Digitoxin promotes apoptosis and inhibits proliferation and migration by reducing HIF-1α and STAT3 in KRAS mutant human colon cancer cells

Colon cancer patients with mutant KRAS are resistant to cetuximab, an antibody directed against the epidermal growth factor receptor. New treatment options are needed to improve survival in patients with KRAS mutated colorectal cancer. Digitoxin is a cardiotonic drug, which has been demonstrated to exhibit anticancer effects in a number of cancers. However, the anticancer mechanisms of digitoxin in KRAS mutant human colon cancer cells remain elusive. Our result demonstrated that digitoxin but not cetuximab markedly decreased the expression of hypoxia-inducible factor-1α (HIF-1α), signal transducer and activator of transcription 3 (STAT3) and p-STAT3 protein in KRAS mutant colon cancer cells. Further analysis revealed that digitoxin inhibited HIF-1α protein synthesis, without affecting the expression level of HIF-1α mRNA or degradation of HIF-1α protein. The phosphorylation levels of ribosomal protein S6 kinase (p70S6K) and eIF4E binding protein-1 (4E-BP1) were significantly suppressed by digitoxin. Digitoxin inhibited the expression and activation of STAT3 through upregulation of phosphatase and tensin homolog deleted on chromosome ten (PTEN), SHP1 and protein inhibitors of activated STAT3 (PIAS3) and direct binding to STAT3. Meanwhile, digitoxin inhibited HIF-1α in STAT3-independent manner in KRAS mutant colon cancer cells. Moreover, digitoxin promoted apoptosis and inhibited proliferation and migration, which was potentially mediated by suppression of HIF-1α and STAT3. We also found that digitoxin administration inhibited tumor growth in a mouse xenograft model. Taken together, our findings highlight the therapeutic potential of digitoxin for the treatment of cetuximab-resistant human colon cancer.