Cell-therapy
Cell-therapy focuses on using the immune system to eliminate cancer cells. This can be done by using the patients own immune system/cells or using donor cells. An example of making use of a patients' own system is the development of a vaccine. The concept of vaccine development in cancer is comparable to vaccine in infectious diseases, where the immune system is stimulated to fight against invaders.
Making use of a donor immune system to fight cancer is possible because the immune system of the donor will recognize the patients cells - including cancer cells - as non-self, a reason to kill them. Both research lines and 2006 activities are described below.
Vaccine development
The focus of this research line is the development of dendritic cells (DC = antigen presenting cell). DC do have the capacity to present antigens to the helper and effector cells of the immune system. They are the central regulators of our immune system. In a normal immune response - like infections - DC pick up these pathogens. After activation of the DC they will migrate to lymfnodes where the immune system is activated.
It is possible to culture DC in the laboratory (in vitro) from peripheral blood monocytes and from bone marrow stem cells. Both techniques are available in our laboratory. The concept of the vaccine is to load DC with tumour-antigens (peptide, protein etc) and to inject the DC into the patient. We have selected Mucine-1 as tumour-antigen since it is present on several malignant cells, including breast cancer, lung cancer, ovarian cancer and M. Myeloma. We have realised loading of the DC with this mucine-1 making use of peptides and/or an adenoviral vector. This second method induces high expression of mucin-1 on the DC including a pattern that is similar to tumour cells (various glycosylation forms). This is an important step in inducing glycoform specific immune responses. It need to be proven however if expression at the membrane correlates to presentation (via HLA-molecules) to immune T cells.
An important aspect of the research is to select the best possible DC. Since DC vaccination in the clinic is not yet successful the method still has to be improved. One of the most challenging issues in DC preparation is how to make a DC that activates the immune system and can travel to the lymph node, as a DC does in a normal immune response. Our latest results - still to be improved - show that we have a DC that meet all the requirements.
In 2006 we also studied the presence of Mucine-1 on M. Myeloma, demonstrating that tumour specific patterns of Mucine-1 are also present on this tumour, making this disease a good candidate for immunotherapy. In contrast, this tumour specific pattern is not present on acute myeloid leukaemia. The fact that we might be able to use tumour specific Mucine-1 in M. Myeloma is challenging. Since Mucine-1 is thought to be a relevant structure for the interaction of malignant myeloma cells and bone marrow stromal cells, an interaction critical for tumour growth.
An analysis was performed on the presence of Mucine-1 in the thymus. Since tolerance is regulated in this organ it is relevant to know if the tumour specific patterns of Mucine-1 can be find in the thymus as well. This is not the case, making it more likely that an adequate immune response against these antigens can be realized. We could demonstrate that Mucine-1 is present in its full glycoform on thymic medullary epithelial cells, but not the tumour specific glycoforms. These medullary epithelials cells play a crucial role for tolerance induction (manuscript accepted in 2006).
Donor transplantations
Donor transplantations do have several limitations. One of them is the availability of donors. By using haploidentical donors (one chromosome with the relevant major histocompatibility antigens in common) nearly all patients will have a donor (figure). A second limitation is toxicity, that can be reduced by using a so called non-myeloablative (or "mini-") transplantations.
We used a combination of these two (mini-transplant and haploidentical donor) in a new mouse model. This transplant procedure is feasible and we could demonstrate anti-tumour activity against lymphoma and breast cancer.
Figure: Nearly every patient has a haploidentical donor (1 chromosome in common).
In the mouse model mice breast cancer and lymphoma could only be cured by using a donor transplantation. In case a syngeneic (syngeneic in inbred strains = autologous (self) in men) transplantation was performed, mice could never be cured. Since there is no immune effect in syngeneic transplantations , this proves that the immune system of the donor plays a crucial role in fighting the cancer!
Interestingly in haploidentical transplantations not only immune T cells might be relevant but also Natural Killer cells. We are presently analysing what cells do cause the anti-tumour effect in breast cancer.
In the clinic we finished a study on non-myeloablative transplantation in solid tumours. Though the method of non-myeloablative transplantation was successful with respect to donor engraftment there was still substantial toxicity. Unfortunately there was no long term effect in patients with solid tumours, including breast cancer.
