Extensive hepatic replacement due to liver metastases has no effect on 5-fluorouracil pharmacokinetics Jan Gerard Maring 1 , Henk Piersma 2 , Albert van Dalen 3 , Harry J.M. Groen 4 , Donald R.A. Uges 5 , Elisabeth G.E. De Vries 6 1 Department of Pharmacy, Diaconessen Hospital Meppel and Bethesda Hospital Hoogeveen; 2 Department of Internal Medicine, Martini Hospital Groningen; 3 Department of Radiology, Diaconessen Hospital Meppel; Departments of 4 Pulmonary Diseases, 5 Pharmacy and 6 Medical Oncology, University Hospital Groningen, The Nether- lands Cancer Chemother Pharmacol 2003;51:167-173
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Extensive hepatic replacement due to liver metastases has no effect on 5-fluorouracil pharmacokinetics
Jan Gerard Maring1, Henk Piersma2, Albert van Dalen3, Harry J.M. Groen4, Donald R.A.
Uges5, Elisabeth G.E. De Vries6
1Department of Pharmacy, Diaconessen Hospital Meppel and Bethesda Hospital
Hoogeveen; 2Department of Internal Medicine, Martini Hospital Groningen; 3Department of Radiology, Diaconessen Hospital Meppel; Departments of 4Pulmonary
Diseases, 5Pharmacy and 6Medical Oncology, University Hospital Groningen, The Nether-
lands
Cancer Chemother Pharmacol 2003;51:167-173
Chapter 4.1
70
Influence of liver metastases on 5-fluorouracil pharmacokinetics
71
Erosion at work. Zabriskie point. Death Valley USA 1995.
Chapter 4.1
70
Influence of liver metastases on 5-fluorouracil pharmacokinetics
71
Abstract
Aim The influence of liver metastases on the pharmacokinetics of 5-fluorouracil (5-FU)
and its metabolite 5,6-dihydrofluorouracil (DHFU) was studied in patients with liver me-
tastases from gastrointestinal cancer and compared with a control group of patients with
non-metastatic gastrointestinal cancer.
Methods Patients were assigned to two different groups based on the presence of liver
metastases. The percentage of hepatic replacement was determined with CT and ultra-
sonography and classified as <25 %, 25-50 or > 50% from the total liver volume. Chemo-
therapy consisted of leucovorin 20 mg/m2/day plus 5-FU 425 mg/m2/day, both during 5
days. Blood sampling was carried out on the first day of the first chemotherapy cycle. 5-FU
and DHFU were quantified by HPLC in plasma. A four compartment parent drug - metabo-
lite model with non-linear Michaelis-Menten elimination from the central compartment
of the parent drug (5-FU) was applied to describe 5-FU and DHFU pharmacokinetics.
Results No effect of liver metastases on 5-FU clearance was observed between the two
groups. The effect of 18 covariables on pharmacokinetic parameters was also studied in
univariate correlation analyses. Body surface area was positively correlated with the dis-
tribution volume of 5-FU in the central compartment and with Vmax
(r = 0.65 and r = 0.54
respectively).
Conclusions There is no need for dose adjustment of 5-FU as standard procedure in
patients with liver metastases and mild to moderate elevations in liver function tests.
Introduction
Fluorouracil (5-FU) is widely used in chemotherapeutic regimens for the treatment
of breast-, colorectal- and head and neck cancer. The cytotoxic mechanism of 5-FU is
complex, requiring intracellular bioconversion of 5-FU into cytotoxic nucleotides. Inhibi-
tion of thymidylate synthase by the metabolite 5-fluoro-2’-deoxyuridine-5’-monophos-
phate (FdUMP) is thought to be the main mechanism of cytotoxicity [1]. The cytotoxicity
is caused by only a small part of the administered 5-FU dose, as the majority of 5-FU is
rapidly metabolised into inactive metabolites (see figure 1). The initial and rate-limiting
enzyme in the catabolism of 5-FU is dihydropyrimidine dehydrogenase (DPD), catalysing
a reduction of 5-FU into 5,6-dihydrofluorouracil (DHFU). Subsequently DHFU is degraded
into fluoro-β-ureidopropionic acid (FUPA) and fluoro-β-alanine (FBAL) [2]. Several groups
have suggested a major role of DPD in the regulation of 5-FU metabolism and thus in the
amount of 5-FU available for cytotoxicity [3-7]. DPD is present in many tissues, but the
highest activity is found in the liver and, since liver blood flow is relatively high, this organ
is considered as major site for 5-FU degradation [2,8]. In the last decades the role of liver
metastases and liver function impairment on 5-FU pharmacokinetics has been subject to
Chapter 4.1
72
Influence of liver metastases on 5-fluorouracil pharmacokinetics
73
debate.
The effects of liver dysfunction on the pharmacokinetics of drugs are generally difficult to
predict. Liver dysfunction is usually diagnosed on the basis of liver function tests, but for
most drugs the correlation between test values and drug metabolism is only weak. Liver
dysfunction due to liver metastases is an even more complicated issue. Liver metastases
displace healthy liver tissue and this may directly affect the liver metabolic capacity due
to tissue loss, but also indirectly due to tissue damage caused by cholestasis as a result of
compression of surrounding structures including bileducts. Furthermore, liver dysfunc-
tion may affect plasma volume, serum albumin, drug protein binding, and hepatic blood
flow, all of which can influence pharmacokinetic parameters in complex ways.
In the last decade, several groups have studied the effects of liver metastases on 5-FU
clearance, in particular during continuous infusion. In an early small study, a reduction of
clearance of continuously infused 5-FU was reported in four patients with gastrointesti-
nal carcinoma and hepatic metastases [9]. Contrary to this, others found no correlation
between drug clearance and hepatic function tests in a larger study with 187 patients
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Figure 1 Metabolism of 5-FU. 5-Fluoro-2’-deoxyuridine-5’-monophosphate (FdUMP) is the cytotoxic product resulting from a multi-step 5-FU activation route . FdUMP inhibits the enzyme thymidylate synthase (TS), which leads to intracellular accumulation of deoxy-uridine-monophospate (dUMP) and depletion of deoxy-thymidine-monophospate (dTMP). This causes arrest of DNA synthesis. The initial and rate-limiting enzyme in the catabolism of 5-FU is dihydropyrimidine dehydrogenase (DPD), catalys-ing the reduction of 5-FU into 5,6-dihydrofluorouracil (DHFU). Subsequently, DHFU is degraded into fluoro-β-ureidopropionic acid (FUPA) and fluoro-β-alanine (FBAL).
