Cyclooxygenase-2 Inhibitors Down-regulate Osteopontin and Nr4a2-New Therapeutic Targets for Colorectal Cancers

Abstract

Background & Aims: Cyclooxygenase-2 inhibitors reduce colon cancer risk by mechanisms that are not fully understood. We performed microarray analysis of adenomas from ApcΔ14/+ mice to identify genes that respond to these drugs. Methods: ApcΔ14/+ mice were given a single daily injection of parecoxib for up to 9 weeks; intestinal tracts of these and control mice were analyzed by microarray analysis, immunohistochemistry, in situ hybridization, and quantitative real-time polymerase chain reaction. Findings were further assessed using Apclox/loxvil-CreERT2 mice, the CT26 cancer cell line, and human colon tumor samples. Results: Microarray analysis revealed that osteopontin, a marker of colon cancer progression, was down-regulated in polyps from ApcΔ14/+ mice given parecoxib compared with controls. ApcΔ14/+ mice given parecoxib had longer survival times and reduced polyp burdens. Osteopontin was quickly down-regulated by parecoxib in intestinal polyps from ApcΔ14/+ mice, and 2 components of the osteopontin regulatory network-the orphan nuclear receptor NR4A2 and Wnt/β-catenin signaling-were sequentially repressed. NR4A2 activated the osteopontin promoter in CT26 cells; this effect was blocked by mutation of the NR4A2 binding response element, cotransfection of a dominant-negative form of NR4A2, and small inhibitory RNA against NR4A2. NR4A2 levels were increased throughout tumor progression in ApcΔ14/+ mice but, unlike osteopontin, did not correlate with tumor stage. NR4A2 levels were reduced in adenomas from patients treated with rofecoxib. Conclusions: Down-regulation of osteopontin, probably through blockade of NR4A2 and Wnt signaling, is an important component of the antitumor activity of cyclooxygenase-2 inhibitors. These factors might be developed as therapeutic targets for intestinal cancers. © 2009 AGA Institute.