Affiliations of authors: A. J. Marrogi, M. A. Khan, J. A. Welsh, H. Rahim, S. P. Hussain, C. C. Harris (Laboratory of Human Carcinogenesis, Center for Cancer Research), H. E. van Gijssel, M. C. Poirier (Laboratory of Cellular Carcinogenesis and Tumor Promotion, Center for Cancer Research), National Cancer Institute, Bethesda, MD; A. J. Demetris, Department of Pathology and Laboratory Medicine, University of Pittsburgh, PA; K. W. Kowdley, Department of Surgery, University of Washington, Seattle; J. Nair, H. Bartsch, German Cancer Center, Heidelberg, Germany; N. Okby, K. G. Ishak, Department of Hepatic and Gastrointestinal Pathology, The Armed Forces Institute of Pathology, Washington, DC.
Correspondence to: Curtis C. Harris, M.D., National Institutes of Health, Bldg. 37, Rm. 2C05, MSC 4255, Bethesda, MD 208924255 (e-mail: Curtis_Harris{at}nih.gov).
The incidence of hepatocellular carcinoma (HCC) in individuals with hereditary hemochromatosis is 200 times greater than in the general population (1). This increased risk of HCC may be the result of deregulation of oxidation reduction and generation of reactive oxygen species from free iron, directly through the Fenton reaction and indirectly through the acceleration of lipid peroxidation [reviewed in (2)]. In an ironnitrilotriacetic acid rat model of hemochromatosis, renal samples showed an increase in the levels of several reactive intermediates, including 4-hydroxy-2-nonenal and malondialdehyde (3,4), both of which are known to be cytotoxic and genotoxic (5,6). These increases were accompanied by decreased availability of systems that protect against oxidation by iron, such as vitamin E levels, the glutathione system, thiol-specific antioxidants, and superoxide dismutase (7,8). In addition, excess iron provides a strong growth-promotion advantage in human hepatoma cell lines and chemically induces carcinomas in experimental animals (9,10)
Previously, we have reported higher frequencies of p53 mutations, including G : C to T : A transversions at codon 249 and C : G to A : T and C : G to T : A changes at codon 250 in liver tissue samples from cancer-free patients with either hemochromatosis or Wilson's disease (11). We present here a detailed analysis of 14 cases of hemochromatosis-associated HCC (Fig. 1, A), including the p53 mutation spectrum, Prussian blue stain for iron (Fig. 1
, B), and immunohistochemical analysis of p53, nitric oxide synthase-2 (NOS-2), and cyclooxygenase-2 (COX-2). The study protocol was submitted to the human subjects committee at our institution but was exempted, because the tissues were obtained in the course of patients' treatment and without the knowledge of patients' identities. Histologically, all tumors, except three, were well differentiated. Nuclear p53 overexpression was observed in eight of 14 HCC cases (Fig. 1
, C) with varying intensity, including two (+1), two (+2), three (+3), and one (+4), according to the following scoring scheme: less than 10% nuclear reactivity = 0, greater than 11% to less than 25% = +1, greater than 25% to less than 50% = +2, greater than 50% to less than 90% = +3, and greater than 90% = +4 (12,13). Cytoplasmic NOS-2 immunoreactivity was seen in one HCC and in eight adjacent non-neoplastic regenerative nodules (Fig. 1
, D). COX-2 overexpression was not seen in these cases of HCC.
|
|
We thank Dorothea Dudek for editorial assistance.