Chapter 4.1
72
Influence of liver metastases on 5-fluorouracil pharmacokinetics
73
receiving 5-FU as continuous infusion [10]. More recently, Etienne et al. studied several co-
variables affecting 5-FU clearance during continuous infusion in 104 patients with various
cancers [11]. They found no effect of liver metastases in a subgroup of seven patients,
but did not further specify the involvement of concurrent liver dysfunction. Although
these studies suggest that liver metastases do not affect 5-FU clearance during continu-
ous infusion, it is important to realise that the pharmacokinetic behaviour of 5-FU after
bolus injection or short time infusion differs from that during continuous infusion. After
rapid infusion, 5-FU displays non-linear pharmacokinetic behaviour [12-15], probably due
to saturation of the DPD enzyme at higher plasma levels, although the exact mechanism
is still unclear [16]. Thus, data on 5-FU clearance obtained during continuous infusion do
not necessarily predict the situation after bolus injection. So far, however, only few reports
have been published regarding the effects of liver metastases and/or liver dysfunction on
the pharmacokinetics of bolus injected 5-FU [12,17,18].
Christophidis et al. studied the bioavailability of 5-FU after oral and intravenous drug ad-
ministration in 12 patients with liver metastases, and concluded that the bioavailability
was not related to liver function test abnormalities or metastatic deposits [12]. Nowa-
kowska-Dulawa also found no effect of liver metastases on 5-FU clearance in 20 patients
with colorectal cancer (compared to 8 controls) [17]. Unfortunately, in both studies patient
characteristics and liver function test results were not further specified. More recently,
Terret et al. studied the dose and time dependencies of 5-FU pharmacokinetics in 21
patients and also included some liver function parameters in their analysis [18]. Their data
suggest that 5-FU clearance might increase with the volume of hepatic replacement.
Since in most studies information regarding the extent of liver metastatic involvement is
concise or lacking, we decided to design a protocol to study the effects of this parameter
on the pharmacokinetics of 5-FU and DHFU after bolus injection. This study should provide
more insight in potential interactions between liver function and 5-FU pharmacokinetics.
Patients and Methods
Patients
Patients, aged 18 years and older, scheduled to receive adjuvant or palliative 5-FU treatment
for gastrointestinal cancer, and formerly chemotherapy naive, were included. Patients with
anaemia (Hb < 6 mmol/l), known disorders of hemostasis (e.g. haemophilia), severe renal
failure (GFR < 30 ml/min) or a history of alcohol or drug abuse were excluded. Patients
were assigned to two different groups based on the presence of liver metastases, that were
identified and measured with ultrasound and/or CT imaging. Other pre-treatment meas-
urements were body weight, height, blood cell counts, standard liver function tests (ALT,
AST, LDH, ALP, bilirubin) and markers for the liver synthesis function (pseudocholineste-
rase, albumin and antithrombin-III). Chemotherapy consisted of leucovorin 20 mg/m2/day,
Chapter 4.1
74
Influence of liver metastases on 5-fluorouracil pharmacokinetics
75
administered as short time infusion, followed by 5-FU 425 mg/m2/day, administered as
bolus intravenous injection during 2 min. Both drugs were given on 5 consecutive days,
in a 28-day cycle (Mayo regimen). On the day of blood sampling, leucovorin was infused
after the last sample. On the following 4 days, the same 5-FU dose was administered as
short time infusion (10-15 min), after the administration of leucovorin. During the first
chemotherapy cycle, toxicity was scored according to the Common Toxicity Criteria. The
study was approved by the Institutional Medical Ethics Review Board in the participating
hospitals and written informed consent was obtained from all patients.
Quantification of liver metastatic involvement
The diameter of the metastases was measured with standard CT and ultrasound imaging
software. From each lesion the maximum diameter was measured. This diameter was
used to calculate the volume of the metastatic lesions in relation to the total liver volume.
Four levels were defined for semi-quantification of the extent of liver metastatic disease,
according to Hunt et al. [19]. Absence of liver metastases was classified as level 0 (control
group), less than 25 % liver metastatic involvement as level 1, 25-50 % as level 2, and more
than 50% as level 3.
Collection of blood samples
For pharmacokinetic sampling, a canule was placed intravenously in the arm of the
patient contralateral to the side of drug administration. Blood samples of 5 ml were
collected in heparinised tubes just before, and 2, 5, 10, 20, 30, 45, 60, 80, 100, 120, 150 and
180 min after 5-FU injection. Collected samples were immediately placed on ice and sub-
sequently centrifuged at 2,500 g for 10 min at 4°C and stored at –80 °C till analysis. The
plasma samples were analysed for 5-FU and DHFU concentrations by high-performance
liquid chromatography (HPLC).
Chemicals
5-FU and chlorouracil were obtained from Sigma Chemical Co (Zwijndrecht, the
Netherlands). 5,6-dihydro-5-fluorouracil was kindly provided by Roche Laboratories
(Basel, Switzerland). Human heparinised plasma was obtained from the Red Cross Blood
Bank (Groningen, the Netherlands). All other chemicals were of analytical grade.
Reversed phase HPLC analysis
5-FU and DHFU concentrations were measured by HPLC analysis using a modification of
the method described by Ackland et al. [20]. Briefly, 100 µl chlorouracil internal standard
solution (80 mg/l in water) was added to 1 ml plasma sample, and this mixture was
vortexed and subsequently deproteinated with 50 µl of a 50% (w/v) trichloracetic acid
solution. After centrifugation at 8,000 g for 2 min the supernatant was transferred into a
20 ml centrifuge tube and neutralised with 1 ml 1 M sodium acetate solution. Then 5 ml
Chapter 4.1
74
Influence of liver metastases on 5-fluorouracil pharmacokinetics
75
ethylacetate was added and the mixture was vortexed during 10 min. After separation of
the organic and aqueous layers by centrifugation at 5,000 g for 5 min, the ethylacetate
layer was transferred into a 10 ml tube and evaporated under a stream of nitrogen at 25 °C.
