1090. Enzyme Replacement Therapy with Pegylated Adenosine Deaminase (PEG-ADA) Does Not Impede Immune Reconstitution Following Transplantation of Gene-Corrected Bone Marrow Cells in the Murine Model of ADA-SCID

Abstract

In humans, the genetic deficiency of adenosine deaminase (ADA) is responsible for approximately 20% of Severe Combined Immune Deficiency (SCID) and results in severe lymphopenia. The current standard is either an allogeneic bone marrow transplant from an MHC-matched donor or enzyme replacement therapy (ERT) with PEG-ADA; BMT from a donor other than a matched sibling has poor outcome. Because of a postulated selected advantage for corrected T cells, ADA-deficient SCID has been an early candidate disease for gene therapy. Recent studies by Aiuti and co-workers using oncoretroviral vectors to transduce CD34+ cells from the bone marrow of ADA-deficient SCID patients, together with cessation of ERT and moderate cytoreduction with Busulfan show clinical benefit, however, the relative roles of the cytoreduction and the cessation of PEG-ADA have yet to be elucidated. A murine model of ADA-deficient SCID was produced by Rodney Kellems and co-workers (U.Texas, Houston) by ADA gene knock-out. These mice die from non-infectious pulmonary insufficiency between days 19-20 post-natal, unless maintained on enzyme replacement therapy (ERT) with PEG-ADA. In these studies, we evaluated the role of ERT as a single variable on the ability of gene-corrected cells to repopulate 10-week old ADA-deficient mice that received a cytoablative dosage of total-body irradiation (900 cGy) prior to receiving ADA-deficient marrow transduced with the retroviral vector MND-MFG-huADA. These mice, previously maintained on ERT, were either continued on the same low dose ERT (n=8) or not (n=8). ADA -/- mice receiving ERT had 89% survival and those not receiving ERT had 75% survival after radiation and BMT. Mice were euthanized between 4-5 months post transplant and were perfused with PBS prior to tissue removal. Proviral marking was assessed by qPCR in the peripheral blood leukocytes, thymus, spleen, bone marrow, lung and liver, as well as in sorted cell populations from the thymus (CD4/8), spleen (CD19) and bone marrow (CD11b). Gene marking was seen in all of the tissues assessed with little difference between those animals receiving ERT and those that did not. Although not significant, marking was slightly higher in the hematopoietic tissues analyzed from mice receiving ERT. Proviral marking (copies/ cell) was highest in the thymus and spleen and lowest in the bone marrow. Cell populations enriched from these organs had marking similar to the organ from which they were isolated. Marking in the lung and liver was similar to that seen in the hematopoietic tissues and may be relevant due to the severe lung and hepatocellular pathology seen in ADA-deficient mice. With or without ERT, ADA- deficient mice receiving gene therapy had improved immune cellularity and function compared to ADA-deficient mice receiving mock transductions and maintained on ERT. In these studies, we have observed no selective lymphoid expansion of the gene-corrected T cells, whether ERT was given or withheld, but nevertheless there was immunologic improvement from host (ADA-deficient) cells.