1,0 g0,9 ivn0,8 i v ruS0,7 n ito0,6 r o rop0,5 P itve0,4 lua0,3 m uC0,2 0,1 0,00 Cumulative Proportion Surviving (Kaplan-Meier) Complete Censored p=0,002 B12<300 B12>300 200 400 600 Time, days 800 1000 1200 Figure 1. Overall survival curve according to base-line B12 status. 5-Fluorouracil (5-FU) has been the mainstay of treatment of patients with ACRC and remains an integral component of modern combination regi- mens. Antifolate chemotherapy agents such as 5-FU reduce proliferation of neoplastic cells by inhibiting DNA synthesis. 5-FU interferes with the folate metabolism by inhibition of thymidylate synthetase. Interrupting the action of this enzyme blocks synth- esis of the pyrimidine thymidine, which is a nuc- leotide required for DNA replication. Thymidylate synthase methylates deoxyuridine monophoshate (dUMP) into thymidine monophosphate (dTMP). It has been postulated that measuring the levels of some of the key components (B12, folate, and homocysteine) in the folate metabolism would be of importance for tailored treatments for our patients. A number of clinical studies have illustrated that pretreatment levels of folate and homocysteine are predictive for both tumour response and toxicity (low folate levels are associated with increased toxicity) in patients treated with either thymidylate synthase (TS) inhibitors or folate antagonists (8,17,18). Pre- vious murine studies have indicated that folate levels may predict for the cytotoxic efficacy of 5-FU (19). It has also been shown that an increased amount of exogenous folate leads to enhanced TS inhibition (20). On the other hand, two recent clinical studies have indicated a relationship between dietary folate intake and fluoropyrimidine toxicity (21,22). In one study, 86 Canadian patients receiving adjuvant Table III. Healthy controls (n 224). S-Cobalamin, pmol/L S-Folate, nmol/L S-Homocysteine, mmol/L CI confidence intervals. Mean 376 11.5 16.8 Median 302 8.8 14.7 95% CI 342 410 10.2 12.8 15.9 17.8 Serum vitamin B12 and folate status with chemotherapy 163 5-FU and leucovorin were prospectively assessed for biomarkers of folate metabolism. Multivariate analyses identified base-line serum folate as an independent positive predictor for grade 3 4 toxicity. A similar result was found in a study of capecitabine monotherapy, in which patients with higher base-line levels of serum folate had a significantly increased incidence of toxic events (23). These conflicting findings have been debated, and one explanation may be the different mechan- isms of action of the antifolates. The antifolates compete with naturally occurring folate for active moieties on folate-dependent enzymes. Depletion in the level of intracellular folate will therefore lead to increased toxicity. Furthermore, the level of intra- cellular folate feeds back to inhibit the polyglutama- tion of antifolates, limiting toxicity. This, however, remains to be further elucidated. In this study we have shown that base-line levels of B12, folate, and homocysteine for a patient popula- tion with advanced ACRC (n 93) were almost the same as for a previous performed cohort study of 224 healthy individuals from the same county. Our study also reveals that most individuals in an unselected population with advanced ACRC, in spite of their possible intestinal problems and earlier operations, still have normal levels of serum cobalamin, folate, and homocysteine. Cobalamin deficiency is rare in this population, and this is most likely explained by the fact that the endogenous stores of B12 last for several years, and the course of the disease normally is shorter. None of the two patients who had B12 levels below the reference experienced increased toxicity. On the contrary, 10 out of 12 patients who suffered from grade 3 4 toxicity had folate levels below the reference before start of treatment. It is somewhat doubtful that this is related to the subclinically low levels of folate before the start of treatment in this population, given the excess amount of folate admi- nistered as part of the treatment. A partial explana- tion might be the rate-limiting transport mechanism from extra- to intracellular space carried out by reduced folate carriers. Another explanation might be that the patients with the lowest levels of folate might be the ones with the most advanced disease and therefore the most undernourished. Interestingly in this study, patients with sub- clinically low cobalamin values had superior overall survival and time to progression than did patients with higher values. It is not possible to offer a good explanation for this, but it might reflect a worse outcome among patients with more liver metas- tases. It is known that patients with liver injuries can have increased serum cobalamin values. The- oretically, patients with lower cobalamin values