456. HOXB4 Overexpression Expands Short-Term Repopulating Cells in Nonhuman Primates and Dogs

Abstract

Human HOXB4 overexpression is able to extensively expand hematopoietic repopulating stem cells in murine models. Here we examined the effect of HOXB4 in two commonly used large-animal models using competitive repopulation assay. Three macaques and five dogs were transplanted with peripheral blood CD34+ cells transduced with a HOXB4/GFP or a control YFP gammaretroviral vector after lethal irradiation. Two dogs and one monkey died from transplant related complications. Follow-ups for the other three dogs and two monkeys are eight months and nine months, respectively. For all animals pretransplant gene transfer efficiencies in CD34+ cells were similar between HOXB4/GFP and YFP transduced cells: 45% (range 36-55%) and 38% (range 36- 40%) in macaque cells, and 48% (range 23-69%) and 51% (range 31-62%) in dog cells. In all 3 monkeys the initial gene transfer levels in vivo one week after transplantation were higher with the control YFP vector (average, 32% vs 25%). A similar trend was observed for three of the five dogs (average, 21% vs 11% for YFP vs GFP). In all the animals, dogs and monkeys, we observed a dramatic increase in HOXB4/GFP marked cells from 5-30% to 10-62% during the early engraftment period resulting in a significant difference between HOXB4/GFP and YFP marked granulocytes. The peak ratio between HOXB4-overexpressing to YFP-expressing in the three surviving dogs was approximately four at 5-6 weeks post-transplant. In the monkeys the peak ratio was up to ten at 5 weeks post-transplant. Interestingly, in all the dogs the HOXB4/GFP transduced cells decreased to within 2 fold of the control vector by week 15. Taqman PCR analysis of samples at the latest time point in all animals confirmed that the decrease in gene-marked cells was not due to transgene silencing. Subset analysis showed that the ratios of HOXB4-overexpressing cells to YFP-expressing cells were obviously higher for granulocytes than for lymphocytes in macaques and dogs, indicating that HOXB4 overexpression may inhibit lymphoid differentiation or/and promote higher level expansion of myeloid progenitors than lymphoid progenitors. In conclusion, our preclinical studies show that HOXB4 overexpression in CD34+ cells can expand hematopoietic cells that boost short-term engraftment.