Coding region-specific destabilization of mRNA transcripts attenuates expression from retroviral vectors containing...

Class 1 aldehyde dehydrogenases (ALDH-1) function as drug resistance gene products by catalyzing the irreversible conversion of aldophosphamide, an active metabolite of cyclophosphamide, to an inert compound. Because the dose-limiting toxicity of cyclophosphamide is myelosuppression, retrovirus-mediated transfer of ALDH-1 to bone marrow cells has been proposed as a protective strategy. Here we show that expression of ALDH-1 vectors was problematic due to low levels of ALDH-1 mRNA accumulation. A number of vectors containing several different ALDH-1 cDNAs were introduced into a variety of different cell lines either by transfection or transduction. Detectable ALDH-1 protein and enzyme activity was only seen in one transfected cell clone. Cells transduced with ALDH-1 retroviral vectors had no detectable protein expression and very low levels of ALDH-1 mRNA. Analogous vectors containing other drug resistance cDNAs led to much higher levels of steady-state mRNA. The mRNA half-life from ALDH-1 vectors was less than 2 hr suggesting that vector-derived mRNAs were destabilized by ALDH-1 coding sequences. These results suggest that methods which increase the stability of ALDH-1 mRNAs will be important for increased drug resistance in retrovirally transduced hematopoietic cells.