U.S. patent application number 12/375182 was filed with the patent office on 2009-12-31 for trauma therapy.
This patent application is currently assigned to Hibernation Therapeutics Limited. Invention is credited to Geoffrey Philip Dobson.
Application Number | 20090324748 12/375182 |
Document ID | / |
Family ID | 38981058 |
Filed Date | 2009-12-31 |
United States Patent
Application |
20090324748 |
Kind Code |
A1 |
Dobson; Geoffrey Philip |
December 31, 2009 |
TRAUMA THERAPY
Abstract
The invention provides a method of reducing injury to cells,
tissues or organs of a body following trauma by administering a
composition to the body following trauma, including: (i) a
potassium channel opener or agonist and/or an adenosine receptor
agonist; and (ii) a local anaesthetic. Also provided is a
composition for reducing injury to cells, tissues or organs of a
body following trauma including: (i) and (ii). The composition may
be hypertonic.
Inventors: |
Dobson; Geoffrey Philip;
(Wulguru, AU) |
Correspondence
Address: |
VENABLE LLP
P.O. BOX 34385
WASHINGTON
DC
20043-9998
US
|
Assignee: |
Hibernation Therapeutics
Limited
Wulguru
AU
|
Family ID: |
38981058 |
Appl. No.: |
12/375182 |
Filed: |
July 25, 2007 |
PCT Filed: |
July 25, 2007 |
PCT NO: |
PCT/AU07/01029 |
371 Date: |
January 26, 2009 |
Current U.S.
Class: |
424/682 ;
514/46 |
Current CPC
Class: |
A61P 17/02 20180101;
A61K 47/02 20130101; A61K 31/167 20130101; A61K 31/167 20130101;
A61K 31/7076 20130101; A61P 23/02 20180101; A61K 38/28 20130101;
A61K 33/06 20130101; A61K 31/4045 20130101; A61K 9/0019 20130101;
A61K 31/7076 20130101; A61K 2300/00 20130101; A61K 45/06 20130101;
A61K 2300/00 20130101 |
Class at
Publication: |
424/682 ;
514/46 |
International
Class: |
A61K 33/06 20060101
A61K033/06; A61K 31/7076 20060101 A61K031/7076; A61P 23/02 20060101
A61P023/02 |
Foreign Application Data
Date |
Code |
Application Number |
Jul 25, 2006 |
AU |
2006904007 |
Jan 19, 2007 |
AU |
2007900283 |
Claims
1. A method of reducing injury to cells, tissues or organs of a
body following trauma by administering a composition to the body
following trauma, including: (i) a potassium channel opener or
agonist and/or an adenosine receptor agonist; and (ii) a local
anaesthetic.
2. A method according to claim 1 wherein a further composition
including (i) and (ii) is administered following administration of
the composition.
3. A method according to claim 1 wherein the composition further
includes divalent magnesium cations.
4. A method according to claim 3 wherein the concentration of
magnesium is up to about 2.5 mM.
5. A method according to claim 1 wherein the composition is
hypertonic.
6. A composition for reducing injury to cells, tissues or organs of
a body following trauma including: (i) a potassium channel opener
or agonist and/or an adenosine receptor agonist; and (ii) a local
anaesthetic.
7. A composition according to claim 6, further including divalent
magnesium cations.
8. A composition according to claim 6 which is hypertonic.
9. A method of treating a trauma victim by administering a
composition, including: (i) a potassium channel opener or agonist
and/or an adenosine receptor agonist; and (ii) a local
anaesthetic.
10. A method of treating a trauma victim by administering a
composition according to claim 6.
11. A composition according to claim 7, wherein the concentration
of magnesium is up to about 2.5 mM.
Description
FIELD OF THE INVENTION
[0001] This invention relates to a method of reducing injury to
cells, tissues or organs of a body following trauma, including
injury to cells, tissues or organs resulting from shock, stroke,
heart conditions or other injuries that may occur as a consequence
of trauma.
BACKGROUND OF THE INVENTION
[0002] In the western world, many deaths occur suddenly and
unexpectedly, particularly as a consequence of trauma. Medically,
"trauma" refers to a serious or critical bodily injury, wound, or
shock which in some cases may require resuscitation therapy. Trauma
is often associated with trauma medicine practiced in hospital
(such as in hospital emergency rooms), in emergency transport
environments (such as in ambulances), or at out-of-hospital
environments where a trauma has occurred, such as domestic or
industrial accidents, transport accidents, the battlefield, and
terrorist attacks.
[0003] Trauma is a leading cause of death among children and all
individuals to age 34 years and a major cause of death in the older
population resulting in loss of productive life-years with
substantial societal costs. This includes deaths resulting from
burns, heart attacks, strokes and other cardiovascular events.
Deaths can also result from shock or other complications that may
occur as a consequence of trauma.
[0004] Less than 3% of these unconscious trauma patients will
advance to acceptable outcomes. Many survivors require
institutional care after 3 months and a significant proportion
remain permanently disabled. About 20% of soldiers injured in the
battlefield will die, and 90% of deaths occur before reaching a
hospital because of shock during emergency transport. More recent
statistics suggest that 50% of deaths in potentially treatable
combat injuries are due to acute blood loss, making it the leading
cause of death on the battlefield.
[0005] Shock is a circulatory dysfunction causing decreased tissue
oxygenation and accumulation of oxygen debt, which can ultimately
lead to multi-organ system failure if left untreated. The most
common form of shock in both paediatric and adult trauma patients
is hemorrhagic or hypovolemic shock (not enough blood volume).
Cardiogenic shock (not enough output of blood by the heart, see
below) is also a common form of shock. Shock as a result of blood
loss is a frequent complication of trauma. About half of trauma
deaths occur during the first hour after injury from a profound
compromise in cardiopulmonary and cerebral function. The signs and
symptoms of shock include low blood pressure (hypotension),
overbreathing (hyperventilation), a weak rapid pulse, cold clammy
greyish-bluish (cyanotic) skin, decreased urine flow (oliguria),
and mental changes (a sense of great anxiety and foreboding,
confusion and, sometimes, combativeness). When blood is lost, the
greatest immediate need is replacing the lost volume with blood or
blood volume expanders. Provided blood volume is maintained by
volume expanders, a trauma patient can safely tolerate very low
blood haemoglobin levels, less than one third of a healthy
person.
[0006] During trauma, the electrical properties of vital organs and
tissues cannot be maintained. Falls in resting cell voltage occur
during trauma and can lead to the triggering of highly injurious
arrhythmias in the heart and activation of systemic inflammatory,
coagulative and free radical generating processes that can lead to
multiple organ failure and death. During severe haemorrhage,
patients become unconscious when the mean arterial perfusion
pressure decreases to about 40 mm Hg and the pulse is no longer
palpable in the large arteries. When breathing stops and pulsations
are no longer palpable, cardiac arrest is assumed. The mortality
rate for trauma patients who become pulseless from massive blood
loss and undergo emergency department thoracotomy is around
97%.
[0007] One form of shock is called "cardiogenic shock". This may be
caused by the failure of the heart to pump effectively due to, for
example, damage to the heart muscle (as may result from a large
myocardial infarction (heart-attack), disorders of the heart muscle
(including rupture), disturbances to the electrical
excitation-relaxation (or conduction) system and tamponade.
Cardiogenic shock may also be caused by arrhythmias (eg ventricular
tachycardia and ventricular fibrillation), cardiomyopathy, cardiac
valve problems, ventricular outflow obstruction and the like.
Cardiogenic shock is a medical emergency requiring immediate
treatment to save the patient's life.
[0008] One cause of cardiogenic shock is a so-called
"heart-attack". This term is used to refer to a number of different
conditions which lead to heart ischaemia, which leads to the death
of heart muscle (typically caused by blockage of a coronary
artery). The muscle death causes chest pain and electrical
instability of the heart muscle tissue. This electrical instability
may manifest as "ventricular tachycardia" and "ventricular
fibrillation". Ventricular tachycardia is a tachydysrhythmia
originating from a ventricular ectopic focus and characterized by a
rate typically greater than 120 beats per minute and must be
treated quickly to avoid morbidity or mortality as it may
deteriorate rapidly into ventricular fibrillation. Ventricular
fibrillation is a condition in which there is chaotic electrical
disturbances of the ventricles, such that they no longer beat
regularly, nor pump blood effectively, but simply quiver. During
ventricular fibrillation the heart muscle is affected by a poor
supply of oxygen or by specific heart disorders and the ventricles
contract independently of the atria, and some areas of the
ventricles contract while others are relaxing, in a disorganized
manner. Ventricular fibrillation leads to widespread ischaemia.
Unless treated immediately, ventricular fibrillation causes death
and is responsible for 75% to 85% of sudden deaths in persons with
heart problems. In the USA alone there are nearly 450,000 sudden
deaths per year, and in the united kingdom around 70,000-90,000
sudden deaths per year. Ventricular tachycardia and ventricular
fibrillation are therefore medical emergencies because if they
persist more than a few seconds, the blood circulation will cease,
there will be no pulse, no blood pressure and no respiration and
death will occur. Typically, medications and procedures at this
time are directed towards stabilising the rhythm of the heart and,
in the case of the unconscious subject with no measurable pulse,
resuscitating the subject by restarting the heart, opening the
airways and restoring spontaneous breathing. Amiodarone can be used
to treat life-threatening heart arrhythmias, however, the drug can
have serious side effects including causing cardiac rhythm
irregularities and cardiac arrest itself. Other side effects of
amiodarone include lung infiltration, neuropathy, tremors, thyroid
disorders, nausea, low blood pressure and liver damage. Effective
medications for stabilizing the heart or restarting the heart and
restoring the spontaneous circulation in these emergency situations
are therefore very limited or non-existent. Noradrenalin or
adrenalin (with or without vasopressin) can be used in conjunction
with cardiopulmonary resuscitation, however, epinephrine can
exacerbate heart contractions and promote heart dysfunction by
increasing myocardial oxygen consumption during ventricular
fibrillation, as well as eliciting microvascular disorders. If the
treatments are successful in stabilising the heart after
ventricular tachycardia or ventricular fibrillation, a number of
medications are then administered such as oxygen (if available to
help breathing), beta-blockers (to help relax the heart),
vasodilators (to help deliver more blood to the heart), blood
agents (anti-coagulants, anti-platelet agents, thrombolytics and
the like) and pain relievers. Apart from a few drugs to treat the
heart as well as other tissues and organs, the medications are not
directed to treating the cardiac tissue specifically. There is no
effective pharmaceutical treatment for the failing heart muscle
itself, nor for common ventricular fibrillation. If treated, this
is usually treated by electrical shock (cardioversion).
[0009] Damage may also be caused to a heart upon reperfusion. One
example of reperfusion damage is when a heart becomes "stunned". In
this condition, the bloodflow has been restored but the heart is
functioning abnormally and may result in a further heart-attack
(such as ventricular fibrillation) if not treated. Cardiac
reanimation inevitably involves reperfusion of the heart with the
consequent dangers associated with reperfusion injury, particularly
to heart muscle. Where the muscle cells die, this is regarded as an
infarction. If blood flow is restored to the cells within a short
period of about 15 to 20 minutes the cells may respond to the
reperfusion and survive (thus not forming an infarction) but may be
"stunned" in the sense that they do not operate normally nor
perform their usual function during reperfusion.
[0010] In patients who survive resuscitation where the initial
event may be less traumatic, they remain at a significant risk from
systemic and local inflammatory and immune activation followed by
multiple organ dysfunction and failure. Multiple organ failure is
believed to be the result of an excessive self-destructive systemic
inflammation and immunologic functions, in which hypoxemia, tissue
hypoxia/nonviable tissue, micro-organisms/toxins and
antigen/antibody complexes may be involved. In particular, the
activation of a number of humoral (e.g. complement, coagulation)
and cellular systems (endothelium activation, neutrophils,
platelets, macrophages) are believed to be involved. Neutrophils
play a key role in injury to the lung, heart, kidney, liver, and
gastrointestinal tract, often seen after major trauma. As a
consequence there is synthesis, expression and release of numerous
mediators (toxic oxygen species, proteolytic enzymes, adherence
molecules, cytokines), which may produce a generalized inflammation
and tissue damage in the body.
[0011] The critical core body temperature also can aggravate many
of these post-traumatic secondary complications. Below 34.degree.
C. mortality increases significantly. Despite this, a number of
investigators have suggested a beneficial effect of deliberate
hypothermia because this may prolong the "golden hour" of trauma
victims by preventing hypoxic organ dysfunction and initiation of
the inflammatory response. Organ failure is also the leading cause
of death in the postoperative phase after major surgery. An
excessive inflammatory response followed by a dramatic depression
of cell-mediated immunity after major surgery appears to be
responsible for the increased susceptibility to subsequent
sepsis.
[0012] Resuscitation therapy is generally regarded as any procedure
which improves the management of sudden states of life-threatening
illnesses or traumatic injuries, such as those from cardiac arrest,
respiratory failure, hemorrhagic blood loss, neurological injury,
and traumatic injuries to the soft tissues and body skeleton.
Generally, resuscitation therapy deals with treating whole body
oxygen deprivation. As such, current resuscitation strategies aim
to optimize tissue supply and demand ratio and avoid complications
of overaggressive volume replacement, which exacerbate haemorrhage,
pulmonary oedema, and intracranial hypertension following brain
injury.
[0013] Resuscitation therapy is very different from treating a
localized "big heart attack" or a localized "big stroke". It
involves a complex interplay between multiple organ-tissue
responses via poorly understood actions, which separates this
science from treatments to preserve particular organs or tissues.
Resuscitation is known to involve a complex biological system, with
many interactions. These cannot be predicted from study of
individual components. Injured organs have secondary effects on
other organs, which affects the whole body and can lead to
debilitating injuries and death.
[0014] Current therapies involve fluid or volume replacement that
can either be crystalloid or colloidal. Crystalloids are commonly
used for resuscitation therapy because they appear to be safe and
help with the negative side effects of coagulation. Crystalloids
have been shown to increase coagulation, an effect which seems to
be independent of the type of crystalloid used. A
crystalloid-induced hypercoagulable state appears to be due to an
imbalance between naturally occurring anticoagulants and activated
procoagulants. Crystalloids used for volume replacement can be
three main types: 1) hypotonic (eg. dextrose in water), 2) isotonic
(normal saline or Ringers solution with lactate or acetate) or 3)
hypertonic (eg 7.5% saline). Since crystalloids are freely
permeable to the vascular membrane, only about 25% remain in the
blood compartment and the remainder in the body's interstitial
and/or intercellular compartment leading to tissue oedema.
Crystalloid resuscitation is therefore less likely to achieve
adequate restoration of microcirculatory blood flow compared to a
colloidal-based volume replacement strategy.
[0015] Colloid replacement therapies employ colloids, such as
dextran-70, dextran-40, hydroxyethyl starch, pentastarch,
lactobionate, sucrose, mannitol and a modified fluid gelatine as
artificial colloids, for this purpose. There is much controversy as
to the most appropriate solution for volume replacement.
[0016] Currently there is no optimal fluid composition or fluid
resuscitation regimen to treat severe hemorrhagic shock in soldiers
on the battlefield or civilians at a natural disaster site or
injured from a terrorist attack. Indeed, the majority of approved
resuscitation fluids have no intrinsic tissue protection and can
trigger life-threatening inflammatory and hypercoagulable
imbalances that negatively impact on the resuscitative outcome. A
further challenge in designing new drug products and resuscitation
therapies, in particular for the military, is hampered by
logistical considerations imposed by the combat conditions
themselves such as weight and practicability to transport, ease of
deployment, administration in low-light environments and stability
of drugs in the field, notwithstanding ensuring their safety and
clinical effectiveness to increase the survival times of wounded
soldiers after prolonged evacuation.
[0017] In warfare, bullets and penetrating fragments from exploding
munitions frequently cause life-threatening hemorrhage. Acute
hemorrhage is the leading cause of mortality in battlefield
injuries and responsible for 50% of deaths in potentially treatable
combat casualties. One major unmet medical need on the battlefield
is how to prevent cardiac destabilization and arrest during severe
hemorrhage before control of bleeding is possible. Stabilizing
heart and circulatory deficiencies before shock is of paramount
importance. Successful treatment of cardiac arrest requires an
electrically stable and mechanically viable heart to be
re-established. Currently there is no clinically effective method
of stabilizing and protecting the heart from fibrillating and
arresting before hemorrhagic shock. Indeed, many pharmacological
interventions employed to convert the heart to sinus rhythm may
unfortunately inflict additional injury and compromise cardiac
resuscitability
[0018] In those severe traumatic hemorrhagic cases where the heart
does not destabilize and arrest, the loss of blood volume, blood
pressure and organ perfusion can lead to severe organ ischemia and
eventually multiple organ dysfunction and failure (MOF) and death.
MOF is the leading cause of mortality secondary to shock
(hemorrhage/trauma) and resuscitation, and involves the lungs,
kidneys, intestinal tract, pancreas, liver, brain and heart.
Importantly, MOF is not an end-point per se but a process involving
an overwhelming self-destructive, local and systemic, inflammatory
responses and immunologic functions. Despite decades of research,
resuscitation fluids restore tissue perfusion, however they have no
specific anti-inflammatory, immunosuppression or pro-survival
properties. Importantly, the activation of shock-induced
inflammatory response occurs during the shock itself, during early
crystalloid or colloid-based resuscitation therapy, and during
final resuscitation efforts with blood replacement.
[0019] It is not known whether protection from injury from trauma
could be elicited by a form of artificial hibernation. Natural
hibernators possess the ability to lower their metabolic energy
demand for days to months. Hibernation, like sleep, is a form of
dormancy and helps to keep the animal's metabolic supply and demand
ratio in balance. Remarkably, no damage occurs during these
prolonged "ischemic" states, nor does the cardiac rhythm
deteriorate into ventricular fibrillation. However, there is no
known method of stimulating a similar response in humans,
particularly trauma patients, despite the potential for substantial
saving of life or minimising injury.
[0020] WO00/56145, WO04/056180 and WO04/056181 describe
compositions useful to limit damage to a cell, tissue or organ by
administering them in a clinical setting prior to a medical
procedure. These compositions are also usually administered
following diagnosis of the patient and directly to the cell, tissue
or organ. However, much damage or injury to cells, tissues or
organs may arise before the patient gets to the hospital and/or at
hospital, for example, before substantive medical attention is
available or a condition can be diagnosed.
SUMMARY OF THE INVENTION
[0021] The present invention is directed toward overcoming or at
least alleviating one or more of the difficulties and deficiencies
of the prior art.
