Rosa Bacchetta
e-mail: bacchetta.rosa AT hsr.it
affiliation: San Raffaele Scientific Institute
research area(s): Immunity And Infection, Genetics And Genomics
Course:
Basic and Applied Immunology
University/Istitution: Università Vita-Salute San Raffaele
University/Istitution: Università Vita-Salute San Raffaele
Education:
1987 - General Medicine, Degree at Medical School, University of Turin, Italy M.D.
1987 - Natl.Board in General Medicine.
1987-1991 - Residency in Pediatrics, University of Turin (Italy)
Specific training in Paediatric Immunology.
Employment and experience:
1985-1987 - Predoctoral Internship. Department of Paediatric Immunology. School of Medicine, University of Turin, Italy.
1988-1989 - Research Fellow at UNICET Laboratory for immunological research, Lyon, France.
1990-1995 - Postdoctoral Fellow at DNAX Research Institute of Molecular and Cellular Biology, Palo Alto CA, USA.
1997-present - Medical Research Associate, Group Leader at TIGET-HSR. Milan, Italy.
Member of the Pediatric Clinical Research Unit, in charge of the outpatients Clinic of Paediatric Immunology.
Patents:
Issued: US Patent 6277635 - Use of interleukin-10 to produce a population of suppressor cells (08/21/2001)
Issued: US Patent 6884410 - Methods for modulating antigen-specific immune responses (26/04/2005)
Filed: WO2007/131575: - Tr1 dendritic cells, method to generate regulatory type 1 T (Tr1) cells and uses thereof (04/01/2007)
Scientific Activities
Published more than 30 papers in international scientific journals and contributed to several reviewers and chapters to books, on the following topics: Transplantation Tolerance, Immunodeficiencies, Cytokines, Regulatory T cells.
Invited every year as a speaker to International Meetings or Workshops in the field of Tolerance, Transplantation and Primary Immunodeficiencies.
Co-Inventor in 3 international patents on topics related to methods to generate regulatory T cells.
Reviewer for several International Scientific Journals in the fields of Immunology.
Director of Studies PhD Students of the HSR- International PhD program
Member of the European Society of Immunodefiency (ESID), American Clinical Immunology Society (CIS) and Italian Society of Pediatric Oncology and Hematology (AIEOP).
1987 - General Medicine, Degree at Medical School, University of Turin, Italy M.D.
1987 - Natl.Board in General Medicine.
1987-1991 - Residency in Pediatrics, University of Turin (Italy)
Specific training in Paediatric Immunology.
Employment and experience:
1985-1987 - Predoctoral Internship. Department of Paediatric Immunology. School of Medicine, University of Turin, Italy.
1988-1989 - Research Fellow at UNICET Laboratory for immunological research, Lyon, France.
1990-1995 - Postdoctoral Fellow at DNAX Research Institute of Molecular and Cellular Biology, Palo Alto CA, USA.
1997-present - Medical Research Associate, Group Leader at TIGET-HSR. Milan, Italy.
Member of the Pediatric Clinical Research Unit, in charge of the outpatients Clinic of Paediatric Immunology.
Patents:
Issued: US Patent 6277635 - Use of interleukin-10 to produce a population of suppressor cells (08/21/2001)
Issued: US Patent 6884410 - Methods for modulating antigen-specific immune responses (26/04/2005)
Filed: WO2007/131575: - Tr1 dendritic cells, method to generate regulatory type 1 T (Tr1) cells and uses thereof (04/01/2007)
Scientific Activities
Published more than 30 papers in international scientific journals and contributed to several reviewers and chapters to books, on the following topics: Transplantation Tolerance, Immunodeficiencies, Cytokines, Regulatory T cells.
Invited every year as a speaker to International Meetings or Workshops in the field of Tolerance, Transplantation and Primary Immunodeficiencies.
Co-Inventor in 3 international patents on topics related to methods to generate regulatory T cells.
Reviewer for several International Scientific Journals in the fields of Immunology.
Director of Studies PhD Students of the HSR- International PhD program
Member of the European Society of Immunodefiency (ESID), American Clinical Immunology Society (CIS) and Italian Society of Pediatric Oncology and Hematology (AIEOP).
