Academic Appointments

All Publications

  • Preclinical Safety and Efficacy Validation of CD4(LVFOXP3) Cells as an Innovative Cell-Based Gene Therapy Approach for IPEX Syndrome Sato, Y., Nathan, A., Wright, J., Tate, K., Wani, P., Fazeli, F., Timnak, A., Bhatia, N., Agarwal-Hashmi, R., Bertaina, A., Roncarolo, M., Bacchetta, R. CELL PRESS. 2021: 340
  • Co-Expression of FOXP3FL and FOXP3Δ2 Isoforms Is Required for Optimal Treg-Like Cell Phenotypes and Suppressive Function. Frontiers in immunology Sato, Y., Liu, J., Lee, E., Perriman, R., Roncarolo, M. G., Bacchetta, R. 2021; 12: 752394


    FOXP3 is the master transcription factor in both murine and human FOXP3+ regulatory T cells (Tregs), a T-cell subset with a central role in controlling immune responses. Loss of the functional Foxp3 protein in scurfy mice leads to acute early-onset lethal lymphoproliferation. Similarly, pathogenic FOXP3 mutations in humans lead to immunodysregulation, polyendocrinopathy, enteropathy, and X-linked (IPEX) syndrome, which are characterized by systemic autoimmunity that typically begins in the first year of life. However, although pathogenic FOXP3 mutations lead to overlapping phenotypic consequences in both systems, FOXP3 in human Tregs, but not mouse, is expressed as two predominant isoforms, the full length (FOXP3FL) and the alternatively spliced isoform, delta 2 (FOXP3Δ2). Here, using CRISPR/Cas9 to generate FOXP3 knockout CD4+ T cells (FOXP3KOGFP CD4+ T cells), we restore the expression of each isoform by lentiviral gene transfer to delineate their functional roles in human Tregs. When compared to FOXP3FL or FOXP3Δ2 alone, or double transduction of the same isoform, co-expression of FOXP3FL and FOXP3Δ2 induced the highest overall FOXP3 protein expression in FOXP3KOGFP CD4+ T cells. This condition, in turn, led to optimal acquisition of Treg-like cell phenotypes including downregulation of cytokines, such as IL-17, and increased suppressive function. Our data confirm that co-expression of FOXP3FL and FOXP3Δ2 leads to optimal Treg-like cell function and supports the need to maintain the expression of both when engineering therapeutics designed to restore FOXP3 function in otherwise deficient cells.

    View details for DOI 10.3389/fimmu.2021.752394

    View details for PubMedID 34737751

    View details for PubMedCentralID PMC8560788

  • Pre-clinical development and molecular characterization of an engineered type 1 regulatory T-cell product suitable for immunotherapy. Cytotherapy Liu, J. M., Chen, P., Uyeda, M. J., Cieniewicz, B., Sayitoglu, E. C., Thomas, B. C., Sato, Y., Bacchetta, R., Cepika, A. M., Roncarolo, M. G. 2021


    Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative therapeutic approach for many hematological disorders. However, allo-HSCT is frequently accompanied by a serious side effect: graft-versus-host disease (GVHD). The clinical use of allo-HSCT is limited by the inability of current immunosuppressive regimens to adequately control GvHD without impairing the graft-versus-leukemia effect (GvL) conferred by transplanted healthy immune cells. To address this, the authors have developed an engineered type 1 regulatory T-cell product called CD4IL-10 cells. CD4IL-10 cells are obtained through lentiviral transduction, which delivers the human IL10 gene into purified polyclonal CD4+ T cells. CD4IL-10 cells may provide an advantage over standard-of-care immunosuppressants because of the ability to suppress GvHD through continuous secretion of IL-10 and enhance the GvL effect in myeloid malignancies through targeted killing of malignant myeloid cells.Here the authors established a production process aimed at current Good Manufacturing Practice (cGMP) production for CD4IL-10 cells.The authors demonstrated that the CD4IL-10 cell product maintains the suppressive and cytotoxic functions of previously described CD4IL-10 cells. In addition, RNA sequencing analysis of CD4IL-10 identified novel transcriptome changes, indicating that CD4IL-10 cells primarily upregulate cytotoxicity-related genes. These include four molecules with described roles in CD8+ T and natural killer cell-mediated cytotoxicity: CD244, KLRD1, KLRC1 and FASLG. Finally, it was shown that CD4IL-10 cells upregulate IL-22, which mediates wound healing and tissue repair, particularly in the gut.Collectively, these results pave the way toward clinical translation of the cGMP-optimized CD4IL-10 cell product and uncover new molecules that have a role in the clinical application of CD4IL-10 cells.