[0022] In one aspect the invention is directed to a method of
reducing injury to cells, tissues or organs of a body following
trauma by administering a composition to the body following trauma,
including: (i) a potassium channel opener or agonist and/or an
adenosine receptor agonist; and (ii) a local anaesthetic.
[0023] According to this aspect, a further composition comprising
components (i) and (ii) may be administered to the body following
administration of the composition.
[0024] Either composition may include Magnesium cations (divalent)
and/or may be hypertonic.
[0025] In another aspect the invention is directed to a composition
for reducing injury to cells, tissues or organs of a body following
trauma including: (i) a potassium channel opener or agonist and/or
an adenosine receptor agonist; and (ii) a local anaesthetic. In one
embodiment of this aspect, the composition may include divalent
magnesium cations and/or may be hypertonic.
DETAILED DESCRIPTION OF THE INVENTION
[0026] The invention is directed to improved resuscitation
therapies for trauma victims in hospital, emergency transport and
out-of-hospital environments. In particular, the invention has
application to minimise life-threatening complications of persons
suffering injury to cells, tissues or organs resulting from burns,
shock, stroke, heart attack or other physical events, including
complications from surgical or clinical interventions, as a
consequence of trauma. Injured soldiers on the battlefield or
civilians at a natural disaster site or injured from a terrorist
attack are situations where such treatment may be useful.
[0027] The invention applies to protecting, preserving or
stabilising key organs such as the heart and brain, other neuronal
tissues and cells, renal tissue, lung tissue, muscle tissue, liver
and other tissues of the body.
[0028] In one form, the invention provides a method of reducing
injury to the cells, tissues or organs of a body following trauma
by administering a composition to the body following trauma
including: (i) a potassium channel opener or agonist and/or an
adenosine receptor agonist; and (ii) a local anaesthetic.
[0029] In another form of the invention, the invention is directed
towards treating tachycardia and/or fibrillation. In one form, the
invention treats heart arrhythmias of atrial or ventricular origin,
especially ventricular fibrillation. The treatment of tachycardia
and/or fibrillation, including ventricular fibrillation and
arrhythmias, comprises administering a composition including: (i) a
potassium channel opener or agonist and/or an adenosine receptor
agonist; and (ii) a local anaesthetic, in amounts effective to
arrest a heart. In one embodiment, the amount administered is
effective to arrest the heart only momentarily. This is often
sufficient to facilitate the heart cardioconverting back to normal
rhythm. In an alternate embodiment, the amount administered is
effective to substantially down-regulate the beating of the heart
for a period of a few beats, before allowing the heart to regain
its usual rhythm. The invention also extends to a method for
treating tachycardia and/or fibrillation accordingly. Preferably,
the composition is administered as a bolus. The administration of
the composition is believed to quell the tachycardia and/or
fibrillation allowing the heart to cardiovert to a normal and
desirable sinus rhythm.
[0030] In a preferred embodiment, the invention comprises the
further step of subsequently administering a second composition
which includes (i) a potassium channel opener or agonist and/or an
adenosine receptor agonist; and (ii) a local anaesthetic, in
amounts below that effective to arrest a heart. The purpose of the
second composition is to protect the heart and other tissues such
as brain, liver, lung and kidney, or assist in doing so. In
particular, this embodiment is directed towards reducing
reperfusion injury or stunning. As outlined above, reperfusion
injury is a common deleterious occurrence upon successfully
converting a tachycardic/fibrillating heart to a normal and
desirable sinus rhythm. In a preferred embodiment, the second
composition is administered as another non-arresting bolus
injection or delivered continuously via an intravenous drip or by
another delivery device or route.
[0031] Without being bound by any theory or mode of action, the
inventor has found that the composition according to the invention
can be used to place the body, in effect, toward a state of
suspended animation like a natural hibernator or to stabilise the
body prior to diagnosis or until suitable medical attention can be
provided to the trauma victim. The overall protection provided by
therapy according to the invention is thought to involve a
multi-tiered system from modulating membrane excitability to a
multitude of intracellular signalling pathways, including heat
shock and pro-survival kinase pathways. A primary focus is on
reducing damage to the brain, heart and lungs, because this has
been correlated with improved recovery and clinical outcomes.
Nonetheless, broad-acting approaches reducing damage throughout the
body in a non-specific way are desirable. Proposed mechanisms of
the composition of the invention include (i) reduced ion
imbalances, in particular sodium and calcium ion loading in the
cells, which may help defend the cell's voltage when stressed; (ii)
attenuation of local and systemic inflammatory response to injury,
which is protective in itself to reduce injury as well as reduce
secondary effects such as free radical production; and (iii)
protection from entering into a hypercoagulable state, ie an
anti-clotting or anti-thrombolytic activity. Moreover, it is
believed that, in respect of the heart, the invention
simultaneously provides improved atrial and ventricular matching of
electrical conduction to metabolic demand, which may involve
modulation of gap junction communication, and, in respect of the
brain, improved brain function. It is also believed that the
composition may reduce the body's demand for oxygen to varying
degrees and thus reduce damage to the body's cells, tissues or
organs. In another form, the invention provides a composition for
reducing injury to cells, tissues or organs of a body following
trauma including (i) a potassium channel opener or agonist and/or
an adenosine receptor agonist; and (ii) a local anaesthetic. The
composition may further include other components as identified
below. In some embodiments, the potassium channel opener or agonist
and/or adenosine receptor agonist is replaced by another component
such as a calcium channel blocker. The composition preferably
contains an effective amount of (i) and (ii) for a single dose to
reduce injury.
[0032] More surprisingly, it has been observed that administration
of a composition with arresting or near-arresting concentrations of
components (i) and (ii) to a subject experiencing ventricular
fibrillation assists the heart to regain normal sinus rhythm
without the requirement for electrical shock treatment.
[0033] The invention may also be used to treat or inhibit
arrhythmias including ventricular fibrillation during or prior to
an angiogram test or an exercise test. Similarly it has application
during emergency transport of an injured patient and for on-site
emergency treatment (ie, at the site of injury or heart-attack such
as an airport, sports stadium, hospital, battlefield or disaster
site). It can also be used before, during and/or after coronary
interventions such as angioplasty, cardiac catheter procedures, or
insertion of a pacemaker or leads or a device, or for surgical
procedures including paediatric or adult heart surgery, hip, knee,
vascular or brain surgery, aortic dissections, carotid
endaterectomy or general surgery.
[0034] In the embodiments of the invention described above and
below, component (i) of the composition may be an adenosine
receptor agonist. While this obviously includes adenosine itself,
the "adenosine receptor agonist" may be replaced or supplemented by
a compound that has the effect of raising endogenous adenosine
levels. This may be particularly desirable where the compound
raises endogenous adenosine levels in a local environment within a
body. The effect of raising endogenous adenosine may be achieved by
a compound that inhibits cellular transport of adenosine and
therefore removal from circulation or otherwise slows its
metabolism and effectively extends its half-life (for example,
dipyridamole) and/or a compound that stimulates endogenous
adenosine production such as purine nucleoside analogue
Acadesine.TM. or AICA-riboside (5-amino-4-imidazole carboxamide
ribonucleoside). Acadesine.TM. is also a competitive inhibitor of
adenosine deaminase (Ki=362 microMolar in calf intestinal mucosa.)
Acadesine.TM. is desirably administered to produce a plasma
concentration of around 50 microM (uM) but may range from 1 microM
to 1 mM or more preferably from 20 to 200 uM. Acadesine.TM. has
shown to be safe in humans from doses given orally and/or
intravenous administration at 10, 25, 50, and 100 mg/kg body weight
doses.
[0035] In one form of the invention, the composition, and
optionally the second composition, also contains divalent magnesium
cations. In one embodiment, the concentration of magnesium is up to
about 2.5 mM and in another embodiment magnesium is present in
higher concentrations, for example up to about 20 mM. The magnesium
is present as a physiologically and pharmaceutically acceptable
salt, such as for example magnesium chloride and magnesium
sulphate.
[0036] In another form the composition according to the invention
is hypertonic. Preferably the composition contains 7.5% NaCl. The
inventor has found that only a small volume of this hypertonic
composition may be administered to the subject in need thereof.
This is particularly advantageous where the composition according
to the invention has application during emergency or for emergency
transport. According to this aspect, only a small amount of the
composition according to the invention needs to be available, for
example, in a medical kit or ambulance. Thus the composition is
easier to store and/or transport. This "low volume" composition has
unique features of fluid replacement and specific
anti-inflammatory, immunosuppression pro-survival properties. The
composition according to this aspect of the invention
pharmacologically "buys" time for wounded soldiers on the
battlefield or civilians in urban "disaster zones" which allow for
safer evacuation, triage, and initiation of supportive therapies.
The ability of a solution to change the shape or tone of cells by
altering their internal water volume is called tonicity
(tono=tension). A Hypertonic solution contains a higher
concentration of electrolytes than that found in body cells and,
therefore, relatively less water in this compartment than inside
the body cells. In such a hypertonic environment, osmotic pressure
causes water to flow out of the cell into the hypertonic
environment. Thus a hypertonic solution creates a hyperosmotic
environment and the higher osmotic pressure in this environment
relative to the surrounding cells in tissues causes fluid to flow
from the cells towards such a system. If too much water is removed
in this way, the cell may have difficulty functioning.
[0037] The invention described in this specification largely
relates to methods of treatment, and methods of manufacturing a
medicament for treatment involving a composition which is described
as containing these components (i) and (ii). For convenience, this
composition will be referred to in this specification as the
"composition of the invention", although there are a number of
combinations of components embodying the invention which are
compositions according to the invention. Moreover, as explained
particularly in WO00/56145, the components (i) and (ii) may be
present in a concentration which arrests, or does not arrest a
heart. These two compositions are used in different ways in the
invention described in the specification, and are referred to
respectively as an "arresting" concentration of the composition and
a "non-arresting" concentration of the composition. In one form,
the arresting composition contains adenosine and lignocaine, each
at greater than 0.1 mM (and preferably below 20 mM). The arresting
composition may in some circumstances be referred to as a
"cardioplegia solution". In one form of the non-arresting
composition, adenosine and lignocaine are both below 0.1 mM and
preferably 50 nM to 95 uM, or more preferably from 1 uM to 90
uM.
[0038] In a further form, the invention provides use of (i) a
potassium channel opener or agonist and/or an adenosine receptor
agonist; and (ii) a local anaesthetic, for the preparation of a
medicament for reducing injury to cells, tissues or organs of a
body following trauma. The use preferably includes administering
the medicament in one or more of the ways set out elsewhere in this
specification.
[0039] In another form, the invention provides a method of, in
effect, placing the body in or toward a hibernating-like state of
suspended animation following trauma. This is achieved by
administering a composition as described above.
[0040] The term "trauma" is used herein in its broadest sense and
refers to a serious or critical injury, wound or shock to the body.
Trauma may be caused by unexpected physical damage (or injury) to
the body as a result of, for example, transport or industrial
accidents, birth, surgery, heart attack, stroke, burns,
complications due to surgery or other medical interventions etc.
Trauma may result from injury to a body, both in a hospital or out
of hospital. Trauma is often associated with trauma medicine
practiced in hospital (such as in hospital emergency rooms), during
emergency transport or at out-of-hospital environments where a
trauma has occurred, such as domestic or industrial accidents,
transport accidents, the battlefield, and terrorist attacks. In
many cases, trauma therapy may also include resuscitation
therapy.
[0041] The term "tissue" is used herein in its broadest sense and
refers to any part of the body exercising a specific function
including organs and cells or parts thereof, for example, cell
lines or organelle preparations. Other examples include circulatory
organs such as the heart, blood vessels and vasculature,
respiratory organs such as the lungs, urinary organs such as the
kidneys or bladder, digestive organs such as the stomach, liver,
pancreas or spleen, reproductive organs such as the scrotum,
testis, ovaries or uterus, neurological organs such as the brain,
germ cells such as spermatozoa or ovum and somatic cells such as
skin cells, heart cells ie, myocytes, nerve cells, brain cells or
kidney cells. The tissues may come from human or animal donors. The
donor organs may also be suitable for xenotransplantation.
[0042] The term "organ" is used herein in its broadest sense and
refers to any part of the body exercising a specific function
including tissues and cells or parts thereof, for example,
endothelium, epithelium, blood brain barrier, cell lines or
organelle preparations. Other examples include circulatory organs
such as the blood vessels, heart, respiratory organs such as the
lungs, urinary organs such as the kidneys or bladder, digestive
organs such as the stomach, liver, pancreas or spleen, reproductive
organs such as the scrotum, testis, ovaries or uterus, neurological
organs such as the brain, germ cells such as spermatozoa or ovum
and somatic cells such as skin cells, heart cells i.e., myocytes,
nerve cells, brain cells or kidney cells.
[0043] It will also be understood that the term "comprises" (or its
grammatical variants) as used in this specification is equivalent
to the term "includes" and should not be taken as excluding the
presence of other elements or features.
[0044] Potassium channel openers are agents which act on potassium
channels to open them through a gating mechanism. This results in
efflux of potassium across the membrane along its electrochemical
gradient which is usually from inside to outside of the cell. Thus
potassium channels are targets for the actions of transmitters,
hormones, or drugs that modulate cellular function. It will be
appreciated that the potassium channel openers include the
potassium channel agonists which also stimulate the activity of the
potassium channel with the same result. It will also be appreciated
that there are diverse classes of compounds which open or modulate
different potassium channels; for example, some channels are
voltage dependent, some rectifier potassium channels are sensitive
to ATP depletion, adenosine and opioids, others are activated by
fatty acids, and other channels are modulated by ions such as
sodium and calcium (ie. channels which respond to changes in
cellular sodium and calcium). More recently, two pore potassium
channels have been discovered and thought to function as background
channels involved in the modulation of the resting membrane
potential.
[0045] Potassium channel openers may be selected from the group
consisting of: nicorandil, diazoxide, minoxidil, pinacidil,
aprikalim, cromokulim and derivative U-89232, P-1075 (a selective
plasma membrane KATP channel opener), emakalim, YM-934,
(+)-7,8-dihydro-6,6-dimethyl-7-hydroxy-8-(2-oxo-1-piperidinyl)-6H-pyrano[-
2,3-f]benz-2,1,3-oxadiazole (NIP-121), RO316930, RWJ29009,
SDZPCO400, rimakalim, symakalim, YM099,
2-(7,8-dihydro-6,6-dimethyl-6H-[1,4]oxazino[2,3-f][2,1,3]benzoxadiazol-8--
yl) pyridine N-oxide,
9-(3-cyanophenyl)-3,4,6,7,9,10-hexahydro-1,8-(2H,5H)-acridinedione
(ZM244085),
[(9R)-9-(4-fluoro-3-125iodophenyl)-2,3,5,9-tetrahydro-4H-pyrano[3,4-b]thi-
eno[2,3-e]pyridin-8(7H)-one-1,1-dioxide] ([125I]A-312110),
(-)-N-(2-ethoxyphenyl)-N'-(1,2,3-trimethylpropyl)-2-nitroethene-1,1-diami-
ne (Bay X 9228), N-(4-benzoyl
phenyl)-3,3,3-trifluoro-2-hydroxy-2-methylpropionamine (ZD6169),
ZD6169 (KATP opener) and ZD0947 (KATP opener), WAY-133537 and a
novel dihydropyridine potassium channel opener, A-278637. In
addition, potassium channel openers can be selected from
BK-activators (also called BK-openers or BK(Ca)-type potassium
channel openers or large-conductance calcium-activated potassium
channel openers) such as benzimidazolone derivatives NS004
(5-trifluoromethyl-1-(5-chloro-2-hydroxyphenyl)-1,3-dihydro-2H-benzimidaz-
ole-2-one), NS1619
(1,3-dihydro-1-[2-hydroxy-5-(trifluoromethyl)phenyl]-5-(trifluoromethyl)--
2H-benzimidazol-2-one), NS1608
(N-(3-(trifluoromethyl)phenyl)-N'-(2-hydroxy-5-chlorophenyl)urea),
BMS-204352, retigabine (also GABA agonist). There are also
intermediate (eg. benzoxazoles, chlorzoxazone and zoxazolamine) and
small-conductance calcium-activated potassium channel openers.
Other compounds that are believed to open KATP channels include
Levosimendan and hydrogen sulphide gas (H.sub.2S) or the H.sub.2S
donors (eg sodium hydrosulphide, NaHS).
[0046] In addition, potassium channel openers may act as indirect
calcium antagonists, ie they act to reduce calcium entry into the
cell by shortening the cardiac action potential duration through
the acceleration of phase 3 repolarisation, and thus shorten the
plateau phase. Reduced calcium entry is thought to involve L-type
calcium channels, but other calcium channels may also be
involved.
[0047] Adenosine (6-amino-9-.beta.-D-ribofuranosyl-9H-purine) is
particularly preferred as the potassium channel opener. Adenosine
is capable of opening the potassium channel, hyperpolarising the
cell, depressing metabolic function, possibly protecting
endothelial cells, enhancing preconditioning of tissue and
protecting from ischaemia or damage. Adenosine is also an indirect
calcium antagonist, vasodilator, antiarrhythmic, antiadrenergic,
free radical scavenger, arresting agent, anti-inflammatory agent
(attenuates neutrophil activation), metabolic agent and possible
nitric oxide donor. More recently, adenosine is known to inhibit
several steps which can lead to slowing of the blood clotting
process. In addition, elevated levels of adenosine in the brain has
been shown to cause sleep and may be involved in different forms of
dormancy. An adenosine analogue, 2-chloro-adenosine, may be
used.
[0048] Suitable adenosine receptor agonists may be selected from:
N.sup.6-cyclopentyladenosine (CPA), N-ethylcarboxamido adenosine
(NECA), 2-[p-(2-carboxyethyl)phenethyl-amino-5'-N-ethylcarboxamido
adenosine (CGS-21680), 2-chloroadenosine,
N.sup.6-[2-(3,5-demethoxyphenyl)-2-(2-methoxyphenyl]ethyladenosine,
2-chloro-N-6-cyclopentyladenosine (CCPA),
N-(4-aminobenzyl)-9-[5-(methylcarbonyl)-beta-D-robofuranosyl]-adenine
(AB-MECA),
([IS-[1a,2b,3b,4a(S*)]]-4-[7-[[2-(3-chloro-2-thienyl)-1-methyl-propyl]ami-
no]-3H-imidazole[4,5-b]pyridyl-3-yl]cyclopentane carboxamide
(AMP579), N.sup.6--(R)-phenylisopropyladenosine (R-PLA),
aminophenylethyladenosine (APNEA) and cyclohexyladenosine (CHA).