From FOXP3 mutation to IPEX syndrome: genotype/phenotype, immune pathogenic mechanisms and therapeutic options.
Immune dysregulation, Polyendocrinopathy, Enteropathy, X-linked (IPEX) syndrome is a genetic autoimmune disease due to mutations of FOXP3 gene, characterized by early onset refractory and life threatening enteropathy, eczema and Type I diabetes.
The transcription factor FOXP3 is the driving force for the differentiation and function of regulatory T (Treg) cells but it is still questionable whether its transcriptional activity can be relevant also for the development of effector T (Teff) cells. Mortality can be high within the first year of life, whereas later on, patients can respond to conventional immunosuppression and show a clinical improvement. The disease course is not always correlated to the type of mutation. Forms that are diagnosed later in life and remain oligo-symptomatic are also observed. In addition, in several patients with IPEX-like symptoms, no FOXP3 mutations can be detected.
At present, the pathogenesis of the disease is not known, although it is clear that Treg cells are dysfunctional in IPEX patients. However, FOXP3 has multiple functions, many of which not well defined yet, and it remains to be demonstrated whether the lack of Treg cells is the only cause of the autoimmune pathology in IPEX. Current therapies for IPEX present several limitations: immunosuppression is aspecific and has only partial effects with relevant toxicity, whereas hematopoietic stem cell transplantation (HSCT) is not always feasible.
The overall objectives of this translational research projects on IPEX are:
1. to define a genotype/phenotype correlation of IPEX, providing precise criteria for diagnosis, investigating the effector and regulatory immune mechanisms altered in IPEX patients, identifying molecular and immunological features relevant for determining the outcome of the natural course of the disease and assessing the responses to different therapeutic approaches, including BMT.
2. to explore different experimental approaches that can lead to innovative more targeted treatments for IPEX. Significant insights have been recently gained on the methods to identify, isolate and expand different types of regulatory T cells and we have developed effective techniques to stably express or knockdown FOXP3 in T cells by LV mediated gene transfer. Using these approaches we will investigate the effects of different FOXP3 mutations into the biology of Treg and Teff cells and we will consider in vitro restoration of suppressive activity by wild type (wt) FOXP3 gene transfer into mutated cells. Ultimately, these cells will be tested in vivo in humanized murine models, with the aim of assessing the feasibility and efficacy of cell/gene therapy approaches for IPEX patients.
Thanks to a multicenter study group (www.ipexconsortium.org), coordinated by Dr.ssa R. Bacchetta, the number of patients with suspected IPEX who were referred to us for a diagnosis has increased during the past two years. Results from these studies will be critical for the identification and cure of patients affected with IPEX and also represent a unique source of knowledge of the genetic and immunological mechanisms responsible for the establishment of tolerance.
Role of regulatory T cells in transplantation tolerance
This area of research is part of the core research at HSR-TIGET, aimed at characterizing different subsets of regulatory T cells and identifying specific molecules involved in their function, and includes the following projects:
1. cellular and molecular characterization of immunological mechanisms, in particular those sustained by T regulatory cells, in patients affected by hematological malignancies or genetic diseases who underwent allogeneic HSC transplantations. Thalassemic patients with post-transplant persistent mixed chimerism and patients who received haploidentical HSCT followed by cellular therapy with IL-10 producing type 1 regulatory T cells, are included in these studies;
2. development and transfer of novel cell/gene therapy approaches for induction of tolerance in HSC or organ transplantation;
3. immunogenetic studies aimed at identifying genetic factors predisposing to transplant rejection, graft-versus-host disease or engraftment.
Immune dysregulation, Polyendocrinopathy, Enteropathy, X-linked (IPEX) syndrome is a genetic autoimmune disease due to mutations of FOXP3 gene, characterized by early onset refractory and life threatening enteropathy, eczema and Type I diabetes.
