In Vitro Cyclooxygenase-2 Protein Expression and Enzymatic Activity in Neoplastic Cells

Background: Cyclooxygenase‐2 (COX‐2) and its principle enzymatic metabolite, prostaglandin E2 (PGE2), are implicated in cancer progression. Based upon immunohistochemical (IHC) evidence that several tumor types in animals overexpress COX‐2 protein, COX‐2 inhibitors are used as anticancer agents in dogs and cats. Hypothesis: IHC is inaccurate for assessing tumor‐associated COX‐2 protein and enzymatic activity. Methods: Five mammalian cell lines were assessed for COX‐2 protein expression by IHC and Western blot analysis (WB), and functional COX‐2 activity was based upon PGE2 production. Results: Detection of COX‐2 protein by IHC and WB were in agreement in 4 of 5 cell lines. In 1 cell line that lacked COX‐2 gene transcription because of promoter hypermethylation (HCT‐116), IHC produced false‐positive staining for COX‐2 protein expression. Functional COX‐2 enzymatic activity was dissociated from relative IHC‐based COX‐2 protein expression in 2 cell lines (RPMI 2650 and SCCF1). The RPMI 2650 cell line demonstrated strong COX‐2 protein expression but minimal PGE2 production. Conclusions and Clinical Importance: Western blot is more accurate than IHC for the detection of COX‐2 protein in the cell lines studied. Furthermore, the semiquantitative identification of COX‐2 protein by IHC or WB does not necessarily correlate with enzymatic activity. Based upon the potential inaccuracy of IHC and dissociation of COX‐2 protein expression from enzymatic activity, the practice of instituting treatment of tumors with COX‐2 inhibitors based solely on IHC results should be reconsidered.