CCPA is a particularly preferred. Others include full adenosine A1
receptor agonists such as N-[3-(R)-tetrahydrofuranyl]-6-aminopurine
riboside (CVT-510), or partial agonists such as CVT-2759 and
allosteric enhancers such as PD81723. Other agonists may include
N.sup.6-cyclopentyl-2-(3 phenylaminocarbonyltriazene-1-yl)
adenosine (TCPA), a very selective agonist with high affinity for
the human adenosine A1 receptor and allosteric enhancers of A1
adenosine receptor includes the 2-amino-3-napthoylthiophenes.
[0049] In one aspect, the composition according to the invention
includes an A1 adenosine receptor agonist and a local anaesthetic.
CCPA is a particularly preferred A1 adenosine receptor agonist.
[0050] Some embodiments of the invention utilise direct calcium
antagonists, the principal action of which is to reduce calcium
entry into the cell. These are selected from at least five major
classes of calcium channel blockers as explained in more detail
below. It will be appreciated that these calcium antagonists share
some effects with potassium channel openers, particularly
ATP-sensitive potassium channel openers, by inhibiting calcium
entry into the cell.
[0051] Calcium channel blockers are also called calcium antagonists
or calcium blockers. They are often used clinically to decrease
heart rate and contractility and relax blood vessels.
[0052] They may be used to treat high blood pressure, angina or
discomfort caused by ischaemia and some arrhythmias, and they share
many effects with beta-blockers, which could also be used to reduce
calcium. Beta-blockers (or beta-adrenergic blocking agents) include
atenolol (Tenormin.TM.), propranolol hydrochloride (such as
Inderal.TM.), esmolol hydrochloride (Brevibloc.TM.), metoprolol
succinate (such as Lopressor.TM. or Toprol XL.TM.), acebutolol
hydrochloride (Sectral.TM.), carteolol (such as Cartrol.TM.),
penbutolol sulfate (Levatol.TM.) and pindolol (Visken.TM.).
[0053] Five major classes of calcium channel blockers are known
with diverse chemical structures: 1. Benzothiazepines: eg
Diltiazem, 2. Dihydropyridines: eg nifedipine, Nicardipine,
nimodipine and many others, 3. Phenylalkylamines: eg Verapamil, 4.
Diarylaminopropylamine ethers: eg Bepridil, 5.
Benzimidazole-substituted tetralines: eg Mibefradil.
[0054] The traditional calcium channel blockers bind to L-type
calcium channels ("slow channels") which are abundant in cardiac
and smooth muscle which helps explain why these drugs have
selective effects on the cardiovascular system. Different classes
of L-type calcium channel blockers bind to different sites on the
alpha1-subunit, the major channel-forming subunit (alpha2, beta,
gamma, delta subunits are also present). Different sub-classes of
L-type channel are present which may contribute to tissue
selectivity. More recently, novel calcium channel blockers with
different specificities have also been developed for example,
Bepridil, is a drug with Na+ and K+ channel blocking activities in
addition to L-type calcium channel blocking activities. Another
example is Mibefradil, which has T-type calcium channel blocking
activity as well as L-type calcium channel blocking activity.
[0055] Three common calcium channel blockers are diltiazem
(Cardizem), verapamil (Calan) and Nifedipine (Procardia).
Nifedipine and related dihydropyridines do not have significant
direct effects on the atrioventricular conduction system or
sinoatrial node at normal doses, and therefore do not have direct
effects on conduction or automaticity. While other calcium channel
blockers do have negative chronotropic/dromotropic effects
(pacemaker activity/conduction velocity). For example, Verapamil
(and to a lesser extent diltiazem) decreases the rate of recovery
of the slow channel in AV conduction system and SA node, and
therefore act directly to depress SA node pacemaker activity and
slow conduction. These two drugs are frequency- and
voltage-dependent, making them more effective in cells that are
rapidly depolarizing. Verapamil is also contraindicated in
combination with beta-blockers due to the possibility of AV block
or severe depression of ventricular function. In addition,
mibefradil has negative chronotropic and dromotropic effects.
Calcium channel blockers (especially verapamil) may also be
particularly effective in treating unstable angina if underlying
mechanism involves vasospasm.
[0056] Omega conotoxin MVIIA (SNX-111) is an N type calcium channel
blocker and is reported to be 100-1000 fold more potent than
morphine as an analgesic but is not addictive. This conotoxin is
being investigated to treat intractable pain. SNX-482 a further
toxin from the venom of a carnivorous spider venom, blocks R-type
calcium channels. The compound is isolated from the venom of the
African tarantula, Hysterocrates gigas, and is the first R-type
calcium channel blocker described. The R-type calcium channel is
believed to play a role in the body's natural communication network
where it contributes to the regulation of brain function. Other
Calcium channel blockers from animal kingdom include Kurtoxin from
South African Scorpion, SNX-482 from African Tarantula, Taicatoxin
from the Australian Taipan snake, Agatoxin from the Funnel Web
Spider, Atracotoxin from the Blue Mountains Funnel Web Spider,
Conotoxin from the Marine Snail, HWTX-1 from the Chinese bird
spider, Grammotoxin SIA from the South American Rose Tarantula.
This list also includes derivatives of these toxins that have a
calcium antagonistic effect.
[0057] Direct ATP-sensitive potassium channel openers (eg
nicorandil, aprikalem) or indirect ATP-sensitive potassium channel
openers (eg adenosine, opioids) are also indirect calcium
antagonists and reduce calcium entry into the tissue. One mechanism
believed for ATP-sensitive potassium channel openers also acting as
calcium antagonists is shortening of the cardiac action potential
duration by accelerating phase 3 repolarisation and thus shortening
the plateau phase. During the plateau phase the net influx of
calcium may be balanced by the efflux of potassium through
potassium channels. The enhanced phase 3 repolarisation may inhibit
calcium entry into the cell by blocking or inhibiting L-type
calcium channels and prevent calcium (and sodium) overload in the
tissue cell.
[0058] Calcium channel blockers can be selected from nifedipine,
nicardipine, nimopidipine, nisoldipine, lercanidipine, telodipine,
angizem, altiazem, bepridil, amlopidine, felodipine, isradipine and
cavero and other racemic variations.
[0059] In a preferred form, the potassium channel opener or agonist
and/or an adenosine receptor agonist has a blood half-life of less
than one minute, preferably less than 20 seconds.
[0060] In some embodiments, the composition may include additional
potassium channel openers or agonists, for example diazoxide or
nicorandil.
[0061] The inventor has also found that the inclusion of diazoxide
with a potassium channel opener or adenosine receptor agonist and a
local anaesthetic reduces injury. Thus in another aspect, the
composition according to the invention further includes
diazoxide.
[0062] Diazoxide is a potassium channel opener and in the present
invention it is believed to preserve ion and volume regulation,
oxidative phosphorylation and mitochondrial membrane integrity
(appears concentration dependent). More recently, diazoxide has
been shown to provide cardioprotection by reducing mitochondrial
oxidant stress at reoxygenation. At present it is not known if the
protective effects of potassium channel openers are associated with
modulation of reactive oxygen species generation in mitochondria.
Preferably the concentration of the diazoxide is between about 1 to
200 uM. Typically this is as an effective amount of diazoxide. More
preferably, the concentration of diazoxide is about 10 uM.
[0063] The inventor has also found that the inclusion of nicorandil
with a potassium channel opener or adenosine receptor agonist and a
local anaesthetic reduces injury. Thus in another aspect, the
composition according to the invention further includes
nicorandil.
[0064] Nicorandil is a potassium channel opener and nitric oxide
donor which can protect tissues and the microvascular integrity
including endothelium from ischemia and reperfusion damage. Thus it
can exert benefits through the dual action of opening KATP channels
and a nitrate-like effect. Nicorandil can also reduce hypertension
by causing blood vessels to dilate which allows the heart to work
more easily by reducing both preload and afterload. It is also
believed to have anti-inflammatory and anti-proliferative
properties which can further attenuates ischemia/reperfusion
injury.
[0065] The composition according to the invention also includes a
compound for inducing local anaesthesia, otherwise known as a local
anaesthetic. The local anaesthetic may be selected from mexiletine,
diphenylhydantoin, prilocalne, procaine, mepivocaine, quinidine,
disopyramide and Class 1B antiarrhythmic agents such as lignocaine
or derivatives thereof, for example, QX-314.
[0066] Preferably the local anaesthetic is Lignocaine. In this
specification, the terms "lidocaine" and "lignocaine" are used
interchangeably. Lignocaine is preferred as it is capable of acting
as a local anaesthetic probably by blocking sodium fast channels,
depressing metabolic function, lowering free cytosolic calcium,
protecting against enzyme release from cells, possibly protecting
endothelial cells and protecting against myofilament damage. At
lower therapeutic concentrations lignocaine normally has little
effect on atrial tissue, and therefore is ineffective in treating
atrial fibrillation, atrial flutter, and supraventricular
tachycardias. Lignocaine is also a free radical scavenger, an
antiarrhythmic and has anti-inflammatory and anti-hypercoagulable
properties. It must also be appreciated that at non-anaesthetic
therapeutic concentrations, local anaesthetics like lignocaine
would not completely block the voltage-dependent sodium fast
channels, but would down-regulate channel activity and reduce
sodium entry. As anti-arrhythmic, lignocaine is believed to target
small sodium currents that normally continue through phase 2 of the
action potential and consequently shortens the action potential and
the refractory period.
[0067] As lignocaine acts by primarily blocking sodium fast
channels, it will be appreciated that other sodium channel blockers
may be used instead of or in combination with the local anaesthetic
in the method and composition of the present invention. It will
also be appreciated that sodium channel blockers include compounds
that act to substantially block sodium channels or at least
downregulate sodium channels. Examples of suitable sodium channel
blockers include venoms such as tetrodotoxin and the drugs
primaquine, QX, HNS-32 (CAS Registry # 186086-10-2), NS-7,
kappa-opioid receptor agonist U50 488, crobenetine, pilsicainide,
phenytoin, tocainide, mexiletine, NW-1029 (a benzylamino
propanamide derivative), RS100642, riluzole, carbamazepine,
flecainide, propafenone, amiodarone, sotalol, bretylium, imipramine
and moricizine, or any of derivatives thereof. Other suitable
sodium channel blockers include: Vinpocetine (ethyl
apovincaminate); and Beta-carboline derivative, nootropic
beta-carboline (ambocarb, AMB).
[0068] In one aspect, the composition according to the invention
consists essentially of (i) a potassium channel opener or agonist
and/or an adenosine receptor agonist; and (ii) a local
anaesthetic.
[0069] In another aspect, the composition according to the
invention may further include an opioid. The further addition of an
opioid may have similar if not improved effect on the reduction of
injury.
[0070] Opioids, also known or referred to as opioid agonists, are a
group of drugs that inhibit opium (Gr opion, poppy juice) or
morphine-like properties and are generally used clinically as
moderate to strong analgesics, in particular, to manage pain, both
peri- and post-operatively. Other pharmacological effects of
opioids include drowsiness, respiratory depression, changes in mood
and mental clouding without loss of consciousness.
[0071] Opioids are also believed to be involved as part of the
`trigger` in the process of hibernation, a form of dormancy
characterised by a fall in normal metabolic rate and normal core
body temperature. In this hibernating state, tissues are better
preserved against damage that may otherwise be caused by diminished
oxygen or metabolic fuel supply, and also protected from ischemia
reperfusion injury.
[0072] There are three types of opioid peptides: enkephalin,
endorphin and dynorphin. Opioids act as agonists, interacting with
stereospecific and saturable binding sites, in the heart, brain and
other tissues. Three main opioid receptors have been identified and
cloned, namely mu, kappa, and delta receptors. All three receptors
have consequently been classed in the G-protein coupled receptors
family (which class includes adenosine and bradykinin receptors).
Opioid receptors are further subtyped, for example, the delta
receptor has two subtypes, delta-1 and delta-2.
[0073] Cardiovascular effects of opioids are directed within the
intact body both centrally (ie, at the cardiovascular and
respiratory centres of the hypothalamus and brainstem) and
peripherally (ie, heart myocytes and both direct and indirect
effects on the vasculature). For example, opioids have been shown
to be involved in vasodilation. Some of the action of opioids on
the heart and cardiovascular system may involve direct opioid
receptor mediated actions or indirect, dose dependent non-opioid
receptor mediated actions, such as ion channel blockade which has
been observed with antiarrhythmic actions of opioids, such as
arylacetamide drugs. It is also known that the heart is capable of
synthesising or producing the three types of opioid peptides,
namely, enkephalin, endorphin and dynorphin. However, only the
delta and kappa opioid receptors have been identified on
ventricular myocytes.
[0074] Without being bound by any mode of action, opioids are
considered to provide cardioprotective effects, by limiting
ischemic damage and reducing the incidence of arrhythmias, which
are produced to counter-act high levels of damaging agents or
compounds naturally released during ischemia. This may be mediated
via the activation of ATP sensitive potassium channels in the
sarcolemma and in the mitochondrial membrane and involved in the
opening potassium channels. Further, it is also believed that the
cardioprotective effects of opioids are mediated via the activation
of ATP sensitive potassium channels in the sarcolemma and in the
mitochondrial membrane. Thus it is believed that the opioid can be
used instead or in combination with the potassium channel opener or
adenosine receptor agonist as they are also involved in indirectly
opening potassium channels.
[0075] It will be appreciated that the opioids include compounds
(natural or synthetic) which act both directly and indirectly on
opioid receptors. Opioids also include indirect dose dependent,
non-opioid receptor mediated actions such as ion channel blockade
which have been observed with the antiarrhythmic actions of
opioids.
[0076] Accordingly, the opioid may be selected from enkephalins,
endorphins and dynorphins. Preferably the opioid is an enkephalin
which targets delta, kappa and/or mu receptors. More preferably the
opioid is a delta opioid receptor agonist. Even more preferably the
opioid is selected from delta-1-opioid receptor agonists and
delta-2-opioid receptor agonists. [D-Pen 2, 5]enkephalin (DPDPE),
is a particularly preferred delta-1-opioid receptor agonist.
[0077] In another aspect the composition of the invention consists
essentially of (i) a potassium channel opener or agonist and/or an
adenosine receptor agonist; (ii) a local anaesthetic and (iii) a
delta-1-opioid. DPDPE is a particularly preferred delta-1-opioid
receptor agonist.
[0078] The inventor has found that the inclusion of a compound for
minimizing or reducing the uptake of water by a cell in a tissue
with a potassium channel opener or adenosine receptor agonist and a
local anaesthetic assists in reducing injury to a body, such as a
composition comprising sucrose, adenosine and lignocaine.
[0079] Thus in a further aspect, the composition according to the
invention may further include at least one compound for minimizing
or reducing the uptake of water by a cell in the cell, tissue or
organ.
[0080] A compound for minimizing or reducing the uptake of water by
a cell in the tissue tends to control water shifts, ie, the shift
of water between the extracellular and intracellular environments.
Accordingly, these compounds are involved in the control or
regulation of osmosis. One consequence is that a compound for
minimizing or reducing the uptake of water by a cell in the tissue
reduces cell swelling that is associated with Oedema, such as
Oedema that can occur during ischemic injury.
[0081] Compounds for minimizing or reducing the uptake of water by
a cell in a tissue are typically impermeants or receptor
antagonists or agonists. An impermeant according to the present
invention may be selected from one or more of the group consisting
of: sucrose, pentastarch, hydroxyethyl starch, raffinose, mannitol,
gluconate, lactobionate, and colloids. Colloids include albumin,
hetastarch, polyethylene glycol (PEG), Dextran 40 and Dextran 60.
Other compounds that could be selected for osmotic purposes include
those from the major classes of osmolytes found in the animal
kingdom including polyhydric alcohols (polyols) and sugars, other
amino acids and amino-acid derivatives, and methylated ammonium and
sulfonium compounds.
[0082] Cell swelling can also result from an inflammatory response
which may be important during organ retrieval, preservation and
surgical grafting. Substance P, an important pro-inflammatory
neuropeptide is known to lead to cell oedema and therefore
antagonists of substance P may reduce cell swelling. Indeed
antagonists of substance P, (-specific neurokinin-1) receptor
(NK-1) have been shown to reduce inflammatory liver damage, i.e.,
oedema formation, neutrophil infiltration, hepatocyte apoptosis,
and necrosis. Two such NK-1 antagonists include CP-96,345 or
[(2S,3S)-cis-2-(diphenylmethyl)-N-((2-methoxyphenyl)-methyl)-1-azabicyclo-
[2.2.2.)-octan-3-amine (CP-96,345)] and L-733,060 or
[(2S,3S).sub.3-([3,5-bis(trifluoromethyl)phenyl]methoxy)-2-phenylpiperidi-
ne]. R116301 or
[(2R-trans)-4-[1-[3,5-bis(trifluoromethyl)benzoyl]-2-(phenylmethyl)-4-pip-
eridinyl]-N-(2,6-dimethylphenyl)-1-acetamide
(S)-Hydroxybutanedioate] is another specific, active neurokinin-1
(NK(1)) receptor antagonist with subnanomolar affinity for the
human NK(1) receptor (K(i): 0.45 nM) and over 200-fold selectivity
toward NK(2) and NK(3) receptors. Antagonists of neurokinin
receptors 2 (NK-2) that may also reduce cell swelling include
SR48968 and NK-3 include SR142801 and SB-222200. Blockade of
mitochondrial permeability transition and reducing the membrane
potential of the inner mitochondrial membrane potential using
cyclosporin A has also been shown to decrease ischemia-induced cell
swelling in isolated brain slices. In addition glutamate-receptor
antagonists (AP5/CNQX) and reactive oxygen species scavengers
(ascorbate, Trolox(R), dimethylthiourea, tempol(R)) also showed
reduction of cell swelling. Thus, the compound for minimizing or
reducing the uptake of water by a cell in a tissue can also be
selected from any one of these compounds.
[0083] It will also be appreciated that the following energy
substrates can also act as impermeants. Suitable energy substrate
can be selected from one or more from the group consisting of:
glucose and other sugars, pyruvate, lactate, glutamate, glutamine,
aspartate, arginine, ectoine, taurine, N-acetyl-beta-lysine,
alanine, proline, beta-hydroxy butyrate and other amino acids and
amino acid derivatives, trehalose, floridoside, glycerol and other
polyhydric alcohols (polyols), sorbitol, myo-innositol, pinitol,
insulin, alpha-keto glutarate, malate, succinate, triglycerides and
derivatives, fatty acids and carnitine and derivatives. In one
embodiment, the at least one compound for minimizing or reducing
the uptake of water by the cells in the tissue is an energy
substrate. The energy substrate helps with recovering metabolism.
