Immunology and Stem Cells Laboratory

“Combining immunology and stem cells to rejuvenate and re-program the immune system for better health”

Group Leaders

Postal
Level 3, STRIP 1 – Building 75
Monash University
Clayton VIC 3800
Australia

Laboratory Members

Management

Mr Mark Malin
Ms Luciana Thompson
Ms Samantha Harris

Research Fellows

Dr Tracy Heng
Dr Natalie Seach

Research Assistants

Ms Maree Hammett
Ms Lisa Spyroglou
Ms Jade Barbuto
Ms Jade Homann

PhD Students

Ms Chew-Li Soh
Mrs Joanna Lim
Mr Marco Barsanti
Ms Danika Khong
Ms Jessica Morison
Ms Adrienne Calder
Ms Kahlia Wong
Mr Anthony Park
Mr Michael Hun
Mr Abdulaziz Alsharif

Research Overview

The Immune regeneration laboratory focuses on the:
• Mechanisms of development and function of the thymus
• Nature of the stromal cells which form specific "niches" for developing cells in the thymus and bone marrow
• Use of stem cells to create the thymus
• Use of donor haemopoietic stem cells to induce transplantation tolerance
• Reversal of age- and chemotherapy induced thymic and bone marrow atrophy
• Thymic rejuvenation to boost immune responses to vaccines and cancer
• Treatment of autoimmune disease by re-educating the immune system
• Use of mesenchymal and other stem cells to suppress aberrant immune responses
• Molecular basis to thymus and bone marrow regeneration and function
• Gene Discovery Program" to identify new therapeutic approaches
• Clinical translation of the research into patients with eg cancer, severe immunodeficiencies (eg HIV AIDS), autoimmunity or foreign transplants

An enigmatic feature of the immune system is that the thymus undergoes profound degeneration from puberty. A loss in the generation of new naive T cells means that the TCR repertoire becomes progressively restricted with age and this is associated with a skewing towards memory cells. This in turn translates to declining immune responsiveness to neoantigens in adults and the aged. Despite the broad acceptance of thymic atrophy and associated severe clinical conditions, there have been remarkably few attempts to reverse this process and none that have been applied clinically (with any success). Our approach to this problem over many years has been to understand how the thymus functions, through exhaustive research on the stromal cells that constitute the specific inductive microenvironment/niche of this organ. We have developed invaluable reagents and techniques to study these complex stromal cells and as a consequence have been able to map its development through ontogeny and its response to physiological signals throughout life. Using our unique technologies we also evaluated the aged thymus and established that although severely atrophic and infiltrated with adipose cells, all the thymocytes and stromal cell subsets are present but the architectural structure is severely disrupted.

Our research programs investigate ways in which the immune system, particularly the thymus, but also more recently the bone marrow, can be functionally restored following their destruction by aging, chemotherapy, radiotherapy and corticosteroids. We have two main approaches to this: utilising the body's own rules to reverse the aging process, and using stem cell research to build a new thymus. We have shown that inhibition of the sex steroids responsible for inducing thymic atrophy induces a dramatic reversal of its degenerative state, fully restoring it to its youthful maximum potential. This occurs equally in males and females. The reduction in sex steroids can be achieved surgically but more practically chemically (reversible) through the hormone Lutenising Hormone Releasing Hormone (LHRH, also known as GnRH). There are several models of immune function being evaluated in the laboratory: response to influenza and hepatitis immunisations, challenge with cancer and chemotherapy/BMT, and treatment of autoimmune disease. The data on sex steroid blockage through LHRH, have collectively resulted in Phase II clinical trials being performed to restore immune capacity in leukaemia patients receiving bone marrow or HSC transplantation. We were invited to extend this trial in the USA as part of the American Society of BMT Clinical Trials Network and with NIH funding through a PO1 grant for a consortium of clinicians headed by Dr. Lee Nadler (Dana Faber Cancer Centre, Boston) at three of the most prominent centres: Dana Faber, MD Anderson, and University of Minnesota Centre for Transplantation. The protocol has received FDA approval. We also have a preclinical large animal study on the induction of transplantation tolerance using thymic regrowth technology combined with donor HSC, with Dr. David Sachs at MGH, Boston.

We are also using sophisticated molecular expression profiling to investigate the factors that cause thymic (and bone marrow) re-growth, which may eventually lead to new therapies. Futhermore, we have identified a population of thymic progenitor epithelial cells with our mAb MTS 24, with the capacity to induce the formation of an entire functioning thymus. We have more recently found this antibody to identify a population of epidermal stem cells in the skin.

Rebuilding the immune system should allow much better defence against infections and cancer, and better responses to vaccines. Importantly by manipulating the restoration of thymic function we can also "educate" the immune system to prevent rejection of foreign transplants - including new stem cell-derived cellular therapies. Re-educating the immune system using a combination of thymus rejuvenation and uptake of HSC together with inhibition of the aberrant immune system, also provides for the first time, a realistic means of overcoming the devastating effects of autoimmune disease.

