Assessment of trans-intestinal cholesterol excretion in patients with total biliary obstruction
(TICE-PTC)

Reverse cholesterol transport (RCT) is an important anti-atherogenic mechanism,
as it mediates the elimination of excess cholesterol out of the body into the
faeces. Recently, the concept of RCT has been revisited. In the *classical*
concept, the liver is considered to be the only organ capable to eliminate
cholesterol via excretion into the bile. However, evidence from animal models
suggests that the intestine is also an important secretory organ for
cholesterol.
In fact, direct trans-intestinal cholesterol excretion (TICE) accounted for 33%
of total faecal sterol excretion in mice. It is unknown whether TICE is present
in humans. However, previous human perfusion studies suggest secretion of
cholesterol in the small intestine. Based on previous calculations, TICE in
humans is estimated to amount to 300mg/day in subjects with an average body
weight of 70kg. This is approximately one third of the amount secreted into
bile. Finally, several pharmacological compounds have been found to stimulate
trans-intestinal cholesterol secretion in mice. This suggests that TICE might
be a novel therapeutic target for cholesterol excretion and thereby for
cardiovascular disease prevention.

In order to establish the existence of such a direct trans-intestinal
cholesterol-excreting pathway, we propose to study faecal recovery of a single
dose of intravenously administered 13C-cholesterol (stable isotope) in patients
with a total biliary occlusion. If 13C-cholesterol can indeed be recovered from
faeces in these patients, the only possible explanation is that it was secreted
directly by the intestine to the faeces, since endogenous biliary secretion is
completely blocked. Hence, this study will provide the proof-of-concept for the
existence of TICE in humans.
If valid, TICE could serve as a novel cholesterol-lowering mechanism, in order
to prevent cardiovascular disease. Further research will be necessary to
evaluate how to stimulate this cholesterol-lowering mechanism best, in order to
accomplish additional reductions in blood cholesterol concentrations and
thereby cardiovascular disease.

To establish the presence of direct trans-intestinal cholesterol excretion
(TICE) in humans by faecal recovery of i.v. administered 13C-cholesterol in
patients with a total biliary obstruction. This is a so-called proof-of-concept
study.

The design of the study is a cross-sectional proof-of-concept study, which
comprises a single measurement of cholesterol excretion in a period of 5 days.
The study flow chart is depicted on page 16 of the study protocol.

Patients will be recruited via the departments of gastroenterology and
intervention radiology of the AMC. Eligible participants will be informed about
the study objective and procedures by the gastroenterologist in the outpatient
clinic, during the visit in which the PTC-procedure is explained. In case
subjects are referred to our hospital from elsewhere, recruitment will take
place by the intervention radiologist.
Eligible subjects will be asked for their permission to be contacted by the
study physician, who will explain the procedures into more detail and who will
obtain informed consent.

Following written informed consent and successful placement of the biliary
PTC-drain, a blood sample (12 ml) will be collected for the measurement of
lipoprotein levels and the background 13C-cholesterol enrichment. This will be
done via the same venous catheter used in the PTC-procedure. Subsequently, at
T=0, an intravenous dose of 30 mg of 13C-cholesterol dissolved in 22 ml Liposyn
III 10% in ethanol will be administered in 30 minutes, again through the same
venous catheter. Four and twelve hours after 13C-cholesterol administration
(T=4h and T=12h), the study physician will obtain a blood and biliary sample
from the drain at the hospital ward.

In the 4 days following i.v. 13C-cholesterol administration, the study
physician will obtain a daily bloodsample in the morning fasting state (T=24h,
T=48h, T=72h, T= 96h, 15ml at each timepoint). This will be done either on the
hospital ward or at the subjects* homes, depending on their clinical course.
Simultaneously, a biliary sample will be obtained from the PTC-drain. Finally,
participants are asked to collect daily faecal samples, using a special
specimen collection system from day 1 until the end of study (day 4).

In case patients are scheduled for removal of the PTC-drain earlier than day 4,
this will be the end of study.

No serious adverse events are expected to occur in this study. Stable isotopes
of cholesterol behave like their natural substrates and therefore carry no
known risks. This has also been established by extensive human and animal
experience. The dose of isotopes administered to the subjects is small compared
to the amount already present in the body. Subjects can experience slight
transient burning in the infusion arm during administration of the
cholesterol-Liposyn solution. This is not harmful and disappears as soon as
administration is cessated.

Although participation to this study does not carry any health risks, it does
not carry any benefits or advantages either, neither to the participants
themselves, nor to the population of patients with total biliary occlusion as a
whole. Participation to this study will only serve a scientific purpose to
develop the cardiovascular research field in the search for new
cholesterol-lowering strategies.

In addition, subjects are subjected to behavioural changes, such as the
collection of faecal stool samples and the subjection to daily blood draws for
5 days. The latter can cause a hematoma at the site of venepuncture.