RANDOMIZED, DOUBLE-BLIND, PLACEBO-CONTROLLED DOSE ESCALATION STUDY TO EVALUATE THE SAFETY AND PHARMACOKINETICS OF SINGLE AND MULTIPLE DOSES OF 2-IMINOBIOTINE (2-IB) IN HEALTHY MALE SUBJECTS

The drug to be given 2-iminobiotin (2-IB), is a new, investigational compound
that may eventually be used for the treatment of perinatal asphyxia (serious
oxygen deprivation in a baby around the birth). The new drug is still under
development.
Perinatal asphyxia is the medical condition resulting from deprivation of
oxygen to a newborn infant long enough to cause apparent permanent harm. It
results most commonly from a interference of the blood flow (ie. When the
umbilical cord is wrapped around the babies neck)..
2-iminobiotin (2-IB) is a selective nitric oxide synthase inhibitor which
inhibits the chemical processes which are triggered after oxygen deprivation.
It is expected that it may contribute to the treatment and prevention of the
devastating effects of perinatal asphyxia.

Primary objectives
- to investigate the safety and tolerability of single and multiple doses of
2-IB pulse iv infusion in healthy male subjects
- to determine the pharmacokinetics after single and multiple doses of 2-IB
pulse iv infusion in healthy male subjects

Secondary objectives
-To investigate the safety and tolerability of multiple doses of a 5% Captisol®
infusion
formulation in healthy male subjects
-To determine the effect of a 5% Captisol® infusion formulation on the safety
and
pharmacokinetics of 2-IB
-To determine the pharmacokinetics after multiple doses of a 5% Captisol®
infusion formulation in healthy male subjects*

Design:
A randomized, double-blind, placebo-controlled, dose escalation study with 2
groups of 9 healthy male subjects each receiving a pulsed intravenous (iv)
infusion of 2-IB or placebo in 3 periods. Treatments will be randomized such
that each subject receives 2 out of 3 foreseen dose levels of 2 IB and once
placebo. There will be a washout period of at least 7 days between each dosing
period. During Period 1 of Group 1, 3 subgroups (Groups 1a, 1b and 1c) of 3
subjects (2 on active and 1 on placebo) will be dosed. after the dosing of
Cohort 1, there will be an interim PK evaluation to reconsider the
dose-escalation and PK sampling for Cohort 2. In Cohort 2 a new formulation of
the compound will be used in order to limit the IV volume as much as possible.

Procedures and assessments
Screening and follow-up:
Clinical laboratory, vital signs, physical examination, ECG; at eligibility
screening: medical history, drug screen, HBsAg, anti HCV, anti-HIV 1/2; drug
screen to be repeated upon each admission.

Observation period:
Group 1: 3 periods in the clinic; first period in clinic from -17 h up to 24 h
after start of first drug administration, second and third period in clinic
from -17 h up to 48 h after start of the first drug administration
Group 2: 3 periods in the clinic; each period in clinic from -17 h up to 48 h
after start of the first drug administration

Blood sampling:
for pharmacokinetics of 2-IB (Groups 1a, 1b, 1c and 2) and Captisol® (Group 2
only)in plasma: 11 time points after the start of the infusion (Cohorts 1a, b,
c * Period 1), 9 time points after the start of the 1st and 3rd infusion
(Cohort 1a, b, c * Periods 2 and 3), 9 time points after the start of the 1st
and 3rd infusion (Cohort 2 * Period 1), 8 time points after the start of the
1stand 3rd infusion (Cohort 2 * Periods 2 and 3) and 11 time points after the
last infusion (Cohort 1a, b, c * Periods 2 and 3 and Cohort 2 * Periods 1, 2
and 3)

Urine sampling:
For pharmacokinetics of 2-IB: Groups 1a, 1b and 1c, Period 1: in intervals of
0-4 h and 4 12 h after the start of the infusion, Groups 1a, 1b and 1c, Periods
2 and 3: in intervals of 20-24 h and 24-32 h after the start of the first
infusion, Group 2: each period in intervals of
20-24 h and 24-32 h after the start of the first infusion.

Pharmacodynamic assessment:
Near infrared spectroscopy (NIRS): at pre-dose of the first infusion and at 15
min and 2.5 h after the start of the third infusion in Period 3 of Group 1 and
in all periods of Group 2 (the timing of the NIRS assessments may be adjusted
for subsequent groups based on the results).

