Purpose: The purpose of our study was to determine the maximum-tolerated dose, dose-limiting toxicity, safety profile, and pharmacokinetics of the polyamine synthesis inhibitor SAM486A given in combination with 5-fluorouracil/ leucovorin (5-FU/LV) in cancer patients. Experimental Design: Patients with advanced colorectal cancer were treated with 5-FU [bolus (400 mg/m2) followed by a 22-h infusion (600 mg/m2)] and LV (200 mg/m 2) and escalating doses of SAM486A, 1-3-h infusion daily for 3 days. Plasma sampling was performed to characterize the pharmacokinetics and pharmacodynamics of the combination Results: Twenty-seven patients with metastatic colorectal cancer and 1 with pseudomyxoma peritonei were treated. Twenty-six patients received SAM486A in the combination at doses ranging from 25 to 150 mg/m2/day. Dose-limiting toxicity consisting of fatigue grade 3 was seen at 150 mg/m2/ day. Other adverse events included neutropenia, hand and foot syndrome, nausea, vomiting, diarrhea, and constipation. Fifteen of 26 patients evaluable for best response according to the Southwest Oncology Group criteria achieved a partial response [8 (30%) of 26] or stable disease [9 (35%) of 26]. SAM486A did not influence the pharmacokinetics of 5-FU, and SAM486A clearance was similar to that when used as a single agent. Conclusions: The novel molecular agent SAM486A is tolerable and safe in combination with a standard 5-FU regimen in patients with advanced colorectal cancer. The dose of SAM486A recommended for additional studies with this combination is 125 mg/m2/day. A disease-directed evaluation of SAM486A using this regimen is warranted.