A Cyclic Phosphoramidate Prodrug of 2’-deoxy-2’-fluoro-2’-C-methylguanosine for the Treatment of Dengue Infection

Abstract

47Monophosphate prodrug analogs of 2'-deoxy-2'-fluoro-2'-C-methylguanosine have been 48 reported as potent inhibitors of hepatitis C virus (HCV) RNA-dependent RNA polymerase. These 49 prodrugs also display potent anti-dengue activities in cellular assays although their prodrug 50 moieties were designed to produce high levels of triphosphate in the liver. Since peripheral blood 51 mononuclear cells (PBMCs) are one of the major targets of dengue virus, different prodrug 52 moieties were designed to effectively deliver 2'-deoxy-2'-fluoro-2'-C-methylguanosine 53 monophosphate prodrugs and their corresponding triphosphates into PBMCs after oral 54 administration. We identified a cyclic phosphoramidate prodrug 17 demonstrating a well-balanced 55 anti-dengue cellular activity and in vitro stability profiles. In dogs, oral administration of 17 56 resulted in high PBMC triphosphate level, exceeding TP50 (the intracellular triphosphate 57 concentration at which 50% of virus replication is inhibited) at 10 mg/kg. Compound 17 58 demonstrated 1.6-and 2.2 log viremia reduction in the dengue mouse model at 100 and 300 mg/kg 59 twice daily, respectively. At 100 mg/kg twice daily, the terminal triphosphate concentration in 60 PBMCs reached above TP50, defining for the first time the minimum efficacious dose for a 61 nucleos(t)ide prodrug. In the two-week dog toxicity studies at 30 to 300 mg/kg/day, no observed 62 adverse effect level (NOAEL) could not be achieved due to pulmonary inflammation and 63 hemorrhage. The preclinical safety results suspended further development of 17. Nevertheless, 64 present work has proven the concept that an efficacious monophosphate nucleoside prodrug could 65 be developed for the potential treatment of dengue infection. 67The mosquito-borne dengue virus is endemic to tropical and sub-tropical regions 68 throughout the world, making dengue fever the most important mosquito-borne viral disease 69 afflicting humans. Its global distribution is comparable to that of malaria, with an estimated 2.5 70 billion people at risk for epidemic transmission (1). There has been steady increases in countries 71 affected and incidence since the 1950s and recent estimates suggest annual rates of 390 million 72 cases accompanied by 20,000 deaths (2). 73 Dengue viruses (DENVs) can be further classified into four different serotypes (DENV-1 74 to -4), all of which can lead to disease symptoms with varying severity. Secondary infection by a 75 different serotype may increase the risk of severe dengue diseases. While diagnosis of dengue 76 infection can be rapid and simple, serotype distinction requires additional instrumentation, usually 77 in a laboratory setting. Thus, the ideal treatment for dengue fever should possess pan-serotype 78 activities (3). Recently, a dengue vaccine was approved in certain countries but is recommended 79 only for individuals with prior DENV exposure. This limits its use as well as necessitating pre-80 vaccination screening (4). No antivirals are currently available for the treatment of d...