The transcription factor FOXP3 is the driving force for the differentiation and function of regulatory T (Treg) cells but it is still questionable whether its transcriptional activity can be relevant also for the development of effector T (Teff) cells. Mortality can be high within the first year of life, whereas later on, patients can respond to conventional immunosuppression and show a clinical improvement. The disease course is not always correlated to the type of mutation. Forms that are diagnosed later in life and remain oligo-symptomatic are also observed. In addition, in several patients with IPEX-like symptoms, no FOXP3 mutations can be detected.
At present, the pathogenesis of the disease is not known, although it is clear that Treg cells are dysfunctional in IPEX patients. However, FOXP3 has multiple functions, many of which not well defined yet, and it remains to be demonstrated whether the lack of Treg cells is the only cause of the autoimmune pathology in IPEX. Current therapies for IPEX present several limitations: immunosuppression is aspecific and has only partial effects with relevant toxicity, whereas hematopoietic stem cell transplantation (HSCT) is not always feasible.
The overall objectives of this translational research projects on IPEX are:
1. to define a genotype/phenotype correlation of IPEX, providing precise criteria for diagnosis, investigating the effector and regulatory immune mechanisms altered in IPEX patients, identifying molecular and immunological features relevant for determining the outcome of the natural course of the disease and assessing the responses to different therapeutic approaches, including BMT.
2. to explore different experimental approaches that can lead to innovative more targeted treatments for IPEX. Significant insights have been recently gained on the methods to identify, isolate and expand different types of regulatory T cells and we have developed effective techniques to stably express or knockdown FOXP3 in T cells by LV mediated gene transfer. Using these approaches we will investigate the effects of different FOXP3 mutations into the biology of Treg and Teff cells and we will consider in vitro restoration of suppressive activity by wild type (wt) FOXP3 gene transfer into mutated cells. Ultimately, these cells will be tested in vivo in humanized murine models, with the aim of assessing the feasibility and efficacy of cell/gene therapy approaches for IPEX patients.
Thanks to a multicenter study group (www.ipexconsortium.org), coordinated by Dr.ssa R. Bacchetta, the number of patients with suspected IPEX who were referred to us for a diagnosis has increased during the past two years. Results from these studies will be critical for the identification and cure of patients affected with IPEX and also represent a unique source of knowledge of the genetic and immunological mechanisms responsible for the establishment of tolerance.
Role of regulatory T cells in transplantation tolerance
This area of research is part of the core research at HSR-TIGET, aimed at characterizing different subsets of regulatory T cells and identifying specific molecules involved in their function, and includes the following projects:
1. cellular and molecular characterization of immunological mechanisms, in particular those sustained by T regulatory cells, in patients affected by hematological malignancies or genetic diseases who underwent allogeneic HSC transplantations. Thalassemic patients with post-transplant persistent mixed chimerism and patients who received haploidentical HSCT followed by cellular therapy with IL-10 producing type 1 regulatory T cells, are included in these studies;
2. development and transfer of novel cell/gene therapy approaches for induction of tolerance in HSC or organ transplantation;
3. immunogenetic studies aimed at identifying genetic factors predisposing to transplant rejection, graft-versus-host disease or engraftment.
1. Roncarolo MG, Gregori S, Lucarelli B, Ciceri F, Bacchetta R. 2011. Clinical tolerance in allogeneic hematopoietic stem cell transplantation. Immunol Rev. May;241(1):145-63.
2. Passerini L, Di Nunzio S, Gregori S, Gambineri E, Cecconi M, Seidel MG, Cazzola G, Perroni L, Tommasini A, Vignola S, Guidi L, Roncarolo MG, Bacchetta R. 2011. Functional type 1 regulatory T cells develop regardless of FOXP3 mutations in patients with IPEX syndrome. Eur J Immunol. Apr;41(4):1120-31.
3. McMurchy AN, Gillies J, Allan SE, Passerini L, Gambineri E, Roncarolo MG, Bacchetta R, Levings MK. 2010. Point mutants of forkhead box P3 that cause immune dysregulation, polyendocrinopathy, enteropathy, X-linked have diverse abilities to reprogram T cells into regulatory T cells. J Allergy Clin Immunol. Dec;126(6):1242-51.
