A Niclosamide Prodrug SSL-0024 with Enhanced Bioavailability Suppresses Hepatocellular Carcinoma via Multi-Pathway Signaling Inhibition

BackgroundHepatocellular carcinoma (HCC) remains a leading cause of cancer-related mortality worldwide, highlighting the urgent need for effective therapies. Niclosamide, an FDA-approved anthelmintic, reverses HCC gene expression profile to that of normal hepatocytes, and exhibits promising anti-tumor activity in HCC in vitro; however, its clinical translation is limited by poor aqueous solubility, low bioavailability, and short systemic exposure, resulting in lack of in vivo activity. We previously used an established phosphate prodrug approach to provide proof-of-concept that increasing oral bioavailability was essential for niclosamide to achieve in vivo anti-tumor activity. MethodsWe designed a panel of novel niclosamide prodrugs and screened eight candidates for water solubility, chemical stability, and in vitro anti-proliferative activity in HCC cell lines. The lead compound, SSL-0024, was further evaluated for its pharmacokinetics and anti-tumor efficacy in immunodeficient mice bearing orthotopic HCC patient-derived xenografts (PDX). Mechanisms underlying its observed activity were assessed through protein-level analysis of AKT-mTOR-STAT3, RAF, Wnt/{beta}-catenin signaling pathways, vasorin-associated pathways, and PD-L1. ResultsSSL-0024 demonstrated markedly improved aqueous solubility and stability in gastric and plasma conditions, supporting oral administration. Pharmacokinetic analyses revealed a plasma half-life of [~]24 hours, dramatically extended relative to native niclosamide. Once daily oral administration of SSL-0024 (100 mg/kg) in orthotopic HCC PDX mice achieved [~]60% tumor growth inhibition at only [~]46.8% of the dose required for the positive control (niclosamide ethanolamine), with minimal systemic toxicity. Mechanistically, SSL-0024 concurrently suppressed AKT-mTOR-STAT3 signaling, RAF kinases, Wnt, and VASN-associated pathways, with additional downregulation of PD-L1, resulting in reduced proliferation, survival, and immune-evasion signaling. ConclusionThrough rational design and systematic screening, we have identified a lead niclosamide prodrug candidate, SSL-0024, which exhibited improved water solubility and stability, extended plasma half-life, enhanced oral bioavailability, and preservation of biological activity in vitro and in vivo. Future studies will include combination therapy with standard-of-care treatments, as well as safety and formulation studies to enable its clinical translation for the treatment of HCC and other solid tumors impacted by the multiple oncogenic pathways modulated by niclosamide.