citations

                    Modeling, optimization, and comparable efficacy of T cell and hematopoietic stem cell gene editing for treating hyper-IgM syndrome
                

Authors:

Valentina Vavassori , Elisabetta Mercuri , Genni E Marcovecchio , Maria C Castiello , Giulia Schiroli , Luisa Albano , Carrie Margulies , Frank Buquicchio , Elena Fontana , Stefano Beretta , Ivan Merelli , Andrea Cappelleri , Paola Mv Rancoita , Vassilios Lougaris , Alessandro Plebani , Maria Kanariou , Arjan Lankester , Francesca Ferrua , Eugenio Scanziani , Cecilia Cotta-Ramusino , Anna Villa , Luigi Naldini , Pietro Genovese

In:

Source: EMBO Mol Med
Publication Date: (2021)
Issue: 13(3): e13545

Research Area:

Immunotherapy / Hematology

Gene Expression

Regenerative medicine

Cells used in publication:

CD34+ cell, human: Species: human; Tissue Origin: blood

Platform:

4D-Nucleofector® X-Unit

Experiment

After 3 days of stimulation, 5 × 105/106 primary T cells derived from both healthy donors and patients were washed with PBS and electroporated with 1.25 µM of ribonucleoprotein (RNP), unless otherwise specified (P3 Primary Cell 4D-Nucleofector X Kit, programs DS-130; Lonza). IDLV transduction was performed 24h before electroporation at MOI 100. AAV6 transduction was performed 150 after electroporation at a dose of 5 × 104 vg/cell, unless otherwise specified. dsDNA donors were electroporated along with RNP at a dose of 150 ng. After 3 days of stimulation, CB CD34+ cells were washed with PBS and electroporated with 2.5 µM of RNP (P3 Primary Cell 4DNucleofector X Kit, program EO-100; Lonza). 3 µg of GSE56 mRNA (Schiroli et al, 2019) was transfected along with RNP when indicated. AAV6 transduction was perform ed 15’ after electroporationprocedure at a dose of 104 vg/cell.

Abstract

Precise correction of the CD40LG gene in T cells and hematopoietic stem/progenitor cells (HSPC) holds promise for treating X-linked hyper-IgM Syndrome (HIGM1), but its actual therapeutic potential remains elusive. Here, we developed a one-size-fits-all editing strategy for effective T-cell correction, selection, and depletion and investigated the therapeutic potential of T-cell and HSPC therapies in the HIGM1 mouse model. Edited patients' derived CD4 T cells restored physiologically regulated CD40L expression and contact-dependent B-cell helper function. Adoptive transfer of wild-type T cells into conditioned HIGM1 mice rescued antigen-specific IgG responses and protected mice from a disease-relevant pathogen. We then obtained ~ 25% CD40LG editing in long-term repopulating human HSPC. Transplanting such proportion of wild-type HSPC in HIGM1 mice rescued immune functions similarly to T-cell therapy. Overall, our findings suggest that autologous edited T cells can provide immediate and substantial benefits to HIGM1 patients and position T-cell ahead of HSPC gene therapy because of easier translation, lower safety concerns and potentially comparable clinical benefits.

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