The Impact of Antibody Developed for the Prevention of Acute GVHD upon B Lymphocyte Function - Research Proposal Example

Bone Marrow Transplant Team MASTERS STUDENT PROJECT PLAN Project Title: The impact of the anti-CD83 antibody developed for the prevention of acute GVHD upon B lymphocyte reconstitution and function. Supervisors: Dr Hannah Cullup Assoc/ Prof Alison Rice Hematopoietic Stem Cell Transplant Stem cell therapy is the use of stem cells to replace the damaged cells to produce new cells that can perform the same functions as the damaged ones. Many diseases can now be treated using stem cell therapy1. It is used to treat diseases such those that destroy the bone marrow and those within the soft tissues of the bone marrow such as multiple myeloma, leukemia and cancer patients who have undergone cancer treatments. Other conditions treated using stem cell transplantation are; aplastic anaemia that causes production of abnormal blood cells2,3,4 acute myeloblastic leukemia and malignant lymphoma, that is for auto-transplant treatment; and non-malignant disorders, chronic myeloblastic leukemia, lymphoid malignancies, dysmyelopoietic syndrome, lymphoblastic leukemia, acute myeloblastic leukemia and refractory to tyrosine kinase inhibitors5. Hematopoietic stem transplantation requires selection of an appropriate donor, conditioning which includes chemotherapy, immunosuppression and radiotherapy then post HSCT care. Donors can be HLA identical, mismatched related donors or mismatched unrelated donors which means HSCT can be allogeneic from a non-identical marrow, autologous from the patient’s own marrow and Syngeneic, from genetically identical marrow. It is evident that the type of transplant depends on the donor6,7,8 Harvesting of Stem Cells: There are three sources of stem cells. These are, the cord blood, peripheral blood and bone marrows. Stem cell harvesting in the case of allogeneic stem cell transplantation occurs after the patient has undergone the conditioning therapy while stem cell harvesting for autologous transplantation is done before the conditioning therapy. This is because the stem cells in autologous transplantation procedure are obtained from the patient while in allogeneic transplantation of stem cells; the stem cells are harvested from a different person (donor)9. This eliminates cord blood stem cells as a source of stem cells for autologous stem cell transplantation. Conditioning: The conditioning regimen aims at eradication of cells that have been infected by disease to ensure that the graft is not rejected by the recipient10,11. There are two types of conditioning done during hematopoietic stem cell transplantation process. Conditioning is either myeloablative (where the aim is to achieve high cytotoxicity against the malignant cells) or non-myeloblative. Myeloablative conditioning however leads to high systemic and hematopoietic toxicity. It is used in malignant conditions as well as non-malignant conditions that have high risk of rejection. The regimens used are those that incorporate Busulfan (Bu) or total body irradiation (TBI) given besides cyclophosphamide. Other high cytotoxic drugs such as melphalan are also sometimes used12,13,14 Non-myeloablative conditioning uses less toxic regimens compared to myeloablative conditioning. In this case, the aim is to reduce regimen related toxicity so that by applying less intensive conditioning (limiting inflammatory cytokine release or reducing direct toxicity). The process ensures immunosuppression which is the main aim of RIC and also killing the residual haematopoeitic system15,16 Allogeneic stem cell transplantation has some side effects which are; infections, impaired reproductive ability, cataracts, veno-occlusive disease, raft failure, lung damage, anemia, mucositis, bleeding and GVHD. GVHD has the highest rate of mortality compared to other effects and different methods have been used to eliminate this side effect. GVHD This is a complication that occurs when a donor’s T cells attack the host tissues after a bone marrow transplant. This condition was first discovered half a century ago when an experiment was conducted on irradiated mice. The mice were infused with allogeneic marrow and spleen cell. The mice recovered from the primary abnormality but died after some time of GVHD. At the time the symptoms of GVHD were; diarrhea, skin changes, weight loses and liver abnormalities17,18. It was also found that for this complication to occur; 1) the recipient must not have the capability of rejecting the transplanted cells 2) the graft must have immuno-competent cells 3) the host must express antigens that will be recognized by the transplanted graft cells as foreign hence initiating an immune reaction19,20. Under the condition that the