Aldo-keto reductase family 1 B10 gene silencing results in growth inhibition of colorectal cancer cells: Implication for cancer intervention

Aldo-keto reductase family 1 B10 (AKR1B10), a member of aldoketo reductase superfamily, is overexpressed in human hepatocelWar carcinoma, lung squamous cell carcinoma and lung adenocarcinoma. Our previous study had demonstrated that the ectopic expression of AKR1B10 in 293T cells promotes cell proliferation. To evaluate its potential as a target for cancer intervention, in the current study we knocked down AKR1B10 expression in HCT-8 cells derived from a colorectal carcinoma, using chemically synthesized small interfering RNA (siRNA). The siRNA 1, targeted to encoding region, downregulated AKR1B10 expression by more than 60%, and siRNA 2, targeted to 3' untranslational region, reduced AKR1B10 expression by more than 95%. AKR1B10 silencing resulted in approximately a 50% decrease in cell growth rate and nearly 40% suppression of DNA synthesis. More importantly, AKR1B10 downregulation significantly reduced focus formation rate and colony size in semisolid culture, indicating the critical role of AKR1B10 in HCT-8 cell proliferation. Recombinant AKR1B10 protein showed strong enzymatic activity to acrolein and crotonaldehyde, with K-m = 110.1 +/- 12.2 pM and V-max = 3,122.0 +/- 64.7 nmol/mg protein/min for acrolein and K-m = 86.7 +/- 14.3 mu M and V-max = 2,647.5 +/- 132.2 nmol/mg protein/min for crotonaldehyde. AKR1B10 downregulation enhanced the susceptibility of HCT-8 cells to acrolein (25 pM) and crotonaldehyde (50 pM), resulting in rapid oncotic cell death characterized with lactate dehydrogenase efflux and annexin-V staining. These results suggest that AKR1B10 may regulate cell proliferation and cellular response to additional carbonyl stress, thus being a potential target for cancer intervention. (c) 2007 Wiley-Liss, Inc.