Preclinical development of T cell products for immunotherapy after allogeneic hematopoietic stem cell transplantation

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Date

2024-05-22

Authors

Κεφάλα, Διονυσία

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Abstract

Graft versus Host Disease (GvHD) is a severe complication of allogeneic hematopoietic cell transplantation caused by donor T-cells attacking the host’s tissues. Conventional immunosuppressive pharmacotherapy remains the mainstay against GvHD, with still suboptimal outcomes. As GvHD is thought to result from dysregulated immunity and an imbalance of effector T cells over regulatory T cells (Tregs), several groups have focused on the alternative strategy of the adoptive Treg immunotherapy. The most common approach is the isolation of natural Tregs with however important limitations including their low frequency in the periphery, lack of specific markers for their purification as well as the high-cost requirements. To address these issues, several groups, including ours, have focused on the ex vivo manufacturing of induced Treg (iTreg) products, with two such products having proceeded to phase I/II trials with promising results despite complex production protocols. Our approach aims to replicate the feto-maternal tolerance mechanism mediated by the immunosuppressive Human Leukocyte Antigen-G (HLA-G) molecule, which is naturally expressed in the placenta. Postnatally, HLA-G is epigenetically silenced, but demethylation can reverse this. We demonstrated that exposing human peripheral T cells to hypomethylating agents (azacitidine or decitabine) induces a de novo and stable HLA-G expression, converting them into iTregs with strong immunosuppressive function. The purpose of this thesis is the clinical translation of these findings developing an iTreg immunotherapy. Initially, we optimized the manufacturing process to produce a large- and clinical-scale HLA-G+-enriched regulatory T-cell product, termed iG-Tregs, using a short, simplified and inexpensive protocol. Subsequent molecular and phenotypic characterization revealed that iG-Tregs exhibit traits of well-established regulatory populations by upregulating several inhibitory molecules. Functional assessments demonstrated that our product is highly suppressive and possesses a favorable safety profile paving the way for its clinical application. To facilitate this, we obtained accreditation by the national regulatory authorities for our laboratory, enabling the on-site Good Manufacturing Practice (GMP)-production of Advanced Therapeutic Medicinal Products such as iG-Tregs. Finally, the iG-Tregs generation protocol was successfully validated under GMP-conditions allowing the initiation of their clinical evaluation in a phase I/II dose-escalating study against GVHD (EudraCT 2021-006367-26). Preliminary results indicated successful production of iG-Tregs with a stable phenotype, characterized by a favorable safety profile and a persistence in the patients’ periphery post-infusion.

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Keywords

Allogeneic hematopoietic stem cell transplantation, Graft versus host disease, Adoptive immunotherapy, Regulatory T cells, HLA-G, Pharmacological hypomethylation, Advanced therapeutic medicinal product (ATMP), Good manufacturing practice (GMP), Clinical trial

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