Ki‐ras point mutation characteristically occurs frequently in human pancreatic cancer. To clarify the effect of antisense Ki‐ras RNA on various pancreatic cancer cell lines, a plasmid expressing an antisense Ki‐ras gene fragment (AS‐K‐ras‐LNSX) was transduced into seven human pancreatic cancer cell lines (AsPC‐1, MIA PaCa‐2, PANC‐1, PSN‐1, BxPC‐3, Hs 700T, and Hs 766T) by liposome‐mediated transfection. Western blot analysis showed that transfection of AS‐K‐ras‐LNSX led to a significant reduction in the amounts of Ki‐ras p21 protein in all the pancreatic cancer cell lines except BxPC‐3. The growth of pancreatic cancer cell lines having Ki‐ras point mutations (AsPC‐1, MIA PaCa‐2, PANC‐1, and PSN‐1) was suppressed after transduction of AS‐K‐ras‐LNSX, whereas the effect of the antisense construct on the growth was not significant in cell lines with a wild‐type Ki‐ras gene (BxPC‐3, Hs 700T, and Hs 766T). These results suggest that the pancreatic cancer cells with activated Ki‐ras depend heavily on a Ki‐ras p21–mediated growth signal pathway for their growth because they were far more susceptible to the suppression of the Ki‐ras p21 protein than the cells with wild‐type Ki‐ras. The remarkably increased dependence of the cancer‐cell growth circuitry on one or a few crucial regulatory molecules may thus be a common feature of the cancer cells and implies a novel rationale for the targeting of cancer therapy. Mol. Carcinog. 20:251–258, 1997. © 1997 Wiley‐Liss, Inc.