Nitric oxide (NO) has been implicated as a potential causative factor for endogenous p53 mutations in gastrointestinal
malignancy. To investigate the role of NO in esophageal
adenocarcinoma (EADC), we studied patterns of p53 mutations, expression of
inducible nitric oxide synthase (iNOS) and the tissue accumulation of
nitrotyrosine (NTS), a stable reaction product of NO and a marker for cellular
protein damage, in human premalignant and malignant esophageal epithelia. Tissues were obtained from patients with
gastroesophageal reflux disease (
GERD)-induced
esophagitis (n = 76),
Barrett's esophagus (BE; n = 119) and primary EADC (n = 54).
DNA sequencing was used to characterize p53 mutations, RT-PCR to study iNOS
mRNA expression, and immunohistochemistry to study NTS. Relative to self-matched normal epithelia, a progressive increase in iNOS
mRNA expression was seen in
GERD (30%; 23/76), BE (48%; 57/119), and EADC (63%; 34/54) tissues (P < 0.001). Among patients with EADC, elevated levels of NTS immunoreactivity were more frequent in
tumors with p53 mutations (11/21; 52%) compared with
tumors with wild-type p53 (9/33; 27%; P = 0.063), and specifically in
tumors with p53 mutations at CpG dinucleotides (10/12; 83%) compared with non-CpG p53 mutations (1/9; 11%; P = 0.008). The increasing frequency of iNOS (
mRNA) overexpression in
GERD, BE and EADC supports the hypothesis that an active inflammatory process, most likely a consequence of
GERD, underlies molecular progression to EADC. The highly significant association between NTS, reflecting chronic NO-induced cellular
protein damage, and endogenous p53 mutations at CpG dinucleotides, provides further evidence for a molecular link between chronic
inflammation and esophageal
malignancy.