4. Gregori S, Roncarolo MG, Bacchetta R. 2011. Methods for in vitro generation of human type 1 regulatory T cells. Methods Mol Biol.;677:31-46.
5. Bacchetta R, Gregori S, Serafini G, Sartirana C, Schulz U, Zino E, Tomiuk S, Jansen U, Ponzoni M, Paties CT, Fleischhauer K, Roncarolo MG. 2010. Molecular and functional characterization of allogantigen-specific anergic T cells suitable for cell therapy. Haematologica. Dec;95(12):2134-43.
6. Di Nunzio S, Cecconi M, Passerini L, McMurchy AN, Baron U, Turbachova I, Vignola S, Valencic E, Tommasini A, Junker A, Cazzola G, Olek S, Levings MK, Perroni L, Roncarolo MG, Bacchetta R. 2009. Wild-type FOXP3 is selectively active in CD4+CD25(hi) regulatory T cells of healthy female carriers of different FOXP3 mutations. Blood. Nov 5;114(19):4138-41.
7. Serafini G, Andreani M, Testi M, Battarra M, Bontadini A, Biral E, Fleischhauer K, Marktel S, Lucarelli G, Roncarolo MG, Bacchetta R. 2009. Type 1 regulatory T cells are associated with persistent split erythroid/lymphoid chimerism after allogeneic hematopoietic stem cell transplantation for thalassemia. Haematologica. Oct;94(10):1415-26.
8. Seidel MG, Fritsch G, Lion T, Jürgens B, Heitger A, Bacchetta R, Lawitschka A, Peters C, Gadner H, Matthes-Martin S. 2009. Selective engraftment of donor CD4+25high FOXP3-positive T cells in IPEX syndrome after nonmyeloablative hematopoietic stem cell transplantation. Blood. May 28;113(22):5689-91.
9. Amendola M, Passerini L, Pucci F, Gentner B, Bacchetta R, Naldini L. 2009. Regulated and multiple miRNA and siRNA delivery into primary cells by a lentiviral platform. Mol Ther. Jun;17(6):1039-52.
10. McMurchy AN, Di Nunzio S, Roncarolo MG, Bacchetta R, Levings MK. 2009. Molecular regulation of cellular immunity by FOXP3. Adv Exp Med Biol.;665:30-46. Review.
11. Gambineri E, Perroni L, Passerini L, Bianchi L, Doglioni C, Meschi F, Bonfanti R, Sznajer Y, Tommasini A, Lawitschka A, Junker A, Dunstheimer D, Heidemann PH, Cazzola G, Cipolli M, Friedrich W, Janic D, Azzi N, Richmond E, Vignola S, Barabino A, Chiumello G, Azzari C, Roncarolo MG, Bacchetta R. 2008. Clinical and molecular profile of a new series of patients with immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome: inconsistent correlation between forkhead box protein 3 expression and disease severity. J Allergy Clin Immunol. Dec;122(6):1105-1112.e1.
12. Allan SE, Broady R, Gregori S, Himmel ME, Locke N, Roncarolo MG, Bacchetta R, Levings MK. 2008. CD4+ T-regulatory cells: toward therapy for human diseases. Immunol Rev. Jun;223:391-421.
13. Passerini L, Allan SE, Battaglia M, Di Nunzio S, Alstad AN, Levings MK, Roncarolo MG, Bacchetta R. 2008. STAT5-signaling cytokines regulate the expression of FOXP3 in CD4+CD25+ regulatory T cells and CD4+CD25- effector T cells. Int Immunol. Mar;20(3):421-31.
14. Allan SE, Alstad AN, Merindol N, Crellin NK, Amendola M, Bacchetta R, Naldini L, Roncarolo MG, Soudeyns H, Levings MK. 2008. Generation of potent and stable human CD4+ T regulatory cells by activation-independent expression of FOXP3. Mol Ther. Jan;16(1):194-202.
15. Bacchetta R, Gambineri E, Roncarolo MG. 2007. Role of regulatory T cells and FOXP3 in human diseases. J Allergy Clin Immunol. Aug;120(2):227-35; quiz 236-7. Review.
