Human colon cancer cell lines express epidermal growth factor (EGF) mRNA, secrete EGF and may respond to it via the cell-surface EGF receptor (EGFR). Expression of these molecules in human colon and colon tumor, however, is not clear. Reverse transcription-polymerase chain reaction (RT-PCR) analyses of RNA prepared from paired normal human colon and colon tumor samples from 12 individuals followed by Southern blotting analyses of the RT-PCR products revealed a major fragment of 527 bp and a minor fragment of 404 bp that hybridized to a human EGF cDNA probe under stringent conditions. Identical results were obtained from 8 human colon cancer cell lines. Cloning and sequencing of PCR products confirmed that both fragments were from the human EGF gene; the 527-bp fragment corresponded exactly to nucleotides 2,891 to 3,417 of the human EGF mRNA reported by others. A deletion of 123 nucleotides (nucleotides 3,172 to 3,294) was found in the 404-bp fragment. Immunohistochemical studies using cyostat sections of human colon specimens showed that EGF was expressed in the human colon and that expression was restricted to the epithelial colonic crypt cells and epithelium-derived cancer cells. Since EGF and EGF-related molecules are potent mitogens that mediated their effect through the EGFR, we also determined the efficacy of anti-sense EGFR RNA in circumventing the EGFR-related pathway of proliferation. Expression of anti-sense EGFR RNA, by transfection with an inducible anti-sense EGFR expression vector, down-regulated cell-surface EGFR expression and proliferation of these cells and their ability to grow in soft agar. Anti-sense EGFR RNA was found to be an anti-proliferative agent in both relatively non-aggressive and highly aggressive human colon cancer cells.