    View details for DOI 10.1016/j.jcyt.2021.05.010

    View details for PubMedID 34404616

  • Human-engineered Treg-like cells suppress FOXP3-deficient T cells but preserve adaptive immune responses in vivo. Clinical & translational immunology Sato, Y. n., Passerini, L. n., Piening, B. D., Uyeda, M. J., Goodwin, M. n., Gregori, S. n., Snyder, M. P., Bertaina, A. n., Roncarolo, M. G., Bacchetta, R. n. 2020; 9 (11): e1214


    Genetic or acquired defects in FOXP3+ regulatory T cells (Tregs) play a key role in many immune-mediated diseases including immune dysregulation polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome. Previously, we demonstrated CD4+ T cells from healthy donors and IPEX patients can be converted into functional Treg-like cells by lentiviral transfer of FOXP3 (CD4LVFOXP3). These CD4LVFOXP3 cells have potent regulatory function, suggesting their potential as an innovative therapeutic. Here, we present molecular and preclinical in vivo data supporting CD4LVFOXP3 cell clinical progression.The molecular characterisation of CD4LVFOXP3 cells included flow cytometry, qPCR, RNA-seq and TCR-seq. The in vivo suppressive function of CD4LVFOXP3 cells was assessed in xenograft-versus-host disease (xeno-GvHD) and FOXP3-deficient IPEX-like humanised mouse models. The safety of CD4LVFOXP3 cells was evaluated using peripheral blood (PB) humanised (hu)- mice testing their impact on immune response against pathogens, and immune surveillance against tumor antigens.We demonstrate that the conversion of CD4+ T cells to CD4LVFOXP3 cells leads to specific transcriptional changes as compared to CD4+ T-cell transduction in the absence of FOXP3, including upregulation of Treg-related genes. Furthermore, we observe specific preservation of a polyclonal TCR repertoire during in vitro cell production. Both allogeneic and autologous CD4LVFOXP3 cells protect from xeno-GvHD after two sequential infusions of effector T cells. CD4LVFOXP3 cells prevent hyper-proliferation of CD4+ memory T cells in the FOXP3-deficient IPEX-like hu-mice. CD4LVFOXP3 cells do not impede in vivo expansion of antigen-primed T cells or tumor clearance in the PB hu-mice.These data support the clinical readiness of CD4LVFOXP3 cells to treat IPEX syndrome and other immune-mediated diseases caused by insufficient or dysfunctional FOXP3+ Tregs.

    View details for DOI 10.1002/cti2.1214

    View details for PubMedID 33304583

    View details for PubMedCentralID PMC7688376

  • Immunoregulatory Cell Therapy with Lentiviral-Mediated FOXP3 Converted CD4+T Cells into Treg Cells: Towards the Proof-of-Concept Application in IPEX Syndrome Sato, Y., Passerini, L., Roncarolo, M., Bacchetta, R. CELL PRESS. 2019: 311
  • Tregopathies: Monogenic diseases resulting in regulatory T-cell deficiency. The Journal of allergy and clinical immunology Cepika, A., Sato, Y., Liu, J. M., Uyeda, M. J., Bacchetta, R., Roncarolo, M. G. 2018; 142 (6): 1679–95


    Monogenic diseases of the immune system, also known as inborn errors of immunity, are caused by single-gene mutations resulting in immune deficiency and dysregulation. More than 350 diseases have been described to date, and the number is rapidly expanding, with increasing availability of next-generation sequencing facilitating the diagnosis. The spectrum of immune dysregulation is wide, encompassing deficiencies in humoral, cellular, innate, and adaptive immunity; phagocytosis; and the complement system, which lead to autoinflammation and autoimmunity. Multiorgan autoimmunity is a dominant symptom when genetic mutations lead to defects in molecules essential for the development, survival, and/or function of regulatory T (Treg) cells. Studies of "Tregopathies" are providing critical mechanistic information on Treg cell biology, the role of Treg cell-associated molecules, and regulation of peripheral tolerance in human subjects. The pathogenic immune networks underlying these diseases need to be dissected to apply and develop immunomodulatory treatments and design curative treatments using cell and gene therapy. Here we review the pathogenetic mechanisms, clinical presentation, diagnosis, and current and future treatments of major known Tregopathies caused by mutations in FOXP3, CD25, cytotoxic Tlymphocyte-associated antigen 4 (CTLA4), LPS-responsive and beige-like anchor protein (LRBA), and BTB domain and CNC homolog 2 (BACH2) and gain-of-function mutations in signal transducer and activator oftranscription 3 (STAT3). We also discuss deficiencies in genesencoding STAT5b and IL-10 or IL-10 receptor aspotential Tregopathies.

    View details for DOI 10.1016/j.jaci.2018.10.026

    View details for PubMedID 30527062

  • FOXP3 Gene Transfer in T cells and FOXP3 Gene Editing in HSC as Novel Treatment Options for IPEX Syndrome Goodwin, M., Sato, Y., Passerini, L., Barzaghi, F., Lee, E., Suzette, S. K., Roncarolo, M., Porteus, M., Bacchetta, R. SPRINGER/PLENUM PUBLISHERS. 2018: 427