IN-VITRO AND IN-VIVO TRANSFER AND EXPRESSION OF HUMAN SURFACTANT SP-A-ASSOCIATED AND SP-B-ASSOCIATED PROTEIN CDNAS MEDIATED BY REPLICATION-DEFICIENT, RECOMBINANT ADENOVIRAL VECTORS

Congenital pulmonary alveolar proteinosis (CPAP) is a fatal disease of full-term infants that is unresponsive to current medical therapy. It is now recognized that at least some forms of this disorder are assoc iated with a deficiency of SP-B, one of the surfactant-associated prot eins, as well as probable aberrations in the surfactant-associated pro teins SP-A and SP-C. Given these developments, it is logical to hypoth esize that CPAP may be amenable to gene therapy, in which the human SP -B cDNA, and possibly the cDNAs of the other surfactant associated pro teins, are transferred to the epithelium of the lower respiratory trac t. We constructed replication-deficient, recombinant adenovirus vector s in which a constitutive viral promoter drives the expression of the DNAs for the surfactant-associated proteins, SP-B (AdCMV.SP-B) and SP- A (AdCMV.SP-A). Following infection of the human lung A549 epithelial cell line with these vectors in vitro, the appropriately sized mRNAs f or these cDNAs were detected, whereas cells infected with a control vi rus or uninfected cells produced none. Western blots demonstrated expr ession of these proteins, including appropriate processing of the hydr ophobic protein, SP-B. Following in vivo intratracheal infection of ra ts with these vectors, Northern analysis of the lungs revealed appropr iately sized mRNAs for these cDNAs whereas rats infected with control virus or uninfected rats show no hybridization with the human surfacta nt-associated protein probes. In the AdCMV.SP-A-infected rats. Western blots confirmed the overproduction of the human SP-A protein in both the bronchoalveolar lavage and lung homogenates compared to controls. Thus, it is feasible to utilize adenovirus vectors to transfer and exp ress the human surfactant associated protein cDNAs in vitro and in viv o, presenting a possible mode of therapy for CPAP, as well as other su rfactant deficiency states such as the neonatal respiratory distress s yndrome and possibly the adult respiratory distress syndrome.