Effect of changes in expression of the amphotropic retroviral receptor PiT-2 on transduction efficiency and viral titer: implications for gene therapy.
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.
Macdonald
C
C
Department of Haematology, University College London, UK.
Walker
S
S
Watts
M
M
Ings
S
S
Linch
D C
DC
Devereux
S
S
D016475:3T3 Cells; D000818:Animals; D018952:Antigens, CD34 / Q000032:analysis; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D017931:DNA Primers; D015316:Genetic Therapy; D006801:Humans; D051379:Mice; D011991:Receptors, Virus / Q000235:genetics*; D012190:Retroviridae / Q000235:genetics*; D014161:Transduction, Genetic
D018952:Antigens, CD34; D017931:DNA Primers; D011991:Receptors, Virus; C067981:leukemia virus receptor, gibbon ape