J Med Chem. 2009, 52(19), 5793-802. Discovery of novel antileishmanial agents in an attempt to synthesize aplysinopsin pentamidine hybrid molecule Sharad Porwal, † Shikha Chauhan ,Prem M. S. Chauhan,* † Nishi Shakya, § Aditya Verma, § Suman Gupta § †Division of Medicinal & Process Chemistry, Central Drug Research Institute, Lucknow, 226001, India, and § Division of Parasitology, Central Drug Research Institute, Lucknow, 226001, India In an attempt to synthesize pentamidine aplysinopsin hybrid molecule (A), a lead molecule 8 (containing Z- configured aplysinopsin moiety) was identified for antileishmanial activity. Optimization of lead 8 provided 24 (containing E- configured aplysinopsin) possessing 10 times more activity and 450 fold less toxicity than drug pentamidine. Synthesis of 24 was possible, surprisingly, due to two innate reactivity of indole-3-carbaldehyde which provided it in diastereo and regioselectively pure form without recourse to protection deprotection and long reaction pathway. Leishmaniasis is a vector born parasitic disease of tropics and subtropics which is manifested in four major clinical forms (cutaneous leishmaniasis, mucocutaneous leishmaniasis, visceral leishmaniasis and post kala-azar dermal leishmaniasis or PKDL) depending on causative species of protozoan Leishmania. In all above forms of leishmaniasis, visceral leishmanaiasis (VL) is lethal, if left untreated. There are ≈ 70 000 deaths and 1.5 million new cases emerges per year due to VL. Majority of VL cases (≥ 90%) occurs in just six countries India, Nepal, Bangladesh, Sudan, Ethiopia and Brazil. Situation has become complicated by emergence of PKDL which appears in 0-6 months after successful cure of VL. 1 The WHO has declared VL a neglected and emerging disease. 2 Antimonials are first line treatment options for VL, which was discovered almost 70 years ago. These suffer from major side effects including cardiac arrhythmia, pancreatitis. Despite their toxicity treatment failure to antimonials treatment has increased; some times as high as 62% in some of the regions. 1 Second line treatment options for VL are pentamidine, miltefosin and amphotericin B. However, all of these drugs suffer from several moderate to sever side effects. Pentamidine, an aromatic diamidine, is not active orally and can lead to renal, pancreatic, and hepatic toxicity along with hypotension and dysglycemia. 3 Miltefosine, an alkylphosphocholine, has long half life (100-200 h) in humans and low therapeutic ratio, characteristics that could encourage development of resistance. It is not suitable for pregnancy due to tetratogenecity and also cause mild to severe gastrointestinal side effect. 4 Liposomal amphotericin B is highly effective option; however this drug formulation is very expensive (US$ 2800 per treatment), limiting its use in most endemic regions. Although recent clinical trial with injectable paromomycin have shown encouraging results, an expanded catalogue of new drugs for VL causing parasite L. donovani is required to tackle the problem of resistance. In spite of some side effects of pentamidine, the broad range of its (dicationic class of molecules in general) biological activities, 5 relatively less propensity towards development of resistance (resistance to pentamidine itself has never been a significant problem in the field, despite its widespread use as a prophylactic) 6 and our previous work on this class on molecules 7 prompted us to develop some novel dicationic class of molecules as potential antileishmanial agents with improved efficacy and selectivity than pentamidine. From literature search we found that aplysinopsins (a class of natural products possessing cyclic guanidine function) 8 acts on similar biological targets (plasmepsin II and serotonin receptors) as dicationic class of molecules. 9 Taking inspiration from fragment based drug discovery (FBDD) approach, 10, 11 we designed a hybrid molecule A (Fig. 1), where one amidinophenoxy function of pentamidine has been replaced with aplysinopsin. Due to their synthesis in biological settings small molecule natural products have in built selectivity and phamacokinetic profile, necessary for a drug molecule. 12 Hence, incorporating a drug fragment with natural product may provide molecules with better activity and less toxicity. 13, 14 O HN NH 2 O NH NH 2 N H N N O NH Pentamidine Aplysinopsin N N N O NH O HN H 2 N Pentamidine-aplysinopsin hybride, A Figure 1: Hybrid of pentamidine and aplysinopsin Herein, we describe our efforts for synthesis of hybrid molecule A and its analogues and discovery of new class of antileishmanial in this endeavor. To synthesize compound A and its analogues we adopted route depicted in scheme 1. Direct synthesis of aplysinposin and its subsequent coupling with pentamidine fragment wasn’t feasible due to poor yield in synthesis of aplysinopsin and lack of chemo selectivity in its subsequent coupling with petamidine fragment [mainly p-(hydroxypentyl)benzamidine]. As we proceeded according to Scheme 1, we encountered a problem in direct condensation of 1-methyl-2-imino- 4-imidazolidinon (creatinine) which occurred in poor yield and provided a mixture of diastereomers (E- & Z-). Therefore, we first condensed 2-thio-4- imidazolidinon (2-thiohydantoin) with intermediate I.1 to provide compounds 8, 13, 14 in its Z- geometry. Conversion of cyano function of comp. 8 to amidine (Pinner synthesis) again proved difficult due to its high insolubility and many NH centers. We tried a novel approach for the conversion of cyano function to amidine; by firstcomplexing it with Zn 2+ (using