Red cell substitute

1. Presented by : Dr. Prakriti Shukla

2. Red blood cell substitutes
The term “blood substitute” was first used for plasma
expanders
Later for blood components
Presently it is applied to ex-vivo produced therapeutic
materials with potential to replace or reduce transfusion
requirement of blood and blood components
Presently it is applied to ex-vivo produced therapeutic
materials with potential to replace or reduce transfusion
requirement of blood and blood components

4. Need for the development
1. With rapid advances in surgical sophistication and increasingly aggressive
protocols of cancer management in the recent past ,the transfusion
requirement of red cells have outgrown human resourse.Transfusion
alternatives and blood substitutes are necessary to narrow the gap between
demand and supply.
2. Risk of transmissible infections with blood transfusion has occupied
frontline attention. Blood substitutes which undergo vigorous in vitro
sterlization can alleviate both risk and cost in this regard.
3. Sudden transfusion requirements triggered by massive blood loss in
accidents, natural disasters etc. cannot be entirely met by allogenic blood
transfusions. Off – the –shelf available blood substitutes can be life saving.

5. Need contd..
4. Blood substitutes that are devoid of immune identity markers are
universally acceptable that help to cut across all immunogenic risks
of allogenic transfusions like incompatibility reactions, graft versus
host reactions, allergic and febrile reactions, and transfusion related
lung injury.
5. Limited shelf life of allogenic components resulting in wastage
beyond the expiry date can be avoided by blood substitutes that
show promise of longer shelf life even for several years if stored in a
lyophilized state.

6. 2 main indications for the transfusion of red cells :
1. severe haemorrhage
2. chronic symtomatic anaemia for which no specific therapy exist.
In both the circumstances , the aim of red cell transfusion is to
improve the oxygen supply to the tissues by raising the oxygen
content of the blood , according to the equations:

7. Development of red cell
substitutes
As a result of massive research and competition in
pharmaceutical companies, three categories of red
cell substitute have emerged

8. Cell free haemoglobin
solutions
The most important function of red blood cell is to
carry oxygen and carbon dioxide by virtue of its
haemoglobin content .Therefore , an alternative to
red blood cell transfusion , RBC free haemoglobin
solutions have been prepared.
Problems encountered :
1. Outside the RBC, the nomal tetrameric Hb molecule
undergoes fragmentation into dimeric forms which
have following effects:

10. 2. Outside the red cell free Hb looses its natural buffering
environment of 2,3 DPG thereby adversely affecting the
binding and release of O2 by free Hb molecule.
3. The Intraerythrocytic environment provides for the
important reductive enzymes and mechanisms which
combat the very oxidative nature of oxyhaemoglobin.
4. RBC free Hb is devoid of red cell enzymes and are
therefore incompetent to prevent reperfusion tissue
injury.

11. Strategies to overcome above
difficulties -
1. Modified Haemoglobins
a) polymerized Hb (cross –
linked)
b) conjugated Hb
2. Recombinant Haemoglobins
3. Encapsulated Haemoglobins
4. Perfluoro chemical emulsion

12. 1. Cross linked hemoglobinCross linked hemoglobin
To produce cross-linked hemoglobin,
small bridges of sugar molecules are
covalently attached to the dimers to
create a stable tetramer.
2. Polymerized hemoglobinPolymerized hemoglobin
To polymerize hemoglobin, surface amino acid
groups are linked by reagents such as glutaraldehyde.
Polymerized hemoglobin is the only product to date
that has not triggered significant vasoconstriction
after infusion.

13. 3. Surface-modified hemoglobin3. Surface-modified hemoglobin
Surface-modified hemoglobin is created by attaching large
molecules, such as polyethylene glycol, to surface lysine
groups.
This modification also increases the viscosity and oncotic
pressure of the solution.

14. 4. Perfluorocarbon-based substitutes
PFCs are synthetic hydrocarbons with halide substitutions
and are about 1⁄100th the size of a red blood cell.
These solutions have the capacity to dissolve up to 50
times more oxygen than plasma.
PFC solutions are modified hydrocarbons,however, they
do not mix well with blood and must be emulsified with
lipids or oils.
The best results are obtained if the patient is breathing
100% oxygen at the time of infusion(PaO2 ≥ 350 mm Hg).

