Centre de Référence Maladies Rares

U1035 INSERM

Biotherapy of genetic diseases, inflammatory disorders and cancers (BMGIC)

FR: Biologie Fondamentale Appliquée à la Médecine

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Normal and Leukemic Hematopoietic Stem Cells team
Project 4.3

Engineering to conceive, develop, upgrade and conserve hematopoietic stem cells to clinical scale

Description:

Our project revolves around three major axis:

  1. to conceive, develop, and upgrade to clinical scale the procedures of ex vivo transformation of grafts based on hematopoietic stem and progenitor cells (HSC and HP) and mesenchymal stem cells (whatever sources : bone marrow, steady state peripheral blood, mobilized peripheral blood, cord blood, fat tissue…),
  2. the subsequent conservation of these grafts,
  3. the possibility to use gamma delta T cells as a potentially effective immunotherapy against leukemias.

Concerning items 1. and 2.

The conception of ex vivo procedure depends of the precise application context. With that respect, for each particular context, the stem and progenitor cell population of interest will be defined as well as strategy to amplify them according to specific needs. This development is related to the fundamental results of our and other groups and, in the first line, related to the “Oxygen Stem Cell Paradigm” including appropriate oxygenation, choice of cytokine cocktail interfering with cell response to hypoxia as well as the anti-oxydant capacity of medium.
In a first approach, the project aims to characterize the molecular mechanisms involved in this maintenance/amplification of HSC at low O2 concentrations with or without MSC using a direct approach of stem cells based on the very HSC-enriched population named “Side Population” (SP). The identification of gene and/or proteins up- or down-regulated in SP cells in hypoxic or anti-oxydant context will then serve to develop specific molecular tools allowing the control of HSC amplification and differentiation ex vivo graft expansion at 20% O2.
In parallel, the metabolic signature of HSC will be evidenced using SeaHorse technology (glucose consumption and type of glycolysis) but also the HRMAS RMN spectroscopy (intracellular metabolites concentration) and will help to determine the specific needs of HSC for their maintenance/amplification and to develop the optimal culture medium according to these needs. A similar approach with MSC will be driven and will enable to identify the type of HSC/MSC interactions indispensable for HSC maintenance in a low oxygen concentration context.
In addition to these ex vivo procedures conception, the graft engineering also concerns the conservation of ex vivo treated cells. This conservation is based both on hypothermia (+4°C) and frozen state (cryopreservation). To develop these complex research subjects, the approaches that we are exploring are largely based on our data concerning the phenomenon of cell hibernation strictly related to “Oxygen Stem Cell Paradigm”. The upgrading of this procedure to clinical scale is accelerated in our conditions, due to: i) fact that we use the clinical scale reagents and materials from the very beginning (conception) whenever possible and ii) the ultimate preclinical development is enabled since the clinical cell therapy capacities are within our reach. In addition, the successful development of ex vivo manipulated grafts is enabled by the possibility to use in vivo models (immune-deficient mice, miniaturized swine…) to engraft human cells.

Concerning item 3.

We explore the gamma delta T cells repertoire in patients with leukemia and the potential mechanisms of immune escape developed by these diseases. In particular, we explore the possibility that soluble TCR ligands might inhibit the recognition of the membrane ligands by the gamma delta T cell receptors. Our work aims to define in which cases it is possible to propose a gamma delta T cell adoptive immunotherapy in the acute myeloid leukemias, which are heterogeneous and evolutive pathologies.

Closely related collaboration with fundamental research scientists and clinicians is necessary for efficient conception, development and clinical up scaling of these procedures.

Related publications:

  • Repopulating hematopoietic stem cells from steady-state blood before and after ex vivo culture are enriched in the CD34+CD133+CXCR4low fraction. Lapostolle V, Chevaleyre J, Duchez P, Rodriguez L, Vlaski-Lafarge M, Sandvig I, Brunet de la Grange P, Ivanovic Z. Haematologica. (2018-10) PMID: 29858385

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INSERM U1035! - University of Bordeaux
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sarah.lesjean@inserm.fr