The energy substrate can be selected from one or more from the
group consisting of: glucose and other sugars, pyruvate, lactate,
glutamate, glutamine, aspartate, arginine, ectoine, taurine,
N-acetyl-beta-lysine, alanine, proline and other amino acids and
amino acid derivatives, trehalose, floridoside, glycerol and other
polyhydric alcohols (polyols), sorbitol, myo-innositol, pinitol,
insulin, alpha-keto glutarate, malate, succinate, triglycerides and
derivatives, fatty acids and carnitine and derivatives. Given that
energy substrates are sources of reducing equivalents for energy
transformations and the production of ATP in a cell, tissue or
organ of the body, it will be appreciated that a direct supply of
the energy reducing equivalents could be used as substrates for
energy production. For example, a supply of either one or more or
different ratios of reduced and oxidized forms of nicotinamide
adenine dinucleotide (e.g. NAD or NADP and NADH or NADPH) or flavin
adenine dinucleotides (FADH or FAD) could be directly used to
supply bond energy for sustaining ATP production in times of
stress. Preferably, beta-hydroxy butyrate is added to the
composition of the invention for treatment of trauma or reducing
injury.
[0084] In addition to providing energy substrates to the whole
body, organ, tissue or cell, improvements in metabolising these
substrates may occur in the presence of hydrogen sulphide
(H.sub.2S) or H.sub.2S donors (eg NaHS). The presence of hydrogen
sulphide (H.sub.2S) or H2S donors (eg NaHS) may help metabolise
these energy substrates by lowering energy demand during arrest,
protect and preserve the whole body, organ, tissue or cell during
periods of metabolic imbalance such ischemia, reperfusion and
trauma. Concentrations of Hydrogen sulfide above 1 microM (10-6 M)
concentration can be a metabolic poison that inhibits respiration
at Respiratory Complex IV, which is part of the mitochondrial
respiratory chain that couples metabolising the high energy
reducing equivalents from energy substrates to energy (ATP)
generation and oxygen consumption. However, it has been observed at
lower concentrations, below 10-.sup.6 M (eg 10-.sup.10 to
10-.sup.9M), hydrogen sulfide may reduce the energy demand of the
whole body, organ, tissue or cell which may result in arrest,
protection and preservation. In other words, very low levels of
sulfide down-regulate mitochondria, reduce O.sub.2 consumption and
actually increase "Respiratory Control" whereby mitochondria
consume less O.sub.2 without collapsing the electrochemical
gradient across the inner mitochondrial membrane. Thus there are
observations that a small amount of sulfide, either directly or
indirectly, may close proton leak channels and better couple
mitochondrial respiration to ATP production more tightly, and this
effect may improve the metabolism of high energy reducing
equivalents from energy substrates. There is also the possibility
that a sulphur cycle exists between the cell cytosol and
mitochondria in mammals, including humans, providing the sulphur
concentration is low. The presence of a vestige sulphur cycle would
be consistent with current ideas on the evolutionary origin of
mitochondria and their appearance in eukaryote cells from a
symbiosis between a sulfide-producing host cell and a
sulfide-oxidizing bacterial symbiont. Thus, hydrogen sulphide
(H.sub.2S) or H.sub.2S donors (eg NaHS) may be energy substrates
themselves in addition to improving the metabolism of other energy
substrates. Accordingly, in one form, the invention provides a
composition as described above further including hydrogen sulphide
or a hydrogen sulfide donor.
[0085] In one embodiment, the at least one compound for minimizing
or reducing the uptake of water by the cells in the tissue is
sucrose. Sucrose reduces water shifts as an impermeant. Impermeant
agents such as sucrose, lactobionate and raffinose are too large to
enter the cells and hence remain in the extracellular spaces within
the tissue and resulting osmotic forces prevent cell swelling that
would otherwise damage the tissue, which would occur particularly
during storage of the tissue.
[0086] In another embodiment, the at least one compound for
minimizing or reducing the uptake of water by the cells in the
tissue is a colloid. Suitable colloids include, but not limited to,
Dextran-70, 40, 50 and 60, hydroxyethyl starch and a modified fluid
gelatin. A colloid is a composition which has a continuous liquid
phase in which a solid is suspended in a liquid. Colloids can be
used clinically to help restore balance to water and ionic
distribution between the intracellular, extracellular and blood
compartments in the body after an severe injury. Colloids can also
be used in solutions for organ preservation. Administration of
crystalloids can also restore water and ionic balance to the body
but generally require greater volumes of administration because
they do not have solids suspended in a liquid. Thus volume
expanders may be colloid-based or crystalloid-based
[0087] Preferably, the concentration of the compound for minimizing
or reducing the uptake of water by the cells in the tissue is
between about 5 to 500 mM. Typically this is an effective amount
for reducing the uptake of water by the cells in the tissue. More
preferably, the concentration of the compound for reducing the
uptake of water by the cells in the tissue is between about 20 and
100 mM. Even more preferably the concentration of the compound for
reducing the uptake of water by the cells in the tissue is about 70
mM.
[0088] In a further embodiment, the composition according to the
invention may include more than one compound for minimizing or
reducing the uptake of water by the cells in the tissue. For
example, a combination of impermeants (raffinose, sucrose and
pentastarch) may be included in the composition or even a
combination of colloids, and fuel substrates may be included in the
composition.
[0089] The composition according to the invention may be hypo, iso
or hyper osmotic.
[0090] The inventor has also found that the inclusion of a compound
for inhibiting transport of sodium and hydrogen ions across a
plasma membrane of a cell in the tissue with a potassium channel
opener or adenosine receptor agonist and a local anaesthetic
assists in reducing injury.
[0091] Thus in another aspect, the composition according to the
invention further includes a compound for inhibiting transport of
sodium and hydrogen ions across a plasma membrane of a cell in the
tissue.
[0092] The compound for inhibiting transport of sodium and hydrogen
across the membrane of the cell in the tissue is also referred to
as a sodium hydrogen exchange inhibitor. The sodium hydrogen
exchange inhibitor reduces sodium and calcium entering the
cell.
[0093] Preferably the compound for inhibiting transport of sodium
and hydrogen across the membrane of the cell in the tissue may be
selected from one or more of the group consisting of Amiloride,
EIPA(5-(N-entyl-N-isopropyl)-amiloride), cariporide (HOE-642),
eniporide, Triamterene (2,4,7-triamino-6-phenylteride), EMD 84021,
EMD 94309, EMD 96785, EMD 85131, HOE 694. B11 B-513 and T-162559
are other inhibitors of the isoform 1 of the Na+/H+ exchanger.
[0094] Preferably, the sodium hydrogen exchange inhibitor is
Amiloride (N-amidino-3,5-diamino-6-chloropyrzine-2-carboximide
hydrochloride dihydrate). Amiloride inhibits the sodium proton
exchanger (Na+/H+ exchanger also often abbreviated NHE-1) and
reduces calcium entering the cell. During ischemia excess cell
protons (or hydrogen ions) are believed to be exchanged for sodium
via the Na+/H+ exchanger.
[0095] Preferably, the concentration of the compound for inhibiting
transport of sodium and hydrogen across the membrane of the cell in
the tissue is between about 1.0 nM to 1.0 mM. More preferably, the
concentration of the compound for inhibiting transport of sodium
and hydrogen across the membrane of the cell in the tissue is about
20 uM.
[0096] The inventor has also found that the inclusion of
antioxidant with a potassium channel opener or adenosine receptor
agonist and a local anaesthetic. Thus in another aspect, the
composition of the present invention may further include an
antioxidant.
[0097] Antioxidants are commonly enzymes or other organic
substances that are capable of counteracting the damaging effects
of oxidation in the tissue. The antioxidant component of the
composition according to the present invention may be selected from
one or more of the group consisting of: allopurinol, carnosine,
histidine, Coenzyme Q 10, n-acetyl-cysteine, superoxide dismutase
(SOD), glutathione reductase (GR), glutathione peroxidase (GP)
modulators and regulators, catalase and the other metalloenzymes,
NADPH and AND(P)H oxidase inhibitors, glutathione, U-74006F,
vitamin E, Trolox (soluble form of vitamin E), other tocopherols
(gamma and alpha, beta, delta), tocotrienols, ascorbic acid,
Vitamin C, Beta-Carotene (plant form of vitamin A), selenium, Gamma
Linoleic Acid (GLA), alpha-lipoic acid, uric acid (urate),
curcumin, bilirubin, proanthocyanidins, epigallocatechin gallate,
Lutein, lycopene, bioflavonoids, polyphenols, trolox(R),
dimethylthiourea, tempol(R), carotenoids, coenzyme Q, melatonin,
flavonoids, polyphenols, aminoindoles, probucol and nitecapone,
21-aminosteroids or lazaroids, sulphydryl-containing compounds
(thiazolidine, Ebselen, dithiolethiones), and N-acetylcysteine.
Other antioxidants include the ACE inhibitors (captopril,
enalapril, lisinopril) which are used for the treatment of arterial
hypertension and cardiac failure on patients with myocardial
infarction. ACE inhibitors exert their beneficial effects on the
reoxygenated myocardium by scavenging reactive oxygen species.
Other antioxidants that could also be used include
beta-mercaptopropionylglycine, O-phenanthroline, dithiocarbamate,
selegilize and desferrioxamine (Desferal), an iron chelator, has
been used in experimental infarction models, where it exerted some
level of antioxidant protection. Spin trapping agents such as
5'-5-dimethyl-1-pyrrolione-N-oxide (DMPO) and
(a-4-pyridyl-1-oxide)-N-t-butylnitrone (POBN) also act as
antioxidants. Other antioxidants include: nitrone radical scavenger
alpha-phenyl-tert-N-butyl nitrone (PBN) and derivatives PBN
(including disulphur derivatives); N-2-mercaptopropionyl glycine
(MPG) a specific scavenger of the OH free radical; lipooxygenase
inhibitor nordihydroguaretic acid (NDGA); Alpha Lipoic Acid;
Chondroitin Sulfate; L-Cysteine; oxypurinol and Zinc.
[0098] Preferably, the antioxidant is allopurinol
(1H-Pyrazolo[3,4-a]pyrimidine-4-ol). Allopurinol is a competitive
inhibitor of the reactive oxygen species generating enzyme xanthine
oxidase. Allopurinol's antioxidative properties may help preserve
myocardial and endothelial functions by reducing oxidative stress,
mitochondrial damage, apoptosis and cell death. Preferably, the
concentration of the antioxidant is between about 1 nM to 100
uM.
[0099] The inventor has also found that the inclusion of particular
amounts of calcium and magnesium ions with a potassium channel
opener or adenosine receptor agonist and a local anaesthetic
reduces injury. The effect of the particular amounts of calcium and
magnesium ions is to control the amount of ions within the
intracellular environment. Calcium ions tend to be depleted,
exported or otherwise removed from the intracellular environment
and magnesium ions tend to be increased or otherwise restored to
the levels typically found in a viable, functioning cell.
[0100] Thus in another aspect, the composition according to the
invention further includes a source of magnesium in an amount for
increasing the amount of magnesium in a cell in body tissue.
Preferably the magnesium is present at a concentration of between
0.5 mM to 20 mM, more preferably about 2.5 mM. It will be
appreciated that these concentrations refer to the effective
concentration of the magnesium in the composition that contacts the
tissue, organ or cell.
[0101] In addition, typical buffers or carriers (which are
discussed in more detail below) in which the composition of the
invention is administered typically contain calcium at
concentrations of around 1 mM as the total absence of calcium has
been found to be detrimental to the cell, tissue or organ. In one
form, the invention also includes using carriers with low calcium
(such as for example less than 0.5 mM) so as to decrease the amount
of calcium within a cell in body tissue, which may otherwise build
up during injury/trauma/stunning. As described in the present
invention, elevated magnesium and low calcium has been associated
with protection during ischemia and reoxygenation of an organ. The
action is believed to be due to decreased calcium loading.
Preferably the calcium present is at a concentration of between 0.1
mM to 0.8 mM, more preferably about 0.3 mM.
[0102] In one embodiment, the composition includes elevated
divalent magnesium ions. Magnesium sulphate and magnesium chloride
is a suitable source.
[0103] In the case of a human subject requiring treatment, the
following alternative compositions with corresponding
concentrations of Adenosine(Ado), Lignocaine (Lido) and magnesium
sulphate are provided, without limitation:
TABLE-US-00001 Ado Lido MgSO4 7 H2O I 2.25 mM 1.844 mM 243.4 mM II
3.74 mM 3.688 mM 243.4 mM III 3.74 mM 7.376 mM 243.4 mM IV 5.61 mM
3.688 mM 243.4 mM V 5.61 mM 7.376 mM 243.4 mM VI 22.5 mM 18.44 mM
243.4 mM VII 37.4 mM 36.88 mM 243.4 mM VIII 37.4 mM 73.76 mM 243.4
mM IX 56.1 mM 36.88 mM 243.4 mM X 56.1 mM 73.76 mM 243.4 mM
[0104] The concentrations of each respective active ingredient in
these compositions refer to the concentrations in the composition
before administration. It will be appreciated that the
concentrations may be diluted by body fluids or other fluids that
may be administered together with the composition. Typically, the
composition will be administered such that the concentration of
these ingredients at the tissue is about 100-fold less than the
concentrations in the table above. For example, containers (such as
vials) of such a composition may be diluted 1 to a 100 parts of
blood, plasma, crystalloid or blood substitute for
administration.
[0105] In one embodiment, the composition according to the
invention includes Adenosine and Lignocaine. Typically, the
concentration of Adenosine and Lidocaine in the composition is
between about 1 mM to 100 mM. The final concentration of these
components once administered may be between about 0.1 mM to 10
mM.
[0106] In another embodiment, the composition includes a cellular
transport enzyme inhibitor, such as dipyridamole, to prevent
metabolism or breakdown of components in the composition.
[0107] In a further aspect, the invention provides a composition
including a local anaesthetic and one or more of: [0108] potassium
channel opener; [0109] adenosine agonist; [0110] opioid; [0111] at
least one compound for reducing uptake of water; [0112] sodium
hydrogen exchange inhibitor; [0113] antioxidant; and [0114] a
source of magnesium in an amount for increasing the amount of
magnesium in a cell in body tissue.
[0115] Preferably, this composition has two, three or four of the
above. Preferred compounds for these components are listed
above.
[0116] In another embodiment, the invention provides a composition
including a potassium channel opener and/or an adenosine agonist
and one or more of: [0117] local anaesthetic; [0118] opioid; [0119]
at least one compound for reducing uptake of water; [0120] sodium
hydrogen exchange inhibitor; [0121] antioxidant; and [0122] a
source of magnesium in an amount for increasing the amount of
magnesium in a cell in body tissue.
[0123] Preferably, this composition has two, three or four of the
above. Preferred compounds for these components are listed
above.
[0124] The processes of inflammation and thrombosis are linked
through common mechanisms. Therefore, it is believed that
understanding of the processes of inflammation will help with
better management of thrombotic disorders including the treatment
of acute and chronic ischaemic syndromes. In the clinical and
surgical settings, a rapid response and early intervention to an
organ or tissue damaged from ischemia can involve both
anti-inflammatory and anti-clotting therapies. In addition to
protease inhibitors which attenuate the inflammatory response,
further anti-inflammatory therapies have included the
administration of aspirin, normal heparin, low-molecular-weight
heparin (LMWH), non-steroidal anti-inflammatory agents,
anti-platelet drugs and glycoprotein (GP) IIb/IIIa receptor
inhibitors, statins, angiotensin converting enzyme (ACE) inhibitor,
angiotensin blockers and antagonists of substance P. Examples of
protease inhibitors are indinavir, nelfinavir, ritonavir,
lopinavir, amprenavir or the broad-spectrum protease inhibitor
aprotinin, a low-molecular-weight heparin (LMWH) is enoxaparin,
non-steroidal anti-inflammatory agent are indomethacin, ibuprofen,
rofecoxib, naproxen or fluoxetine, an anti-platelet drug is
Clopidogrel or aspirin, a glycoprotein (GP) IIb/IIIa receptor
inhibitor is abciximab, a statin is pravastatin, an angiotensin
converting enzyme (ACE) inhibitor is captopril and an angiotensin
blocker is valsartin.
[0125] Accordingly, in another embodiment of the invention, a
selection of these agents is added to a composition according to
the invention to deliver improved management of inflammation and
clotting. Alternatively, the composition according to the invention
may be administered together with any one or more of these
agents.
[0126] In particular, protease inhibitors attenuate the systemic
inflammatory response in patients undergoing cardiac surgery with
cardiopulmonary bypass, and other patients where the inflammatory
response has been heightened such as AIDS or in the treatment of
chronic tendon injuries. Some broad spectrum protease inhibitors
such as aprotinin are also reduce blood loss and need for blood
transfusions in surgical operations such as coronary bypass.
[0127] Compounds that substantially prevent the breakdown of
adenosine in the blood such as nucleoside transport inhibitors,
such as dipyridamole could be are used as additives in the
composition of the invention. The half life of adenosine in the
blood is about 10 seconds so the presence of a medicament to
substantially prevent its breakdown will maximise the effect of the
composition of the present invention.
[0128] Optionally, the composition according to the invention may
also include Dipyridamole is advantageously included in a
concentration from about 0.01 microM to about 10 mM, preferably
0.05 to 100 microM., Dipyridamole and has major advantages with
respect to cardioprotection. Dipyridamole may supplement the
actions of adenosine by inhibiting adenosine transport and
breakdown leading to increased protection of cells, tissues and
organs of the body during times of stress. Dipyridamole may also be
administered separately for example by 400 mg daily tablets to
produce a plasma level of about 0.4 microgram/ml, or 0.8 microM
concentration.
[0129] The composition according to the present invention is highly
beneficial at about 10.degree. C. but can also be used to prevent
injury over a wider temperature range up to about 37.degree. C. The
composition according to the invention may be used at a temperature
range selected from the following: 0.degree. C. to 5.degree. C.,
5.degree. C. to 20.degree. C., 20.degree. C. to 32.degree. C. and
32.degree. C. to 38.degree. C.
[0130] The composition may be administered intravenously or be
administered both intravenously and intraperitoneally or in special
circumstances directly accessing a major artery such as the femoral
artery or aorta in patients who have no pulse from massive
exsanguination. In one embodiment, the composition of the invention
may be administered intravenously and intraperineally
simultaneously, the perineum acting as, in effect, a reservoir of
composition for the bloodstream as well as acting on organs in the
vicinity with which it comes into contact. This is particularly
suitable for a trauma victim, such as one suffering shock.
[0131] As described herein, in particular embodiments of the
invention, the composition of the present invention protects and
preserves tissue of a body after trauma, such as heart attacks,
strokes etc, with good to excellent recoveries of function or
viability of body tissue after reperfusion.
