Raltitrexed is a specific, folate-based inhibitor of thymidylate synthase with activity in advanced colorectal cancer comparable with that of fluorouracil (5-fluorouracil) plus folinic acid. Its activity is enhanced by rapid cellular entry and polyglutamation, with the polyglutamated derivatives having approximately 100-fold greater inhibitory potency than the parent compound. A number of phase I/pharmacokinetic studies have been performed, including schedules involving a 15-minute infusion every 3 weeks, weekly x 6 every 8 weeks, and every 2 weeks. The maximum tolerated dose (MTD) for the 3-weekly schedule was 3.5 to 4.5 mg/m2 in adults and 6 mg/m2 in a paediatric population. The MTDs for the other schedules have not yet been reported. The disposition of raltitrexed in patients is best described by a 3-compartment model with a terminal half-life (t1/2gamma) of 260 hours, the latter being subject to significant interpatient variability. A similar protracted t1/2gamma has been detected in all of the animal species studied. Together with evidence from the mass-balance studies performed, this delayed elimination suggests considerable sequestration of raltitrexed in tissues, predominantly as polyglutamate forms. Nevertheless, there has been no pharmacokinetic evidence of drug accumulation in plasma following repeated administration. On the basis of animal experiments, the oral bioavailability and penetration of raltitrexed into cerebrospinal fluid are both likely to be limited in the clinical setting. Raltitrexed is over 90% bound to plasma protein over the concentration range of 20 to 100 micromol/L. Apart from poly-glutamation, raltitrexed does not appear to be metabolised to a significant extent, and most of the excreted drug (approximately 20% of the administered dose) is recovered unchanged in the urine within the first 24 hours post-administration. The average clearance of raltitrexed is 2.4 L/h (40 ml/min), and this value is significantly reduced in patients with compromised renal function (glomerular filtration rate of 25 to 65 ml/min). These patients are more likely to experience severe antiproliferative toxicity with raltitrexed. A careful evaluation of renal function, particularly in the elderly, is warranted. It has not been possible to establish strong correlations between the plasma pharmacokinetics of raltitrexed and toxicity, and the cellular pharmacokinetics of raltitrexed may be more predictive. Studies in mice have demonstrated that delayed administration of folinic acid can assist in the recovery of animals from antiproliferative toxicity, possibly by promoting the release of polyglutamated drug from tissues. This approach should be evaluated as a rescue regimen in patients with severe proliferative toxicity.