Folate levels measured by LC–MS/MS in patients with colorectal cancer treated with different leucovorin dosages

Abstract

PurposeCalcium folinate (leucovorin), which is converted in vivo into biologically active folate, enhances the potency of 5-fluorouracil (5-FU)-based chemotherapy in colorectal cancer. A common dosage of leucovorin in adjuvant and palliative settings is 60 mg/m2. The aim was to determine the levels of tetrahydrofolate (THF), 5,10-methylenetetrahydrofolate (methyleneTHF), and 5-methyltetrahydrofolate (methylTHF) in tumour and mucosa of colorectal cancer patients who received different dosages of leucovorin intravenously at time of surgery.MethodsEighty patients scheduled for colorectal resection with indication of colorectal cancer were randomised into four groups to receive leucovorin at 0, 60, 200, or 500 mg/m2, respectively. Blood samples were taken 10 and 30 min after leucovorin administration. Biopsy samples from tumour and mucosa were collected and snap-frozen at surgery. The levels of THF, methyleneTHF, and methylTHF in tumour and mucosa were assessed by liquid chromatography electrospray ionisation tandem mass spectrometry (LC–MS/MS) and the results were related to clinical diagnosis and therapeutic regimens.ResultsThe folate levels in tissue revealed extensive inter-individual variability. The mean methyleneTHF value for the four treatment groups were 880, 1,769, 3,024 and 3,723 pmol/gww. Only half of the patients who received 60 mg/m2 leucovorin had higher levels of methyleneTHF in tumour than patients who received 0 mg/m2 leucovorin. Rectal cancer patients had significantly lower levels of methyleneTHF compared with colon cancer patients.ConclusionsThere was a large inter-patient variability of tissue folate levels in colorectal cancer patients after supplementation with leucovorin at standardised dosage. High leucovorin doses were needed to exceed baseline methyleneTHF values, especially in rectal cancer patients. The results indicate that the standardised leucovorin dose may be insufficient to attain the full antitumour effect of 5-FU. Further studies are needed to establish whether higher dosage yields a better treatment response.

Relationship between folate concentration and expression of folate-associated genes in tissue and plasma after intraoperative administration of leucovorin in patients with colorectal cancer

“…This finding is in agreement with previous results published by our group [18] as well as by others [23]. Higher concentrations were found in tumour tissue compared to mucosa in all patient groups.…”

“…A recent article published by our group presented data showing a large inter-individual variation of tissue folate concentration in patients with CRC after supplementation with LV at standardized dosage [18]. The concentrations of tetrahydrofolate (THF), 5,10-MeTHF and 5-MTHF in tumour and mucosa were assessed by liquid chromatography electrospray ionization tandem mass spectrometry (LC–MS/MS).…”

An LC-MS chemical derivatization method for the measurement of five different one-carbon states of cellular tetrahydrofolate

“…The concentration of cellular THF species is in the nM to μM range (2, 3), and these species exist in multiple polyglutamated forms (30). This polyglutamation creates analytical challenges by decreasing the signal intensity for each functional THF species in mass spectrometry.…”

Multiplexed Dosing Assays by Digitally Definable Hydrogel Volumes

“…As reported in literature, the in vitro cytotoxic effect of camptothecins against HT29 human colorectal adenocarcinoma cells varies widely, and SN‐38, the active metabolite of the prodrug irinotecan, is 100–1000 fold more potent than other camptothecins in inhibiting topoisomerase I. Folinic acid as single compound enhances cell metabolism in a dose‐dependent manner (Figure S6b, Supporting Information), while it potentiates the cytotoxicity of 5FU when used in combination, also in a dose‐dependent manner in the tested range from 25 × 10 −9 m to 7.41 × 10 −6 m (Figure S7a, Supporting Information). Pharmacokinetical studies in patients treated with intravenous administration of folinic acid alone or 5FU and folinic acid in combination at clinically relevant concentrations showed in vivo plasma concentrations as well as tumor tissue concentrations of folinic acid and its metabolites in the range of 1–10 × 10 −6 m . Thus, the highest tested concentration of 7.41 × 10 −6 m folinic acid in the present study was used in the further in vitro studies of the FOLFOX and FOLFIRI combination regimens.…”