[0132] Affecting viability of a tissue during preservation and
recovery of the body tissue, such that affected tissue remains
viable or living during those processes and is capable of returning
to its function, particularly after the tissue has been subject to
shock, is crucial.
[0133] Preferably, reducing injury to a body relates to maintaining
affected tissue in a viable state, such that the tissue is capable
of returning to its function, after trauma. Maintaining or
stabilising the tissue in a viable state includes maintaining the
membrane potential of tissue cells at or around resting level, so
as to reduce sodium or calcium loading of the cell which is a cause
of injury during ischaemia and reperfusion. Preservation is known
as the act or process of preserving the tissue or keeping from
injury, destruction or decay. In this application, the composition
according to the invention acts to minimise any potential injury,
destruction or decay of the tissue of a body which may be caused by
trauma.
[0134] Injury can be broadly characterised as reversible and
irreversible cell injury. For example, reversible cell injury can
lead to heart dysfunction usually from arrhythmias and/or stunning.
Stunning is normally characterised as loss of left pump function
during restoration of blood flow following periods of ischemia. If
severe, it can lead to the death of the heart, usually from
arrhythmias, even though the heart cells themselves are not
initially dead. Irreversible injury by definition arises from
actual cell death which may be fatal depending upon the extent of
the injury. The amount of cell death can be measured as infarct
size. During recovery from cardioplegic arrest, if the conditions
are adequate, the heart can be restored substantially to normal
function of the tissue by reperfusion, with minimal infarct size.
The most common ways to assess return of function of a heart are by
measuring pressures that the heart can generate:
heart pump flow; and the electrical activity of the heart.
[0135] This data is then compared to data measured from pre-arrest
conditions.
[0136] The composition of the present invention is particularly
useful in reducing injury to heart tissue during heart surgery
(open-heart or robotic heart surgery), including heart transplants,
and neonate/infant hearts. Other applications include reducing
heart damage before, during or following cardiovascular
intervention which may include a heart attack, angioplasty or
angiography. For example, the composition may be administered to
subjects who have suffered or are developing a heart attack and
used at the time of administration of blood clot-busting drugs such
as streptokinase. As the clot is dissolved, the presence of the
composition may protect the heart from further injury such as
reperfusion injury. The composition may be particularly effective
as a cardioprotectant in those portions of the heart that have been
starved of normal flow, nutrients and/or oxygen for different
periods of time. For example, the pharmaceutical composition may
also be used to treat heart ischaemia which could be pre-existing
or induced by cardiovascular intervention. Other applications
include assisting in diagnostic procedures such as assessment of a
subject's health while exercising on a treadmill or, if subjects
cannot exercise on a treadmill, to assist in visualising areas of
the body such as the heart that may have partially or fully blocked
blood vessels, or damaged heart cells. In addition, the invention
may be used during different visualization procedures such as X-ray
(routine and computerized tomography) or magnetic resonance imaging
(MRI) of a subject's body or organs and tissues within the body or
isolated from the body. In addition to providing better
visualisation of potential areas of injury or damage, the invention
may be used to temporarily lower the heart rate of a subject and
thereby reduce movement (ie. from increasing heart relaxation) and
permit faster scan times during the diagnostic assessment of
potential injury in a blood vessel, tissue or organ of the body,
particularly in the heart. Lowering heart rate and permitting
faster scan times may also lower the doses of radiation required to
visualize the potential areas of injury or damage.
[0137] Accordingly, in another embodiment of the invention, there
is provided a method of preserving a vessel, tissue or organ of the
body, such as a heart, comprising administering a composition as
described above before, during or after medical intervention
affecting the vessel, tissue or organ of the body, such as a heart.
The composition used in this embodiment of the invention may have
an arresting or a non-arresting concentration of active components
in it. In one form, the method includes administering a
non-arresting concentration of the composition and, in another
form, it has an arresting concentration of the composition
(preferably as a bolus) followed by a non-arresting concentration
of the composition.
[0138] In another embodiment, the present invention may be
administered with or contain blood or blood products or artificial
blood or oxygen binding molecules or solutions to improve the
body's oxygen transport ability and survival by helping to reduce
hypoxic and ischemic damage from blood loss. The oxygen-containing
molecules, compounds or solutions may be selected from natural or
artificial products. For example, an artificial blood-based product
is perfluorocarbon-based or other haemoglobin-based substitute.
Some of the components may be added to mimic human blood's oxygen
transport ability such Hemopure.TM., Gelenpol.TM., Oxygent.TM., and
PolyHeme.TM.. Hemopore is based on a chemically stabilized bovine
hemoglobin. Gelenpol is a polymerized hemoglobin which comprises
synthetic water-soluble polymers and modified heme proteins.
Oxygent is a perflubron emulsion for use as an intravenous oxygen
carrier to temporarily substitute for red blood cells during
surgery. Polyheme is a human hemoglobin-based solution for the
treatment of life-threatening blood loss.
[0139] It is believed that the oxygenation of the body from a
variety of ways including but not limited to oxygen gas mixture,
blood, blood products or artificial blood or oxygen binding
solutions maintains mitochondrial oxidation and this helps preserve
the myocyte and endothelium of the organ. Without being bound by
any particular mode or theory, the inventor has found that gentle
bubbling with 95% 0.sub.2/5% CO.sub.2 helps maintains mitochondrial
oxidation which helps preserve the myocyte and coronary
vasculature.
[0140] In one preferred embodiment of this aspect of the present
invention with respect to whole body or organs outside the body,
the composition is aerated with a source of oxygen before and/or
during use. The source of oxygen may be an oxygen gas mixture where
oxygen is the predominant component. The oxygen may be mixed with,
for example, CO.sub.2. Preferably, the oxygen gas mixture is 95%
O.sub.2 and 5% CO.sub.2.
[0141] In another aspect of the present invention there is provided
a method for reducing injury including: [0142] providing in a
suitable container a composition according to the invention; [0143]
providing one or more nutrient molecules selected from the group
consisting of blood, blood products, artificial blood and a source
of oxygen; [0144] optionally aerating the composition with the
oxygen (for example, in the case of isolated organs) or combining
the nutrient molecules with the composition, or both; and [0145]
placing the tissue in contact with the combined composition under
conditions sufficient to reduce injury.
[0146] Preferably the oxygen source is an oxygen gas mixture.
Preferably oxygen is the predominant component. The oxygen may be
mixed with, for example CO.sub.2. More preferably, the oxygen gas
mixture is 95% O.sub.2 and 5% CO.sub.2. Preferably the composition
is aerated before and/or during contact with the tissue.
[0147] The composition according to this aspect of the invention
may be in liquid form. Liquid preparations of the pharmaceutical
composition may take the form of, for example, solutions, syrups,
or suspensions, or may be presented as a dry product for
constitution with water or other suitable vehicle. Such liquid
preparations may be prepared by conventional means with
pharmaceutically acceptable additives such as suspending agents,
emulsifying agents, non-aqueous vehicles, preservatives and energy
sources.
[0148] In another form, the invention comprises a composition in
tablet form and in another form, the invention comprises an aerosol
which could be administered via oral, skin or nasal routes.
[0149] In another aspect of the invention, there is provided a
method of protecting heart tissue from reperfusion injury,
including inflammatory and blood clotting and coagulation effects
often experienced during reperfusion following an ischaemic event,
such as in the post-operative period or longer-term recovery. The
method comprises administering a solution comprising a
non-arresting form of the composition according to the present
invention, optionally following a bolus of an arresting form.
[0150] The invention also provides a method for reducing infarction
size and/or reducing inflammation and blood coagulation responses
in heart tissue during ischaemia and/or reperfusion comprising
administration of the same solution.
[0151] The body may be a human or an animal such as a livestock
animal (eg, sheep, cow or horse), laboratory test animal (eg,
mouse, rabbit or guinea pig) or a companion animal (eg, dog or
cat), particularly an animal of economic importance. Preferably,
the body is human.
[0152] The invention also provides a method for managing pain,
including neuropathic pain, including administering an effective
amount of a composition according to the invention described
above.
[0153] The present invention is particularly advantageous in
reducing injury in the body, for example in the treatment of the
heart in circumstances of myocardial infarction or heart attack, or
during surgical procedures, for example during open-heart
surgery.
[0154] The method of the present invention involves contacting a
tissue with the composition according to the invention, for a time
and under conditions sufficient for the tissue to be
preconditioned, arrested, protected and/or preserved. The
composition may be infused or administered as a bolus intravenous,
intracoronary or any other suitable delivery route as pre-treatment
for protection during a cardiac intervention such as open heart
surgery (on-pump and off-pump), angioplasty (balloon and with
stents or other vessel devices) and as with clot-busters
(anti-clotting drug or agents).
[0155] The composition may be administered intravenously or be
administered both intravenously and intraperitoneally or in special
circumstances directly accessing a major artery such as the femoral
artery or aorta in patients who have no pulse from massive
exsanguination, or in the carotid artery or another artery during
aortic dissection to protect the brain from hypoxia or ischemia. In
one embodiment, the composition of the invention may be
administered intravenously and intraperineally simultaneously, the
perineum acting as, in effect, a reservoir of composition for the
bloodstream as well as acting on organs in the vicinity with which
it comes into contact. This is particularly suitable for a trauma
victim, such as one suffering shock. Moreover, where the
composition contains two or more components, these may be
administered separately but simultaneously. Substantially
simultaneous delivery of the component to the target site is
desirable. This may be achieved by pre-mixing the components for
administration as one composition, but that is not essential. The
invention is directed towards the simultaneous increase in local
concentration (for example an organ such as the heart) of the
components of a composition according to the invention (for
example, where a first component is (i) a potassium channel opener
or agonist and/or an adenosine receptor agonist; and (ii) a local
anaesthetic). One preferred form of the composition is a
combination of adenosine and lignocaine.
[0156] The invention may be practised by administering the compound
using a perfusion pump, often associated with a procedure known as
"miniplegia" or "microplegia", in which minimal amount of actives
are titrated by means of a finely adjustable pump directly via a
catheter. In the invention, a protocol utilises miniplegia as
described above, where micro amounts are titrated directly to the
heart, using the patient's own oxygenated blood. The reference to a
"setting" is a measure on the pump, such as a syringe pump, of the
amount of substance being delivered directly to the organ, such as
a heart.
[0157] The composition can also be infused or administered as a
bolus intravenous, intracoronary or any other suitable delivery
route for protection during cardiac intervention such as open heart
surgery (on-pump and off-pump), angioplasty (balloon and with
stents or other vessel devices) and as with clot-busters to protect
and preserve the cells from injury.
[0158] The composition may also be infused or administered as a
bolus intravenous, intracoronary or any other suitable delivery
route for protection following a cardiac intervention such as open
heart surgery (on-pump and off-pump), angioplasty (balloon and with
stents or other vessel devices) and as with clot-busters to protect
and preserve the cells from injury.
[0159] Accordingly, the tissue may be contacted by delivering the
composition according to the invention intravenously to the tissue.
This involves using blood as a vehicle for delivery to the tissue.
In particular, the composition according to the invention may be
used for blood cardioplegia. Alternatively, the composition may be
administered directly as a bolus by a puncture (eg, by syringe)
directly to the tissue or organ, particularly useful when blood
flow to a tissue or organ is limiting. The composition for
arresting, protecting and preserving a tissue may also be
administered as an aerosol, powder, solution or paste via oral,
skin or nasal routes.
[0160] Alternatively, the composition may be administered directly
to the tissue, organ or cell or to exposed parts of the internal
body to reduce injury. In particular, the composition according to
the invention may be used for crystalloid cardioplegia.
[0161] The composition according to the invention may be delivered
according to one of or a combination of the following delivery
protocols: intermittent, continuous and one-shot.
[0162] Accordingly, in another aspect of the invention, there is
provided a composition for arresting, protecting and preserving a
tissue of a body upon administration of a single dose of the
composition, the composition including a primary potassium channel
opener or agonist and/or adenosine receptor agonist and a local
anaesthetic. The invention also provides a method for arresting and
protecting an tissue comprising administering as a single dose an
effective amount of that composition.
[0163] In another aspect of the invention, there is provided a
composition for arresting, protecting and preserving a tissue by
intermittent administration of the composition, the composition
including an effective amount of a primary potassium channel opener
or agonist and/or adenosine receptor agonist and a local
anaesthetic. A suitable administration schedule is a 2 minute
induction dose every 20 minutes throughout the arrest period. The
actual time periods can be adjusted based on observations by one
skilled in the art administering the composition, and the
animal/human model selected. The invention also provides a method
for intermittently administering a composition for arresting,
protecting and preserving a tissue.
[0164] The composition can of course also be used in continuous
infusion with both normal and injured tissues or organs, such as
heart tissue. Continuous infusion also includes static storage of
the tissue, whereby the tissue is stored in a composition according
to the invention, for example the tissue may be placed in a
suitable container and immersed in a solution according to the
invention for transporting donor tissues from a donor to
recipient.
[0165] The dose and time intervals for each delivery protocol may
be designed accordingly. For example, a composition according to
the invention may be delivered as a one-shot to the tissue to
initially arrest of the tissue. A further composition according to
the invention may then be administered continuously to maintain the
tissue in an arrested state. Yet a further composition according to
the invention may be administered continuously to reperfuse the
tissue or recover normal function.
[0166] As mentioned previously, the composition according to the
invention may be used or contact the tissue at a temperature range
selected from one of the following: from about 0.degree. C. to
about 5.degree. C., from about 5.degree. C. to about 20.degree. C.,
from about 20.degree. C. to about 32.degree. C. and from about
32.degree. C. to about 38.degree. C. It is understood that
"profound hypothermia" is used to describe a tissue at a
temperature from about 0.degree. C. to about 5.degree. C. "Moderate
hypothermia" is used to describe a tissue at a temperature from
about 5.degree. C. to about 2.degree. C. "Mild hypothermia" is used
to describe a tissue at a temperature from about 20.degree. C. to
about 32.degree. C. "Normothermia" is used to describe a tissue at
a temperature from about 32.degree. C. to about 38.degree. C.,
though the normal body temperature is around 37 to 38.degree.
C.
[0167] While it is possible for each component of the composition
to contact the tissue alone, it is preferable that the components
of the pharmaceutical composition be provided together with one or
more pharmaceutically acceptable carriers, diluents, adjuvants
and/or excipients. Each carrier, diluent, adjuvant and/or excipient
must be pharmaceutically acceptable such that they are compatible
with the components of the pharmaceutical composition and not
harmful to the subject. Preferably, the pharmaceutical composition
is prepared with liquid carriers, diluents, adjuvants and/or
excipients.
[0168] The composition according to the invention may be suitable
for administration to the tissue in liquid form, for example,
solutions, syrups or suspensions, or alternatively they may be
administered as a dry product for constitution with water or other
suitable vehicle before use. Such liquid preparations may be
prepared by conventional means.
[0169] The composition according to the invention may be suitable
for topical administration to the tissue. Such preparation may be
prepared by conventional means in the form of a cream, ointment,
jelly, solution or suspension.
[0170] The composition may also be formulated as depot
preparations. Such long acting formulations may be administered by
implantation (eg, subcutaneously or intramuscularly) or by
intramuscular injection. Thus, for example, the composition
according to the invention may be formulated with suitable
polymeric or hydrophobic materials (eg, as an emulsion in an
acceptable oil or ion exchange resins, or as sparingly soluble
derivatives, for example, as a sparingly soluble salt.
[0171] Accordingly, this aspect of the invention also provides a
method for reducing injury, which includes providing the
composition together with a pharmaceutically acceptable carrier,
diluent, adjuvant and/or excipient. A preferred pharmaceutically
acceptable carrier is a buffer having a pH of about 6 to about 9,
preferably about 7, more preferably about 7.4 and/or low
concentrations of potassium. For example, the composition has a
total potassium concentration of up to about 10 mM, more preferably
about 2 to about 8 mM, most preferably about 4 to about 6 mM.
Suitable buffers include Krebs-Henseleit which generally contains
10 mM glucose, 117 mM NaCl, 5.9 mM KCl, 25 mM NaHCO.sub.3, 1.2 mM
NaH.sub.2PO.sub.4, 1.12 mMCaCl.sub.2 (free Ca.sup.2+=1.07 mM) and
0.512 mM MgCl.sub.2 (free Mg.sup.2+=0.5 mM), Tyrodes solution which
generally contains 10 mM glucose, 126 mM NaCl, 5.4 mM KCl, 1 mM
CaCl.sub.2, 1 mM MgCl.sub.2, 0.33 mM NaH.sub.2PO.sub.4 and 10 mM
HEPES (N-[2-hydroxyethyl]piperazine-N'-[2-ethane sulphonic acid],
Fremes solution, Hartmanns solution which generally contains 129
NaCl, 5 mM KCl, 2 mM CaCl.sub.2 and 29 mM lactate and
Ringers-Lactate. Other naturally occurring buffering compounds that
exist in muscle that could be also used in a suitable ionic
environment are carnosine, histidine, anserine, ophidine and
balenene, or their derivatives. One advantage of using low
potassium is that it renders the present composition less injurious
to the subject, in particular paediatric subjects such as
neonates/infants. High potassium has been linked to an accumulation
of calcium which may be associated with irregular heart beats
during recovery, heart damage and cell swelling. Neonates/infants
are even more susceptible than adults to high potassium damage
during cardiac arrest. After surgery a neonate/infant's heart may
not return to normal for many days, sometimes requiring intensive
therapy or life support.
[0172] It is also advantageous to use carriers having low
concentrations of magnesium, such as, for example up to about 2.5
mM, but it will be appreciated that high concentrations of
magnesium, for example up to about 20 mM, may be used if desired
without substantially affecting the activity of the
composition.
[0173] In another embodiment of the present invention there is
provided use of a composition according to the present invention
for reducing injury.
[0174] Preferably the composition is aerated before and/or during
administration or contact with the tissue.
IN THE FIGURES
[0175] FIG. 1 shows ECG trace of rat heart (A) prior to hemorrhagic
shock (B) during shock and (C) after bolus administration of 0.5 ml
Adenosine/Lignocaine solution directly into the heart of the
rat.
[0176] FIG. 2 shows in more detail the ECG trace of the rat heart
from FIG. 1 (A) during hemorrhagic shock and after injection of
Adenosine/Lignocaine solution directly into the heart of the rat
and (B) 10 seconds following injection. The time of injection of
the solution is indicated by the arrow (I). Arrow (II) denotes the
proposed time at which further treatment may be required.
[0177] FIG. 3 shows an ECG trace of normal rat heart prior to
commencement of hemorrhagic shock.
