Mutations in the K-ras gene preventing GTPase-dependent down-regulation of the protein are prevalent in a variety of carcinomas (including lung, colon and pancreatic). Despite the implication that sustained activity of K-ras is oncogenic, the expression of oncogenic ras alleles has been shown to induce growth arrest and senescence in human and mouse primary cells, and wild-type K-ras is a tumor suppressor in mouse and probably human lung. Previous work in our laboratory with immortalized, non-transformed lung epithelial cells (mouse E10 and C10, and human HPL lines) discerned an increase in total K-ras protein at growth arrest (induced by cell confluence in culture), and an even greater corresponding increase in activated K-ras. In our current line of investigation, we have observed that conditioned media from lung cells, that have been growth arrested at confluence, upregulate K-ras activity in proliferating cells of the same type by an average of 15-fold. This establishes involvement of a secreted factor. Furthermore, we have demonstrated that this increase in K-ras activity is linked to the EGF receptor (EGFR) through a series of experiments utilizing EGFR inhibitors. The EGFR inhibitor, PD 153035 (1 μM, Calbiochem) reduced K-ras activity by 73 (+/–2)% (P=0.0008) in confluent growth arrested cells. K-ras activity stimulated in proliferating E10 cells by conditioned media from growth arrested cells was similarly inhibited, 69 (+/−4)% (P=0.0028), by the EGFR inhibitor. The low K-ras activity in untreated proliferating E10 cells was inhibited to an insignificant extent by PD 153035, 22(+/−6)% (P=0.071). Currently we are fractionating the conditioned media to discover the factor(s) responsible for EGFR-dependent activation of K-ras in these lung cells. Definition of the control of K-ras in peripheral lung epithelial cells is critical to therapeutic targeting of this critical oncogene/tumor suppressor gene.
[Proc Amer Assoc Cancer Res, Volume 46, 2005]