The research from the laboratory has resulted in a suite of Patents. These have been licensed to a Melbourne based biotech company Norwood Immunology, which was successfully listed on the UK Alternative Investment stock Market (AIM) in 2004. Monash University is an Equity holder in Norwood Immunology. Norwood Immunology provides a large part of the funding for the laboratory and facilitates the clinical translation of the resulting technology.

The laboratory has also formed a strong alliance with the Australian Stem Cell Centre (Boyd directs the Immunology Platform) and receives substantial funding for its program on immune tolerance, which has an important application in enabling patients to accept foreign stem cell-based grafts. We, together with Professors Alan Trounson, Claude Bernard and Ban Hock, were awarded a ~$5million, 5 year NH&MRC program grant on "Innovative stem cell based strategies to establish immune tolerance and tissue repair", from 2007 - 2011.

Thymic and bone marrow niche in aging and regeneration - a molecular analysis

Thymic stromal cells form the complex microenvironment with which bone marrow derived haematopoietic precursors interact as they differentiate and mature, migrating in a directed fashion between niches. Thymic epithelial cells (TECs) form an essential component of the thymic stroma. Once thought to be a static population, TECs have been found by our laboratory to be a dynamic population comprised of precursor cells, transit amplifying cells and terminally differentiated cells. These populations change with age - primarily due to influences from the neuroendocrine system, leading to massive thymic involution from puberty to only about 1-5% of its full capacity. The bone marrow niche also changes with age, likely influencing the numbers and lineage potential of haematopoietic stem cells. By removing the influence of the neuroendocrine axis, for example by sex steroid ablation (surgical or reversible chemical), we are able to regenerate the immune system, both at the level of the bone marrow and the thymus. The involuted thymus returns to its young potential, seeding the periphery with a new repertoire of mature, functional T cells.

The main focus of our research in the Gene Discovery Program is to investigate the mechanisms of aging and regeneration in the thymus and bone marrow niche, including the factors influencing the migration of haematopoietic precursors into the thymus. We are also investigating the influence of sex steroid ablation in niche ‘protection’ and regeneration after chemotherapy. We predominantly use flow cytometry and immunohistology techniques to investigate cellular changes in sub-populations of stromal cells, such as epithelial cells (cortical, medullary), non-epithelial stromal cells (endothelium, fibroblasts) and extracellular matrix from the thymus, and endosteal and vascular niche stromal cells from bone marrow (osteoblasts, osteoclasts, fibroblasts, adipocytes, endothelial cells, etc). To analyse the molecular changes in purified populations of stromal cells, we use real-time PCR and microarrays and proteomics techniques to analyse serum proteins.

Our laboratory has previously produced an antibody (MTS24) that identifies a thymic epithelial precursor. When isolated, reaggregated and grafted under the kidney capsule, as few as 2,500 cells from the E15 embryonic thymus, can form a fully functional thymic organ that attracts haematopoietic precursors from the bone marrow and supports full T cell development. These cells do not appear to have the same capacity in the adult. We are thus further defining the precursor ability of MTS24+ cells at different stages in the embryo and adult and investigating whether growth factors provided by embryonic neural crest derived mesenchyme or the use of 3D-biomatrices, can assist in the survival, differentiation and proliferation of the epithelial precursors in the adult. We are also investigating differentially expressed molecules in embryonic thymic precursor cells, compared to non-precursor cells, to identify molecules specific to epithelial progenitor capacity.

Selected Publications

Adrienne E. Calder, Melanie N. Hince, Jarrod A. Dudakov, Ann P. Chidgey and Richard L. Boyd. Thymic involution: where endocrinology meets immunology. Neuroimmunoendocrinology (2011, in press)

Anne L. Fletcher1,2, Adrienne Calder1, Melanie N. Hince1, Richard L. Boyd and Ann P. Chidgey (2011) The contribution of thymic stromal abnormalities to autoimmune disease. Critical Reviews in Immunology 31(3): 171-187

Morison, J., Heng, T., Chidgey, A., Boyd, R. (2011) The Immunogenicity of stem cells and thymus-based strategies to minimise immune rejection. In, Dr P. Fairchild (ed) Immunological Barriers to Regenerative Medicine, Springer Verlag, UK.

Boyd, N.R., Boyd, R.L, Simon, G.P. and Nisbet, D.R. (2011) Synthetic multi-level matrices for bone regeneration. In: "Tissue Engineering in Regenerative Medicine", Springer, (in Press)

Chen, X.T., Chan, S.T., Hosseini, H., Layton, D., Boyd, R.L., Alderuccio, F., Toh, B.H., and Chan, J. (2011) Transplantation of retrovirally transduced bone marrow prevents autoimmune disease in aged mice by peripheral tolerance mechanisms. Autoimmunity (in press)

Hirakata, A., Okumi, M., Griesemer, A.D., Shimizu, A., Nobori, S., Tena, A., Moran, S., Arn, S., Boyd, R.L., Sachs D.H., and Yamada, K. (2010) Reversal of age-related thymic involution by an LHRH agonist in miniature swine. Transplant Immunology.24(1): 76-81

Heng, T.S.P, Chidgey, A.P., and Boyd, R.L., (2010) Getting back at nature: understanding thymic development and overcoming its atrophy. Curr Opin Pharmacol, 10: 1-9

Dudakov, J.A., Khong, D.M.P., Boyd, R.L., and Chidgey, A.P. (2010) Feeding the fire: the role of defective bone marrow function in exacerbating thymic involution. Trends Immunol, 31(5):191-198.

