The investigators plan to investigate the mechanisms of glucocorticoid (GC) induced growth inhibition of lung cancer, and whether GCs may be effective chemopreventive agents for lung cancer. They hypothesize that GCs, acting through the GC receptor (GCR), inhibit the extracellular signal-regulated kinase (ERK)/mitogen activated protein (MAP) pathway, which in turn suppresses c-Fos gene expression, and thus down-regulates the activity of the transcription factor AP-1, and inhibits cell proliferation. They further propose that GCs may inhibit malignant transformation. Therefore, GCs may be useful in chemoprevention of lung cancer.
Specific Aim 1 is to determine the mechanisms of GC-induced inhibition of c-Fos expression in lung cancer. Using a human non-small cell lung carcinoma cell line, they will first confirm the effect of GCs on c-Fos expression by Northern analysis. They will then determine whether this effect is at the level of transcription using nuclear run-off assay and a fos- luciferase reporter construct. To evaluate the role of the MAP kinase pathways, they will use inhibitors to the ERK and JNK pathways, and transfection of a plasmid containing a constitutively active mutant ERK. A GCR antagonist will help determine that the GC-induced changes in c-Fos expression are mediated by the GCR. Finally, they will confirm that GC inhibits AP-1 activity using an AP-1-luciferase construct, and then investigate the mechanism of this inhibition using an electrophoretic mobility shift assay.
Specific Aim 2 is to determine whether GCs can inhibit malignant transformation. They will test whether GC can inhibit benzo(a)pyrene (BaP) induced transformation of normal (immortalized) human bronchial epithelial (BEAS-21B) cells. They will use anchorage independent growth, colony forming efficiency and tumorigenicity in nude mice to assess for transformation.
Specific Aim 3 is to determine whether GCs may be effective chemopreventive agents for lung cancer. They will investigate whether GCs produce the same effects on the ERK/MAP kinase pathway in vivo as they do in vitro, and whether these affects correlate with decreased proliferation and morphological changes. They will recruit cigarette smokers and perform bronchoscopy with endobronchial biopsies pre- and post-treatment with GC. They will evaluate the affect of GC on cell morphology, a proliferation index and the expression of the MAP kinases and c-Fos. If their studies in human subjects confirm their in-vitro data, they would like to pursue a clinical chemoprevention trial using long-term inhaled GCs in patients at high-risk for lung cancer.
