Lentiviral vectors for precise expression to treat X-linked lymphoproliferative disease

Paul G Ayoub; Julia Gensheimer; Lindsay Lathrop; Colin Juett; Jason Quintos; Kevin Tam; Jack Reid; Feiyang Ma; Curtis Tam; Grace E McAuley; Devin Brown; Xiaomeng Wu; Ruixue Zhang; Kathryn Bradford; Roger P Hollis; Gay M Crooks; Donald B Kohn

doi:10.1016/j.omtm.2024.101323

Lentiviral vectors for precise expression to treat X-linked lymphoproliferative disease

Mol Ther Methods Clin Dev. 2024 Aug 20;32(4):101323. doi: 10.1016/j.omtm.2024.101323. eCollection 2024 Dec 12.

Authors

Paul G Ayoub¹, Julia Gensheimer^{2

3}, Lindsay Lathrop¹, Colin Juett⁴, Jason Quintos⁴, Kevin Tam⁴, Jack Reid⁴, Feiyang Ma⁴, Curtis Tam⁴, Grace E McAuley⁴, Devin Brown⁴, Xiaomeng Wu⁴, Ruixue Zhang⁴, Kathryn Bradford², Roger P Hollis⁴, Gay M Crooks^{2

5

6

7

3}, Donald B Kohn^{1

4

2

5

6}

Affiliations

¹ Department of Molecular & Medical Pharmacology, University of California, Los Angeles, Los Angeles, CA 90095, USA.
² David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA 90095, USA.
³ Department of Pathology & Laboratory Medicine, University of California, Los Angeles, Los Angeles, CA 90095, USA.
⁴ Department of Microbiology, Immunology & Molecular Genetics, University of California, Los Angeles, Los Angeles, CA 90095, USA.
⁵ Division of Pediatric Hematology-Oncology, University of California, Los Angeles, Los Angeles, CA 90095, USA.
⁶ Eli & Edythe Broad Center for Regenerative Medicine & Stem Cell Research, University of California, Los Angeles, Los Angeles, CA 90095, USA.
⁷ Jonsson Comprehensive Cancer Center, University of California, Los Angeles, Los Angeles, CA 90095, USA.

Abstract

X-linked lymphoproliferative disease (XLP1) results from SH2D1A gene mutations affecting the SLAM-associated protein (SAP). A regulated lentiviral vector (LV), XLP-SMART LV, designed to express SAP at therapeutic levels in T, NK, and NKT cells, is crucial for effective gene therapy. We experimentally identified 34 genomic regulatory elements of the SH2D1A gene and designed XLP-SMART LVs to emulate the lineage and stage-specific control of SAP. We screened them for their on-target enhancer activity in T, NK, and NKT cells and their off-target enhancer activity in B cell and myeloid populations. In combination, three enhancer elements increased SAP promoter expression up to 4-fold in on-target populations in vitro. NSG-Tg(Hu-IL15) xenograft studies with XLP-SMART LVs demonstrated up to 7-fold greater expression in on-target cells over a control EFS-LV, with no off-target expression. The XLP-SMART LVs exhibited stage-specific T and NK cell expression in peripheral blood, bone marrow, spleen, and thymic tissues (mimicking expression patterns of SAP). Transduction of XLP1 patient CD8+ T cells or BM CD34+ cells with XLP-SMART LVs restored restimulation-induced cell death and NK cytotoxicity to wild-type levels, respectively. These data demonstrate that it is feasible to create a lineage and stage-specific LV to restore the XLP1 phenotype by gene therapy.

Keywords: HSC; SAP; SH2D1A; XLP; enhancer; gene therapy; lentiviral vector; regulated; stem cell.