The residue was dissolved in 100 µl ultrapure water and 20 µl was injected. 5-FU and DHFU
standards ranging from 0.5 to 20 mg/l were prepared in human plasma. The chromato-
graphic system consisted of a Waters 616 pump equipped with a Waters 717+ autosam-
pler. The separation of 5-FU and DHFU was accomplished by gradient elution at ambient
temperature on a Phenomenex Prodigy ODS 3 column (I.D. 250 x 4.6 mm, 5µm) equipped
with a guard column (30 x 4.6 mm) of the same material (both purchased from Bester,
Amstelveen, The Netherlands). Mobile phase A consisted of 1.5 mM K3PO
4 and 1% (v/v)
methanol in water (pH=6.0) and mobile phase B of 1.5 mM K3PO
4 and 5% (v/v) methanol
in water (pH=6.0). The gradient was programmed as follows: 100% A during 2 min; 100%
A → 100% B in 0.5 min; 100% B during 7 min; 100% B → 100% A in 0.5 min; 100% A during
10 min. Drug detection was performed using a Waters 996 Photo Diode Array UV detector
interfaced with a Millenium 2010 Chromatography Manager Workstation. Spectra were
acquired in the 201-300 nm range. 5-FU was monitored at 266 nm and DHFU at 205 nm.
The internal standard chlorouracil was monitored at both wavelengths. The limit of quan-
tification in plasma was 0.1 mg/l for both 5-FU and DHFU.
Pharmacokinetic analysis
The pharmacokinetic analyses were performed in the ADAPT II Maximum Likelihood
Parameter Estimation program (version 4.0; University of Southern California, Los Angeles,
Ca). The pharmacokinetic data of the first 15 patients were tested in 8 different parent drug-
metabolite pharmacokinetic models, characterised by linear or non-linear (Michaelis-Menten)
parent drug (5-FU) elimination from a central compartment and distribution of 5-FU and me-
tabolite (DHFU) over one or two compartments. Variance for the observations was assumed to
be proportional to the measured values and set at 10%. In each model the patient’s data were
fitted individually and for each data set the Akaike Information Criterion (AIC) was calculated
[21]. The model with the lowest summarised AIC value was selected as the better one. The
area under the curve (AUC 0→3h
) of 5-FU and DHFU was calculated using the trapezoidal
rule. The total clearance of 5-FU was calculated by dividing the administered dose by the
AUC.
Statistical analysis
Patient data were analysed as two groups, based on the presence of liver metastases.
Clinical chemistry and pharmacokinetic data in both groups were compared with a two
sided Student’s t-test. In case of unequal variances as indicated by the Kolmogorov-
Smirnov test, the log transformed data were used, or data were tested with the non-para-
metric Mann-Whitney U-test. The study was powered (>80%) to detect a 25% difference in
population means, assuming a standard deviation in pharmacokinetic parameters of 35%.
Chapter 4.1
76
Influence of liver metastases on 5-fluorouracil pharmacokinetics
77
Correlations between clinical chemistry, demographic and pharmacokinetic data were
tested by Spearman correlation analysis. Statistical significance was at p<0.05. Analyses
were performed with the SYSTAT 7.0 statistical program (SPSS inc. 1997).
Results
Patients
Patients in this study were treated in the Martini Hospital Groningen, the Bethesda
Hospital Hoogeveen or the Diaconessen Hospital Meppel. Between December 1997 and
January 2001, 18 patients were included in the control group and 16 in the liver metas-
tases group. In one patient belonging to the control group, largely reduced clearance of
5-FU was observed. DNA sequence analysis of the gene encoding DPD revealed that this
patient was heterozygous for a G→A point mutation in the DPYD gene. The resulting
protein from the mutated allele is inactive and total DPD activity in this patient was
low. Data on this patient were published elsewhere [22]. The statistical analyses were
performed both with and without inclusion of this patient in the control group. The liver
metastatic involvement of 9 patients in the metastases group was classified as level 1, that
of 4 patients as level 2 and that of 3 patients as level 3. An overview of the patient char-
acteristics is represented in table 1. An overview of treatment related toxicity, observed
during the first cycle, is shown in table 2.
Pharmacokinetics
The mean pharmacokinetic curves of 5-FU and DHFU as measured in both treatment
groups are represented in figure 2. The model, selected for calculating 5-FU and DHFU phar-
macokinetics of the patients in this study, is a four-compartment parent drug-metabolite
model with Michaelis-Menten elimination from the first towards the third compartment
(see figure 3). The model is described by four differential equations:
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Chapter 4.1
76
Influence of liver metastases on 5-fluorouracil pharmacokinetics
77
live
r m
etas
tase
sco
ntr
ols
Su
bg
rou
ps
on
per
cen
tag
e o
f li
ver
met
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tic
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ent
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25
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par
amet
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orm
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ang
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=1
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(n=
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gen
der
M/F
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13/5
4/5
4/0
1/2
age
(yr)
45-
78
(64)
4
5-80
(66)
57-
78(6
4) 4
5-76
(64)
47-
70(6
4)
wei
gh
t(k
g)
60-
94(7
8)
56-
85(7
3) 6
3-90
(78)
60-
94(7
9) 7
5-79
(76)
S-cr
eati
nin
e(4
0-10
0 µm
mo
l/l)
53
-131
(77)
62-1
84(7
7)
53-
131
(75)
71-
94(8
8) 6
1-81
(71)
AST
(<48
U/l
)
12-2
01(4
2) §
1
3-24
(20)
12-
60(3
4) §
26
-137
(59)
§
89-2
01(1
11) §
ALT
(<42
U/l
)
8-
234
(56)
§
5-
44(3
1)
8-58
(37)
§
38-2
02(7
4) §
50
-234
(66)
§
LDH
(200
-500
U/l
)
234-
2662
(5
54) §
17
3-39
6(2
72)
2
34-1
035
(449
) §
348-
1165
(689
) §
839-
2662
(235
7) §
ALP
(<12
5 U
/l)
65-1
221
(146
) §
54
-108
(79)
6
5-16
3(1
07) §
121
-266
(1
81) §
34
4-12
21
(454
) §
Bili
rub
in to
tal
(<17
µm
mo
l/l)
3-29
3(1
3)
5-
14(9
)
3-1
3(1
1)
8-59
(13)
31
-293
(40)
§
alb
um
in(3
5-55
g/l
) 2
4-39
(36)
2
5-43
(36)
31-
39(3
7) 3
6-38
(37)
24-
30(2
6) §
Ach
E-as
e(5
.4-1
3.2
x103 U
/l)
1.2
-8.3
(4.4
)
1.9-
6.5
(3.9
) 3
.0-8
.3(4
.9) §
1.2
-6.3
(3.7
) 1
.5-2
.8(2
.1)
AT-3
(80-
120
%)
5
2-13
7(9
2)
82
-126
(96)
83
-137
(96)
81
-100
(90)
52
-119
(57)
§ Sta
tistic
ally
diff
eren
t fro
m co
ntro
ls at
p<0
.05
(Man
n W
hitn
ey U
test
)
Tabl
e 1
Patie
nt C
hara
cter
istic
s
Data
are
pre
sent
ed a
s ran
ge. T
he m
edia
n va
lue
is pl
aced
bet
wee
n br
acke
ts.
