Investigating the C2 Modulation of the Imidazo[1,2-a]pyrazine-Based Hit Compound CTN1122: Synthesis, in vitro Antileishmanial Activity, Cytotoxicity and Casein Kinase 1 Inhibition

Our research group previously discovered CTN1122, an imidazo[1,2-a]pyrazine compound with promising antileishmanial activity against intramacrophage amastigotes of Leishmania major and L. donovani strains. CTN1122 effectively targets Leishmania casein kinase 1 (L-CK1.2) and exhibits a favorable safety profile. To further explore its chemical space, we developed a convergent strategy to modify the C2 position of the imidazo[1,2-a]pyrazine core using Suzuki-Miyaura coupling of the corresponding triflate intermediate. Among 15 newly synthesized analogs, seven derivatives featuring variously substituted phenyl rings at C2 demonstrated L-CK1.2 inhibition within micromolar to submicromolar ranges and antileishmanial activity in vitro with low cytotoxicity in macrophages. Compounds 7 d and 7 l were particularly potent, with IC₅₀ values of 1.25 μM and 0.92 μM against L. major, and 1.44 μM and 2.34 μM against L. donovani, respectively. They showed IC₅₀ L-CK1.2=0.30 μM and 0.57 μM with enhanced selectivity indices (SI=3.8 and 1.6) over the human CK1ϵ ortholog. Additionally, four C2 analogs and two C5 isomers exhibited notable antiparasitic effects without strongly inhibiting L-CK1.2, indicating a possible alternative mechanism of action. Compound 7 k displayed the highest general activity, with IC₅₀ values of 0.31 μM on L. major and 0.27 μM on L. donovani, coupled with favorable selectivity indexes.