16. Roncarolo MG, Gregori S, Battaglia M, Bacchetta R, Fleischhauer K, Levings MK. 2006. Interleukin-10-secreting type 1 regulatory T cells in rodents and humans. Immunol Rev. Aug;212:28-50. Review.
17. Bacchetta R, Passerini L, Gambineri E, Dai M, Allan SE, Perroni L, Dagna-Bricarelli F, Sartirana C, Matthes-Martin S, Lawitschka A, Azzari C, Ziegler SF, Levings MK, Roncarolo MG. 2006. Defective regulatory and effector T cell functions in patients with FOXP3 mutations. J Clin Invest. Jun;116(6):1713-22.
2. Passerini L, Di Nunzio S, Gregori S, Gambineri E, Cecconi M, Seidel MG, Cazzola G, Perroni L, Tommasini A, Vignola S, Guidi L, Roncarolo MG, Bacchetta R. 2011. Functional type 1 regulatory T cells develop regardless of FOXP3 mutations in patients with IPEX syndrome. Eur J Immunol. Apr;41(4):1120-31.
3. McMurchy AN, Gillies J, Allan SE, Passerini L, Gambineri E, Roncarolo MG, Bacchetta R, Levings MK. 2010. Point mutants of forkhead box P3 that cause immune dysregulation, polyendocrinopathy, enteropathy, X-linked have diverse abilities to reprogram T cells into regulatory T cells. J Allergy Clin Immunol. Dec;126(6):1242-51.
4. Gregori S, Roncarolo MG, Bacchetta R. 2011. Methods for in vitro generation of human type 1 regulatory T cells. Methods Mol Biol.;677:31-46.
5. Bacchetta R, Gregori S, Serafini G, Sartirana C, Schulz U, Zino E, Tomiuk S, Jansen U, Ponzoni M, Paties CT, Fleischhauer K, Roncarolo MG. 2010. Molecular and functional characterization of allogantigen-specific anergic T cells suitable for cell therapy. Haematologica. Dec;95(12):2134-43.
6. Di Nunzio S, Cecconi M, Passerini L, McMurchy AN, Baron U, Turbachova I, Vignola S, Valencic E, Tommasini A, Junker A, Cazzola G, Olek S, Levings MK, Perroni L, Roncarolo MG, Bacchetta R. 2009. Wild-type FOXP3 is selectively active in CD4+CD25(hi) regulatory T cells of healthy female carriers of different FOXP3 mutations. Blood. Nov 5;114(19):4138-41.
7. Serafini G, Andreani M, Testi M, Battarra M, Bontadini A, Biral E, Fleischhauer K, Marktel S, Lucarelli G, Roncarolo MG, Bacchetta R. 2009. Type 1 regulatory T cells are associated with persistent split erythroid/lymphoid chimerism after allogeneic hematopoietic stem cell transplantation for thalassemia. Haematologica. Oct;94(10):1415-26.
8. Seidel MG, Fritsch G, Lion T, Jürgens B, Heitger A, Bacchetta R, Lawitschka A, Peters C, Gadner H, Matthes-Martin S. 2009. Selective engraftment of donor CD4+25high FOXP3-positive T cells in IPEX syndrome after nonmyeloablative hematopoietic stem cell transplantation. Blood. May 28;113(22):5689-91.
9. Amendola M, Passerini L, Pucci F, Gentner B, Bacchetta R, Naldini L. 2009. Regulated and multiple miRNA and siRNA delivery into primary cells by a lentiviral platform. Mol Ther. Jun;17(6):1039-52.
10. McMurchy AN, Di Nunzio S, Roncarolo MG, Bacchetta R, Levings MK. 2009. Molecular regulation of cellular immunity by FOXP3. Adv Exp Med Biol.;665:30-46. Review.
