Endogenous stimulation of the EGF receptor massively increases c-fos and c-jun mrna in human pancreatic cancer cells

Using a model to demonstrate active autocrine growth, we have shown that EGF receptor-mediated pathways are critical in pancreatic cancer cells. In the present study, a human pancreatic cancer cell line (CAPAN-2) was used to investigate whether endogenous activation of the EGF receptor affected mRNA levels for c-fos and c-jun. mRNA detection was performed using quantitative RT-PCR (QRT-PCR). RNA competitors (cRNA) for c-fos and c-jun messages were synthesized. Co-reverse transcription and amplification of the appropriate cRNA and mRNA was performed using a single primer set. CAPAN-2 cells were grown in 25 cnr culture flasks in serum-free media. At different times, RNA was extracted and subsequently quantified using QRT-PCR. In parallel experiments, DNA synthesis was determined using thymidine incorporation and How cytometry. mRNA levels for c-fos and c-jun increased 20-fold after 1 hour of serum free culture. After 6 hours. mRNA levels returned to basal. Thymidine incorporation also increased transiently under these conditions, peaking after 6 hours and returning to basal after 12 hours. Cell cycle analysis revealed that the number of cells undergoing DNA replication doubled after 6 hours. These studies suggest that Fos and Jun, acting as transcription factors, may modulate the activity of particular genes following autocrine stimulation in pancreatic cancer cells. Identification of these genes may give clues as to the aggressive nature of this cancer.