Design and synthesis of tenofovir monophenyl ester phosphonoamidate prodrugs with improved anti-hepatitis B virus activity and intrahepatic tenofovir enrichment.

Tenofovir Alafenamide (TAF, formerly known as GS-7340, Fig. 1-C) is a prodrug of Tenofovir (TFV) that contains a phosphonoamidate group. Compared to Tenofovir Disoproxil Fumarate (TDF), TAF exhibits enhanced stability in non-target tissues, promoting selective accumulation of Tenofovir (TFV) in target cells (e.g., PBMCs and hepatocytes). Subsequent phosphorylation convert TFV to its active form, Tenofovir Diphosphate (TFV-DP), thereby potentiating therapeutic efficacy. However, detectable levels of TFV in whole blood following TAF administration underscore potential safety risks, highlighting the need for novel TFV prodrugs with improved whole-blood stability. However, detectable levels of TFV in whole blood following TAF administration underscore potential safety risks, highlighting the need for novel TFV prodrugs with enhanced whole-blood stability and therapeutic efficacy. In this study, a series of TFV phosphonoamidate prodrugs incorporating disubstituted amino acids were designed and synthesized. Compared with TAF, compound 3a1 exhibited exceptional stability, with no detectable metabolite-TFV found in human whole blood. Furthermore, PBMC-based TFV quantification was employed as a clinically relevant measure of intracellular prodrug delivery and metabolic activation, consistent with the FDA's evaluation framework for tenofovir alafenamide in HBV-infected patients. Under this framework, compound 3a1 demonstrated significantly enhanced intracellular TFV accumulation in PBMCs compared with TAF. Collectively, these results position 3a1 as a promising next-generation TFV prodrug with potential for improved efficacy and safety.