11. Gambineri E, Perroni L, Passerini L, Bianchi L, Doglioni C, Meschi F, Bonfanti R, Sznajer Y, Tommasini A, Lawitschka A, Junker A, Dunstheimer D, Heidemann PH, Cazzola G, Cipolli M, Friedrich W, Janic D, Azzi N, Richmond E, Vignola S, Barabino A, Chiumello G, Azzari C, Roncarolo MG, Bacchetta R. 2008. Clinical and molecular profile of a new series of patients with immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome: inconsistent correlation between forkhead box protein 3 expression and disease severity. J Allergy Clin Immunol. Dec;122(6):1105-1112.e1.
12. Allan SE, Broady R, Gregori S, Himmel ME, Locke N, Roncarolo MG, Bacchetta R, Levings MK. 2008. CD4+ T-regulatory cells: toward therapy for human diseases. Immunol Rev. Jun;223:391-421.
13. Passerini L, Allan SE, Battaglia M, Di Nunzio S, Alstad AN, Levings MK, Roncarolo MG, Bacchetta R. 2008. STAT5-signaling cytokines regulate the expression of FOXP3 in CD4+CD25+ regulatory T cells and CD4+CD25- effector T cells. Int Immunol. Mar;20(3):421-31.
14. Allan SE, Alstad AN, Merindol N, Crellin NK, Amendola M, Bacchetta R, Naldini L, Roncarolo MG, Soudeyns H, Levings MK. 2008. Generation of potent and stable human CD4+ T regulatory cells by activation-independent expression of FOXP3. Mol Ther. Jan;16(1):194-202.
15. Bacchetta R, Gambineri E, Roncarolo MG. 2007. Role of regulatory T cells and FOXP3 in human diseases. J Allergy Clin Immunol. Aug;120(2):227-35; quiz 236-7. Review.
16. Roncarolo MG, Gregori S, Battaglia M, Bacchetta R, Fleischhauer K, Levings MK. 2006. Interleukin-10-secreting type 1 regulatory T cells in rodents and humans. Immunol Rev. Aug;212:28-50. Review.
17. Bacchetta R, Passerini L, Gambineri E, Dai M, Allan SE, Perroni L, Dagna-Bricarelli F, Sartirana C, Matthes-Martin S, Lawitschka A, Azzari C, Ziegler SF, Levings MK, Roncarolo MG. 2006. Defective regulatory and effector T cell functions in patients with FOXP3 mutations. J Clin Invest. Jun;116(6):1713-22.
Project Title:
Project Title:
Preclinical studies of FOXP3 gene transfer for cell therapy with regulatory T cells in genetic autoimmune diseases.
Regulatory T (Treg) cells comprise different type of cells that share the ability to regulate immune responses. Their role has been recognized as essential in the maintenance of immunological tolerance, especially in regulating immune responses towards self-antigens (Ags). Indeed, autoimmune manifestations characterize diseases in which Treg-cell function is impaired. The best example is Immune dysregulation, Polyendocrinopathy, Enteropathy, X-linked (IPEX) syndrome, a genetic autoimmune disease characterized by early onset refractory and life-threatening enteropathy, eczema and Type I diabetes. IPEX is caused by mutations in FOXP3, a transcription factor driving the fate and function of naturally occurring CD4+CD25+Treg cells.
Cell-based therapies using in vitro generated/expanded Treg cells for the cure of autoimmune pathologies have been proposed. However, many questions on the safety, efficacy and mechanisms of action of Treg cells used for cell therapy, still have to be addressed. These include important issues, such as Treg survival in vivo, the risk of detrimental general immunosuppression, the requirement for Ag-specificity to obtain a therapeutic effect. The present project aims at investigating those topics, using lentiviral-vector (LV) mediated FOXP3 gene transfer. We have already demonstrated the feasibility of Treg generation by FOXP3 gene transfer. We will extend our investigation to preclinical studies in animal models of xeno-GvHD, and using suitable in vivo murine or humanized-murine models, we will assess safety and efficacy of polyclonal and Ag specific LV-FOXP3 cells, and whether the efficacy of CD4FOXP3 cells is maintained once injected in an inflammatory and lymphoproliferative environment. Results obtained by these studies will be instrumental to define clinical protocol suitable for the use of the gene modified Treg cells in IPEX patients. However, induction of regulatory function by FOXP3 gene transfer can be considered a useful experimental approach also for other autoimmune disorders in which pathogenic autoreactive T cells can be converted into Treg cells.