[0178] FIG. 4 shows ECG trace of rat at end of bleed period prior
to commencement of "shock period"
[0179] FIG. 5 shows ECG trace of rat heart at the end of first 60
mins shock period
[0180] FIG. 6 shows ECG trace of rat heart at the end of 120 mins
shock period
[0181] FIG. 7 shows an ECG trace of rat heart at the end of 3 hour
shock period
[0182] FIG. 8 shows an ECG trace of rat heart 10 mins after bolus
administration of ALM (Adenosine;Lignocaine;Magnesium)
[0183] FIG. 9 shows an ECG trace of rat heart 30 mins after bolus
administration of ALM
[0184] FIG. 10 shows an ECG trace of rat heart 60 mins after bolus
administration of ALM
[0185] FIG. 11 shows an ECG trace of rat heart 90 mins after bolus
administration of ALM
[0186] FIG. 12 shows ECG trace of rat heart (A) prior to
Hemorrhagic shock (45% blood loss); (B) 60 min following
hemorrhagic shock and intravenous administration of
Adenosine/Lignocaine resuscitation fluid (C) 180 mins following
hemorrhagic shock and intravenous administration of
Adenosine/Lignocaine resuscitation fluid.
[0187] FIG. 13 shows in more detail the ECG monitoring of the rat
heart from FIG. 12 following hemorrhagic shock (A) after
administration of 0.5 ml 7.5% saline and (B) after administration
of 0.5 ml Adenosine/Lignocaine resuscitation fluid.
EXAMPLES
[0188] The following are provided as non-limiting examples of
suitable compositions of the invention for the purpose of
illustrating the invention.
Animals and Reagents:
[0189] Male Sprague Dawley rats (300-350 g) from the James Cook
University Breeding Colony are fed ad libitum and housed in a
12-hour light/dark cycle. On the day of the experiment rats are
anesthetized with an intraperitoneal injection of Nembutal (Sodium
Thiopentone (Thiobarb); 100 mg/kg) and the anaesthetic administered
as required throughout the protocol. Animals are treated in
accordance with the Guide for the Care and Use of Laboratory
Animals published by the US national Institutes of Health (NIH
Publication No. 85-23, revised 1996).
[0190] Lignocaine hydrochloride is sourced as a 2% solution (ilium)
from the local Pharmaceutical Suppliers (Lyppard, Queensland). All
other chemicals, including adenosine (A9251>99% purity), are
sourced from Sigma Aldrich (Castle Hill, NSW).
Surgical Protocol:
[0191] Anesthetized non-heparinized animals are positioned in a
specially designed plexiglass cradle. A tracheotomy is performed
and the animals artificially ventilated at 75-80 strokes per min on
humidified room air using a Harvard Small Animal Ventilator
(Harvard Apparatus, Mass., USA) to maintain blood pO.sub.2,
pCO.sub.2 and pH in the normal physiological range (Ciba-Corning
865 blood gas analyzer).
[0192] Body temperature is maintained at 37.degree. C.
(Homeothermic Blanket Control Unit, Harvard Apparatus, Mass., USA).
A rectal probe is used to measure core body temperature. The left
femoral vein is cannulated using PE-50 tubing for drug withdrawal
and infusions while the right femoral artery is cannulated for
blood collection and blood pressure monitoring (UFI 1050 BP coupled
to a MacLab). All cannulae contains heparinized saline (100 U/ml
saline). Electrocardiogram (ECG) leads are implanted subcutaneously
in a lead II ECG configuration. Rats are stabilized for 15-20
minutes prior to blood withdrawal. Any animal that had dysrhythmias
and/or a sustained fall in mean arterial blood pressure below 80
mmHg are discarded from the study.
Hemorrhagic Shock:
[0193] The following examples are directed to hemorrhagic shock.
Hemorrhagic shock is induced by withdrawing blood from the femoral
vein or artery at a rate of 3 ml/100 g rat over 10 min to lower the
mean arterial blood pressure (MAP) to between 30 and 35 mmHg. For a
300 g rat the total blood volume is estimated to be
0.06.times.300+0.77=18.77 ml. Withdrawing 9 ml over a 10 min period
(0.9 ml/min) would result in a blood volume loss of about 50%.
[0194] For experiments involving 60% blood loss, 11.2 ml is
withdrawn over a 20 min period (0.56 ml/minute). The withdrawn
blood is then kept in a glass syringe that had been pre-rinsed with
0.02 ml heparin (1000 U/ml). MAP is maintained between 30 and 35
mmHg by blood withdrawal or re-infusion as needed for three shock
periods (1 hr or 2 hr or 3 hr, n=6 each shock period) prior to
crystalloid resuscitation.
[0195] At the end of the hemorrhagic shock period, rats receive the
resuscitation solutions outlined in each of the experiments below
to achieve a MAP of 80-90 mmHg (Note: in some experiments the MAP
is kept low to around 40-60 mmHg from the hypotensive effect of
adenosine and lignocaine to better balance the body's energy supply
and energy demand index).
[0196] Survival is assessed from haemodynamics (MAP, Heart rate)
and ECG following resuscitation, which is monitored for up to 6
hours. Death is recognized by the disappearance of MAP, HR and loss
of sinus rhythm, and verified by examination of the heart.
Example 1
Intravenous Administration of Adenosine/Lignocaine Resuscitation
Solution
[0197] Rats are randomly assigned into 4 groups (10 rats per group,
n=10) and prepared and subjected to hemorrhagic shock as described
above. After 60 min shock, the rats are resuscitated as
follows:
[0198] 1.1 Large Volume Fluids: Slow Intravenous Fluid Augmentation
[0199] Group 1: 10 minute infusion of 9 ml/100 g of 0.9% NaCl (3
times the volume of shed blood) containing 10 uM adenosine (or
adenosine analogues or agonists) and 30 microM lignocaine. [0200]
Group 2: 10 minute infusion of 9 ml/100 g of 0.9% NaCl
[0201] 1.2 Small Volume Fluids: Rapid Intravenous Fluid
Augmentation [0202] Group 3: bolus of 0.4 ml/100 g (1.2 ml for a
300 g rat) of 7.5% NaCl/6% dextran-70 containing adenosine (or
adenosine analogues or agonists) and lidocainelignocaine [0203]
Group 4: bolus of 0.4 ml/100 g (1.2 ml for a 300 g rat) of 7.5%
NaCl/6% dextran-70.
Example 2
Intraperitoneal Support of Intravenous Fluid Augmentation
[0204] Rats are randomly assigned into the same number of groups as
in Example 1 above with 10 rats in each group (n=10). Rats are
prepared and subjected to hemorrhagic shock as described above.
After 60 min shock, the rats are resuscitated as described in
Example 1 above plus an intraperitoneal bolus of 5 ml of 0.2 mM
adenosine (or adenosine analogues or agonists) and 0.5 mM
lignocaine.
Example 3
Slow Intravenous Administration of Resuscitation Solution
Containing Adenosine/Lignocaine Plus Additional Component
[0205] Rats are randomly assigned into 18 groups with 10 rats in
each group (n=10). Rats are prepared and subjected to hemorrhagic
shock as described above. After 60 min shock, the rats are
resuscitated using a 10 minute infusion of 9 ml (3 times the volume
of shed blood) of the following solutions: [0206] Group 1: 10 uM
adenosine (or adenosine analogues or agonists) and 30 uM lignocaine
plus 50 uM diazoxide [0207] Group 2: 10 uM adenosine (or adenosine
analogues or agonists) and 30 uM lignocaine plus 1 uM dipyridamole
(MW 504.6), [0208] Group 3: 10 uM adenosine (or adenosine analogues
or agonists) and 30 uM lignocaine plus 1 uM [D-Pen 2,5]enkephalin
(DPDPE) [0209] Group 4: 10 uM adenosine (or adenosine analogues or
agonists) and 30 uM lignocaine plus high magnesium sulphate (5 mM),
[0210] Group 5: 10 uM adenosine (or adenosine analogues or
agonists) and 30 uM lignocaine plus low magnesium sulphate (0.5 mM)
[0211] Group 6: 10 uM adenosine (or adenosine analogues or
agonists) and 30 uM lignocaine plus substrates/fuels (10 mM
glucose, 1 mM pyruvate) [0212] Group 7: 10 uM adenosine (or
adenosine analogues or agonists) and 30 uM lignocaine plus
antioxidant (1 mM allopurinol) [0213] Group 8: 10 uM adenosine (or
adenosine analogues or agonists) and 30 uM lignocaine plus 10 uM
amiloride [0214] Group 9: 10 uM adenosine (or adenosine analogues
or agonists) and 30 uM lignocaine plus 50-100 mM raffinose. [0215]
Group 10: 10 uM adenosine (or adenosine analogues or agonists) and
30 uM lignocaine plus 50-100 mM sucrose. [0216] Group 11: 10 uM
adenosine (or adenosine analogues or agonists) and 30 uM lignocaine
plus 50-100 mM pentastarch. [0217] Group 12: 10 uM adenosine (or
adenosine analogues or agonists) and 30 uM lignocaine plus
Dextran-30 at physiological pH. [0218] Group 13: 10 uM adenosine
(or adenosine analogues or agonists) and 30 uM lignocaine plus
Dextran-40 at physiological pH. [0219] Group 14: 10 uM adenosine
(or adenosine analogues or agonists) and 30 uM lignocaine plus
Dextran-50 at physiological pH. [0220] Group 15: 10 uM adenosine
(or adenosine analogues or agonists) and 30 uM lignocaine plus
Dextran-60 at physiological pH. [0221] Group 16: 10 uM adenosine
(or adenosine analogues or agonists) and 30 uM lignocaine plus
hydroxyethyl starch at physiological pH. [0222] Group 17: 10 uM
adenosine (or adenosine analogues or agonists) and 30 uM lignocaine
plus modified fluid gelatin at physiological pH. [0223] Group 18:
10 uM adenosine (or adenosine analogues or agonists) and 30 uM
lignocaine plus 50 uM diazoxide, 1 uM dipyridamole (MW 504.6), 1 uM
[D-Pen 2,5]enkephalin (DPDPE), high and low magnesium sulphate (5
and 0.5 mM), substrates/fuels (10 mM glucose, 1 mM pyruvate),
antioxidant (1 mM allopurinol), NaH inhibitor (10 uM amiloride),
50-100 mM sucrose and Dextran-40 at physiological pH.
Example 4
Rapid Intravenous Administration of Resuscitation Solution
Containing Adenosine/Lignocaine Plus Additional Component
[0224] Rats are randomly assigned into 16 groups (n=10) and
prepared and subjected to hemorrhagic shock as described above.
After 60 min shock, the rats are resuscitated using a bolus of 0.4
ml/100 g (1.2 ml for a 300 g rat) of the following solutions:
[0225] Group 1: 7.5% NaCl/6% dextran-70 containing 0.2 mM adenosine
(or adenosine analogues or agonists) and 0.5 mM lignocaine plus 50
uM nicorandil. [0226] Group 2: 7.5% NaCl/6% dextran-70 containing
0.2 mM adenosine (or adenosine analogues or agonists) and 0.5 mM
lignocaine plus 1 uM dipyridamole (MW 504.6). [0227] Group 3: 7.5%
NaCl/6% dextran-70 containing 0.2 mM adenosine (or adenosine
analogues or agonists) and 0.5 mM lignocaine plus 1 uM [D-Pen
2,5]enkephalin (DPDPE). [0228] Group 4: 7.5% NaCl/6% dextran-70
containing 0.2 mM adenosine (or adenosine analogues or agonists)
and 0.5 mM lignocaine plus high magnesium sulphate (5 mM). [0229]
Group 5: 7.5% NaCl/6% dextran-70 containing 0.2 mM adenosine (or
adenosine analogues or agonists) and 0.5 mM lignocaine plus low
magnesium sulphate (0.5 mM). [0230] Group 6: NaCl/6% dextran-70
containing 0.2 mM adenosine (or adenosine analogues or agonists)
and 0.5 mM lignocaine plus substrates/fuels (10 mM glucose, 1 mM
pyruvate). [0231] Group 7: NaCl/6% dextran-70 containing 0.2 mM
adenosine (or adenosine analogues or agonists) and 0.5 mM
lignocaine plus 1 mM allopurinol. [0232] Group 8: NaCl/6%
dextran-70 containing 0.2 mM adenosine (or adenosine analogues or
agonists) and 0.5 mM lignocaine plus 10 uM amiloride. [0233] Group
9: NaCl/6% dextran-70 containing 0.2 mM adenosine (or adenosine
analogues or agonists) and 0.5 mM lignocaine plus impermeants
(50-100 mM raffinose, sucrose, pentastarch). [0234] Group 10: 10 uM
adenosine (or adenosine analogues or agonists) and 30 uM lignocaine
plus 50-100 mM sucrose. [0235] Group 11: 10 uM adenosine (or
adenosine analogues or agonists) and 30 uM lignocaine plus 50-100
mM pentastarch. [0236] Group 12: NaCl/6% dextran-70 containing 0.2
mM adenosine (or adenosine analogues or agonists) and 0.5 mM
lignocaine plus colloids (Dextran-30, 40, 50 and 60, hydroxyethyl
starch and a modified fluid gelatin) at physiological pH. [0237]
Group 13: 10 uM adenosine (or adenosine analogues or agonists) and
30 uM lignocaine plus Dextran-40 at physiological pH. [0238] Group
14: 10 uM adenosine (or adenosine analogues or agonists) and 30 uM
lignocaine plus Dextran-50 at physiological pH. [0239] Group 15: 10
uM adenosine (or adenosine analogues or agonists) and 30 uM
lignocaine plus Dextran-60 at physiological pH. [0240] Group 16: 10
uM adenosine (or adenosine analogues or agonists) and 30 uM
lignocaine plus 50 uM diazoxide, 1 uM dipyridamole (MW 504.6), 1 uM
[D-Pen 2,5]enkephalin (DPDPE), high and low magnesium sulphate (5
and 0.5 mM), substrates/fuels (10 mM glucose, 1 mM pyruvate),
antioxidant (1 mM allopurinol), NaH inhibitor (10 uM amiloride),
impermeants (50-100 mM sucrose and Dextran-40 at physiological
pH.
Example 5
Intraperitoneal Support of Slow Intravenous Fluid Augmentation
[0241] Rats are prepared, subjected to hemorrhagic shock and
resuscitated as described in Example 3 together with an
intraperitoneal bolus of 5 ml of 0.2 mM adenosine (or adenosine
analogues or agonists) and 0.5 mM lignocaine.
Example 6
Intraperitoneal Support of Rapid Intravenous Fluid Augmentation
[0242] Rats are prepared, subjected to hemorrhagic shock and
resuscitated as described in Example 4 together with an
intraperitoneal bolus of 5 ml of 0.2 mM adenosine (or adenosine
analogues or agonists) and 0.5 mM lignocaine.
Example 7
The Effect of Lowering Body Temperature on the Different
Resuscitation Strategies
[0243] The above examples (examples 1 to 6) are repeated at 35, 33,
20, and 4.degree. C. The formulations are equilibrated with air or,
if found to be efficacious in preliminary testings, may be aerated
or have an oxygen containing perfluorocarbon based, or haemoglobin
based substitute present or blood, a blood product or artificial
blood. Components may be added to mimic human blood's oxygen
transport ability such as Hemopure.TM., Gelenpol.TM., Oxygent.TM.,
PolyHeme.TM..
Example 8
Treating VF During Cardiac Surgery (on-Pump)
[0244] Different ways of utilising the invention are illustrated in
5 groups labelled A-E of patients as follows: [0245] Group A:
Patients receiving standard local hospital hypothermic
(cardioplegic delivery temp 10.degree. C.) high potassium
cardioplegia plus potassium "hot shot" (arresting dose). [0246]
Group B: Patients receiving standard local hospital warm
(cardioplegic delivery temp 33.degree. C.) high potassium
cardioplegia plus potassium "hot shot" (arresting dose). [0247]
Group C: Patients receiving hypothermic adenosine and lignocaine
(cardioplegic delivery temp 10.degree. C.) cardioplegia (normal
potassium 5 mM) plus HiberStart (non arresting dose) to reanimate
the heart. [0248] Group D: Patients receiving warm adenosine and
lignocaine (cardioplegic delivery temp 33.degree. C.) cardioplegia
(normal potassium 5 mM) plus a non-arresting dose of adenosine and
lignocaine to reanimate the heart. L or ALM in Group B and D to
study the effect of pretreating the heart before arrest. [0249]
Group E: Hearts are pretreated/preconditioned using a solution of
adenosine and lignocaine with or without magnesium (1.0-20 mM) and
then the heart is arrested and reanimated as in Group D. Hearts may
also be postconditioned following reperfusion in combination with
an arresting or non-arresting dose of adenosine and lignocaine
solution with or without magnesium (1.0-20 mM).
[0250] The cardioplegia composition and protocol for human patients
in Groups A-E are as follows.
[0251] 1) Composition of high potassium cardioplegia solution for
Groups A & B:
[0252] Induction cardioplegia 20 mM K.sup.+ solution (final):
BAXTER (Code AHK5524). Each 500 ml contains: Sodium Chloride BP 4.5
g, Potassium Chloride BP 3 g, Magnesium Chloride BP 2.6 g,
Lignocaine HCl BP 250 mg. Before use, Sodium Bicarbonate (25
mmol/500 ml) and monosodium Aspartate (14 mmol/500 ml) added, with
pH .about.3.7 and osmolality .about.547 mOsm.
[0253] Maintenance cardioplegia 9 mM K.sup.+ solution (final):
BAXTER (Code AHK5525). Each 500 ml contains: Sodium Chloride BP 4.5
g, Potassium Chloride BP 1 g, Magnesium Chloride BP 2.6 g. Before
use, Sodium Bicarbonate (25 mmol/500 ml) and monosodium Aspartate
(14 mmol/500 ml) added, with pH .about.3.7 and osmolality
.about.547 mOsm.
[0254] During reanimation, the arrest solution is same as K+
maintenance but the myocardial heart temperature during induction,
maintenance and terminal shot is 32 to 38.degree. C. The heart
remains arrested at this time.
[0255] 2) Composition of adenosine and lignocaine ("AL")
cardioplegia solution for Groups C, D and E:
[0256] The optimal concentrations of AL will be found from a dose
response curve. A is about 0.2 to 2 mM and L is about 0.2 to 4 mM.
These concentrations have been shown to be safe in humans.
Magnesium may be 1.0-20 mM. The arresting induction is at higher
levels of A and L and maintenance dose may be lower e.g. at half
the concentration to induce arrest. Final K+ infused into the heart
around final 3-6 mM (normally around 5 mM). The temperature
profiles of the induction and maintenance volumes are similar to
the temperature protocol described for Group A & B.
