Gemcitabine (2',2'-difluorodeoxycytidine) is a nucleoside analog with antitumor activity against a variety of malignancies. The critical enzyme cytidine kinase is saturated at plasma concentrations achieved after a 30-min infusion at conventional doses. Prolonged infusion time may yield higher intracellular dFdCTP concentrations. A phase I study was designed to determine the maximum tolerated dose (MTD) of gemcitabine, given by infusion for 3 h, in heavily pretreated patients. Twenty-seven patients (13 head and neck cancer, seven sarcoma, three esophageal cancer, three non-small-cell lung cancer and one ovarian cancer) were enrolled. Twenty patients were defined as refractory at first- or second-line chemotherapy. Four different entry dose levels (300, 400, 450 and 500 mg/m(2)) were evaluated for gemcitabine administered on days 1, 8 and 15 of a 28-day cycle. The MTD was defined as 450 mg/m(2), with granulocytopenia, thrombocytopenia and asthenia being dose limiting. The maximum grade III/IV patient toxicities for hemoglobin, leukocytes, neutrophils and platelets for all doses were 7, 19, 19 and 11%, respectively. Non-hematological toxicities included asthenia, nausea/vomiting and diarrhea. Thus, gemcitabine administered at a fixed 3-h infusion was well tolerated up to 450 mg/m(2) in heavily pretreated patients. Myelosupression and asthenia were dose-limiting toxicities.