Bacchetta R., L. Passerini, E.Gambineri, M. Dai, S.E. Allan, L. Perroni, F. Dagna-Bricarelli, C. Sartirana, S. Matthes-Martin, A. Lawitschka, C. Azzari, S.F. Ziegler, M.K. Levings, and M.G. Roncarolo. Defective regulatory and effector T cell functions in patients with FOXP3 mutations. J Clin. Invest. 116 (2006), pp. 1713-1722
Allan S.E., A.N. Alstad, N. Merindol, N.K. Crellin, M. Amendola, R. Bacchetta, L. Naldini, M.G. Roncarolo, H. Soudeyns and M.K. Levings .Generation of Potent and Stable Human CD4(+) T Regulatory Cells by Activation-independent Expression of FOXP3. Mol. Ther. 16 (2008), pp. 194-202
Gambineri E., L. Perroni, L. Passerini, L. Bianchi, C. Doglioni, F. Meschi et al. Clinical and molecular profile of a new series of patients with immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome: Inconsistent correlation between forkhead box protein 3 protein expression and disease severity. J. Allergy Clin. Immunol. 122 (2008), pp. 1105-1112
Cell-based therapies using in vitro generated/expanded Treg cells for the cure of autoimmune pathologies have been proposed. However, many questions on the safety, efficacy and mechanisms of action of Treg cells used for cell therapy, still have to be addressed. These include important issues, such as Treg survival in vivo, the risk of detrimental general immunosuppression, the requirement for Ag-specificity to obtain a therapeutic effect. The present project aims at investigating those topics, using lentiviral-vector (LV) mediated FOXP3 gene transfer. We have already demonstrated the feasibility of Treg generation by FOXP3 gene transfer. We will extend our investigation to preclinical studies in animal models of xeno-GvHD, and using suitable in vivo murine or humanized-murine models, we will assess safety and efficacy of polyclonal and Ag specific LV-FOXP3 cells, and whether the efficacy of CD4FOXP3 cells is maintained once injected in an inflammatory and lymphoproliferative environment. Results obtained by these studies will be instrumental to define clinical protocol suitable for the use of the gene modified Treg cells in IPEX patients. However, induction of regulatory function by FOXP3 gene transfer can be considered a useful experimental approach also for other autoimmune disorders in which pathogenic autoreactive T cells can be converted into Treg cells.
Bacchetta R., L. Passerini, E.Gambineri, M. Dai, S.E. Allan, L. Perroni, F. Dagna-Bricarelli, C. Sartirana, S. Matthes-Martin, A. Lawitschka, C. Azzari, S.F. Ziegler, M.K. Levings, and M.G. Roncarolo. Defective regulatory and effector T cell functions in patients with FOXP3 mutations. J Clin. Invest. 116 (2006), pp. 1713-1722
Allan S.E., A.N. Alstad, N. Merindol, N.K. Crellin, M. Amendola, R. Bacchetta, L. Naldini, M.G. Roncarolo, H. Soudeyns and M.K. Levings .Generation of Potent and Stable Human CD4(+) T Regulatory Cells by Activation-independent Expression of FOXP3. Mol. Ther. 16 (2008), pp. 194-202
Gambineri E., L. Perroni, L. Passerini, L. Bianchi, C. Doglioni, F. Meschi et al. Clinical and molecular profile of a new series of patients with immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome: Inconsistent correlation between forkhead box protein 3 protein expression and disease severity. J. Allergy Clin. Immunol. 122 (2008), pp. 1105-1112
Project Title:
Thymic differentiation of human naturally occurring regulatory T cells: FOXP3 does it all?
