Antithrombin III Supplementation for Cardiopulmonary Bypass in Neonates

Hypothesis:

ATIII supplementation prior to initiation of cardiopulmonary bypass (CPB) to neonates will
be safe and result in less postoperative bleeding and transfusion of packed red blood cells
(pRBC) as compared to placebo controls.

Specific Aims:

1. To determine the safety and feasibility of ATIII supplementation prior to initiation of
CPB in neonates.

2. To determine the pharmacokinetics of ATIII supplementation prior to initiation of CPB
in neonates

3. To compare the postoperative blood loss and pRBC transfusion requirements in an
experimental group of neonates undergoing surgery requiring CPB who receive ATIII prior
to initiation of CPB to that of a placebo control group that did not receive ATIII.

Inclusion Criteria:

All sequential neonates (4-30 days of age) undergoing surgeries that require CPB at CHW are
eligible to be included in the study.

Exclusion Criteria:

Patients with a prior operation utilizing CPB, weight less than 2 kilograms, prematurity
less than 37 weeks estimated gestational age, previously diagnosed pro-thrombotic or
hemorrhagic disorder, known intracranial hemorrhage, prior ATIII supplementation, and prior
therapeutic anticoagulant use.

Protocol:

After informed consent, patients will be risk stratified into two groups based on whether
the planned surgery will result in a cyanotic or acyanotic palliation. The patients in
these two groups will then be randomized to either the ATIII group or the placebo group. A
preoperative ATIII blood level will be obtained. This level will guide the dosing of ATIII
or placebo, as neonatal congenital heart disease patients have variable baseline ATIII
levels. The dose of ATIII or placebo will be determined by the following formula, in order
to obtain ATIII blood levels similar to adults and older children (1.0 U/mL):

ATIII dose given to patient (U) = [(1.0 U/mL - patient's measured ATIII concentration in
U/mL) X weight (kg) X 80 mL/kg]

This formula is derived by taking into account the volume of distribution of ATIII in the
patient's blood volume. 80 mL/kg is an average blood volume for an infant. 1.0 U/ml is the
normal level for older children and adults, and the target level we are trying to achieve.
The patient's baseline level is subtracted from the target level to determine the relative
ATIII deficiency in terms of U/mL. That amount is then multiplied by the estimated blood
volume of 80 mL/kg and the patient's weight in kg to determine an ATIII dose in Units. The
CHW pharmacy will use the above formula to obtain the appropriate dose of ATIII or placebo
for each patient. ATIII or placebo will be administered after induction of general
anesthesia and prior to administration of heparin and initiation of CPB. All treating
physicians and study personal except for the preparing pharmacist will be blinded.

Heparin management during CPB will be accomplished according to the current standard
practice at CHW. Anticoagulation during bypass will be monitored and titrated by use of the
Medtronic Hepcon® HMS Plus Hemostasis Management System. A 3mL sample of the patient's
blood is drawn pre-incision, after ATIII or placebo dosing, and is used by the HMS device to
determine the in vitro individual response to heparin. A patient-specific heparin dose of
between 300 and 800 units/kg will be administered before initiation of CPB, with the
appropriate level of anticoagulation maintained at a minimum activated clotting time (ACT)
of 480 seconds. The level of anticoagulation will be measured every half hour during CPB,
with additional doses of heparin periodically administered as dictated by the HMS System
measurements. At the completion of CPB, the heparin will be reversed with a
patient-specific protamine dose, as calculated by the HMS System. Repeat measurements of
ACT and heparin levels with the HMS device will be done after protamine administration to
insure adequate reversal of the heparin effect.

ATIII levels will be measured within 5-10 minutes after administration (prior to start of
CPB), within 30 minutes of start of CPB, just prior to discontinuation of CPB (prior to
protamine dosing), and upon admission to the ICU. These levels will be kept blinded to all
treating physicians and study personal.

An additional 2 mL of blood will be drawn at each time an ATIII level is obtained (total of
10 mL) to measure markers of activation of the coagulation and inflammatory systems, in
order to better characterize the derangements in this patient population over time. We
intend to use these results to develop future studies.

Intraoperative blood loss while on the CPB machine will be collected and placed back in the
CPB circuit as per standard practice at CHW. Blood loss (mL/kg) will be determined
intraoperatively from 10 minutes after protamine administration until admission to the
intensive care unit (ICU). Chest tube output will then be recorded as a measure of blood
loss for the first 24 hours after admission to the ICU.

After admission to the ICU, a transfusion protocol will then be used to ensure common
indications for blood product replacement. Patients will be transfused with pRBC (10-15
mL/kg) if their hematocrit is less than 42% (higher target for evidence of inadequate oxygen
delivery such as low SvO2, decreased cerebral NIRS, rising lactate, etc.). This protocol is
consistent with our current standard practice at CHW.

If the bedside clinician decides a transfusion is/is not indicated outside of these
guidelines, it will be recorded as a protocol variation and will be analyzed inclusively and
separately. Volume of whole blood transfused will be multiplied by 0.5, as the
concentration of red blood cells in whole blood is about 0.5 times the concentration present
in packed red blood cells.

