Background aims: Drug-resistance genes have been explored as powerful in vivo selection markers in hematopoietic cell gene therapy, and cytidine deaminase (CDD) represents a particularly attractive candidate given the virtual absence of non-hematopoietic side-effects after low/intermediate dose application of the associated drug cytosine-arabinoside (Ara-C).
Methods: We investigated the in vivo selection potential of CDD overexpression and prolonged low/intermediate-dose Ara-C application in a murine model. Furthermore, non-transplanted mice were utilized to study Ara-C toxicity in different hematopoietic cell compartments.
Results: Significant protection of myelo- and thrombopoiesis and up to 6-fold in vivo enrichment of CDD-transduced hematopoietic cells was observed. Enrichment was most robust early after Ara-C application and was correlated with dosage and duration of chemotherapy. Enrichment remained significant for several weeks, indicating selection at the level of a progenitor population. This notion was supported by Ara-C toxicity studies, demonstrating profound hematotoxicity and a marked delay in hematopoietic recovery, specifically in the progenitor/stem cell compartment after low/intermediate-dose Ara-C.
Conclusions: These data support the concept of CDD/Ara-C as a clinically applicable in vivo selection system in hematopoietic gene therapy. The data also demonstrate marked differences in hematotoxicity between alternative Ara-C dosing schemes and suggest thorough in vivo toxicity studies to optimize further Ara-C dosing en route to safe and stable enrichment of gene-corrected hematopoiesis.