Chapter 4.1
78
Influence of liver metastases on 5-fluorouracil pharmacokinetics
79
The compartments 1 and 2 represent the central and peripheral compartment for 5-FU
(parent drug) pharmacokinetics, compartments 3 and 4 are the central and peripheral
compartment for DHFU (metabolite) pharmacokinetics. The X values indicate the amount
of drug in each compartment respectively. Data are imported in the model as plasma drug
concentrations, measured in compartment 1 (5-FU) and compartment 3 (DHFU) respec-
tively. The volume of compartment 1 and 3 is calculated by dividing the drug amount
by drug concentration. The k-values represent linear distribution- and elimination rate
constants, and the Vmax
and Km
values represent Michaelis-Menten constants for non-linear
elimination from the first compartment. The Km
value was kept constant during fitting of
patient data. To determine the best fitting value, this parameter was varied between 0.5
and 15 mg/l. Km
=5 mg/l was selected as most optimal value, based upon the lowest sum-
marised AIC. Rinf
represents the infusion rate of 5-FU in mg/h. No differences existed in
model parameters between the 2 treatment groups and no correlations were measured
between individual model parameters and patient characteristics. Therefore, the calcu-
lated model parameters are listed in table 3 as mean values obtained from all 33 patients
in this study. The interindividual variation in 5-FU clearance was considerable, resulting in
metastases (n=15) no metastases (n=18)
CTC grade I II III IV I II III IV
gastrointestinal
nausea 6 0 0 0 5 0 0 0
vomiting 3 0 0 0 2 0 0 0
diarrhea 1 0 0 0 4 2 0 0
mucositis 3 4 0 0 2 3 0 0
Flu-like symptoms
fever 1 1 0 0 1 0 1 0
malaise 1 1 0 0 6 1 0
Others
dermatological 1 0 0 0 2 0 0 0
eyes 1 0 0 0 2 0 0 0
Toxicity of any kindb 10 (66%) 13 (72%)
aThe figures represent the number of patients suffering from a particular type of toxicity (graded ac-cording to the Common Toxicity Criteria version 2.0) bTotal number of patients suffering from toxicity of any kind at any grade.
Table 2 Overview of side effects during the first cyclea.
Chapter 4.1
78
Influence of liver metastases on 5-fluorouracil pharmacokinetics
79
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Figure 2 Pharmacokinetics of 5-FU. Shown are 5-FU ( ) and DHFU ( ) plasma levels observed in control patients (n=18) and 5-FU ( ) and DHFU ( ) plasma levels observed in patients with liver metastases (n=16). All values are depicted as mean ± SD.
Figure 3 Four compartiment parent drug - metabolite pharmacokinetic model describing 5-FU and DHFU pharmacokinetics.
Chapter 4.1
80
Influence of liver metastases on 5-fluorouracil pharmacokinetics
81
5-FU AUCs ranging from the lowest to the highest value over a factor 3 (not including the
patient with DPD deficiency).
Correlation between patient covariables and pharmacokinetic parameters
We identified 18 patient covariables that were each tested in a univariate Spearman cor-
relation analysis. Tested covariables were sex, age, height, weight, body surface area (BSA),
lean body mass (LBM), creatinine, urea, level of liver metastatic involvement, AST, ALT, ALP,
LDH, bilirubin, albumin, AT-III, pseudocholinesterase and 5-FU dose. Positive correlations
were found between the BSA and the Vmax
and V1 values, regardless the level of liver meta-
static involvement (r = 0.65 and r = 0.54 respectively)
Discussion
The persistent uncertainty regarding the effects of liver metastases and liver dysfunction
on the pharmacokinetics of bolus injected 5-FU- made us to design the current study. We
decided to target the study on the extent of liver metastatic involvement and also decided
to include a detailed characterisation of the liver function in the study design.
We eventually included 34 patients from which 16 had liver metastases. We choose to
classify the extent of liver metastatic involvement in these patients as categorical rather
than continuous variable, according to Hunt et al.[19], since accurate measurement of the
percentage hepatic replacement is difficult to perform. More subtle differences between
patients cannot be detected in this approach, but this was not considered disadvanta-
Model parameter mean (SD)
Vmax (h-1) 1472 (356)
V1 (l) 15.5 (5.3)
K12 (h-1) 7.73 (4.13)
K21 (h-1) 6.24 (2.77)
V3 (l) 97 (42)
K34 (h-1) 6.18 (12.83)
K43 (h-1) 4.91 (7.38)
K30 (h-1) 1.80 (1.71)
AUC 5-FU (mg.h/l) 10.1 (3.7)
Cl 5-FU (ml/min) 1485 (537)
AUC DHFU (mg.h/l) 5.6 (2.0)
Table 3 Pharmacokinetic parameters
Chapter 4.1
80
Influence of liver metastases on 5-fluorouracil pharmacokinetics
81
geous, since small differences will probably be clinically irrelevant.
All patients received treatment according to the Mayo Clinics scheme. The 5-FU dose was
administered as short-time infusion (5-10 min), according to common practice in the
participating hospitals, but on the day of blood sampling, 5-FU was given as 2 min bolus
injection to warrant precise and standardised drug administration. The short half-life of
5-FU necessitated this standardisation, since uncertainty in this parameter can hamper
pharmacokinetic calculations.
To describe the non-linear pharmacokinetics, we applied a relatively complex model
with Michaelis-Menten elimination from the central compartment, based on the model
proposed by Collins et al [13]. Our model was selected on the basis of ‘best fit’ (objectified
by the Akaike Information Criterion) from a series of 8 different variants on the Collins
model. The final model that we used is almost identical to the model proposed by Terret et
al. [18] in their analysis of 5-FU pharmacokinetics. We also included the metabolite DHFU
in our model, but the additional value of this seemed limited, since large 95% confidence
intervals were found around the calculated K34
, K43
and K30
values.
The consequences of non-linear Michaelis-Menten pharmacokinetics are most profound
at plasma levels exceeding the Km
value. In the case of 5-FU, such plasma levels are
reached after bolus injection but not during continuous infusion. A reduction of liver DPD
capacity, due to liver metastases and/or liver dysfunction, might result in lower Vmax
values
and, thus, in a lower 5-FU clearance immediately after bolus injection. Our results do not
confirm this, as patients in both treatment groups displayed similar pharmacokinetics.