Safety assessments:
Adverse events: throughout the study; vital signs: each period pre-first dose
and at approximately 30 min after each infusion; ECG: each period pre-first
dose and 6, 10, 24 (Cohorts 1 and 2), 30, 34 and 48 h (Cohort 2 only) after the
start of the first infusion; clinical laboratory: 24 (Cohorts 1 and 2) and 48 h
(Cohort 2 only) after the start of the first infusion; NIRS: training session
on Day -1 (first period only) and pre-dose and 30 min and 4 h after start of
first and last infusion, infusion site inspection at least twice after eacht
infusion.

Bioanalysis:
analysis of plasma and urine samples for 2-IB (Groups 1a, 1b, 1c and 2) and
Captisol® (Group 2 only) by the Sponsor using a validated liquid
chromatography-mass spectrometry/mass spectrometry method*

Study Medication
Active substance : 2-IB
Activity : unknown
Indication : unknown
Strength : for Groups 1a, 1b and 1c: 1.0 mg/mL in pH 4
citrate buffer
for Group 2:
4.0 mg/mL in pH 4 citrate buffer and 5% Captisol® *
Dosage form : i.v. infusion

Treatments
Group 1a
Period 1: a single iv infusion of 0.6 mg/kg 2-IB (n=2) or placebo (n=1) over 4
h on Day 1
Period 2: a 15-min iv infusion of 0.6 mg/kg 2-IB (n=2) or placebo (n=1) every 4
h on Day 1 (6 infusions in total)
Period 3: a 15-min iv infusion of 2 mg/kg 2-IB (n=2) or placebo (n=1) every 4 h
on Day 1 (6 infusions in total)

Group 1b
Period 1: a single iv infusion of 0.6 mg/kg 2-IB (n=2) or placebo (n=1) over 1
h on Day 1
Period 2: a 15-min iv infusion of 0.6 mg/kg 2-IB (n=2) or placebo (n=1) every 4
h on Day 1 (6 infusions in total)
Period 3: a 15-min iv infusion of 2 mg/kg 2-IB (n=2) or placebo (n=1) every 4 h
on Day 1 (6 infusions in total)

Group 1c
Period 1: a single iv infusion of 0.6 mg/kg 2-IB (n=2) or placebo (n=1) over 15
min on Day 1
Period 2: a 15-min iv infusion of 0.6 mg/kg 2-IB (n=2) or placebo (n=1) every 4
h on Day 1 (6 infusions in total)
Period 3: a 15-min iv infusion of 2 mg/kg 2-IB (n=2) or placebo (n=1) every 4 h
on Day 1 (6 infusions in total)

Dosing starting at t = 0 h, t = 4 h, t = 8 h, t = 12 h, t = 16 h and t = 20 h
in Periods 2 and 3.

Group 2
Period 1 : a 15-min iv infusion of 2 mg/kg 2-IB (n=6) or placebo (n=3) every 4
h on Day 1 (6 infusions in total)
Period 2 : a 15-min iv infusion of 6 mg/kg 2-IB (n=6) or placebo (n=3) every 4
h on Day 1 (6 infusions in total)
Period 3: a 30-min iv infusion of 12 mg/kg 2-IB (n=6) or placebo (n=3) every 4
h on Day 1 (6 infusions in total)*

Dosing starting at t = 0 h, t = 4 h, t = 8 h, t = 12 h, t = 16 h and t = 20 h
in Periods 1, 2 and 3.

Procedures:
Pain, light bleeding, heamatoma, possibly an infection.

Medication:
As 2-iminobiotin (2-IB), will be administered to man for the first time in this
study, to date adverse effects in man have not been reported. However, there
have been extensive studies on the safety of 2-IB in rats, dogs and mini pigs.
In these studies (very) high doses of 2-IB have been administered, following a
similar scheme as in the present study, over 24 to 96 hour periods. These
studies have shown that 2-IB was well tolerated up to the highest dose levels.
Only mild adverse effect have been observed in the test animals that were most
likely not related to 2-IB, and the results justify the administration of 2-IB
in humans. Captisol is an adjuvant that has been administered in preclinical
and clinical studies before and is also used in a number of already available
drugs. From these studies it was concluded that Captisol is safe and has no
significant biological effects.. The highest daily doses of Captisol
administered in humans thus far was 35 gram (4 administrations of 5 minutes
within 24 hours). Although the highest dose of Captisol used in this study will
exceed the highest daily dose administered in humans thus far, no safety issues
are expected of the new formulation since Captisol leaves the body quickly.