S‐adenosylmethionine synthetase (SAMS) catalyzes the formation of S‐adenosylmethionine (SAM) and is essential to normal cell function. There are two forms of SAMS, liver‐specific and nonliver‐specific (often referred to as “kidney”), which are products of two different genes. SAMS isoenzymes differ greatly in kinetic parameters and sensitivity to inhibition by methionine analogs. The current work studied changes in SAMS and their significance in liver cancer. Northern blot analysis showed that while normal liver expresses only liver‐specific SAMS, both HepG2 and HuH‐7 cells express only nonliver‐specific SAMS. Absence of liver‐specific SAMS messenger RNA (mRNA) was not because of gene deletion or rearrangement but complete lack of gene transcription. Reverse‐transcription polymerase chain reaction (RT‐PCR) with liver‐and kidney‐specific SAMS primers showed that liver‐specific SAMS mRNA was absent with only kidney SAMS mRNA present in HepG2, HuH‐7, Hep3B, and HuH‐1 cells, and four consecutive hepatocellular carcinoma (HCC) specimens. Normal liver tissues from the same patients express both forms of SAMS mRNA. As a result of the change in SAMS expression, SAMS activity was higher in HepG2 and HuH‐7 cells at physiologically relevant methionine concentrations but lower at high (mmol/L) methionine concentrations than rat hepatocytes. Treatment with ethionine and seleno‐D,L‐ethionine, two inhibitors known to have I50 values 50 to 60 times lower against SAMS purified from Novikoff hepatoma cells as compared with SAMS purified from normal rat liver, resulted in increased cell lysis in HepG2 and HuH‐7 cells but not cultured rat hepatocytes. These agents did not affect cellular adenosine triphosphate (ATP) levels but inhibited SAMS activity in HepG2 and HuH‐7 cells when added to their protein extracts. In summary, expression of SAMS is altered in human liver cancer. This occurrence may provide a potentially exploitable target for cancer chemotherapy.