REFERENCES
1 Hsing AW, McLaughlin JK, Olsen JH, Mellemkjar L, Wacholder S, Fraumeni JF Jr. Cancer risk following primary hemochromatosis: a population-based cohort study in Denmark. Int J Cancer 1995;60:1602.[Medline]
2 Meneghini R. Iron homeostasis, oxidative stress, and DNA damage. Free Radic Biol Med 1997;23:78392.[Medline]
3 Toyokuni S, Uchida K, Okamoto K, Hattori-Nakakuki Y, Hiai H, Stadtman ER. Formation of 4-hydroxy-2-nonenal-modified proteins in the renal proximal tubules of rats treated with a renal carcinogen, ferric nitrilotriacetate. Proc Natl Acad Sci U S A 1994;91:261620.[Abstract]
4 Fukuda A, Osawa T, Oda H, Tanaka T, Toyokuni S, Uchida K. Oxidative stress response in iron-induced acute nephrotoxicity: enhanced expression of heat shock protein 90. Biochem Biophys Res Commun 1996;219:7681.[Medline]
5 Esterbauer H, Eckl P, Ortner A. Possible mutagens derived from lipids and lipid precursors. Mutat Res 1990;238:22333.[Medline]
6 Esterbauer H, Schaur RJ, Zollner H. Chemistry and biochemistry of 4-hydroxynonenal, malonaldehyde and related aldehydes. Free Radic Biol Med 1991;11:81128.[Medline]
7
Cheng KC, Cahill DS, Hasai H, Nishimura S, Loeb LA. 8-Hydroxyguanine, an abundant form of oxidative DNA damage, causes CT and A
C substitutions. J Biol Chem 1992;267:16672.
8 Dabbagh AJ, Mannion T, Lynch SM, Frei B. The effect of iron overload on rat plasma and liver oxidant status in vivo.Biochem J 1994;300:799803.[Medline]
9 Hann HW, Stahlhut MW, Hann CL. Effect of iron and desferoxamine on cell growth and in vitro ferritin synthesis in human hepatoma cell lines. Hepatology 1990;11:5669.[Medline]
10 Smith AG, Carthew P, Clothier B, Constantin D, Francis JE, Madra S. Synergy of iron in the toxicity and carcinogenicity of polychlorinated biphenyls (PCBs) and related chemicals. Toxicol Lett 1995;8283:94550.
11
Hussain SP, Raja K, Amstad PA, Sawyer M, Trudel LJ, Wogan GN, et al. Increased p53 mutation load in nontumorous human liver of Wilson disease and hemochromatosis: oxyradical overload diseases. Proc Natl Acad Sci U S A 2000;97:127705.
12
Marrogi AJ, Travis WD, Welsh JA, Khan MA, Rahim H, Tazelaar H, et al. Nitric oxide synthase, cyclooxygenase 2, and vascular endothelial growth factor in the angiogenesis of non-small cell lung carcinoma. Clin Cancer Res 2000;6:473944.
13 Przygodzki RM, Bennett WP, Guinee DG Jr, Khan MA, Freedman A, Shields PG, et al. p53 mutation spectrum in relation to GSTM1, CYP1A1 and CYP2E1 in surgically treated patients with non-small cell lung cancer. Pharmacogenetics 1998;8:50311.[Medline]
14
Ahrendt SA, Halachmi S, Chow JT, Wu L, Halachmi N, Yang SC, et al. Rapid p53 sequence analysis in primary lung cancer using an oligonucleotide probe array. Proc Natl Acad Sci U S A 1999;96:73827.
15 Vautier G, Bomford AB, Portmann BC, Metivier E, Williams R, Ryder SD. p53 mutations in British patients with hepatocellular carcinoma: clustering in genetic hemochromatosis. Gastroenterology 1999;117:15460.[Medline]
16 Hussain SP, Harris CC. p53 mutation spectrum and load: the generation of hypotheses linking the exposure of endogenous or exogenous carcinogens to human cancer. Mutat Res 1999;428:2332.[Medline]
17 Hachiya M, Chumakov A, Miller CW, Akashi M, Said J, Koeffler HP. Mutant p53 proteins behave in a dominant, negative fashion in vivo.Anticancer Res 1994;14:18539.[Medline]
18
Murphy KL, Dennis AP, Rosen JM. A gain of function p53 mutant promotes both genomic instability and cell survival in a novel p53-null mammary epithelial cell model. FASEB J 2000;14:2291302.
19
Ambs S, Bennett WP, Merriam WG, Ogunfusika MO, Oser SM, Harrington AM, et al. Relationship between p53 mutations and inducible nitric oxide synthase expression in human colorectal cancer. J Natl Cancer Inst 1999;91:868.
