DESIGN AND DEVELOPMENT OF POTENTIAL ANTISICKLING AGENTS

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The long range goals of this research are to discover a potential therapeutic agent to treat Sickle Cell anemia, develop allosteric inhibitors of hemoglobin for ischemic diseases and for radiation oncology, and to advance the fundamental understanding of small molecule - large molecule interactions. A combination of research fields will continue to be employed in our studies with extensive use of X-ray crystallographic and molecular modeling techniques to discover lead molecules and to elucidate mechanisms of action of active compounds at the molecular level. The corresponding molecules will be synthesized and solution and cellular studies will be conducted to evaluate the in vitro activity and potential toxicity problems of new molecules. We have advanced one compound from basic laboratory studies to the clinic testing phase with an IND from the FDA. Our specific aims include the: synthesis and testing of pro-drug like analogues of antisickling agents; synthesis and testing of other aromatic aldehydes and derivatives of non-toxic starting materials or natural products as potential antisickling agents; design, synthesize and testing of new allosteric modulators of hemoglobin; determination of the deoxy- incubated Hb binding sites for the antisickling drug 12C79 (the first therapeutic agent designed de novo from the 3D structure of hemoglobin to reach clinical trials) and solution binding measurements; determination of the solution binding constants of active antisickling agents and four classes of hemoglobin allosteric effectors; determination of the effects of antisickling agents and allosteric inhibitors on the oxygen dissociation curve of hemoglobin solutions; comparison of the solution binding constants with the P50 values for allosteric inhibitors to ascertain whether the intrinsic activity can be observed with a protein system; determination of the binding sites and interactions of active molecules crystallographically; evaluation of the binding sites for positive and negative polar and hydrophobic interactions using the new computer software (HINT) developed during the previous proposal; further development of the HINT software; measurement of the rheological effect of active compounds on erythrocytes; evaluation of allosteric inhibitors an anti-ischemic agents in a variety of in vitro and in vivo systems and measurement of blood levels and the % hemoglobin reaction in human subjects.