Additional serum blood samples will be drawn preoperatively and postoperatively (on the day
the last invasive line is in place or 28 days postoperatively whichever is first) to
determine antibody formation to recombinant antithrombin and goat milk proteins as a measure
of safety of the specific ATIII product used.

Data Collection, Analysis, and Power Calculation:

Data will be recorded on a standardized data collection forms and stored in a RedCap data
base which allows for menu driven options, inbuilt checks and generation of reports.

Safety (Aim 1): We hypothesize that measurements of safety in the experimental group will
be the same or less than in the placebo controls We will record the mortality rate,
incidence of extracorporeal membrane oxygenation (ECMO) support within 24 hours
postoperatively, incidence of mediastinal exploration for bleeding and other indications
within 24 hours postoperatively, incidence of thrombotic disease at discharge (ultrasound or
other radiographic evidence if obtained for routine patient care), incidence of intracranial
hemorrhage (ultrasound or computed tomography if obtained for routine patient care), days to
delayed sternal closure, days to cessation of mechanical ventilation, and days to hospital
discharge in the experimental and control groups. We will compare these data using a
chi-square for categorical data, a t-test for continuous data, and Kaplan Meir curves with a
Wilcoxon rank sum test for censored data. We will also compare serum samples at baseline
and postoperatively for antibody formation to recombinant antithrombin or goat milk
proteins. An internal data safety monitoring board (DSMB) will be in place to review the
safety of the study after every patient for the first five, and then after every subsequent
five patients. In addition, the DSMB will convene to review any serious adverse events
potentially related to ATryn administration. A serious adverse event will be defined as
death, intracranial hemorrhage, or >60 ml/kg/day in pRBC transfusion volume and/or 120
ml/kg/day of chest tube output (2 standard deviations above the mean in historical
controls). At all points when the DSMB is planned to meet, the study will be placed on
temporary hold, and no subsequent patients will be enrolled until after the DSMB convenes.
If the DSMB concludes that ATryn administration contributed to a serious adverse event, or
that the rates of safety measures described above are significantly worse in the
ATryn-treated group at the time of a review, the study will be stopped.

Feasibility (Aim 1): We hypothesize that recruitment will be at a rate of 5 enrolled
patients per month.

During the course of this project, the number of subjects approached for consent, the number
consented, and difficulties in measurement (including unobtainable measures) will be
summarized overall and by demographics. Associations will be investigated with scatter
plots, Loess smooth curves, and Pearson correlation coefficients.

ATIII Pharmacokinetics (Aim 2): ATIII levels measured within 5-10 minutes after
administration (prior to start of CPB), within 30 minutes of start of CPB, just prior to
discontinuation of CPB (prior to protamine dosing), and upon admission to the ICU will be
assessed to determine if the single dose of ATIII sustained normal ATIII levels in the
experimental group throughout these time periods, and will be compared to placebo control
patients. If any ATIII level is above 1.2 (20% higher than anticipated based on dose
calculation) at the time of a review by the DSMB, the study will be placed on temporary hold
in order to review the dose calculation protocol.

Postoperative Blood Loss and pRBC Transfusion Volume (Aim 3): Our primary efficacy
hypothesis for this study is that blood loss and pRBC transfusion volumes will be less than
placebo. Blood loss will be measured in terms of volumes (mL/kg) of blood loss and chest
tube output from 10 minutes after protamine administration to 24 hours after ICU admission.
If a patient requires re-exploration of the chest in the first 24 hours postoperatively, the
volume of chest tube output for the duration of exploration will be recorded as a
time-weighted average equal to the chest tube loss for the 2 hours prior to and 2 hours
after the exploration. Volumes (mL/kg) of packed red blood cells and 0.5 X volume (mL/kg)
of whole blood transfused from 10 minutes after protamine administration to 24 hours after
ICU admission will be recorded. Patients who require ECMO within the first 24 hours
postoperatively will be analyzed separately, as this is a known risk factor of increased
bleeding and transfusion support. We will compare the blood loss and pRBC transfusion
volumes in the two groups using a t-test or a Mann Whitney test. Further, we will allow for
demographics such as sex and race, using a regression model with blood loss or pRBC volume
as the outcome and group, and demographics as the independent variables.

Power analysis: Since there are two primary outcomes of blood loss volume and pRBC
transfusion volume, we will use a Bonferroni adjustment and an alpha of 0.05/2= 0.025. With
30 patients in the experimental group and 30 controls, we will have at least 80% power to
detect a difference of 0.81 standard deviations.

Inclusion Criteria:

all sequential neonates (4 - 30 days of age) undergoing surgery requiring cardiopulmonary
bypass are eligible to be included in the study.

Exclusion Criteria:

- prior operation utilizing cardiopulmonary bypass

- weight less than 2 kilograms

- prematurity less than 37 weeks estimated gestational age

- previously diagnosed pro-thrombotic or hemorrhagic disorder

- known intracranial hemorrhage

- prior ATIII supplementation

- prior therapeutic anticoagulant use

- known hypersensitivity to goat and goat milk proteins.