No significant correlations were found between pharmacokinetic parameters, including
overall 5-FU clearance, and liver function parameters.
The fact that most patients with liver metastases had mild to moderate liver dysfunction
might explain our observations. However, in three patients with extensive (level 3) meta-
static disease, attended by cholestase as indicated by high bilirubin and ALP levels, 5-FU
clearance also was unchanged. These patients further displayed high transaminase levels
(indicating cell damage) and low albumin and AT-III levels (indicating loss of function).
This observation suggests that the influence of extensive liver metastatic disease,
including liver damage, on 5-FU pharmacokinetics is at leas not dramatic. Recently, Terret
et al. performed a NONMEM analysis to identify covariables that affect 5-FU model pa-
rameters and they observed that their Vmax
values tended to increase with the volume of
liver metastatic involvement [18]. In our study this correlation was only weak (r = 0.21, see
figure 4) and at least clinically irrelevant. These results suggest that the metabolism of 5-
FU in metastatic tumour tissue at least equals that in healthy liver tissue. Extensive 5-FU
uptake in metastatic tissue indeed has been demonstrated during 18F-fluorouracil labelled
positron emission tomography in patients with liver metastases from colorectal cancer
[23]. Although the DPD activity in liver metastases from colon cancer seems to be lower
than in adjacent normal liver tissue [24], hepatic arterial blood flow is generally increased
in liver metastatic disease [19,25]. Since the elimination rate of drugs with a very large
Chapter 4.1
82
Influence of liver metastases on 5-fluorouracil pharmacokinetics
83
extraction ratio is strongly depending on the hepatic blood flow, an increase of liver blood
flow in metastatic disease might compensate for a reduced DPD activity in parts of the
liver replaced by metastases.
More or less expected on the basis of the almost identical pharmacokinetic profiles,
treatment related toxicity was comparable in both treatment groups. The incidence of
diarrhoea and malaise was somewhat higher in the control group, but this is probably
accidental. One patient in the control group experienced excessive toxicity as a result of
decreased 5-FU clearance due to DPD deficiency. A full description of the pharmacoki-
netics of 5-FU in this patient was published elsewhere [22]. Interestingly, we found that
irrespective of metastatic status 5-FU clearance was significantly lower in patients experi-
encing mucositis (1696 ± 640 vs. 1209 ± 290 ml/min, p<0.05, see figure 5). Such a trend was
not observed for nausea. This finding might be a coincidence, but it might also be possible
that late toxic effects such as mucositis are more related to pharmacokinetics than direct
toxic effects such as nausea. It has been shown that the extent of salivary excretion is a
predictor for the development of mucositis and salivary excretion is generally higher in
patients with low drug clearance [26].
Based on current data we conclude that there is no need for dose adjustment of 5-FU in
patients with liver metastases and mild to moderate elevations in liver function tests. Both
the extent of liver metastatic involvement and liver function parameters were included
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Figure 4 Correlation between level of hepatic replacement and calculated Vmax (h-1) value.
Chapter 4.1
82
Influence of liver metastases on 5-fluorouracil pharmacokinetics
83
as parameters in this study to this allowed a more comprehensive evaluation of liver
metastatic disease on 5-FU pharmacokinetics. We believe that there is no reason to expect
increased 5-FU toxicity in patients with liver metastases due to a reduced 5-FU clearance.
Therefore we do not recommend 5-FU dose reduction as standard procedure in these
patients.
Acknowledgement
We would like to thank Roche Basel for providing dihydrofluorouracil chemical standard, Dr.
Hans Proost (Department of Pharmacokinetics and Drug Delivery, University Groningen,
NL) for his advice on pharmacokinetic modelling, Barbara Bong and Dr. Robert de Jong
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Figure 5 Correlation between 5-FU clearance (ml/min) and the occurrence of mucositis (upper panel) and nausea (lower panel).
Chapter 4.1
84
Influence of liver metastases on 5-fluorouracil pharmacokinetics
85
(Martini Hospital Groningen, NL), Henk de Korte (Diaconessen Hospital Meppel, NL) and
Janny Haasjes (Bethesda Hospital Hoogeveen, NL) for patient inclusions, the departments
of radiology of the participating hospitals for help on calculations regarding the extent of
liver metastatic involvement, and last but not least all nurses of the oncology wards of the
participating hospitals for their assistance during blood sampling.
Chapter 4.1
84
Influence of liver metastases on 5-fluorouracil pharmacokinetics
uracil and its metabolites in plasma, urine and bile. Cancer Res 1987;47:2203-2206
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7. Lu Z, Zhang R, Diasio RB. Population characteristics of hepatic dihydropyrimidine dehydrogenase
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8. Ho DH, Townsend L, Luna MA, Bodey GP. Distribution and inhibition of dihydrouracil dehydroge-
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9. Floyd RA, Hornbeck CL, Byfield JE, Griffiths JC, Frankel SS. Clearance of continuously infused 5-flu-
orouracil in adults having lung or gastrointestinal carcinoma with or without hepatic metastases.
Drug Intell Clin Pharm 1982;16: 665-667
10. Fleming RA, Milano GA, Etienne MC, Renee N, Thyss A, Schneider M, Demard F. No effect of dose,
hepatic function, or nutritional status on 5-FU clearance following continuous (5-day) 5FU infu-
sion. Br J Cancer 1992;6: 668-672
11. Etienne MC, Chatelut E, Pivot X, Lavit M, Pujol A, Canal P, Milano G. Co-variables influencing 5-fluo-
rouracil clearance during continuous venous infusion. A NONMEM analysis. Eur J Cancer 1998;34:
92-97
12. Christophidis N, Vajda FJE, Lucas I, Drummer O, Moon WJ, Louis WJ. Fluorouracil therapy in pa-
tients with carcinoma in the large bowel: a pharmacokinetic comparison of various rates and
routes of administration. Clin Pharmacokinet 1978;3:330-336
13. Collins JM, Dedrick RL, King FG, Speyer JL, Myers CE. Nonlinear pharmacokinetic models for 5-flu-
orouracil in man: intravenous and intraperitoneal routes. Clin Pharmacol Ther 1980;28:235-246
14. McDermot BJ, van den Berg HW, Murphy RF.Nonlinear pharmacokinetics for the elimination of
5-fluorouracil after intravenous administration in cancer patients. Cancer Chemother Pharmacol
