To elucidate the prevalence and biologic significance of the c-myc gene in human hepatocellular carcinoma (HCC), DNA samples were taken from the paired tumorous and nontumorous tissues of 77 cases of resected primary HCC and were analyzed by Southern blot hybridization. We demonstrated modest, but significant c-myc amplification (group A) in 28 (36.4%) of the cases: 1.6- to 2.0-fold in 18, 2.1- to 3.0-fold in four, and > 3.0-fold in six. Compared to HCC without c-myc amplification (group B), group A HCC occurred more often in patients < 50 years old (54.5% vs 29.1%, p < 0.02) with serum alpha-fetoprotein (AFP) levels > 320 ng/mL (61.1% vs 14.6%, p < 0.00002). Group A HCC occurred more frequently in patients with hepatitis B virus infection than in those with hepatitis C virus infection (p < 0.03). Group A HCC was more likely to be poorly differentiated (44.8% vs 10.5%, p < 0.004) and associated with intrahepatic portal vein spread (57.1% vs 28.6%, p < 0.02). The c-myc amplification did not correlate with sex or tumor size. For small HCC, group A had a worse one-year survival rate than group B (72.2% vs 90.9%, p < 0.04). These findings suggest that c-myc amplification is not an uncommon event in human hepatocarcinogenesis, occurs more frequently in young patients who have an elevated serum AFP level or HBV infection, and is related to the biologic behavior of HCC.