20 Wilson KT, Fu S, Ramanujam KS, Meltzer SJ. Increased expression of inducible nitric oxide synthase and cyclooxygenase-2 in Barrett's esophagus and associated adenocarcinomas. Cancer Res 1998;58:292934.[Abstract]
21
Goto T, Haruma K, Kitadai Y, Ito M, Yoshihara M, Sumii K, et al. Enhanced expression of inducible nitric oxide synthase and nitrotyrosine in gastric mucosa of gastric cancer patients. Clin Cancer Res 1999;5:14115.
22
Marrogi A, Pass HI, Khan M, Metheny-Barlow LJ, Harris CC, Gerwin BI. Human mesothelioma samples overexpress both cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (NOS2): in vitro antiproliferative effects of a COX-2 inhibitor. Cancer Res 2000;60:3696700.
23
Majano PL, Garcia-Monzon C, Lopez-Cabrera M, Lara-Pezzi E, Fernandez-Ruiz E, Garcia-Iglesias C, et al. Inducible nitric oxide synthase expression in chronic viral hepatitis. Evidence for a virus-induced gene upregulation. J Clin Invest 1998;101:134352.
24
Vickers SM, MacMillan-Crow LA, Green M, Ellis C, Thompson JA. Association of increased immunostaining for inducible nitric oxide synthase and nitrotyrosine with fibroblast growth factor transformation in pancreatic cancer. Arch Surg 1999;134:24551.
25
Hussain SP, Amstad P, Raja K, Ambs S, Nagashima M, Bennett WP, et al. Increased p53 mutation load in noncancerous colon tissue from ulcerative colitis: a cancer-prone chronic inflammatory disease. Cancer Res 2000;60:33337.
26 Tatemichi M, Ogura T, Nagata H, Esumi H. Enhanced expression of inducible nitric oxide synthase in chronic gastritis with intestinal metaplasia. J Clin Gastroenterol 1998;27:2405.[Medline]
27 Zhuang JC, Wright TL, deRojas-Walker T, Tannenbaum SR, Wogan GN. Nitric oxide-induced mutations in the HPRT gene of human lymphoblastoid TK6 cells and in Salmonella typhimurium.Environ Mol Mutagen 2000;35:3947.[Medline]
28
Gal A, Wogan GN. Mutagenesis associated with nitric oxide production in transgenic SJL mice. Proc Natl Acad Sci U S A 1996;93:151027.
29 Alagol H, Erdem E, Sancak B, Turkmen G, Camlibel M, Bugdayci G. Nitric oxide biosynthesis and malondialdehyde levels in advanced breast cancer. Aust N Z J Surg 1999;69:64750.[Medline]
30 Wink DA, Vodovotz Y, Laval J, Laval F, Dewhirst MW, Mitchell JB. The multifaceted roles of nitric oxide in cancer. Carcinogenesis 1998;19:71121.[Abstract]
31
Kim YM, Chung HT, Simmons RL, Billiar TR. Cellular non-heme iron content is a determinant of nitric oxide-mediated apoptosis, necrosis, and caspase inhibition. J Biol Chem 2000;275:1095461.
32 Wink DA, Kasprzak KS, Maragos CM, Elespuru RK, Misra M, Dunams TM, et al. DNA deaminating ability and genotoxicity of nitric oxide and its progenitors. Science 1991;254:10013.[Medline]
33 Nair J, Carmichael PL, Fernando RC, Phillips DH, Strain AJ, Bartsch H. Lipid peroxidation-induced ethenoDNA adducts in the liver of patients with the genetic metal storage disorders Wilson's disease and primary hemochromatosis. Cancer Epidemiol Biomarkers Prev 1998;7:43540.[Abstract]
34 Niemela O, Parkkila S, Britton RS, Brunt E, Janney C, Bacon B. Hepatic lipid peroxidation in hereditary hemochromatosis and alcoholic liver injury. J Lab Clin Med 1999;133:45160.[Medline]
35 Wright DK, Manos MM. Sample preparation from paraffin-embedded tissues. In: Innis MA, Gelfand DH, Sninsky JJ, White TJ, editors. PCR protocols. A guide to methods and applications. New York (NY): Academic Press; 1990. p. 1538.
Manuscript received December 26, 2000; revised August 24, 2001; accepted September 4, 2001.
This article has been cited by other articles in HighWire Press-hosted journals:
![]() |
||||
|
Oxford University Press Privacy Policy and Legal Statement |