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1982;9:173-178
15. Van Groeningen CJ, Pinedo HM, Heddes J, Kok RM, De Jong AP, Wattel E, Peters GJ, Lankelma J.
Pharmacokinetics of 5-fluorouracil assessed with a sensitive mass spectrometric method in pa-
tients on a dose escalation schedule. Cancer Res 1988;48:6956-6961
16. Gamelin E, Boisdron-Celle M. Dose monitoring of 5-fluorouracil in patients with colorectal or
head and neck cancer - status of the art. Crit Rev Oncol Hematol 1999;30: 71-79
17. Nowakowska-Dulawa E. Circadian rhythm of 5-fluorouracil pharmacokinetics and tolerance.
Chronobiologica 1990;17:27-35
18. Terret C, Erdociain E, Guimbaud R, Boisdron-Celle M, McLeod HL, Fety-Deporte R, Lafond T, Game-
lin E, Bugat R, Canal P, Chatelut E. Dose and time dependencies of 5-fluorouracil pharmacokinet-
ics. Clin Pharmacol Ther 2000;68:270-279
19. Hunt TM, Flowerdew ADS, Britten AJ, Fleming JS, Karran SJ, Taylor I. An association between
parameters of liver blood flow and percentage hepatic replacement with tumour. Br J Cancer
1989;59:410-414
20. Ackland SP, Garg MB, Dunstan RH. Simultaneous determination of dihydrofluorouracil and 5-
fluorouracil in plasma by high-performance liquid chromatography. Anal Biochem 1997;246:
79-85
21. Bourne DWA. Evaluation of program output. In: Bourne DWA (ed) Mathematical modelling of
pharmacokinetic data. Technomic publishing Co inc. 1995 Lancaster Basel p.107-109
22. Maring JG, Van Kuilenburg ABP, Piersma H, Groen HJM, Uges, DRA, De Vries EGE. Reduced 5-FU
clearance in a patient with low DPD activity due to heterozygosity for a mutant allele of the
DPYD gene. Br J Cancer 2002;86;1028-1033
23. Moehler M, Dimitrakopoulou-Strauss A, Gutzler F, Raeth U, Strauss LG, Stremmel W.18F-labeled
fluorouracil positron emission tomography and the prognoses of colorectal carcinoma patients
with metastases to the liver treated with 5-fluorouracil. Cancer 1998;83:245-253
24. Johnston SJ, Ridge SA, Cassidy J, McLeod HL. Regulation of dihydropyrimidine dehydrogenase in
colorectal cancer. Clin Cancer Res 1999;5:2566-2570
25. Leen E, Goldberg JA, Robertson J, Angerson WJ, Sutherland GR, Cooke TG, McArdle CS. Early de-
tection of occult colorectal hepatic metastases using duplex colour Doppler sonography. Br J
Surg 1993; 80:1249-1251
26. Joulia JM, Pinguet F, Ychou M, Duffour J, Astra C, Bresolle F. Plasma and salivary pharmacokinetics
of 5-fluorouracil (5-FU) in patients with metastatic colorectal cancer receiving 5-FU bolus plus
continuous infusion with high-dose folinic acid. Eur J Cancer 1999;35:296-301
Reduced 5-FU clearance in a patient with low DPD activity due to heterozygosity for a mutant allele of the DPYD gene
Jan Gerard Maring1, André B.P. van Kuilenburg2, Janet Haasjes2, Henk Piersma3, Harry J.M.
Groen4, Donald R.A. Uges5, Albert H. van Gennip2, Elisabeth G.E. de Vries6
1Department of Pharmacy, Diaconessen Hospital Meppel and Bethesda Hospital
Hoogeveen; 2Department of Clinical Chemistry, Academic Medical Center Amsterdam; 3Department of Internal Medicine, Martini Hospital Groningen; Departments of 4Pulmonary Diseases, 5Pharmacy and 6Medical Oncology, University Hospital Groningen,
The Netherlands
Br J Cancer 2002;86;1028-1033
Chapter 4.2
88
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
89
One out of billions. Lago di Garda. Italy 2000.
Chapter 4.2
88
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
89
Abstract
Aim 5-FU pharmacokinetics, DPD-activity and DNA sequence analysis were compared
between a patient with extreme 5-FU induced toxicity and six control patients with
normal 5-FU related symptoms.
Methods Patients were treated for colorectal cancer and received chemotherapy consist-
ing of folinic acid 20 mg/m2 plus 5-FU 425 mg/m2 . Blood sampling was carried out on day
1 of the first cycle.
Results The 5-FU AUC in the index patient was 24.1 mg.h/l compared to 9.8 ± 3.6 (range
5.4-15.3) mg.h/l in control patients. The 5-FU clearance was 520 ml/min versus 1293 ± 302
(range 980-1780) ml/min in controls. The activity of DPD in mononuclear cells was lower
in the index patient (5.5 nmol/mg/h) compared to the 6 controls (10.3 ± 1.6, range 8.0-
11.7 nmol/mg/h). Sequence analysis of the DPD gene revealed that the index patient was
heterozygous for a IVS14+1G>A point mutation.
Conclusions Our results indicate that the inactivation of one DPYD allele can result in a
strong reduction in 5-FU clearance, causing severe 5-FU induced toxicity.
Introduction
Fluorouracil (5-FU) is widely used in chemotherapeutic regimens for the treatment
of breast-, colorectal- and head- and neck cancer. The cytotoxic mechanism of 5-FU is
complex, requiring intracellular bioconversion of 5-FU into cytotoxic nucleotides (see
figure 1). Inhibition of thymidylate synthase by the metabolite 5-fluoro-2’-deoxyuridine-
5’-monophosphate is thought to be the main mechanism of cytotoxicity [1]. The cytotox-
icity is caused by only a small part of the administered 5-FU dose, as the majority of 5-FU is
rapidly metabolised into inactive metabolites. The initial and rate-limiting enzyme in the
catabolism of 5-FU is dihydropyrimidine dehydrogenase (DPD), catalysing the reduction
of 5-FU into 5,6-dihydrofluorouracil (DHFU). Several groups have suggested a major role
of DPD in the regulation of 5-FU metabolism and thus in the amount of 5-FU available for
cytotoxicity [2-5]. Indeed, in patients with DPD enzyme deficiency, 5-FU chemotherapy
is associated with severe, life-threatening toxicity [6]. Moreover, a markedly prolonged
elimination half-life of 5-FU has been observed in a patient with complete deficiency
of DPD enzyme activity [7]. Several mutations in the dihydropyrimidine dehydrogenase
gene (DPYD), which encodes for the DPD enzyme have recently been identified [6,8].