Fletcher, A.L., Lukacs-Kornek, V., Reynoso, E.D., Pinner, S.E., Bellemare-Pelletier, A., Curry, M.S., Collier, A., Boyd, R.L., and Turley, S.J. (2010) Lymph Node Fibroblastic Reticular Cells Directly Present Peripheral Tissue Antigen Under Steady-State and Inflammatory Conditions. J Exp Med, 207(4): 689-697.

van Dommelen S.L., Rizzitelli A., Chidgey A, Boyd R., Shortman K., Wu L. (2010) Regeneration of dendritic cells in aged mice. Cell Mol Immunol. 7(2): 108-15.

Goldberg G.L., Dudakov J.A., Seach N., Reiseger J., Ueno T., Vlahos K, Hammett M., Young L., Boyd R.L., Chidgey A.P. (2010) Sex steroid ablation enhances thymic recovery following anti-neoplastic therapy in young mice. J Immunol. 184(11): 6014-24.

Heng, T.S.P., Dudakov, J.A., Khong, D.M.P., Chidgey, A.P., and Boyd, R.L. (2009) Stem cells - meet immunity. J Mol Med, 87: 1061-1069.

Seach, N., Mattesich, M., Abberton, K., Matsuda, K., Tilkorn, D.J., Rophael, J., Boyd, R.L., and Morison, W.A. (2009) Vascularized Tissue Engineering Mouse Chamber Model Supports Thymopoiesis of Ectopic Thymus Tissue Grafts. Tissue Engineering, Part C: Methods. 16(3): 543-551.

Dudakov, J.A., Goldberg, G.L., Reiseger, J.J., Vlahos, K., Chidgey, A.P., and Boyd, R.L. (2009) Sex steroid ablation enhances hematopoietic recovery following cytotoxic antineoplastic therapy in aged mice. J Immunol. 183: 7084-7094.

Lynch, H.E., Goldberg, G.L., Chidgey, A.P., Van den Brink, M.R.M., Boyd, R., and Sempowski, G.D. (2009) Thymic involution and immune reconstitution. Trends Immunol. 30 (7): 366-373.

Fletcher, A.L., Lowen, T.E., Sakkal, S., Reiseger, J.J., Hammett, M.V., Seach, N., Scott, H.S., Boyd, R.L., and Chidgey, A.P. (2009) Ablation and Regeneration of Tolerance-Inducing Medullary Thymic Epithelial Cells after Cyclosporine, Cyclophosphamide, and Dexamethasone Treatment. J Immunol 183: 823-831

Dudakov, J.A., Goldberg, G.L., Reiseger, J.J., Chidgey, A.P., and Boyd, R.L. (2009) Withdrawal of Sex Steroids Reverses Age- and Chemotherapy-Related Defects in Bone Marrow Lymphopoiesis. J Immunol 182: 6247-6260

Seach, N., Shannon, V., Boyd, R. (2009) Ethics versus patient needs - the delicate ‘rheostat' for stem cell research. Chemistry in Australia, 76 (2): 16-19 Reproduced in "Issues" magazine, March 2009 (86), 46-48.

Cuddihy, A.R., Ge, S., Zhu, J., Jang, J., Chidgey, A., Thurston, G., Boyd, R., and Crooks, G.M. (2009) VEGF-mediated cross-talk within the neonatal murine thymus. Blood, 113:2723-2731.

Cheong, W., Reiseger, J., Turner, S.J., Boyd, R.L. Netter, H. (2009) Chimeric virus- like particles for the delivery of an inserted conserved influenza A-specific CTL epitope. Antiviral Research 81: 113-122

Goldberg, G.L., King, C.G., Suh, D.Y., Smith, O.M., Ajodan, R.H., Samstein, R.M., Dudakov, J.A., Chidgey, A.P., Boyd, R.L., van den Brink, M.R.M. (2009) Luteinizing hormone releasing hormone enhances T cell recovery following allogeneic bone marrow transplantation. J. Immunol. 182: 5846 - 5854.

Fletcher, A.L., Seach, N., Reiseger, J.J., Lowen, T.E., Hammett, M.V., and Boyd, R.L. (2009) Reduced Thymic Aire expression and Abnormal NF-κB2 Signalling in a Model of Systemic Autoimmunity. Journal of Immunology. 182 (5): 2690 - 2699

Research Support

• Australian Stem Cell Centre
• NH&MRC
• Monash University Strategic (collaborative) Grant