (antigen-presenting cells) which process the invader and present it for recognition by a roaming T-cell.

Our bodies maintain a huge and changing repertoire of T-cells, each one uniquely able to recognise a tiny but unique bit from the invaders.  Having made this recognition, T-cells ‘talk’ to B-cells and the B-cells attempt to make antibodies that also recognise the invader.  However, the immune system must not let the response get out of hand, so all sorts of regulatory systems come into play.  Whilst this is highly effective team work against bacteria and other germs, cancer cells are sometimes too clever for the immune system.

There are, in fact, many different ways that the successful tumour cell uses to evade the immune response; it is now clear that we do actually recognize most malignant cells with our own T-cells, yet the immune system itself prevents that response from becoming effective - it has been vetoed. 

Another sort of lymphocyte is also very important in fighting cancer - the natural killer or NK cell.  This is a very special cell that can recognize tumours directly without having antigen presented by a special cell.  Unfortunately, these too can be switched off by malignant cells and become inactive. Waves of new NK cells emerge very early after a transplant (even an autologous transplant) and these sometimes overcome the tumour for a while.

How did ‘minitransplants’ come about? 

Growing cells of different immune systems together was originally devised as a rather sophisticated model in the laboratory to study how immune systems either react positively or negatively to different antigen challenges. These studies demonstrate the phenomenon of immune tolerance.  This search has fascinated immunologists for many years and has come to be something of a holy grail for them.  There are in fact all sorts of reasons to want to be able to manipulate immune responses to specific antigenic stimuli, some of which have nothing to do with transplantation.  The treatment of autoimmune disorders, where the patient's immune system reacts against self-antigens, would be one important example. 

In the transplant setting, reactivity towards a different immune system is known as alloreactivity and the antigenic challenges are the tissue type molecules themselves – the human leucocyte antigens (HLA). It should be explained straight away that, unless transplanting from a genetically identical twin (or using an individual’s own cells), there are always HLA differences, although in a ‘matched’ sibling these will be at multiple minor sites and not the major sites. This reaction is responsible for the graft-versus-host disease that is seen, sometimes quite severely after transplant. Alloreactivity of the host cells against the incoming donor cells can also result in rejection of the marrow with consequent marrow failure.

Some history...

In conventional stem cell transplants, performing an immunological balancing act between donor driven graft-versus-host disease and host driven rejection (both alloreactive responses), yet still allowing an anti-tumour response - graft-versus-tumour reaction (see more about this below) has been a real challenge.  A breakthrough came when scientists in Boston in the late 1980's (David Sachs and Megan Sykes) found they could induce tolerance after transplant by mixing host and donor grafts together, and in some cases doing an autologous transplant followed by an allogeneic transplant a week or more later. They found that certain host cells were able to seek out and destroy the alloreactive T-cells of the donor that were responsible for initiating graft-versus-host disease. The action of these cells is now known as veto activity and refers to not just one type of T-cell but a group of different types of cells all possessing veto activity (including some stem cells).  These cells switch off the T-cells causing graft-versus-host disease. The beauty of their work was that this did not prevent the development of an otherwise perfectly functioning new graft and immune system – and there was no graft-versus-host-disease.

This work was taken further by several other groups and eventually developed into the clinical application we know as minigrafting by Professor Shimon Slavin and others.  It swiftly became apparent that some of the old notions that we had in transplantation were incorrect.  There was no need to create space for a new bone marrow by eliminating all traces of the old one.  Not only that, but retention of some parts of the patient's old (host) immune system could actively aid in the induction of tolerance if left behind and allowed to proliferate. If we didn’t have to get rid of the patient’s marrow altogether we could theoretically reduce the intensity of the preparative regimen that is given before transplant.  So now there was finally an opportunity to do away with the rather toxic ablative chemotherapy and total body radiotherapy that is often used in CST. This advantage has come to be viewed as a major advance in extending transplants to older patients.

Why do donor transplants kill tumour?

Since the mechanisms used by malignant cells exploit the body’s own HLA recognition mechanisms, one very successful way of getting T-cells to kill the tumour is to use someone else’s T-cells.  The problem is that, if the HLA differences are too great, then too many of the  incoming donor T-cells are alloreactive (since they come from a random population of circulating cells from a donor) and begin to attack the body’s normal cells as well - the phenomenon of graft-versus-host disease. 

A balance had been struck in conventional stem cell transplants when using fully HLA matched donors (generally siblings).  In this situation there are enough differences in the HLA system that the malignant cell is often unable to disarm the donor T-cell in the same way as it can one of its 'own' T-cells.  This is all fine but the problem arises that for some (as yet unknown) reason we all maintain a surprisingly large number of T-cells whose function is to recognise HLA (major and minor) differences.  Why we would need these at all is not quite clear. 

In the early 1980's when the major threat to transplant success was graft-versus-host disease and its consequences, we tried removing all the T-cells from the donor allograft (the major thrust of research at that time) but it soon became clear that we were also removing the graft’s anti-tumour effect.  The result was relapse, as the malignant cells just came back at a higher rate. In addition, the body’s residual immune system was then able to reject the graft more easily.  What we need to be able to do is just remove the alloreactive T-cells (against the HLA differences) and still infuse the T-cells that recognise the malignant cells, since the patient’s own have been switched off by the malignant cells themselves.

