Scaling up for preclinical study of hematopoietic stem cells from patients with Immune Aplastic Anemia expanded with UM171

Abstract

Bone marrow failure syndromes are a group of heterogeneous diseases that are characterized by the failure of hematopoietic stem and progenitor cells (HSCs), which can be caused by multiple factors that cause a reduction in HSCs, several factors can lead to such a reduction, among them the immune attack through cytotoxic T cells that attack HSCs and induce apoptosis. This attack is responsible for the development of acquired immune aplastic anemia (AA). During immune AA, cytotoxic Th1 cells release cytokines that trigger changes in cell cycle and cell transcription, leading to HSC apoptosis, causing pancytopenia. Current treatments are limited and are hampered by the lack of compatible donors and the side effects caused. The current standard treatment is allogeneic hematopoietic stem cell transplantation, however the complications related to such transplantation are graft rejection and graft-versus-host disease (GVHD), in addition to the fact that the risk of death increases with age. of the patient. Another treatment alternative would be immunosuppressive therapy (IST), which has a high rate of response and long-term survival, however, approximately one-fifth of patients do not respond and almost half of patients with the disease ends up returning. Therefore, our group has been studying a new therapeutic alternative for patients with bone marrow failure, through the development of ex vivo CD34+ cell expansion processes from the patients themselves. The UM171 molecule, a pyrimidoindole derivative, is an alternative, as it is capable of stimulating the expansion of human HSCs, in addition to inhibiting their differentiation, allowing the long-term repopulation of these cells when transplanted in animal models. It is known that UM171 blocks epigenetic changes in the cell through the potentiation of complexes responsible for degrading the CoREST protein complex, thus allowing the HSC self-renewal process. Preliminary data from our group demonstrate the potential of UM171 in the expansion of HSCs in patients with acquired immune AA, suggesting that UM171 is capable of promoting in vitro expansion of HSCs without causing genetic changes in the cells. In this project, we propose to evaluate the capacity of UM171 in the escalation of HSCs in patients with immune AA. The bone marrow of seven patients with immune AA will be collected, and HSCs will be isolated and cultured in the presence of UM171 for expansion. We will verify the phenotypic and functional expansion of HSCs in open and closed systems, genomic stability through next-generation sequencing, and analyze the engraftment potential of these expanded HSCs. Cells will be subjected to long-term culture to determine their ability to form colonies. The safety of the process will be investigated through the analysis of telomere length, molecular cytogenetics, the emergence of somatic mutations, and long-term xenotransplantation. The results may contribute to the conduction of human clinical studies using UM171. (AU)