Surfactant Protein A-Directed Toxin Gene Kills Lung Cancer Cells In Vitro

Abstract

Human surfactant protein A (SPA) expression is considered a marker of respiratory epithelial differentiation. Non-small cell lung cancers (NSCLC) are respiratory epithelial derivatives, and it was previously shown that a minority of these cancers expressed SPA, presumably a consequence of their respiratory epithelial origin. In the studies reported here, SPA-I gene transcriptional regulatory sequences were localized to a 2.75-kb genomic 5′-flanking region fragment obtained by screening a human genomic library. The 2.75-kb fragment was used to direct a luciferase coding sequence transcriptionally within a plasmid construct. In plasmid transduction experiments, the SPA-directed luciferase plasmid produced significant luciferase activity in the SPA-expressing NSCLC cell line, H441, but only background levels in the non-SPA-expressing A549 cells. Because Northern blot analysis of resected NSCLC showed that the majority expressed SPA, an SPA-transcriptional targeting strategy was investigated using chimeric toxin genes comprising the coding sequence for herpes simplex virus thymidine kinase (HSV-TK) under transcriptional control of SPA or SV40 regulatory sequences. As expected, transduction of the constitutive, SV40-directed plasmid followed by ganciclovir treatment reduced numbers of both the A549 and H441 cells. In contrast, the SPA-directed plasmid reduced only the SPA-expressing H441 cells and had no significant effect on the A549 cells. The results of these in vivo experiments suggest the concept of transcriptionally directing toxin genes with SPA can produce targeted toxicity in NSCLC.

Overview summary

Smith et al. have found that the majority of resected lung cancers express the surfactant protein A (SPA) gene. The regulatory sequences of the human SPA-1 gene were localized to the 5′-flanking region of the gene by reporter assays. These regulatory sequences were used to direct transcriptionally the expression of herpes simplex virus thymidine kinase (HSV-TK) coding sequence within plasmid constructs. Using the adenovirus polylysine-mediated gene transfer method under conditions that transduced a small minority of the cells, significant numbers of the unselected, SPA-expressing, cancer cells were killed in contrast to non-SPA-expressing cells that were not significantly affected by similar treatment. These results suggest that regulatory sequences from respiratory epithelial gene expressed in the majority of lung cancers can be used to target toxicity transcriptionally to those cancers.