Furthermore, the frequency of DPD deficiency has been estimated to be as high as 2-3%
[5,9,10]. To date, a direct correlation between DPD gene mutation and decreased 5-FU
clearance has only been suggested but never been proven. In this study, we provide the
first detailed analysis of 5-FU pharmacokinetics in a patient with low DPD-activity due to
heterozygosity for a mutant allele of the gene encoding DPD.
Chapter 4.2
90
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
91
Methods
Chemicals
5-FU was obtained from Sigma Chemical Co. (Zwijndrecht, the Netherlands). 5,6-dihydro-
5-fluorouracil was kindly provided by Roche Laboratories (Basel, Switzerland). AmpliTaq
Taq polymerase and BigDye-Terminator-Cycle-Sequencing-Ready-Reaction kit were
supplied by Perkin Elmer (San Jose, CA, USA). A Quaquik Gel Extraction kit was obtained
from Qiagen (Hilden, Germany). Human heparinised plasma was obtained from the Red
Cross Blood Bank (Groningen, the Netherlands). [4-14C ]Thymine (1.85-2.22 GBq/mmol) was
obtained from Moravek Biochemicals (CA, USA) and Lymphoprep (spec.gravity 1.077 g/ml,
280 mOsm) was from Nycomed Pharma AS (Oslo, Norway). Leucosep tubes were supplied
by Greiner (Frickenhausen, Germany). All other chemicals were of analytical grade.
Patient and controls
All patients were treated for colorectal cancer and participated in a protocol that had
been designed to study 5-FU and DHFU pharmacokinetics. The protocol was approved
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Figure 1 Metabolism of 5-FU. 5-Fluoro-2’-deoxyuridine-5’-monophosphate (FdUMP) is the cytotoxic product resulting from a multi-step 5-FU activation route . FdUMP inhibits the enzyme thymidylate synthase (TS), which leads to intracellular accumulation of deoxy-uridine-monophospate (dUTP) and depletion of deoxy-thymidine-monophospate (dTMP). This causes arrest of DNA synthesis. The initial and rate-limiting enzyme in the catabolism of 5-FU is dihydropyrimidine dehydrogenase (DPD), catalys-ing the reduction of 5-FU into 5,6-dihydrofluorouracil (DHFU). Subsequently, DHFU is degraded into fluoro-β-ureidopropionic acid (FUPA) and fluoro-β-alanine (FBAL).
Chapter 4.2
90
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
91
by the Medical Ethics Review Committee of the Martini Hospital Groningen and written
informed consent was obtained from all patients. All patients who entered this protocol
were chemotherapy naive. Chemotherapy consisted of folinic acid 20 mg/m2 combined
with 5-FU 425 mg/m2 , both on 5 successive days, in a 28-day cycle. Blood sampling was
carried out on the first day of the first chemotherapy cycle immediately following the 5-FU
dose, administered as bolus intravenous injection over 2 min. Folinic acid was infused after
the end of blood sampling. On the following 4 days, the same 5-FU dose was administered
as short time infusion, after folinic acid administration.
One patient experienced severe toxicity during the first chemotherapy cycle and,
therefore, a screening on DPD deficiency was initiated. Data from seventeen patients who
participated in the same study protocol were analysed for reference pharmacokinetics.
Six patients, who showed no signs of severe toxicity, were randomly selected for reference
DPYD genotyping and DPD enzyme activity. These patients served as controls.
Collection of blood samples
For pharmacokinetic sampling, a canule was placed in the arm of the patient contralateral
from drug administration. Blood samples (5 ml) were collected in heparinised tubes just
before, and 2, 5, 10, 20, 30, 45, 60, 80, 100, 120, 150 and 180 min postinjection. Collected
samples were immediately placed on ice and subsequently centrifuged at 2,500 g for 10
min. The plasma samples were analysed for 5-FU and DHFU concentrations by high-per-
formance liquid chromatography (HPLC) on the day of collection.
Blood samples for DPD analysis were collected 5 to 23 months after blood sampling for
5-FU pharmacokinetics, which corresponds to intervals ranging from 2 to 17 months after
the last 5-FU dose. None of the patients received chemotherapy at that moment.
Reversed phase HPLC analysis
5-FU and DHFU concentrations were measured by HPLC analysis using a modification of
the method described by Ackland et al [11]. Briefly, 100 µl chlorouracil internal standard
solution (80 mg/l in water) was added to 1 ml plasma sample, and this mixture was
vortexed and subsequently deproteinated with 50 µl of a 50% (w/v) trichloracetic acid
solution. After centrifugation at 8,000 g for 2 min the supernatant was transferred into a
20 ml centrifuge tube and neutralised with 1 ml 1M sodium acetate solution. Then 5 ml
ethylacetate was added and the mixture was vortexed during 10 min. After separation of
the organic and aqueous layers by centrifugation at 5,000 g for 5 min, the ethylacetate
layer was transferred into a 10 ml tube and evaporated under a stream of nitrogen at 25
°C. The residue was dissolved in 100 µl ultrapure water and 20 µl was injected. 5-FU and
DHFU standards ranging from 0.5 to 20 mg/l were prepared in human plasma.
The chromatographic system consisted of a Waters 616 pump equipped with a Waters
717+ autosampler. The separation of 5-FU and DHFU was accomplished by gradient
elution at ambient temperature on a Phenomenex Prodigy ODS 3 column (I.D. 250x4.6
Chapter 4.2
92
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
93
mm, 5µm) equipped with a guard column (30x4.6 mm) of the same material. Mobile phase
A consisted of 1.5 mM K3PO
4 and 1% (v/v) methanol (pH=6.0) and mobile phase B of 1.5
mM K3PO
4 and 5% methanol (pH=6.0).
The gradient was programmed as follows: 100% A during 2 min; 100% A → 100% B in 0.5
min; 100% B during 7 min; 100% B → 100% A in 0.5 min; 100% A during 10 min. Detection
was performed using a Waters 996 Photo Diode Array UV detector interfaced with a
Millenium 2010 Chromatography Manager Workstation. Spectra were acquired in the 201-
300 nm range. 5-FU was monitored at 266 nm and DHFU at 205 nm. The internal standard
chlorouracil was monitored at both wavelengths.
