Prolonged storage of red blood cells

Details Prolonged storage of red blood cells Prolonged storage of red blood cells

2000-09-19

2002-09-10

Saucier, Sandra E. (Department: 1651)

Chemistry: molecular biology and microbiology

Maintaining blood or sperm in a physiologically active state...

Reexamination Certificate

active

06447987

ABSTRACT:

BACKGROUND OF THE INVENTION
1. Field of Invention
The invention generally relates to methods and materials associated with the storage of whole blood and red blood cells (RBC).
2. Description of Related Art
Whole blood storage was first demonstrated by Rous & Turner in 1916 and Robertson in 1917. Acid-citrate-dextrose (ACD, 1943) and Citrate-phosphate-dextrose solution (CPD, 1957) were subsequently approved for 21-day storage of whole blood. CPD with adenine (CPDA-1, 1979) was later introduced and used for extending the shelf-life of stored whole blood and packed RBC for up to 5 weeks. Additive Solutions extended RBC storage to 6 weeks in 1981. Red blood cells (RBCs) stored in these solutions have shown steady deterioration after about 6 weeks as determined by the inability of 75% of such cells to survive in the circulation for 24 hours after reinfusion back into the human donor. It has been observed that during continued refrigerated storage, glucose is consumed at a decreasing rate, as the concentration of metabolic waste, i.e. lactic acid and hydrogen ions, increases. Such a decrease in the rate of glucose metabolism leads to depletion of adenosine triphosphate (ATP) which directly correlates to the recovery of RBCs when the cells are returned to the circulation.
The development of additive solutions for the preservation of red blood cells (RBCs) after their separation from whole blood has allowed the design of formulations which are specifically tailored to the needs of RBCs. Additive solutions such as Adsol® (AS-1), Nutricel® (AS-3), Optisol® (AS-5), and ErythroSol® were designed to extend the storage of RBCs at 1-6° C.
Almost all of the whole blood collected now is made into components, and the RBC fraction is stored as packed RBCs. For blood drawn into the additive solution systems, RBCs are packed by centrifugation, plasma is removed so that RBCs make up 80% of the volume, and then 100 ml of additive solution is added sterilely. The resulting suspensions have a RBC volume fraction of approximately 55%. RBCs stored in the conventional FDA-approved additive solutions can be stored for only 6 weeks with an acceptable 24-hour in vivo recovery.
To increase the time of acceptable in vivo recovery of RBCs in liquid storage, attempts have been made to improve additive solutions and storage processes. In “Studies In Red Blood Cell Preservation-7. In vivo and in vitro Studies With A Modified Phosphate-Ammonium Additive Solution,” by Greenwalt et al.,
Vox. Sang.
65:87-94 (1993), the authors determined that the experimental additive solution (EAS-2) containing in mM: 20 NH
4
Cl, 30 Na
2
HPO
4
, 2 adenine, 110 dextrose, 55 mannitol, pH 7.15, is useful in extending the storage shelf-life of human RBCs from the current standard of 5-6 weeks to an improved standard of 8-9 weeks. However, packed RBCs stored in the medium were not directly infusible but required the removal of the supernatant with a washing step prior to transfusion due to the presence of ammonium in the additive solution.
In “Studies in Red Blood Cell Preservation-8. Liquid Storage of Red Cells in Glycerol-Containing Additive Solution,”
Vox. Sang.
67:139-143 (1994), Greenwalt et al. described an Experimental Additive Solution 25 (EAS-25) that allowed 73 percent recovery of packed red cells at nine weeks. However, the resulting RBC units contained about 1 percent glycerol and thus, are not safe for transfusion in humans in massive amounts.
In “Extending the Storage of Red Cells at 4° C.,”
Transfus. Sci.
15:105-115 (1994) by Meryman et al., acceptable viability of RBCs stored in very dilute suspensions at low hematocrit for as long as 27 weeks were demonstrated. However, such stored RBC suspensions were not acceptable for direct infusion due to their high content of potassium and ammonia and their low volume fraction of RBCs.
Consequently, there remains a need for improved additive solutions and processes which increase the storage time of human RBCs over that of conventional solutions and processes while allowing the RBC storage suspension to be directly tranfusable into humans and maintaining an acceptable in vivo recovery of RBCs.
SUMMARY OF THE INVENTION
The present invention relates to a novel additive solution useful for the storage of human RBCs under refrigerated conditions using an additive solution to preserve RBCs at about 1 to 6° C. for up to about 11 weeks or more.
Additive solutions and processes in accordance with the present invention allow the viable storage of human RBCs for an extended period of time in a solution which is directly infusible in humans.
It is, therefore, an object of the present invention to provide an additive solution for storage of human RBCs which solution substantially increases the storage time of the RBCs at about 1 to about 6° C. while maintaining an acceptable recovery of the RBCs.
It is also an object of the present invention to provide an additive solution for storage of human RBCs which is physiologically safe and suitable for direct infusion into humans in massive amounts.
It is yet another object of the present invention to provide a method of storing human RBCs for about 11 weeks or more at about 1 to about 6° C. with an acceptable 24-hour in vivo fractional recovery of the RBCs.
It is also another object of the present invention to provide novel RBC storage suspensions which are directly infusible into humans following about 11 weeks or more storage at about 1 to about 6° C.
To achieve the foregoing and other objects in accordance with the purposes of the present invention, a novel additive solution for preserving RBCs was developed. The aqueous solution contains adenine, dextrose, Na
2
HPO
4
, mannitol, and at least one physiologically acceptable sodium salt in amounts sufficient to preserve RBCs which amount includes a buffering amount of sodium bicarbonate and or trisodium citrate to maintain the pH at or above about 8, preferably at about 8.4.
The additive solutions are useful in a method for storing RBCs, which method include the steps of:
(a) mixing a sample of whole blood containing the RBCs and plasma with an anticoagulant solution, forming thereby a suspension of whole blood;
(b) treating the whole blood suspension to separate the RBCs from the plasma, forming thereby packed RBCs;
(c) mixing the packed RBCs with an appropriate amount of an additive solution in accordance with the invention thereby forming a suspension of RBCs;
(d) cooling said suspension of RBCs to about 1 to about 6° C.; and
(e) storing said cooled suspension of RBCs according to standard blood bank procedures for a period of about 11 weeks or more.
RBC suspensions produced in accordance with the invention after about 11 weeks or more of storage provide a sufficiently therapeutic amount of recoverable RBCs and are directly infusible into humans without further processing in accordance with known standards established for transfusion of RBCs.


REFERENCES:
patent: 5789151 (1998-08-01), Bitensky et al.
patent: 5906915 (1999-05-01), Payrat et al.
patent: 6150085 (2000-11-01), Hess et al.
Mazor et al., “Prolonged Storage of Red Cells: The Effect of pH, Adenine and Phosphate”, Vox Sang. 66:264-269 (1994).

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