TRANSFER OF A CONSTITUTIVE VIRAL PROMOTER CYSTIC-FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR CDNA TO HUMAN EPITHELIAL-CELLS CONVEYS RESISTANCE TO DOWN-REGULATION OF CAMP-REGULATED CL- SECRETION IN THE PRESENCE OF INFLAMMATORY STIMULI

The expression of the cystic fibrosis transmembrane conductance regula tor (CFTR) gene can be downregulated by inflammatory stimuli such as p horbol myristate acetate (PMA). Since the respiratory manifestations o f cystic fibrosis (CF) are characterized by intense chronic airway inf lammation very early in life, successful gene therapy far CF will requ ire that expression of the transferred normal CFTR gene be resistant t o down-regulation by inflammatory mediators. To evaluate the concept t hat a viral promoter - human CFTR cDNA unit would be resistant to this form of down-regulation, a retrovirus promoter (5' long terminal repe at of the Moloney murine leukemia virus)- human CFTR cDNA unit was tra nsferred to T84 human colon carcinoma cell line using a retrovirus vec tor. Exposure of the retrovirus-modified T84 cells to PMA resulted in down-regulation of the endogenous CFTR mRNA transcripts (6.5 kb), but did not affect the level of exogenous CFTR transcripts (8.0 kb). Impor tantly, in parallel with the persistence of the exogenous CFTR transcr ipts, the modified cells still maintained cAMP-regulated Cl- secretion in the presence of PMA. These in vitro data suggest that a constituti ve viral promoter- CFTR cDNA unit should be resistant to modulation by inflammatory stimuli, a likely requirement for successful gene therap y for CF.