Studies on the mode of action of two antisickling agents, glyceraldehyde and methyl acetyl phosphate, will be continued from three perspectives. Studies aimed at the chemical mechanism of the reaction of each of these compounds with hemoglobin S will be pursued. With glyceraldehyde we are especially interested in uncovering the mechanism of the Amadori rearrangement. For this purpose we will be studying model peptide systems in which we have already separated the intermediate aldimine and product ketoamine by mild chromatographic procedures. The stability of each of these adducts will be evaluated under a variety of conditions. For methyl acetyl phosphate we will continue to study the reason for the lowered oxygen affinity when this acetylating agent reacts with hemoglobin S in the deoxy state. Studies on cell density and oxygen affinity will also be performed. The second part of these studies will evaluate the effects of each of these compounds on intact sickled cells. This is made possible because each compound can readily enter the sickle cell and react with intracellular hemoglobin. We plan to study the kinetics of deformability of the sickle cell after treatment with either glyceraldehyde or methyl acetyl phosphate. Studies on cell density using the phthalate ester method will also be performed, as well as studies on the viscosity of suspensions of sickled cells. Studies on hemoglobin biosynthesis in a reticulocyte system will be performed in the presence of glyceraldehyde or methyl acetyl phosphate. The third aspect of these studies will involve preclinical studies. For either glyceraldehyde or methyl acetyl phosphate, studies in animals will be carried out to determine how much of a given dose administered by various routes will be necessary to achieve the desirable amount of reaction with circulating red cells in vivo. In humans, studies on glyceraldehyde will be pursued in order to determine whether or not there is any new immunological determinant which is conferred upon the cells after the treatment with glyceraldehyde. For these purposes we will perform Coombs tests as well as a third chromate survival time in vivo. Finally, we will continue to assist Dr. Samuel Charache of Johns Hopkins in his studies on the treatment of sickle cells with glyceraldehyde.
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