graft must have immuno-competent cells, it is now known after scientific research that it is the mature T lymphocytes that are responsible for inducing GVHD. Further research on T cells indicates that the number of infused mature T lymphocytes corresponds to the severity of GVHD after an Allogeneic Hematopoietic Stem cell Transplant.21,22 There is a noticeable difference between GVHD incidence and HSC source, BM, PBSC and cord blood due to the presence and maturity of the T cells.23,24 The second condition for GVHD to occur is if the recipient is immunocompetent which means the patient’s immune system has to be suppressed. In most Haematopoietic Stem cell Transplantation (HSCT) cases, the patients have undergone some chemotherapy or radiation before stem cell infusion making their immune system immunosuppressed. In a normal immune system, the T cells from a foreign donor would be rejected. The third condition is when a recipient has some tissue antigens that the donor does not have. This is sometimes referred to as MHC differences between the donor and the recipient. MHC produces HLA (Human Leucocyte Antigens) as its gene products and these are expressed in every cell surface of a human being. They are also used in activation of T cells. T cells have their own specific MHC molecules so that when non self MHC molecules are recognized, the activation of T cells leads to destruction of allogeneic tissues. There are also polymorphic genes expression products such as minor histocompatibility antigens that can also cause the induction of GVHD25. Types of GVHD: There are two types of GVHD, one that occurs immediately after the transplant (within 100 days) known as acute GVHD and the other that occurs late after some time known as chronic26. Acute GVHD: The risk of this kind of GVHD occurring depends on the patient’s age and the donor’s age (increasing age increases the risk), on if the donor had been pregnant in the earlier period, when there is less exact tissue match and when reduced doses of immunosuppressive drugs are used after the transplant27. Symptoms: Acute GVHD mostly involves the liver, the skin and the intestines. The patient experiences skin rash in various parts of the body, which may become severe and cause peeling of the skin or formation of blisters. The patient also experiences cramping in the abdomen, blood in the stools or diarrhoea, liver infection results to jaundice28 (). Chronic GVHD: This kind of GVHD presents itself in so many ways. It affects the skin in most cases but also affects the liver, the lungs, and the immune system sometimes. The difference as noted earlier is in the time of appearance. Chronic GVHD can appear three months to six months after the immune-suppressive post transplant treatment. It occurrence is also affected by age just like acute GVHD, infection by the Herpes Zoster Virus (HZV), preceding acute GVHD and donor lymphocyte infusion. 29 GVHD Therapies that have been used Preventing GVHD requires determination of a well matched donor (HLA identical), but this requirement limits the number of donors available for patients. According to Cant, Galloway and Jackson, only 25-30% of the UK population has full HLA-matched donors30. It is therefore essential to find other donors but be able to prevent the occurrence of GVHD or its severance. If a fully matched donor cannot be found, then donors that have a few mismatches at the HLA loci can be used, but the doctors know this gives a greater risk of developing GVHD- therefore they automatically would increase the level of immunosuppression given to the patient to prevent severe Acute GVHD from occurring. There are three methods that have been used to ensure prevention of GVHD and its severity which are; pre and peri-HSCT Serotherapy, In-vitro Tcell depletion and Post HSCT Immunosuppression31. In Vitro T cell Depletion: In acute GVHD, the T lymphocytes act against the recipient’s antigens expressed in the host’s tissues. This can be prevented by serotherapy which involves the use of antibodies such as ATG (Anti-Thymocyte Globulin) or Campath (alemtuzumab) against the lymphocytes to reduce the number of allospecific T-cells32. T cells are involved in the reaction against the recipient’s tissue cells and a reduction in these T cells can reduce the severity of or occurrence of GVHD. The sources of the stem cells such as the bone marrow and PBSC are depleted of T cells through in vitro means. This can be done by immunological or physical techniques33. The physical techniques include; Counterflow centrifugal clutriation Sheep red blood cell resetting Fractionation on a density gradient Immunological techniques include; Immuno-magnetic negative selection, for example of CD3-positive cells, to remove T cells Antibody mediated purging, for