[0257] During reanimation, the adenosine and lignocaine solution
does not arrest the heart but protects and preserves the heart
during reanimation. It may beat prior to release of cross clamp.
Concentrations of AL and M are A 10-40 micromolar, L: 30-50
micromolar and magnesium sulphate of 10-20 mM and the temperatures
32 to 38.degree. C.
[0258] 3) Composition of cardioplegia solution for Group E: Same as
Group D above but with pretreatment/preconditioning doses of
adenosine and lignocaine concentrations with or without magnesium
sulphate during reperfusion or during postconditioning +adenosine
and lignocaine concentrations with or without magnesium
sulphate.
[0259] If a MPS Quest cardioplegia perfusion pump system using
microplegia (1 part and 9 parts of blood) is available, the
following adenosine and lignocaine concentrations with and without
magnesium can be used to test the arresting and maintenance doses
to be used in the study.
[0260] Inducing Arrest: 54 mg A+132 mg L in the 50 ml cassette (0.5
mM and 1.0 mM final concs in the blood hitting the heart). The
studies by Mentzer et al have shown that 2 mM ado is safe in
cardioplegia in humans.
[0261] Maintenance: 26 mg A+66 mg L in the 50 ml cassette (0.2 mM
ado and 0.5 mM lido).
[0262] Reanimation: AL plus Mg++(also called "ALM") used at 10 uM
A, 30 uM L and 16 mM MgSO4 cassette.
[0263] Treatment of patients enrolled in the study who suddenly
experience life-threatening arrhythmias (ventricular tachycardia
and/or fibrillation): If a patient has a sudden cardiac event such
as a heart attack and the heart's beating rhythm abruptly changes
(eg. converts to ventricular tachycardia or ventricular
fibrillation) in hospital prior to or after surgery, a bolus dose
of adenosine and lignocaine with or without magnesium is given
intravenously (or intracardiac) to resuscitate the heart prior to
using the defibrillator (if a defibrillator is required). If the
patient has life-threatening severe arrhythmias (ventricular
fibrillation or ventricular tachycardia) while in the operating
room or intensive care ward a bolus dose of adenosine and
lignocaine with or without magnesium is given intravenously (or
intracardiac) to resuscitate the heart prior to using the
defibrillator (if a defibrillator is required).
Example 9
Protecting Against Heart Arrhythmias and Treating VF During and
Following Off-Pump Cardiac Surgery in Humans
[0264] Preclinical studies showed that an intravenous infusion
adenosine and lignocaine with or without magnesium is highly
protective to the heart during myocardial ischemia in the in vivo
rat and canine model. A three-pronged attack is envisaged: 1)
maintain the resting cell's membrane potential or voltage during
times of ischemia, 2) down-regulate metabolism, and 3) blunt the
inflammatory and hypercoagulable responses. Defending the membrane
potential close to the resting polarized state reduces ionic and
metabolic imbalances; down regulating the cell's metabolism lowers
the demand, and attenuating the inflammatory and blood clotting
responses, reduces further damage during reperfusion. Targeting all
three provides greater protection from life-threatening arrhythmias
and other ischemia-related damage to both the myocardium and
coronary vasculature. (Canyon, S and Dobson, G P 2004 "Protection
against ventricular arrhythmias and cardiac death using adenosine
and lignocaine during regional ischemia in the in vivo rat",
American Journal of Physiology, 287: H1286-H1295).
[0265] Intravenous infusion of adenosine and lignocaine using a
lower lignocaine dose was highly cardioprotective. Adenosine and
lignocaine with lower lignocaine concentrations resulted in no
deaths, virtually abolished severe arrhythmias and decreased
infarct size in the rat model of acute ischemia.
[0266] A composition of the invention administered in patients
undergoing beating-heart surgery will be via an intravenous route
through a dedicated port on a central venous line. The composition
comprises 305 .mu.g/kg/min adenosine plus 60 .mu.g/kg/min
lignocaine and is administered intravenously 5 min before and
during each coronary artery anastomosis. A single lignocaine bolus
(1 mg/kg) is injected for 3 min immediately before the first
administration of the AL solution.
[0267] For example, for a 70 kg patient: 0.305.times.70=21.35 mg
adenosine per min and 0.06.times.75=4.2 mg Lignocaine per minute is
administered. Prior to adenosine and lignocaine solution 70 mg of
lignocaine-HCL is given as a bolus.
[0268] Limits of infusion: If constantly infused for 1 hour: total
amount of adenosine and lignocaine administered if constantly
infused for 60 min equates to 60.times.21.35=1281 mg adenosine and
60.times.4.2 mg=252 mg lignocaine (+70 mg bolus=322 mg). Half-life
of adenosine is 4 to 10 seconds in human blood. We envisage a total
adenosine and lignocaine infusion time of 30 min to complete 3
anastomoses, average time 10 min each Another example is at an
infusion rate of 0.3 mg/kg/min in a 70 Kg subject, we need to
infuse 21 mg/min. If the infusion mixture is 300 mg in 540 ml (ie
0.56 mg/ml), for this subject we need to run at 37.5 ml/min or
2.250 l/hr.
[0269] Pre-clinical studies: In a rat study, 0.0567 g Adenosine and
0.565 ml lignocaine-HCl (20 mg/ml) to 10 ml saline were infused IV
1 ml/hour into a 300 g rat. So for our 35 min pretreatment and
ischemia period we only use 35/60.times.1 ml=0.58 ml of 10 ml
solution we make.
[0270] Start with a 1 mg/kg bolus of lido-HCl followed by a
infusion of the adenosine and lignocaine solution. The adenosine
infusion rate translates to 0.311 mg/kg/min (ie 1 ml/hour infusion
rate or 1/60 ml per min or 1/60.times.5.6 mg/ml (in 10 ml we
make).times.1000/300 (for 300 g rat)=0.311 mg/kg/min). The
lignocaine infusion rate translates to 0.0627 mg/kg/min (ie
1/60.times.1.13 mg/ml (in 10 ml we make).times.1000/300=0.0627
mg/kg/min).
Human Study:
[0271] For the 70 kg human study make a 500 ml (50 times) bag with
0.0567.times.70/0.3=13.23 g Adenosine and 0.565.times.70/0.3=132 ml
lignocaine-HCl (20 mg/ml) and deliver the solution at an IV rate of
50 ml per hour. This delivers the same amount of drug per unit
mass. For 5 anastomoses and pretreating each for 5 min before and
say each anastomoses takes 20 min (MAX) then that is 5.times.(20+5)
min=125 min (MAX)
[0272] Therefore at 50 ml/hour, infuse about 50/60.times.125=104 ml
per patient for 125 min total anastomoses time. Example of IV
infusion Protocol: To make 300 ml (not 500 ml) it would be
3/5.times.13.23 g Ado and 3/5.times.132 ml lignocaine HCl and
infuse iv at 50 ml per hour. For a 70 kg human, Adenosine infusion
rate: 0.311 mg/kg/min or 21.77 mg/human patient/min and
Lignocaine-HCl infusion rate: 0.0627 mg/kg/min or 4.39 mg/human
patient/min.
[0273] 6 min before first anastomoses inject bolus of lignocaine (1
mg/kg) followed by AL solution for 5 min. Stop infusion after each
anastomoses has been completed. When the surgeon is ready for the
next anastomoses, begin infusion IV 5 min before. And repeat the
same for each anastomoses.
[0274] Timing of administration: 5 min before surgery, continued
during regional ischaemia and stop following completion of the
anastomosis.
Example 10
Treating VF and Using Adenosine and Lignocaine with and without
Magnesium During `On-Pump` Cardiac Surgery
[0275] Currently over 99% of all surgical cardioplegia solutions
contain high potassium (15-20 mM), which arrests the heart
unnaturally by depolarising the membrane potential from -83 mV to
about -50 mV. At these depolarizing potentials sodium can increase
inside cells via the Na.sup.+`window` current, which, in turn,
leads to a rise in intracellular Ca.sup.2+ through the reversal of
the Na/Ca.sup.2+ exchanger. The potentially damaging accumulation
of Ca.sup.2+ may occur during the cardioplegic period (induction
and maintenance phase) and/or during the reanimation-reperfusion
phase following arrest. High potassium-linked Ca.sup.2+ loading has
been linked to myocardial stunning, ventricular arrhythmias,
ischaemic injury, microvascular injury, tissue oedema, free radical
production and functional loss during the reperfusion period.
Depolarising potassium is also a potent coronary vasoconstrictor
and this may further compound any antecedent vulnerability of the
heart to injury during cardioplegic arrest, maintenance and
recovery.
Example 11
Treatment for Life-Threatening Ventricular Tachycardia and/or
Fibrillation
[0276] A large number of sudden deaths are caused by acute
ventricular tachyarrhythmias (ventricular tachycardia and/or
fibrillation) and often triggered by acute coronary events in
association with heart disease or in persons without known cardiac
disease. The most common pathophysiological cascade in the
appearance of fatal arrhythmias is that ventricular tachycardia
degenerates to ventricular fibrillation and later to asystole or
cardiac arrest and death. If a patient experiences a sudden cardiac
event such as a heart attack and the heart's beating rhythm
abruptly changes (eg. converts to ventricular tachycardia or
ventricular fibrillation) in hospital prior to or after surgery, a
bolus dose of adenosine and lignocaine with or without magnesium is
given intravenously (or intracardiac) to resuscitate the heart
prior to using the defibrillator (if a defibrillator is required).
If the patient has life-threatening severe arrhythmias (ventricular
fibrillation or ventricular tachycardia) while in the operating
room or intensive care ward a bolus dose of adenosine and
lignocaine with or without magnesium is given intravenously (or
intracardiac) to resuscitate the heart prior to using the
defibrillator (if a defibrillator is required).
[0277] The method to treat human subjects suffering from an
unexpected cardiac event leading to irregular arrhythmias such as
acute ventricular tachyarrhythmias and pharmacologically convert
the heart to normal beating or sinus rhythm using adenosine and
lignocaine with and without magnesium is as follows.
[0278] Hearts will be arrested using a microplegia method of
hyperkalemic cardioplegia induction, maintenance and reanimation.
Microplegia is an alternative method to infusing the myocardium
with the standard 4:1 mixture of blood and cardioplegia to arrest
the heart. Microplegia aims to induce and maintain aerobic arrest
of the heart by delivering continuous oxygen rich blood coupled
with micro titrations of potassium (arrest) and magnesium
(additive) solutions. Aerobic arrest offers superior myocardial
protection over that of standard 4:1 cardioplegia regimens and
tighter control of blood glucose levels. Most importantly, with the
addition of adenosine and lignocaine to the additive mixture, an
even higher level of myocardial protection is expected to produce
long-lasting perioperative benefits to the patient. The composition
of adenosine and lignocaine makes cardiac surgery safer for the
patient and more predictable for the surgeon.
[0279] This example compares potassium arrest induction and
maintenance cardioplegia and a non-arrest reanimation solution
using adenosine and lignocaine with and without magnesium. The
maintenance solution may also contain adenosine and lignocaine but
the principal mode of arrest in these groups will be high
potassium. In another separate group, adenosine and lignocaine will
be the principal mode of arrest, protection and preservation for
induction and maintenance cardioplegia and reanimation will be
compared to the potassium arrest, maintenance and reanimation
groups. In cases, where the heart does not return to proper
function after reanimation a bolus of adenosine and lignocaine with
or without magnesium will be given in the perfusion line (or
intracardiac) to resuscitate the heart prior to using the
defibrillator (if a defibrillator is required).
[0280] Microplegia Delivery Protocol: Patients scheduled for
on-pump coronary artery bypass surgery, valve surgery or combined
procedure; or re-operations of the same. Patients receive
anaesthesia and cardiac surgery as per usual practice. The use of
inotropes, vasoconstrictors is "protocol-driven" and based on
criteria for use as agreed upon by the surgical/anaesthesia team.
The following are cassette formulations designed for arrest,
maintenance and reanimation of the heart in, for example, the Quest
MPS.RTM. Microplegia System.
[0281] The Arrest Cassette: [0282] 1. 80 mEq of undiluted
Potassium=40 mL. [0283] 2. High Setting: 25 mEq/L. [0284] 3. Low
Setting: 10 mEq/L.
[0285] The Additive Cassette: [0286] 1. 12 mg Adenosine=4 mL.
[0287] 2. 25 mg Lignocaine=1 mL. [0288] 3. 5 gm Magnesium
Sulfate=10 mL. [0289] 4. 30 mL crystalloid prime (e.g. Plasmalyte).
[0290] 5. Total Volume in Additive Cassette: 45 mL. [0291] 6.
Additive Setting: 10 mL/L.
[0292] Upon heparinization, fill the ice reservoir to the top with
ice. Refill reservoir as needed. Delivery temp .about.8-12.degree.
C. Temperature setting for warm induction is 37 degrees. [0293] 1.
Cardiac arrest is induced with normothermic hyperkalemic blood
microplegia. Patients receive hyperkalemic blood microplegia with
1) adenosine and lignocaine and magnesium in the additive cassette
as a pre-treatment regimen, 2) lower adenosine and lignocaine,
magnesium and potassium levels during the delivery of maintenance
cardioplegia, and 3) adenosine and lignocaine and magnesium without
potassium as a warm reperfusion dose. [0294] 2. Upon application of
the cross clamp for the induction phase, ramp up flow for antegrade
quickly to 500 mL/min then immediately back down to 320 to 350
mL/min. This ensures closure of the aortic valve. A total of
700-1000 mL of warm blood cardioplegia with high (25 mEq) potassium
is delivered in antegrade fashion. Upon achieving quiescence,
switch to retrograde warm and deliver an additional 700 mL. [0295]
3. Switch the water bath to cold (4.degree. C.; the delivery
temperature is between 8 and 12.degree. C.). Lower additive setting
(saline or adenocaine) to 2 mL/L. Whenever possible, continue
microplegia administration throughout the case. [0296] 4. When
approaching the last ten minutes of cross clamp, preparations are
made to deliver the warm reperfusion dose by switching the water
bath to warm at 37-38 degrees. [0297] 5. Warm reperfusion dose:
Start or continue delivery of warm blood with or without adenosine
and lignocaine in retrograde fashion. Administer warm retrograde
microplegia until grafts are completed and then switch to antegrade
delivery mode. This facilitates de-airing of the grafts and allow
the right side of the heart to be perfused. The antegrade modality
of delivery ensures that microplegia is adequately delivered and
distributed to the myocardium (when all grafts are completed).
After three to five minutes in the antegrade mode, when de-airing
is complete, continue with warm antegrade until remaining volume in
additive cassette is given (usually 500-1000 cc).
[0298] A number of end points and measurable outcomes are assessed
for comparison: [0299] 1. Patient demographics and history
including: age, gender, co-morbidities (i.e. diabetes,
hypercholesterolemia, hypertension, smoking, COPD, renal failure).
[0300] 2. Pre-op data including: diagnosis, BSA and lab values
(BUN/Creatinine, INR, PT/PTT, INR, PLT. count, post-heparin glucose
and HCT), number of intended grafts. [0301] 3. Intra-op data
including: [0302] 1) number of vessels bypassed [0303] 2) length of
bypass (start to wean time) [0304] 3) cross-clamp time [0305] 4)
blood glucose levels, insulin dosing [0306] 5) HCT during the case
[0307] 6) total volume of microplegia delivered, amount of additive
and potassium given [0308] 7) duration of warm reperfusion dose,
volume of hotshot [0309] 8) number of breakthrough events
(re-animation of myocardium) and potassium level needed to
re-arrest [0310] 9) pre- and post-CPB ejection fraction [0311] 10)
return to sinus rhythm before cross-clamp removal (yes/no);
incidence of ventricular fibrillation [0312] 11) Incidence of
atrial fibrillation [0313] 12) Need for cardioversion (number of
shocks, energy level of each) [0314] 13) Need for any rearrest
protocol [additional or extended hotshot (at surgeon's request),
need to convert to adenocaine at the point of hotshot] [0315] 14)
Urine output from O.R. until first 24 hours post-op. [0316] 15)
Blood product usage (FFP, Platelets, RBCs and Cryo.) from O.R.
until discharge. [0317] 16) Blood glucose levels and insulin dosing
(first 24 hours post-op). [0318] 17) Plasma Troponin I levels at 6,
12, 24 hours post-op. [0319] 18) Clinical evidence of acute
myocardial infarction (Q-waves, arrythmias). [0320] 4. Time on
ventilator (time to extubation). [0321] 5. Length of stay in ICU
and total length of stay in hospital (time to discharge). [0322] 6.
Treatment of post-op atrial fibrillation, counter-shocks (number
and joules), and use of pacemakers measured according to the
following schedule: [0323] 1) From X-clamp removal to exit of O.R.
[0324] 2) First 24 hours post-op in TICU. [0325] 3) From exit of
TICU until discharge. [0326] 7. Use of inotropes and
vasoconstrictors protocol driven and measured according to the
following schedule: [0327] 1) Rate of each upon leaving the O.R.
[0328] 2) Rate of each after first 24 hours post-op in TICU.
[0329] In some cases during reanimation of the heart after surgery,
the heart will not respond and it fibrillates. A bolus dose of
adenosine and lignocaine with or without magnesium will be given in
the perfusion line or suitable entry point to the heart muscle (or
intracardiac) to resuscitate the heart prior to using the
defibrillator (if a defibrillator is required). If the patient has
life-threatening severe arrhythmias in the intensive care ward a
bolus dose of adenosine and lignocaine with or without magnesium
will be given intravenously (or intracardiac) to resuscitate the
heart prior to using the defibrillator (if a defibrillator is
required).
Example 12
Treatment During Surgery
[0330] The compositions and methods of the invention can also be
used during periods of reduced metabolic activity to reduce damage,
such as cell quiescence (medically induced or otherwise). Cardiac
surgery is one example. In this example, a known hyperkalemic
cardioplegic is used, and the composition of the present invention
is administered to reduce tissue damage during the operation.
[0331] This protocol uses miniplegia as described above, where
micro amounts of the composition of the invention are mixed at
various proportions with the patient's own oxygenated blood and
perfused into the heart at different settings. The reference to a
"setting" is a measure on the pump, such as a syringe pump, of the
amount of substance being mixed in blood and delivered directly to
the organ, in this example a heart.
[0332] Two cassettes were prepared as follows.
