Lentivector Iterations and Pre-Clinical Scale-Up/Toxicity Testing: Targeting Mobilized CD34+ Cells for Correction of Fabry Disease

Ju Huang; Aneal Khan; Bryan C Au; Dwayne L Barber; Lucía López-Vásquez; Nicole L Prokopishyn; Michel Boutin; Michael Rothe; Jack W Rip; Mona Abaoui; Murtaza S Nagree; Shaalee Dworski; Axel Schambach; Armand Keating; Michael L West; John Klassen; Patricia V Turner; Sandra Sirrs; C Anthony Rupar; Christiane Auray-Blais; Ronan Foley; Jeffrey A Medin

doi:10.1016/j.omtm.2017.05.003

Lentivector Iterations and Pre-Clinical Scale-Up/Toxicity Testing: Targeting Mobilized CD34⁺ Cells for Correction of Fabry Disease

Mol Ther Methods Clin Dev. 2017 May 12:5:241-258. doi: 10.1016/j.omtm.2017.05.003. eCollection 2017 Jun 16.

Authors

Ju Huang¹, Aneal Khan², Bryan C Au¹, Dwayne L Barber^{1

3

4}, Lucía López-Vásquez^{1

5}, Nicole L Prokopishyn⁶, Michel Boutin⁷, Michael Rothe⁸, Jack W Rip⁹, Mona Abaoui⁷, Murtaza S Nagree^{1

4}, Shaalee Dworski^{1

5}, Axel Schambach^{8

10}, Armand Keating¹, Michael L West¹¹, John Klassen¹², Patricia V Turner¹³, Sandra Sirrs¹⁴, C Anthony Rupar⁹, Christiane Auray-Blais⁷, Ronan Foley¹⁵, Jeffrey A Medin^{1

4

5

16}

Affiliations

¹ University Health Network, Toronto, ON M5G 1L7, Canada.
² Department of Medical Genetics, Alberta Children's Hospital Research Institute, Cumming School of Medicine, University of Calgary, Calgary, AB T2N 4N1, Canada.
³ Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON M5S 1A8, Canada.
⁴ Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada.
⁵ Institute of Medical Science, University of Toronto, Toronto, ON M5S 1A8, Canada.
⁶ Department of Pathology and Laboratory Medicine, University of Calgary and Cellular Therapy Laboratory, Calgary Lab Services, Calgary, AB T2N 1N4, Canada.
⁷ Division of Medical Genetics, Department of Pediatrics, Université de Sherbrooke, Sherbrooke, QC J1K 2R1, Canada.
⁸ Institute of Experimental Hematology, Hannover Medical School, 30625 Hannover, Germany.
⁹ Department of Pathology and Laboratory Medicine, Western University, London, ON N6A 5C1, Canada.
¹⁰ Division of Hematology/Oncology, Boston Children's Hospital, Harvard Medical School, Boston, MA 02115, USA.
¹¹ Division of Nephrology, Department of Medicine, Dalhousie University, Halifax, NS B3H 1V8, Canada.
¹² Department of Hematology, University of Calgary, Foothills Hospital, Calgary, AB T2N 2T9, Canada.
¹³ Department of Pathobiology, University of Guelph, Guelph, ON N1G 2W1, Canada.
¹⁴ Division of Endocrinology, Department of Medicine, University of British Columbia, Vancouver, BC V5Z 1M9, Canada.
¹⁵ Juravinski Hospital and Cancer Centre, Hamilton, ON L8V 5C2, Canada.
¹⁶ Medical College of Wisconsin, Milwaukee, WI 53226, USA.

Abstract

Fabry disease is a rare lysosomal storage disorder (LSD). We designed multiple recombinant lentivirus vectors (LVs) and tested their ability to engineer expression of human α-galactosidase A (α-gal A) in transduced Fabry patient CD34⁺ hematopoietic cells. We further investigated the safety and efficacy of a clinically directed vector, LV/AGA, in both ex vivo cell culture studies and animal models. Fabry mice transplanted with LV/AGA-transduced hematopoietic cells demonstrated α-gal A activity increases and lipid reductions in multiple tissues at 6 months after transplantation. Next we found that LV/AGA-transduced Fabry patient CD34⁺ hematopoietic cells produced even higher levels of α-gal A activity than normal CD34⁺ hematopoietic cells. We successfully transduced Fabry patient CD34⁺ hematopoietic cells with "near-clinical grade" LV/AGA in small-scale cultures and then validated a clinically directed scale-up transduction process in a GMP-compliant cell processing facility. LV-transduced Fabry patient CD34⁺ hematopoietic cells were subsequently infused into NOD/SCID/Fabry (NSF) mice; α-gal A activity corrections and lipid reductions were observed in several tissues 12 weeks after the xenotransplantation. Additional toxicology studies employing NSF mice xenotransplanted with the therapeutic cell product demonstrated minimal untoward effects. These data supported our successful clinical trial application (CTA) to Health Canada and opening of a "first-in-the-world" gene therapy trial for Fabry disease.

Keywords: GMP production; hematopoietic stem/progenitor cell; lysosomal storage disease; xenotransplantation; α-galactosidase A.