NADPH oxidase overexpression in human colon cancers and rat colon tumors induced by 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)

NADPH oxidase/dual-oxidase (Nox/Duox) family members have been implicated in nuclear factor kappa-B (NF kappa B)-mediated inflammation and inflammation-associated pathologies. We sought to examine, for the first time, the role of Nox/Duox and NF kappa B in rats treated with the cooked meat heterocyclic amine carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP). In the PhIP-induced colon tumors obtained after 1 year, Nox1, Nox4, NF kappa B-p50 and NF kappa B-p65 were all highly overexpressed compared with their levels in adjacent normal-looking colonic mucosa. Nox1 and Nox4 mRNA and protein levels also were markedly elevated in a panel of primary human colon cancers, compared with their matched controls. In HT29 human colon cancer cells, Nox1 knockdown induced G1 cell cycle arrest, whereas in Caco-2 cells there was a strong apoptotic response, with increased levels of cleaved caspase-3, -6, -7 and poly(ADP-ribose) polymerase. Nox1 knockdown blocked lipopolysaccharide-induced phosphorylation of I kappa B kinase, inhibited the nuclear translocation of NF kappa B (p50 and p65) proteins, and attenuated NF kappa B DNA binding activity. There was a corresponding reduction in the expression of downstream NF kappa B targets, such as MYC, CCND1 and IL1 beta. The results provide the first evidence for a role of Nox1, Nox4 and NF kappa B in PhIP-induced colon carcinogenesis, including during the early stages before tumor onset. Collectively, the findings from this investigation and others suggest that further work is warranted on the role of Nox/Duox family members and NF kappa B in colon cancer development.