In thalassemia patients, iron overload can stimulate lipid peroxidation (LPO), thereby generating miscoding DNA adducts. Adducted DNA was measured in the lymphocytes of β‐Thal/Hb E patients and healthy controls and in the organs of thalassemic mice. εdA, εdC and M₁dG residues were quantified by ³²P‐postlabeling‐TLC/HPLC. M₁dG levels in lymphocyte DNA from patients were 4 times as high as in controls, while the increase in εdA and εdC was not significant. Adducted DNA accumulated in the liver of thalassemic mice having >2.7 mg Fe/g tissue dry weight; DNA adducts and iron were highly correlated. εdA was not specifically generated in certain mouse liver cell types as revealed by immunohistochemical staining. We found elevated LPO‐induced DNA damage in the liver of thalassemic mouse and in lymphocytes, implicating that massive DNA damage occurs in the liver of thalassemia patients. We conclude that promutagenic LPO‐derived DNA lesions are involved in the onset of hepatocellular carcinoma in these patients. © 2009 UICC