Pharmacokinetic analysis
The pharmacokinetic analyses were performed in the ADAPT II computer program (version
4.0; USC Los Angeles). The pharmacokinetic data of both the index patient and seventeen
reference patients (among which the six controls) were tested in 8 different models. In each
model the patient’s data were fitted individually and for each data set the Akaike Information
Criterion (AIC) was calculated. The model with the lowest summarised AIC value was selected
as the better one (data not shown). The model used for calculating 5-FU pharmacokinetics is a
two-compartment model with Michaelis-Menten elimination from the first compartment
and is described by two differential equations:
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X1
and X2
indicate the amount of drug in each compartment, respectively. The k-values
represent linear distribution- and elimination rate constants, and the Vmax
and Km
values
represent Michaelis-Menten constants for non-linear elimination from the first compartment.
Rinf
represents the infusion rate of 5-FU.
The area under the curve of 5-FU and DHFU was calculated using the trapezoid rule. The
average systemic clearance of 5-FU was calculated by dividing the administered dose by the
area under the curve (AUC).
Determination of dihydropyrimidine dehydrogenase activity
To investigate whether the 5-FU toxicity might have been caused by a partial deficiency of
DPD, we determined the activity of DPD in peripheral blood mononuclear (PBM) cells.
Therefore PBM cells were isolated from 15 ml EDTA anticoagulated blood and the activity
of DPD was determined according previously decribed methods [12]. In brief, the sample
was incubated in a reaction mixture containing 35mM potassium phosphate pH 7.4, 1mM
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
93
After an appropriate incubation time, the reaction catalysed by DPD was terminated by
adding 10 % (v/v) perchloric acid. The reaction mixture was centrifuged at 11,000 g for 5
min to remove protein. The separation of radiolabelled thymine and the reaction products
was performed by reversed phase HPLC. Protein concentrations were determined with a
copper-reduction method using bicinchoninic acid, as decribed by Smith et al. [13].
PCR amplification of coding exons
The DNA from the index and control patients was isolated from PBM cells as previously
described [14]. PCR amplification of exon 14 and flanking intronic regions was carried out
according to Van Kuilenburg et al. [6]. PCR products were separated on 1% agarose gels,
visualised with ethidium bromide and purified using a Qiaquick Gel Extraction kit and
used for direct sequencing.
Sequence analysis
Sequence analysis was carried out on a Applied Biosystems model 377 automated DNA
sequencer using Dye-Terminator method for the DPD cDNA and genomic fragments.
Statistical analysis
Each value, measured in the index patient, was compared to the mean ± 2 SD range of the
corresponding parameter in the control group . Values outside this range were considered
abnormal (p<0.05). We did not match our control patients for age and gender.
Results
Clinical evaluation
Patient characteristics from the index and control patients, as measured before 5-FU ad-
ministration on the first day of the first chemotherapy cycle, are listed in table 1. The index
patient is a 60-year-old white female who received adjuvant chemotherapy for Dukes C
colon carcinoma. She was known with a chronic moderate renal function impairment as
a result of a double sided nephrolithotomy at age 40. The first two injections with a total
dose of 800 mg 5-FU/day were tolerated well by the patient without complications. On the
third day of chemotherapy she experienced nausea and cold shivers. The nausea was suc-
cessfully treated with metoclopramide. The cold shivers remained on days 4 and 5. Twelve
days after administration of the first 5-FU injection, leukopenia (1.5x109 leukocytes/l) and
thrombocytopenia (26x109 platelets/l) developed along with nausea, diarrhoea, stomati-
tis, fever and hair loss. The next day leukocytes and platelets decreased to 0.5x109/l and
12x109/l (both nadir values respectively). During this period the patient developed leuko-
penic fever (40 °C) for which antibiotics were administered. Until day 20 the leucocytes
and platelets remained low (1x109/l and 13x109/l respectively). During the subsequent
Chapter 4.2
94
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
95
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Figure 2 Pharmacokinetics of 5-FU. Shown are 5-FU plasma levels observed in a patient with a IVS14+1G→A mutation in the DPYD gene ( ) and the 5-FU plasma levels resulting from simulation of a normal renal function in the same patient. 5-FU plasma levels from control patients are depicted as mean ± SD ( ; n=6).
Index patient Controls (n=6)
mean ± 2 SD
Gender M/F F 4/2
Age (yr) 60 64 ± 12
Weight (kg) 73 77 ± 12
Serum creatinine (µmmol/L) 184 § 81 ± 30
Aspartate aminotransferase (U/L) 23 18 ± 8
Alanine aminotransferase (U/L) 41 29 ± 26
LDH (U/L) 379 302 ± 152
Alkaline phosphatase (U/L) 95 72 ± 24
Bilirubin total (µmmol/L) 7 10 ± 6
Albumin (g/L) 32 37 ± 8
§ outside 95% control range, p<0.05
Table 1 Patient Characteristics
Chapter 4.2
94
Influence of DPD deficiency on 5-fluorouracil pharmacokinetics
95
week the clinical picture and hematological parameters gradually improved and normal-
ised. On day 34 the patient was discharged from the hospital.
The toxicity observed in the six control patients was limited to mild nausea (n=4), vomiting
(n=2) and CTC grade 1 stomatitis (n=1).
Pharmacokinetic analysis
The clearance of 5-FU was considerable slower in the index patient than in the six control
patients. In all control patients the plasma level at t=90 min was below 0.1 mg/l, whereas
in the index patient the plasma level was still 3.8 mg/l at this time point (see figure 2).
The AUC in the patient suffering from toxicity was 24.1 mg.h/l compared to 15.3 mg.h/l
as highest AUC value in control patients. We calculated an average systemic clearance of
only 520 ml/min versus 980-1780 ml/min in controls. The Vmax value, calculated by phar-
macokinetic modelling was 548 mg/h, while the Vmax values of control patients ranged
from 984 to 1772 mg/h (see table 2). The pharmacokinetic data of the six control patients
did statistically not differ from data of the reference group. The effect of the impaired
renal function of the index patient on 5-FU clearance was studied by pharmacokinetic
modelling.
The excretion of 5-FU in urine was measured in five patients of the reference group
(including two control patients) and kurine
was estimated 0.5 ± 0.08 h-1. This kurine
value, in-
dividually normalised on calculated GFR, was used during subsequent modelling of other
patient data. A normal renal function was simulated in the index patient by replacing the
GFR related kurine
by kurine
=0.6. Renal function impairment appeared to have only a slight
effect on 5-FU clearance. In the index patient, we estimated an additional 18% increase of