FOXP3 gene is known as the main transcription factor for the development and function of CD4+CD25+ naturally occurring regulatory T (nTreg) cells, which are essential for the maintenance of peripheral tolerance. Patients with mutations in FOXP3, indeed develop in early life a severe form of autoimmune disease (IPEX). However, we recently observed that, although dysfunctional, nTreg cells are present in the peripheral blood of IPEX patients, thus suggesting that thymic development can occur in the presence of FOXP3 mutations. On the other hand, we also reported that in healthy carriers of FOXP3 mutations the nTreg cell compartment only comprises nTreg cells expressing wtFOXP3. Both findings question whether and how FOXP3 influences the development of nTreg cells.
The present project aims at shedding light on the FOXP3-dependence/independence of human Treg cell development and function, using recently reported in vitro culture systems that mimic thymic development or ad hoc humanized-mouse models suitable for terminal differentiation of nTreg cells.
To this aim healthy donor CD34+ cells from cord blood or bone marrow will be used to first evaluate, hu-myeloid and T/B cell engraftment in the mice. We will study thymic development of Treg cells by evaluating their number, phenotype, gene profile and function at different stages of thymic development. To investigate how mutated FOXP3 in CD34+ cells can alter Treg cell development and lead to pathology, we plan to infuse mut CD34+ cells from BM of patients with FOXP3 mutations and carrier mothers, or as an alternative, we will transduce healthy donor CD34+ cells with LV encoding for siRNA targeting FOXP3-expression, resulting in knock-down of FOXP3 expression in the CD34+ cells that will be used to reconstitute the hu-mice. Results from this work will also provide a new in vivo hu-mice model of human autoimmune pathology that could be used for further studies aimed at modulation or gene correction of nTreg cell development or function.
Gavin MA, Rasmussen JP, Fontenot JD, Vasta V, Manganiello VC, Beavo JA,
Rudensky AY. 2007. Foxp3-dependent programme of regulatory T-cell
differentiation. Nature 445:771-775.
Di Nunzio S., M. Cecconi, L. Passerini, A. McMurphy, U. Baron, I. Turbachova, S. Vignola, E. Valencic, A. Tommasini, A. Junker, G. Cazzola, S. Olek, M.K. Levings, L. Perroni, M.G. Roncarolo and R. Bacchetta. Wild-Type FOXP3 is selectively active in CD4+CD25hi regulatory T cells of healthy female carriers of different FOXP3 mutations. Blood 114 (2009), pp. 4138-4141
Amendola M., L. Passerini, F. Pucci, B. Gentner, R. Bacchetta, and L. Naldini. Regulated and multiple miRNA and siRNA delivery into primary cells by a lentiviral platform. Mol. Ther. 17 (2009), pp. 1039-1052
Billerbeck E, Barry WT, Mu K, Dorner M, Rice CM, Ploss A. 2011. Development of human CD4+FoxP3+ regulatory T cells in human stem cell factor, GM-CSF and interleukin 3 expressing NOD SCID IL2R{gamma}NULL humanized mice. Blood.
The present project aims at shedding light on the FOXP3-dependence/independence of human Treg cell development and function, using recently reported in vitro culture systems that mimic thymic development or ad hoc humanized-mouse models suitable for terminal differentiation of nTreg cells.
To this aim healthy donor CD34+ cells from cord blood or bone marrow will be used to first evaluate, hu-myeloid and T/B cell engraftment in the mice. We will study thymic development of Treg cells by evaluating their number, phenotype, gene profile and function at different stages of thymic development. To investigate how mutated FOXP3 in CD34+ cells can alter Treg cell development and lead to pathology, we plan to infuse mut CD34+ cells from BM of patients with FOXP3 mutations and carrier mothers, or as an alternative, we will transduce healthy donor CD34+ cells with LV encoding for siRNA targeting FOXP3-expression, resulting in knock-down of FOXP3 expression in the CD34+ cells that will be used to reconstitute the hu-mice. Results from this work will also provide a new in vivo hu-mice model of human autoimmune pathology that could be used for further studies aimed at modulation or gene correction of nTreg cell development or function.
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Billerbeck E, Barry WT, Mu K, Dorner M, Rice CM, Ploss A. 2011. Development of human CD4+FoxP3+ regulatory T cells in human stem cell factor, GM-CSF and interleukin 3 expressing NOD SCID IL2R{gamma}NULL humanized mice. Blood.