example the use of Campath 1M, activated through complement Immuno-magnetic positive selection for example of CD 34-positive cells, to give a stem cell population34. This type of therapy however has another problem. Depletion of T-cells have been found to weakens problem the graft-versus-leukaemia (GVL) effect and relapse may occur. Post HSCT Immunosuppression: This includes the use of drugs such as methotraxate, ciclosporin A, Corticosteroids such as methylprednisolone, tacrolimus (FK 506) sometimes used as an alternative to ciclosporin and Mycophenolate mofetil (MMF) sometimes used in combination with ciclosporin. The most commonly used drug is ciclosporin which is sometimes combined with methotraxate during the first 6-18 days of post HSCT35. Novel GVHD therapies – anti-CD83 Several approaches to prevention of GVHD have been developed but most of them target T cells. This has proven inefficient since T cells are important in the engrafting process (they interfere with the post transplant immunity of the patient)36. Other methods have been researched and recently, a test of “CD83 antibody in the human T cell–dependent peripheral blood mononuclear cell transplanted SCID (hu-SCID) mouse model of GVHD”37 showed that CD83 antibody is an effective treatment of GVHD without compromising the patients’ immunity after a transplant38. Wilson et al found out that activated Dentritic cell depleting CD83 antibody mixed with allo lymphocyte cultures suppressed alloproliferation of the T cells without reducing the number of T cells plus those of CMV. CD83 antibody was also found not to prevent engraftment of human T cells CTL which are responsive to malignant cells and viruses included39. Breloer also indicated that CD83 is a regulator of peripheral immune response and central maturation of T cells40. B Cell Development The development of B cells begins in the liver and goes on to the bone marrow where they are produced throughout an individual’s life. Stromal cells of the bone marrow send signals to the lymphoid progenitor cells to initiate B cell development. CD34+ lymphoid progenitor also synthesize TdT and RAG-2 & RAG-1 in response to cytokines induction. The cells form early pro-B cells by joining the D-Js of the H chain chromosome and a lso start expressing class II MHC and CD45 (B220). The next step in this process is joining of the already formed D-JH chain to a V segment which forms late pro-B cell stage. The cells become pre B cells which then develop to mature B cells when m chains with surrogate light chains in the pre-B- receptor are expressed on their surface41. CD83 and B Cells CD83 is considered an immune system regulator. It has so many functions in the immune system. In B cells, CD83 activates peripheral B cells and is involved in activation of antigen-specific humoral immune responses. According to Fujimoto & Tedder, deficiency in CD83 leads to an inhibition of antigen specific humoral immune responses and reduces the activation of peripheral B cells42. The antigen however does not affect any inherent proliferative function of B cells. Kretschmer also showed in a study that CD83 antibody modulates the function of B cells43. CD83 and Its Relationship to the Project The aim of this project is to find out the impact of CD83 antibodies on B lymphocytes. Studies on prevention of GVHD have improved to a level that strategies dealing with inactivation of host-antigen presenting cells are now being investigated. A study still under investigation by Shlomchik showed that even though after a transplant, the donor antigen-presenting cells are also available; it is always the host’s antigen-presenting cells that initiate a graft Vs host disease44. From this finding, a proposal was made that strategies which focus on inactivation of the host antigen-presenting cells be used in studying prevention and treatment of GVHD. One such strategy is the use of CD83 antibodies.45. CD83 antibodies in treatment of GVHD have been found to be an effective treatment of control of GVHD as tested in hu-SCID mouse model of GVHD46. It is however expressed in so many cells from DCs, CD4+ blasting T cells, B lymphocytes and monocytes. Recent studies have found out the effect of CD83 antibodies on Dendritic Cells and also found its effectiveness because of its large expression in DCs cell surfaces, but no studies have shown the effect of CD83 antibodies on other cells that express CD83. It is for this reason that the study of the impact of CD83 antibodies on B Lymphocytes is proposed47. Project Aims: 1) To fully characterize CD83 expression on the B lymphocyte lineage: bone marrow, spleen and lymph node resident B cells, circulating B lymphocytes and resting versus activated B lymphocytes. 2) To determine whether CD83 antibody suppresses B cell proliferation in vitro. 