Synthesis and Biological Evaluation of Small Molecule Inhibitors of Immune Cytopenias.

Immune cytopenias are a group of autoimmune disorders where patients develop autoantibodies against certain types of blood cells such as red blood cells (RBCs) or thrombocytes. We investigated small molecules as potential inhibitors of phagocytosis of blood cells that are prevalent in immune thrombocytopenia (ITP) and warm autoantibody immune hemolytic anemia (wAIHA). Upon screening a chemical library of over 13,000 compounds in silico, followed by evaluating 80 compounds in vitro as inhibitors of phagocytosis of opsonized RBCs by monocytes, we identified four hit molecules. These compounds contain a pyrazole moiety as a key structural feature. Here, we reveal the independent synthesis and re-evaluation of these hits, as well as revalidate the biological activities and the synthesis of their analogs to understand the structure-activity relationships. Two of the resynthesized compounds showed up to a 9-fold difference in their inhibitory activities between the commercial and synthesized batches, and the analogs exhibited either equal or weaker potency than the parent compounds targeting phagocytosis of RBCs. The role of regioisomers and the importance of an ester moiety are revealed as important structural features through these analogs. The pharmacokinetics of the promising compound suggested that this compound shows significant efficacy in restoring platelet counts in the mouse model of ITP, despite the rapid hydrolysis of its methyl ester moiety.