Abstract
The aim of this study was to quantify the impact of amphotropic retroviral receptor (PiT-2) levels on susceptibility to transduction and to determine whether the low level of PiT-2 found on CD34+ hematopoietic cells is within the range likely to compromise gene transfer. Receptor-deficient Chinese hamster ovary (CHO) cells were transfected with a PiT-2 construct that could be induced by the removal of tetracycline. The level of PiT-2 expression measured by virus binding in uninduced and in fully and partially induced transfectants correlated with the efficiency of transduction by an amphotropic retroviral reporter vector. Fully induced CHO-PiT-2 cells gave apparent viral titers similar to NIH 3T3 fibroblasts while addition of tetracycline reduced titers by up to 140-fold. Binding of the same vector preparation to purified CD34+ peripheral blood stem cells (PBSCs) was always less than to uninduced CHO-PiT-2 transfectants even after preincubation in 10-ng/ml concentrations of IL-3, IL-6, and stem cell factor, which increased retroviral binding by an average of 35%. The level of expression of the amphotropic retroviral receptor PiT-2 thus significantly limits transduction efficiency within the range observed in target cells of importance in human gene therapy.
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