Enhancement of Cerebral pO2 by RSR13, a Synthetic Allosteric
Modifier of Hemoglobin
1Oleg Y. Grinberg, 1Minoru Miyake, 1Huagang Hou, 2Hisham Elkadi, 3Robert
Steffen,
and 1Harold M. Swartz
1EPR Center for the Study of Viable Biological Systems, Dartmouth Medical
School, Hanover,
NH; 2Boston Medical Center, Boston, MA; 3Allos Therapeutics, Inc., Denver,
CO.
INTRODUCTION: Oxygen plays an essential role in brain functioning. It
well known that ischemia
and subsequent reperfusion cause irreversible injury of brain cells. RSR13
(2-[4-[[(3,5-
dimethylanilino) carbonyl] methyl] phenoxyl] -2-methylproprionic acid"Na), a
synthetic
allosteric modifier of hemoglobin decreases oxygen affinity of blood.
Therefore one may expect
that this drug could be useful for improving oxygen tension in the brain and
there is a growing
body of indirect evidence that this is the case. By use of the unique
capabilities of Electron
Paramagnetic Resonance (EPR) oximetry we have been able to measure directly
in vivo for the
first time whether this allosteric modifier does improve brain tissue
oxygenation in normal
functioning brain and following the severe stress of acute hemorrhage shock
in rats.
Physiological parameters also were monitored in order to examine the
physiological effect of
RSR13 other than the hemoglobin affinity.
METHODS: EPR oximetry utilizes an oxygen sensitive material LiPc
(lithium phthalocyanine)
whose spectroscopic characteristics vary with the oxygen level. These
crystals were placed into
the cerebral cortex of rats one week before the experiment using
stereotactic techniques. During
the experiment the rats were anesthetized with 1.2-1.5% isofluorane.
Respiratory support under
controlled respiration (FIO2: 25-28%) was provided by a volume cycled
respirator after
endotracheal intubation. In addition two polyethylene catheters were placed
in the left femoral
artery and vein for continuous monitoring of BP and periodic blood gas
measurements and
administration of RSR13. Using the data from a rectal probe, a heated pad
maintained body
temperature of the rats.
RESULTS: The experiment in normal functioning brain was performed
using three group of rats
(six animals per group): high-dose RSR13 (300mg/kg I.V.), low dose RSR13
(150 mg/kg I.V.),
and control group (with saline). Arterial blood gas was taken twice before
RSR13 administration
in order to find the optimal setting for the ventilator, and then blood
sampling was carried out at
15 min, 120min, 210 min, and 300 min after the start of the injection of
RSR13 or saline. The
RSR13 or saline was given I.V. over 15 min through the catheter to the
femoral vein using an
infusion pump Brain tissue pO2, blood pressure (systolic, diastolic, and
mean), heart rate, and
core body temperature were recorded every 3 min during the administration of
RSR13 or saline
and then every 15 min for 300 min. RSR13 significantly increased the pO2 in
the brain by about
6 torr in the lower-dose group and 12 torr in the high dose group.
The effect of RSR13 on brain tissue pO2 following severe hemorrhagic shock
in rats was
studied in a group of six controls and seven experimental rats. Hemorrhagic
shock was induced
by withdrawing blood (2.7-2.8 mL/100 g/15 min). Brain tissue pO2 was
monitored by EPR
oximetry throughout the experiment. Following a 30 min shock period,
resuscitation was
performed by infusion with Ringer lactate at 8.1-8.7 ml/100 g. Animals
received either RSR13
(150 mg/kg) or saline (control). Blood pressure (from the femoral artery:
systolic, diastolic, &
mean) and heart rate were periodically obtained. Core body temperature was
monitored
continuously by a rectal probe every 3 min during the first 15 min after
administration of RSR13
or saline and then every 15 min for 180 min. Following hemorrhage, brain pO2
decreased by
15.8±3.1 mm Hg and 13.3±1.9 mm Hg in the animals that would later receive
RSR13 or saline
(control), respectively. In the control group, following crystalloid
resuscitation, brain pO2
remained depressed. In the rats that received RSR13 with resuscitation
(experimental group), the
pO2 in the brain returned to the pre-hemorrhage values or slightly above.
Arterial pO2 (PaO2)
post-RSR13 administration also immediately increased and was maintained at
about 140 torr
from the post-hemorrhage value of about 90 torr. The controls had a very
gradual increase in
PaO2 to about 105 torr at 120 minutes. There was no difference in the blood
pressure or heart
rate between groups.
CONCLUSIONS: These effects have potentially important clinical
applications. RSR13 reversed the
cerebral hypoxia associated with acute blood loss, in a time period and
under conditions that
appear to be relevant to the clinical circumstances in which RSR13 might be
utilized as part of
the emergency treatment of traumatic hemorrhage. The resulting increase and
maintenance of
brain pO2 by RSR13 following severe hemorrhagic shock potentially may reduce
the neuronal
damage that can result from an insufficient supply of oxygen to the brain
induced by severe
hemorrhagic shock.
Presented at the Annual Meeting of the International Society for Oxygen
Transport to
Tissue and manuscript published in the conference proceedings.
M. Miyake, O. Y. Grinberg, H. Hou, R.P. Steffen, H. El-Kadi, and H.M.
Swartz, “The Effect of RSR13, a
Synthetic Allosteric Modifier of Hemoglobin, on Brain Tissue pO2 (Measured
by EPR Oximetry)
Following Severe Hemorrhagic Shock in Rats,” Oxygen Transport to Tissue
XXII. (Dunn, J.F. and H.M.
Swartz eds). Pabst Science Publishers. Lengerich. (2001).