(1) The Arrest Cassette:
[0333] 1. 40 mls of undiluted Potassium having 80 mEq--thus, 2
mEq/ml [0334] 2. High Setting: 25 mEq's per litre [0335] 3. Low
Setting: 10 mEq's per litre [0336] The potassium in item 1 above
was the primary cardioplegic agent. High potassium is the most well
known and used cardioplegic, despite its known disadvantages and
deleterious side-effects. An alternative cardioplegic is disclosed
in WO 00/56145 (GP Dobson) comprising a potassium channel
opener/agonist and/or an adenosine receptor agonist (eg. adenosine)
together with a local anaesthetic (eg. lignocaine) in mM amounts.
The contents of this specification are incorporated herein by
reference in entirety. Although not exemplified here, the high
potassium cardioplegic of item 1 above could be replaced by such a
cardioplegic.
(2) The Additive Cassette:
[0336] [0337] 1. 4 ml Adenosine having 12 mg--thus, 3 mg/ml [0338]
2. 10 mls Magnesium Sulfate=5 g (or a vial of MgSO.sub.4 to equal 5
g) [0339] 3. 30 mls--whatever crystalloid prime is in a pump can be
used (e.g. L/R, Plasmalyte.TM., Normosol.TM.) [0340] 4. Total
Volume in Additive Cassette: 44 mls [0341] 5. Additive Setting: 10
mls per litre [0342] This cassette is suitable for machines which
support 50 ml cassettes.
[0343] Lignocaine is added to this cassette as described below to
deliver the improved results. Lignocaine is added at a
concentration of 0.1 to 10 times that of adenosine, preferably 0.5
to 2 times.
[0344] The data below is from experiments where no lignocaine was
added to this cassette until the recovery phase shortly before
cross-clamp removal. However, in another embodiment of the
invention, lignocaine is added to this cassette from its first use
so that a combination of adenosine and lignocaine is administered
during the maintenance or quiescent phase of a procedure. It is
found that this further improves the prospects of heart recovery
and/or reduced post-operative complications.
[0345] The procedure used to administer the composition in this
example was as follows, with an overall objective of creating
aerobic arrest, not ischemic arrest. [0346] 1. Upon heparinization,
fill the ice reservoir to the top with ice. Reservoir need not be
filled again unless x-clamp time exceeds 3 hours. Delivery temp
will be about 12.degree. C. Towards the last third of the x-clamp
period, some metabolism of oxygen rich blood should occur. [0347]
2. Temperature setting is for warm induction: Warm (37.degree. C.)
[0348] 3. High setting for arrest: 25 mEq/litre of the hyperkalemic
Arrest cassette induces a rapid arrest [0349] 4. Setting for
Additive: 10 ml/litre before cross-clamp
Upon Application of Cross-Clamp:
[0349] [0350] 1. Increase flow for antegrade quickly to 500 mls
then immediately back down to 320 to 350 mls/min so as to ensure
closure of the aortic valve. [0351] 2. Give 700 mls warm antegrade.
Once quiescence achieved, give 300 mls more and then switch to low
K+ setting (ie 10 ml/litre). [0352] 3. Give 700 mls warm
retrograde. [0353] 4. Switch water temp to cold. Administer cold
retrograde for as long as possible. Lower arrest setting
empirically the longer flow continues. [0354] 5. Lower additive
setting to 2 ml/litre. Most preparation of the heart has occurred.
[0355] 6. If you are doing a CABG and distals are performed first:
after the first graft, hook up the graft to the pump via
multi-catheter lines. The flow is then increased very slowly to
achieve a pressure of 150 Torr and the flow is noted, which is
useful information for the surgeon. This will accomplish several
things: [0356] controlled mechanical device to determine patency of
the graft utilizing the gold standard of pressure to flow ratio;
[0357] surgeon has a means to check hemostasis of the anastomotic
site; and [0358] capability to deliver antegrade to the target site
and retrograde simultaneously if desired. [0359] 7. If the
procedure involves work on a valve and coronaries, perform the
coronaries first. This way a sick heart is provided with the
nutrients it needs while the valve is being worked on. [0360] 8.
Monitor K+ according to usual SOP and adjust potassium
concentration to meet desired level.
[0361] When approaching the last 10 minutes of x-clamp,
preparations are made for the warm shot. These include: [0362] 1
Water setting: Warm (37 degrees) [0363] 2 Arrest setting: 0--to
wash out the K+ and other metabolites [0364] 3 25 mg. Lignocaine is
injected into Additive bag (in this embodiment being described, it
has not been added earlier) to accomplish target delivery of the
prophylactic antiarrhythmic composition--typically there is about
18-35 ml left in the Additive bag at this point depending on the
length of time for the procedure, which provides a lignocaine
concentration of about 1 mg/ml. [0365] 4 Additive setting: 15 to
18--the goal is to empty the Additive bag prior to removal of
cross-clamp.
[0366] For warm shot: usually started 5 to 10 minutes prior to
x-clamp removal [0367] 1 Start retrograde warm. Zero potassium,
additive setting at 15. Make sure retrograde pressure is maintained
at highest level (35 to 40 Torr) [0368] 2 When electrical activity
begins, continue retrograde for another minute. [0369] 3 Switch to
antegrade for 2 to 3 minutes (when not obscuring surgeons' vision).
This will facilitate de-airing grafts, allowing the right side of
the heart to be perfused and, usually, will achieve a stable heart
rate. [0370] 4 Switch back to retrograde for duration of x-clamp.
[0371] 5 If additive setting runs out, continue with pure warm
blood through x-clamp removal.
[0372] With microplegic techniques, the more volume you give, the
better the heart likes it as it is aerobic arrest. In many
instances, if administered properly, the oxygen supply/demand ratio
is reversed. Administration of over 1 and up to 6 litres is
associated with the greatest reduction in post-operative
fibrillation.
[0373] The clinical results attained with warm blood cardioplegia
have suggested that earlier observations on impairment of some cell
functions by hypothermia may be more relevant than previously
thought. These include reduced: [0374] 1 Membrane stability [0375]
2 Ability to utilize glucose and fatty acids [0376] 3 Mitochondrial
generation of adenosine tri-phosphate leading to depressed Cell
membrane function [0377] 4 Activity of adenosine tri-phosphatase
system, leading to impaired cell volume regulation [0378] 5
Decreased ability of the sarcoplasmic reticulum to bind calcium
[0379] 6 Mitochondrial state respiration and activity of citrate
synthetase [0380] 7 Control of intracellular pH [0381] 8 Activity
of the sarcoplasmic reticulum with regard to calcium uptake
[0382] Coupling warm induction with cold maintenance and warm shot
towards the end of cross clamp provides superior results. Warm
induction, especially with the addition of adenosine (a very
powerful vasodilator, among other functions), opens up all the
collaterals and provides the necessary conduit for arrest and
additives to reach the myocyte and endothelium. With cold induction
comes constriction and the inability to globally distribute
cardioplegia down to the myocyte and endothelium.
[0383] Cold maintenance provides a reduction in metabolic uptake
with the slow increase in temperature occurring during the natural
course of cross clamp due to ice melting. Average temperature will
drift to around 12 to 14.degree. C. The warm shot at the end is a
most important aspect of myocardial protection. By allowing the
heart to experience warm blood (32 to 37.degree. C.) as long as is
possible, can mean the difference in regaining most of the heart's
functional recovery as opposed to a flaccid, lifeless heart,
requiring inotropes and electrical support. There is also evidence
that subjecting a cold, flaccid, non-beating heart to the trauma of
high flow warm blood, such as experienced when the cross clamp is
removed, sets the heart up for sure fire reperfusion injury.
[0384] Over the course of the last 30 years, surgeons and
perfusionists have refined their operative techniques, allowing
them to "customize" how they approach each patient's particular
needs and demands. The only area that has essentially remained a
"cookie cutter" approach has been myocardial protection;
essentially "one size fits all". Without being bound by any
particular theory or mode of action, it is believed that the method
of this preferred embodiment is more sensitive to not
over-hemodiluting the patient and thus results in improved
outcomes.
[0385] In one experiment, 2688 patients undergoing cardiac surgery
using cardioplegia were assessed at 6 different hospitals using
different surgeons and their different techniques to assess for
variability in this delicate environment. All patients were treated
with a standard hyperkalemic cardioplegic solution to induce
arrest. Of the patients, 1279 were in the group subjected to
typical standard crystalloid-cardioplegic protocol ("Standard").
1409 were subjected to a microplegia protocol (ie one using minimal
amounts of cardioplegic directly administered to the heart) using
the same hyperkalemic cardioplegic and with a warm ALM Additive
cassette as described above, ie having a composition according to
the invention. The invention is not specific or limited to this
form of cardioplegia, but it forms application of the method of the
invention and is discussed here to assess and illustrate the effect
of the invention.
[0386] The Additive cassette was used as described above, such that
during the recovery phase it contained Adenosine, Lignocaine and
Magnesium (hence the label "ALM"). The method of the invention is
referred to as "ALM" as a convenient abbreviation only. ALM was
administered at cross-clamp removal in accordance with the protocol
described above.
[0387] Table 1 sets out the characteristics of the 2688 patients
and Table 2 sets out the occurrence of different post-operative
complications measured.
TABLE-US-00002 TABLE 1 Patient Groups Standard ALM Number of
Patients 1279 1409 Age (Years) 62 .+-. 10 65.7 .+-. 10 Weight (kg)
89 .+-. 16 79.5 .+-. 16 Height (cm) 174 .+-. 9 168 .+-. 10 Body
mass index 30 .+-. 5 29 .+-. 5 Male (%) 53 60 Peripheral vascular
disease (%) 18 21 Diabetes mellitus (%) 36 35 Emergency surgery (%)
8 10 Extra corporeal bypass time (min) 87 .+-. 29 110 .+-. 37
[0388] In Table 2, the clinical outcomes are tabulated for the
patients identified in Table 1. The third column represents the ALM
proportion of patients as a percentage of the proportion of
standard cardioplegia patients for each outcome (ie second column
as a percentage of the first column). All of the outcomes in the
left column are negative outcomes, and thus their minimisation is
desired.
TABLE-US-00003 TABLE 2 Clinical Observations ALM as % Standard ALM
of standard Intra-operative inotropes (%) 93% 13% (14%)
Intra-operative pacing (%) 86% 33% (38%) Intra-operative
transfusions (%) 43% 24% (56%) Length of Stay post-op (days) 7 6
(79%) Post-op atrial fib. (%) 34% 3% (9%)
[0389] It can be seen that there was a substantial reduction in
complications following the above protocol, especially in
post-operative atrial fibrillation and the need for intra-operative
inotropes. In particular, the reductions in these negative outcomes
are: 86% reduction of intraoperative inotropes; 64% reduction in
intraoperative pacing; 44% reduction in intraoperative
transfusions; 21% reduction in length of stay post-operative days
and 91% reduction in post-operative atrial fibrillation.
Example 13
Administration of Adenosine/Lignocaine Solution Following Shock
[0390] The following Adenosine/Lignocaine (AL) solution(s) were
used in this example: AL solution=200 microM Adenosine, 500 microM
Lignocaine in Krebs Henseleit solution
[0391] Rats were subjected to hemorrhagic shock for 2 hrs and 10
mins as described above.
[0392] FIG. 1 shows the ECG monitoring of the rat heart during this
experiment. FIG. 1A shows the rat heart as normal prior to
hemorrhagic shock (Heart rate (HR)=375 bpm and MAP 114 mmHg).
Following shock, the HR was reduced to 35 bpm BP<10 mmHg (FIG.
1B). 0.5 mL bolus of the AL solution was administered directly into
the heart. FIG. 1C shows that the HR increased to 207 bpm 1.5
seconds following administration of the solution.
[0393] FIG. 2A shows in more detail the cardioversion of the rat
heart during this experiment. In particular, 1.5 seconds following
administration of the solution the rat heart rate increased from 35
bpm to 207 bpm. The point of administration of the solution is
denoted as (I). FIG. 2B shows the heart rate of this rat slowing
again 10 seconds after the administration of the solution.
[0394] Without being bound by any particular mode of action or
theory, these results show that since the animal has very little
blood volume a bolus of the AL solution can return the heart rate
for an initial period. Further intervention as shown in FIG. 2B at
(II), such as chest compressions and/or further shot of AL
solution, would then be required to keep the subject alive,
preferably with a blood volume replacement as well.
[0395] This example aims to pharmacologically induce a
hypometabolic `hibernating-like` state during resuscitation to
better balance the whole body oxygen supply-demand ratio and to
aggressively attenuate the inflammatory and hypercoagulable
imbalances associated with traumatic hemorrhagic shock and
resuscitation with particular emphasis on reducing damage to the
vital organs such as brain, heart, lung and gut. The inflammatory
state and edematous nature of the lung, the so-called "wet-lung",
"shock lung", "Da-nang lung" or "acute respiratory distress
syndrome" can occur in up to 50% of severely traumatized
patients.
Example 14
Intravenous Therapy with Adenosine/Lignocaine Resuscitation Fluid
Following Hemorrhagic Shock
[0396] The following Adenosine/Lignocaine solution(s) were used in
this example:
[0397] ALM (resuscitation solution)=10 uM Adenosine, 30 uM
Lignocaine and 2.5 mM MgSO.sub.4 in 7.5% NaCl solution.
[0398] FIG. 3 shows the ECG trace of the rat during normal period.
The MAP and HR measured at this time are shown in Table 3
below.
[0399] Rats were subjected to hemorrhagic shock involving
approximately 45% blood loss as described above until MAP drops to
around 30 to 35 mmHg. The maximum blood withdrawn was 8.6 ml over
the course of the shock period.
[0400] Total blood volume estimated to be 0.06.times.304+0.77=19.01
ml. Therefore, % blood volume
lost=8.6/19.01.times.100=.about.45%.
[0401] FIG. 4 shows the ECG monitoring of the Rat heart at the end
of the bleed period prior to the commencement of the "shock
period". The MAP and HR measured at this time are shown in Table 3
below.
[0402] The animal was kept in shock for 180 mins then either ALM or
7.5% saline is administered via iv bolus. The HR and MAP were
measured during each 60 mins shock period, ie (i) 0-60 mins, (ii)
60-120 mins, (iii) 120-180 mins, and shown in Table 3 below. The
ECG monitoring was continued (FIG. 5, FIG. 6 and FIG. 7).
[0403] At the end of the 3 hr shock period 11.0 ml bolus ALM (7.5%
NaCl, 2.5 mM MgSO4, 10 uM Adenosine, 30 uM Lidocaine) was infused
slowly into the femoral vein.
[0404] ECG trace 10 mins after infusion is shown in FIG. 8. The HR
and MAP measurements taken at this time are shown in Table 3
below.
[0405] ECG trace 30 mins after infusion is shown in FIG. 9. The HR
and MAP measurements taken at this time are shown in Table 3
below.
[0406] ECG trace 60 mins after infusion is shown in FIG. 10. The HR
and MAP measurements taken at this time are shown in Table 3
below.
[0407] ECG trace 90 mins after infusion is shown in FIG. 11. The HR
and MAP measurements taken at this time are shown in Table 3
below.
TABLE-US-00004 TABLE 3 Mean systolic Mean diastolic pressure
pressure Mean HR (mmHg) (mmHg) MAP (bpm) Normal (pre- 119.69 89.77
350 hemorrhagic shock) Bleed Period End of bleed -- -- 40 350
period (pre- hemorrhagic shock) Hemorrhagic shock period Shock
66.48 33.01 -- 281.58 (0-60 mins)* Shock 69.77 29.53 -- 250.39
(60-120 mins)* Shock 67.41 28.33 -- 245.80 (120-180 mins)* Shock
65.58 27.46 40.17 239 (at 180 mins) Recovery 10 mins after 91.66
36.62 268.0 ALM infusion 30 mins after 79.59 28.31 276.0 ALM
infusion 30-60 mins after 73.82 28.38 43.53 263.30 ALM infusion* 60
mins after 71.35 28.13 249 ALM infusion 60-90 mins after 65.81
25.52 252.96 ALM infusion* 90 mins after 58.72 24.05 228 ALM
recovery *the mean values of each of the measurements taken over
the indicated time period are shown
Example 15
Comparative Example of Intravenous Therapy with
Adenosine/Lignocaine Resuscitation Fluid and 7.5% Saline Following
Hemorrhagic Shock
[0408] The following Adenosine/Lignocaine solution(s) were used in
this example:
[0409] ALM (resuscitation solution)=10 uM Adenosine, 30 uM
Lignocaine and 2.5 mM MgSO.sub.4 in 7.5% NaCl solution.
[0410] Rats were subjected to hemorrhagic shock involving
approximately 45% blood loss as described in the previous example
until MAP drops to around 30 to 35 mmHg.
[0411] FIG. 12 shows the ECG trace of the rat during this
experiment. FIG. 12A shows the rat heart as normal prior to
hemorrhagic shock (HR approx 350 bpm; MAP 100 mmHg). FIG. 12B shows
the ECG monitoring 60 mins after shock. The MAP and HR were
measured at this time (MAP 44 mmHg; HR increased approx 280 bpm)
(FIG. 12B). ECG monitoring is continued for a further 120 mins.
FIG. 12C shows that the MAP remains relatively stabile after 180
mins of shock at 40 mmHg (HR approx 239 bpm).
[0412] FIG. 13A shows the ECG trace of the rat following
administration of 0.5 mL 7.5% saline after 180 mins shock. The HR
dropped to around 39 bpm (MAP 30 mmHg). This was maintained for
about 10 mins after administration of the 7.5% saline solution. The
heart rate then increased to 270 bpm
[0413] FIG. 13B shows the ECG trace of the rat following
administration of 0.5 ml bolus of ALM after 180 mins shock. The HR
increased to 261 bpm (MAP 35 mmHg) immediately.
[0414] This (and the previous example) shows that heart function
can be maintained by periodic bolus administration of ALM to a
subject that has suffered hemorrhagic shock. Without being bound by
any particular theory or mode of action, this low volume solution
could be used in situations where sufficient medical assistance is
delayed. For example, the solution could be administered at
periodic intervals by field medics at the site of an accident or in
the battlefield to provide intraperitoneal support during
complicated or prolonged evacuations or transport of the patient to
a hospital. This example demonstrates that an intravenous (iv)
bolus of the solution could be deployed immediately after severe
blood loss to stabilize and protect the heart from ischemic
depolarization and arrhythmias and to pharmacological down-regulate
the major organs of the body before resuscitation. This possible
battlefield scenario assumes a military medic or combat life-saver
is able to assist the wounded soldier near the scene of
trauma/injury.
[0415] It will be understood that the invention disclosed and
defined in this specification extends to all alternative
combinations of two or more of the individual features mentioned or
evident from the text or drawings. All of these different
combinations constitute various alternative aspects of the
invention.
* * * * *