15. Oxygent- The product is currently in phase III
clinical trials for use in cardiac and general surgical
patients
Fluosol (Japanese) - withdrawn c/o complement
activation & side effects
Pertorfan - licensed in Russia and Mexico

16. New Perfluoro compound !!!
POLYFLUORO-OCTOBROMIDE (PERFLUBRON)POLYFLUORO-OCTOBROMIDE (PERFLUBRON)
This new perfluoro compound, which is radioopaque,
has two advantages over its predecessors.
First, higher concentrations of perfluoro compound
may be administered, because a 100% (w/v) emulsion
with phospholipid has a sufficiently low viscosity to
be infused without dilution.
Second, oxygen is more soluble in polyfluoro-
octobromide than in anyother perfluoro compound
introduced to date

17. 5. Microencapsulated haemoglobin
 An alternative to polymerisation of haemoglobin to keep it for
longer in the circulation is to incorporate it in a lipid membrane or
liposomes which will act as “pseudoerythrocytes”
 This prevents the escape of the molecule through the glomeruli, and
the oxygen affinity of the solution containing the microspheres is
similar to that of whole blood;
 If pyridoxine 5-phosphate is added to the haemoglobin first a stable
product is created.
 The half life in the circulation is about 20 hours, which is similar
to that of a polymerised haemoglobin solution.

18. 6.6. Recombinant haemoglobin/ genetically engineeredRecombinant haemoglobin/ genetically engineered
haemoglobinhaemoglobin
ItIt is obtained by inserting the gene for human
hemoglobin into bacteria and then isolating the
hemoglobin from the culture.
This process allows for the manipulation of the gene itself
to create variant forms of hemoglobin.
One unit of hemoglobin solution can be produced from
750 L of Escherichia coli culture.
Optro - recombinant human Hb ( -linked)developed by
Baxter (discontinued)

19. Hemoglobin Based Blood Products:
Current Status
Hb Source TreatmentHb Source Treatment ProductProduct CompanyCompany
Human Hb, outdated blood, x-Human Hb, outdated blood, x-
linkedlinked
HemHem
AssistAssist
BaxterBaxter
Bovine Hb, heterogeneous MWBovine Hb, heterogeneous MW Hemo PureHemo Pure BiopureBiopure
Human Hb, outdated blood, x-Human Hb, outdated blood, x-
linked, polymerizedlinked, polymerized
HemolinkHemolink HemosolHemosol
Human Hb, outdated blood,Human Hb, outdated blood,
chemically polymerizedchemically polymerized
PolyHemePolyHeme NorthfieldNorthfield
Genetically engineered HbGenetically engineered Hb OptroOptro SomatogenSomatogen

20. Ideal red cell substitute
Easily picks up oxygen in the lungs
Easily delivers oxygen to the tissues
Does not need crossmatching
Long shelf life
Stable at room temperature

23. How Far How Near !!
Different materials are being used to encapsulate hemoglobin, one
being a biodegradable polymer called polyactide.
Incorporating nanotechnology : An example is the use of dendrimer
polymers to transport dissolved oxygen via intramolecular void spaces
and on its surface
Exploring other carriers: other oxygen carriers such as hemerythrin are
being explored-less prone to oxidative and nitrosative stress which
would alleviate problems like NO scavenging and methemoglobin
production
Applying HBOCs in other areas : cross-linking hemoglobin with
tyrosinase which can increase the efficacy of chemotherapy and
radiation therapy in tumor tissue

24. Unable to duplicate immunological and clotting
properties of blood
Hemopure is one of the farthest along
 Prior preparation not required
 Stable for three years at room temperature
 Reported compatible
with all blood
groups

25. TAKE HOME MESSAGE !!!
The red cell substitutes have far too short a survival time
in circulation ,therefore, it is much easier to envisage a
role in short term procedures such as immediate
resuscitation of military or civilian casualties.
Avoidance of allogeneic blood transfusion would not only
be beneficial for the individual patient, but if the practice
became widespread it would help relieve pressure on
transfusion services for ever increasing quantities of
allogenic blood

26. THANK
YOU !!