3) To determine whether CD83 antibody administration affects B lymphocyte specific antibody response to Thymus dependent and independent mitogens in vivo. 4) To show that CD83 antibody administration does not affect B lymphocyte lineage reconstitution in mice following myeloablative conditioning and syngeneic (Balb/c → Balb/c and C57BL6 → C57BL6) transplant. 5) To show that CD83 antibody administration does not affect B lymphocyte lineage reconstitution in mice following myeloablative conditioning and full MHC mismatch (BL6 → Balb/c) transplant. Bibliography American Cancer Association (ACA), Types of Stem Cell Transplants, 2009. Retrieved on 22nd, March 2010 from: 05/27/2009http://www.cancer.org/docroot/ETO/content/ETO_1_4X_Stem_Cell_Transplant_Basics.asp. Breloer Minka, CD83: Regulator of Central T Cell Maturation and Peripheral Immune Response, Immunology Letters, 115(1): January 2008, 16-17. Campbell Ken, Bone Marrow and Stem Cell Transplantation, Stem Cell Research, 2006, 1-30. Retrieved on 20th, March, 2010 from: http://www.ebmt.org/10.1Patients&Families/Patient&DonorMaterials/10.%20LRF_BMT07_4687.pdf. Cant Andrew J., Galloway Angela and Jackson Graham, Practical Hematopoietic Stem Cell Transplantation, 2007, Oxford, England: Wiley-Blackwell. DBA Fact Sheet, Stem Cell (Bone Marrow) Transplant: Information for people with Diamond Blackfan Anemia and their Families, 1-3. Retrieved on 20th, March 2010 from: http://www.cdc.gov/ncbddd/hbd/dba/documents/508%20DBA%20Stem%20Cell%20Transplant%20Fact%20Sheet.pdf. Ezzone Susan and Schmit-Pokorny Kim, Blood and Marrow Stem Cell Transplantation: Principles, Practice, and Nursing Insights, 2006, Sudbury, Massachusetts: Jones & Bartlett Publishers, 329-410. Fairview Health Services, Blood and Marrow Transplantation: A Patient's Guide to Hematopoietic Stem Cell Transplantation, 2005, Minneapolis, Minnesota: Fairview Press, 56-61. Ferrara, James L. M., Cooke, Kenneth R. and Deeg Hans Joachim, Graft-vs.-Host Disease, 3rd Ed., 2004, Informa Health Care. Fujimoto Yoko and Tedder Thomas F., CD83: A Regulatory Molecule of the Immune System with Great Potential for Therapeutic Application. Journal of Med Dent Sci, 2006, 53: 85-91. Hołowiecki Jerzy, Indications for Hematopoietic Stem Cell Transplantation, Abstract, Polskie Archiwum Medycyny Wewnetrznej, 2008, 118(11), 658-663. Janeway Charles and Travers Paul, Immunobiology: The Immune System in Health and Disease, 2nd Ed., 1996, London, UK: Current Biology, 78-79. Kretschmer Birte, Engagement of CD83 on B Cells Modulates B Cell Function In Vivo, The Journal of immunology, American Association of Immunologists. 2009, 182 (5): 2827-2834. http://cat.inist.fr/?aModele=afficheN&cpsidt=21228469. Lazarus Hillard and Laughlin Mary, Allogeneic Stem Cell Transplantation, 2nd Ed., 2009, Massachusets, Boston: Springer, 450-502. Lüthje Katja, Kretschmer Birte, Fleischer Bernhard and Breloer Minka, CD83 Regulates Splenic B Cell Maturation and Peripheral B Cell Homeostasis, International Immunology, 2008 20(8):949-960. http://intimm.oxfordjournals.org/cgi/content/abstract/20/8/949. Neumann Karl B., Hematopoietic Stem Cell Transplantation Research Advances, 2008, Toronto, Canada; Nova Publishers, 120-129. Powell Jonathan L, Hematopoietic Stem Cell Transplantation, 2009, Retrieved on 22nd, March, 2010 from: http://emedicine.medscape.com/article/991032-overview. Shlomchik, Warren D., Couzens Matthew S., Tang Cheng Bi, McNiff, Jennifer, Robert Marie E., Liu Jinli, Shlomchik Mark J., Emerson Stephen G. Prevention of Graft Versus Host Disease by Inactivation of Host Antigen-Presenting Cells, Science 16 July 1999: 285(5426), 412 – 415. Retrieved on 20th, March, 2010 from: http://www.sciencemag.org/cgi/content/abstract/285/5426/412?maxtoshow=&HITS=10&hits=10&RESULTFORMAT=&fulltext=GVHD+APC&searchid=1136515408707_19296&FIRSTINDEX=0&journalcode=sci. Soiffer Robert J., Hematopoietic Stem Cell Transplantation, 2nd Ed., 2008, New York, US: Springer, 178-208. Soiffer, Robert J. Stem Cell transplantation for Hematologic Malignancies Contemporary Hematology,2004, Sydney Australia: Humana Press, 320-325. The University of Arizona, Immunology Tutorials: B Cells,2007, Retrieved on 28th, March, 2010 from: http://microvet.arizona.edu/Courses/MIC419/Tutorials/Bcelldevelopment.html Wilson John, Cullup Hannah, Lourie Rohan, Sheng Yonghua, Palkova Anna, Radford Kristen J., Dickinson Anne M., Rice1 Alison M., Hart Derek N.J. and Munster David J., Antibody to the dendritic cell surface activation antigen CD83 prevents acute graft-versus-host disease, The Journal of Experimental Medicine, 2009, Vol. 206, No. 2, 387-398. Retrieved on 20th, March, 2010 from: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2646577/pdf/JEM_20070723.pdf. Read More