Page 257 - Personalised medicine of fluoropyrimidines using DPYD pharmacogenetics Carin Lunenburg

Phenotyping assays for predicting DPD deficiency
status, adequate renal and liver biochemistry and haematological values, and no prior treatment with fluoropyrimidines.
Study procedures
One blood draw for the endogenous DHU/U ratio and endogenous uracil level assays was taken prior to start of treatment. For patients participating in three or four DPD phenotyping assays the study scheme was as follows. During two random days prior to start of fluoropyrimidine- based treatment, all three or four phenotyping assays were performed in each patient (study scheme in Figure 1). On the first day, blood draws for the DPD enzyme activity in PBMCs and two DPD phenotyping assays (endogenous DHU/U ratio and endogenous uracil levels) were taken prior to 9 am. Immediately thereafter, the third phenotyping assay (oral uracil loading dose of 1000 mg uracil) was performed. The U/DHU ratio was assessed at 120 minutes after administration of uracil. At least one day later, but prior to start of fluoropyrimidines, the fourth phenotyping assay (2-13C-uracil breath test with 6 mg/kg 2-13C-uracil) was performed including blood draws for 13C-uracil and 13C-dihydrouracil plasma measurements. The DOB50 value from breath samples was correlated to 13C-dihydrouracil plasma levels and the 13C-DHU/U ratio. The oral uracil loading dose and 2-13C-uracil breath test were performed on two separate days to exclude any interference, as uracil was administered orally for both assays. Also, a minimum time interval of 24 hours between the phenotyping assays and start of fluoropyrimidine treatment was taken into account as a safety precaution, although it was expected that the administered uracil would not affect the efficacy and safety of patients when starting their fluoropyrimidine-based treatment, since uracil has a very short half-life of around 40 minutes.31
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Timeline
≥24 hours interval ≥24 hours interval
Intention to start with FP
DPD enzyme activity Endogenous DHU/U ratio Endogenous uracil levels Oral uracil loading dose
2-13C-uracil breath test
Start with FP
Figure 1. Study scheme
The study scheme per patient. Minimum interval between the tests and between tests and start of fluoropyrimidine-therapy was 24 hours. There was no predefined maximum number of days between assays as patients usually started relatively quickly with therapy when the decision to start was made. Abbreviations: FP: fluoropyrimidines; DHU: dihydrouracil; U: uracil.
DPD phenotyping assays
Patients underwent the DPD phenotyping assays in the hospital of their recruitment. Protocols for each DPD phenotyping assay were made available and discussed with executive personnel. In four hospitals, trained personnel was available to execute three or four DPD phenotyping assays. Each DPD phenotyping assay is described in more detail in the
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