Until very recently the techniques enabling us to look at an individual T-cell recognition apparatus have not been nearly sensitive enough, and we still have a long way to go in this type of research. Besides which, we still do not really understand which cells are needed to achieve specific tolerance through this veto mechanism.

Tolerance induction - the key to success...

What Professor Slavin and others have been very interested in for a number of years, is how to induce a specific tolerance in the immune system.  If we could just specifically delete recognition of foreign HLA molecules and leave all the other T-cells in the population alone, we could end up just recognising and destroying the lymphoma.

The separate, but important issue, in a conventional stem cell transplant is the toxicity from the heavy 'conditioning' regimen of radiotherapy and/or chemotherapy.  Not only does this cause damage to the natural defences of the body (eg gut, lung and skin) to infection, but these infections cause the release of all sorts of inflammatory molecules that disturb the delicate balance between host and donor and promote graft-versus-host disease.  Viruses such as cytomegalovirus (CMV) are particularly troublesome in disturbing this delicate balance.  Reducing the intensity of the pre-transplant preparation helps in this regard as well.

So what are we left with as the central problem?  We need to solve how to reduce the intensity of conditioning therapy to allow the host and donor marrows to develop together for long enough to induce the alloreactive veto effect, without one rejecting the other.

How do we prepare a patient for a RIST?

This breakthrough really came with the introduction of the drug fludarabine.  This drug was originally developed as a chemotherapy for chronic lymphocytic leukaemia and lymphoma but, as had been observed in clinical trials, it was also highly immunosuppressive, particularly destroying certain T-cells.  Since these sorts of T-cells give a lot of help to the development of alloreactivity, suppressing them by giving fludarabine to the patient prior to transplant prevents the host rejecting the donor marrow.  As a result, the donor and host marrow can live together, veto each other’s alloreactivity and preserve activity against the malignant cells.

Well, that was the intention anyway.  A variety of combinations of antibodies and chemotherapies (almost always including fludarabine) have now been tried.  Without doubt the intensity of the regime has been lower (hence “RIST”) and this has been a major benefit in allowing patients of an older age group to have minitransplants successfully. 

However, it has already become clear that we have not conquered the alloreactivity problem since graft-versus-host disease still occurs with an unacceptably high frequency (especially chronic graft-versus-host disease).

Some of the problem may relate to the dominance in most minitransplants of donor cells over host cells, so that at day 30 after the re-infusion of stem cells, no host marrow has survived (in most reports). In the original model, central to the success of a RIST was the preservation of some host immunity. So far it has been difficult to individualise the balance for each patient when using a standard regime.

New models of tolerance are now focussing upon precisely what elements of an immune system are able to act in a veto capacity.  It now appears that the stem cells themselves have powerful veto activity. It now appears that the stem cells themselves have powerful veto activity. Early clinical results suggest that by giving huge numbers of donor stem cells to a patient it is possible to reduce the intensity of the preparative regime even further without the danger of rejection.  This added veto activity in the graft then allows tolerance to occur even faster and thus hopefully more successfully.  Naturally we shall eagerly await a new term to be applied to these new high dose stem cell grafts!

The final step in the model...

Tolerance, having been successfully induced, is generally long lasting.  We can then move to a second step in fighting a patient’s malignancy, the use of donor lymphocyte infusions.  Currently infusing large numbers of donor T-cells into a patient after transplant results in severe graft-versus-host disease because within this huge repertoire of T-cells will be graft-versus-host disease causing cells that will survive.  However, in a patient who has developed long-term tolerance, these donor T-cells that cause graft-versus-host disease would be immediately vetoed out, yet the T-cells with anti-tumour activity in the repertoire would survive.

It all sounds too good to be true and as we have already seen there are lots of pitfalls.  

As yet no clear guidelines have emerged on how to condition a patient for a minitransplant, how many stem cells to give, how many T-cells to give and how to manage the donor host balance afterwards.

There is much work still to be done to exploit this new technique to the full, but visible progress is now taking place, progress that we hope will not only improve the prospects for lymphoma patients but from which we may learn a lot about how lymphomas interact with the immune response as well.

Glossary

Allogeneic describes a transplant using someone else’s tissue or organ. Hence an allograft is a transplant using matched donated tissue.

Alloreactivity the ability of an individual’s immune system to recognise a genetically different individual from the same species – in this case human. This is done through major (HLA) and minor (lots of them) tissue typing molecules uniquely present in an individual.

Antigen challenges the sort of challenges against foreign organisms the immune system of the body recognises and responds to every day.

Autologous use of a person’s own tissue (for example stem cells or bone marrow).

Graft-versus-host disease the process by which T-cells from the donor attack the body’s normal cells.

Immune tolerance the process that prevents us from making an immune response against ourselves (self-tolerance - which is programmed in foetal life).  We can also sometimes acquire tolerance against other selected antigens/organisms/cells but as yet we don't know how to reproducibly switch this on and off.