Present research on transplantation program (started 2006)
One of the major limitations - in fact the only one - in clinical haploidentical transplantation is the high rate of infections, because of slow immune reconstitution after the conditioning and because of T cell depletion of the graft.
Most challenging would be if a transplant could be performed without a condition regimen, therefore preventing morbidity and mortality. Therefore we started a collaboration with the research group in Bergamo, Italy, to see if we can develop a cell based tolerance-inducing regimen in haploidentical transplantations. The scope of this program is to induce tolerance of the recipient against the donor to realise engraftment of the donor stem cells.
Also with the aim to restore immunity after transplantation we started a program on de novo T cell production from bone marrow stem cells (Funded by Senter-Novem; partners: Pharmacell, Riken Institute in Japan and Maia scientific in Belgium (2006-2009).
In this project we study if T cells with can be produced de novo, making use of bone marrow derived hematopoietic stem cells. For this an artificial thymus system is used. The expectation is that T cells can be made with antigen specific activity. These T cells could then be used for immune recovery after bone marrow stem cell transplantation. Also tumour specific T cells could be made.
Haploidentical tranplantations clinically
In 2005 and 2006 the first 3 haploidentical transplantations have been performed. Unfortunately, two of three patients died because of infections (in the published literature treatment related mortality is about 30 %). The anti-leukemia effect is however very good. None of the 3 patients showed any evidence of disease, and one patient seems to be cured more than one year after transplantation. This haploidentical transplantation program is based on the observation that Natural Killer Cells are relevant for the anti-tumour effects, especially in Acute Myeloid leukaemia. These cells seem to be far more effective than the immune T cells, that are thought to be the effector cells after HLA matched donor transplantation.
This observation induces a new paradigma in bone marrow stem cell transplantation and some haematologist expect that focusing on Natural Killer cell activity in donor stem cell transplantation will be the standard in a few years from now. This would mean for clinical practice that the now used HLA matched transplantation (only family transplants in Maastricht) will move to haploidentical transplantation. For patients this would mean a 2 fold higher change of finding an adequate donor in the family.
The haploidentical transplantation program is presently extended clinically for acute leukaemia and is in line with the preclinical research line. At present we do analyse if patients with other diseases, like multiple myeloma and breast cancer, might also profit from this new concept in cancer immunotherapy.
Selected publications
S Cloosen, EBM van Leeuwen, BLMG Senden-Gijsbers, EBH Oving, JW Gratama, WTV Germeraad, and GMJ Bos.
Different mucin-1 glycoforms are expressed on multiple myeloma and acute myeloid leukemia cells.
Br J Hematology 2006 135:513-516
VanClee Ariana, Michel van Gelder, Harry schouten, Gerard MJ Bos.
Murine mammary carcinoma can be cured in a model of non-myeloablative haploidentical stem cell stransplantation.
Bone Marrow Transplant. 2006 Jun;37(11):1043?9
EBM van Leeuwen, S Cloosen, BLMG Senden-Gijsbers, WTV Germeraad and GMJ Bos. Transduction with a fiber-modified adenoviral vector is superior to non-viral nucleofection to express tumour-associated antigen Mucin-1 in human dendritic cells.
Journal of Immunotherapy. Cytotherapy 2006 Feb; 8 (1): 24-35
EBM van Leeuwen, S Cloosen, BLMG Senden-Gijsbers, U Mandel, H Clausen, JJ García-Vallejo, WTV Germeraad and GMJ Bos.
Expression of aberrantly glycosylated tumour Mucin-1 on human dendritic cells after transduction with a fiber-modified adenoviral vector.
Cytotherapy 2006 Feb; 8 (1): 36-46
Dr. G. Bos, project leader
Prof.dr. H. Schouten
Dr. M. van Gelder
Dr. W. Germeraad
Dr. P. Hupperets
Post-doctoral fellows
Dr. A. Vanclee
Dr. S. Cloosen
Dr. B. Meek
Dr. J. Vanderlocht
PhD students
M. Huls
J. v.d. Elssen
P. Frijns
Technicians
B. Senden
Students
P. Frings
M. Head
J. Van Elssen
J. Bogie
K. Hoeben