Project Summary Hit-to-Lead Development of the Kalihinol Scaffold for Malaria Treatment The ultimate goal of this collaborative research program is to identify antimalarial clinical candidates among analogues of the kalihinol family of isocyanoterpenes, an understudied class of natural products with potent activity against Plasmodium falciparum, the causative agent of the deadliest form of human malaria. Drug resistance remains the leading factor hampering global efforts aimed at controlling malaria infection, lowering mortality rates and reducing the cost of treatment. Countering malaria drug resistance requires development of novel classes of chemicals not previously used in malaria therapy, and implementation of novel therapeutic strategies for optimal use of these chemicals to prevent drug resistance. Preliminary data generated in our laboratories support the premise of this research that the kalihinols could be developed as novel antimalarial agents. Our data demonstrate that (i) kalihinol natural products have potent activity against blood stages of both drug-sensitive and drug-resistant P. falciparum strains with IC50 values in the low nanomolar range, (ii) these compounds are amenable to rapid and simplified synthesis routes producing analogues that retain potent antimalarial activity, (iii) they are safe, with high therapeutic indices, (iv) their isonitrile functional groups are relatively stable to metabolism, and (v) they may exert their antimalarial activity through a novel mode of action. Building upon this body of data, we propose to delve deeply into the structure-activity relationship of these compounds, characterize their in vitro and in vivo efficacy and safety, and unravel their mode of action. In Aim 1, we will further characterize the biological activity and pharmacological properties of lead kalihinol analogues already in hand, including their ability to inhibit growth of drug-sensitive and drug-resistant malaria parasites within human red blood cells, to block sexual differentiation and transmission to mosquitoes, and to eliminate lethal malaria infection in mice. In Aim 2, we will embrace a general chemical synthesis design that permits access to many diverse kalihinol-type compounds, with the goal of optimizing potency and pharmacological properties. Compounds with excellent potency, selectivity and safety profiles will be further evaluated in vivo for efficacy and safety. In Aim 3. both the mode of action of the drugs and the parasite's possible mechanisms of resistance against them will be further elucidated using state-of-the-art cellular, metabolic, chemical biology and genomics approaches. This collaborative and multidisciplinary project is of relevance to human health because of its potential to produce new preclinical antimalarial leads based on a novel class of chemicals never before used in malaria therapy.