Investigating Fn14 as a cell surface portal for targeted nanotherapeutic delivery to triple-negative breast cancer (TNBC) cells

Abstract

Metastatic triple-negative breast cancer (TNBC) is associated with a dismal prognosis due to a lack of known therapeutic targets and several difficulties in treating disseminated disease. As a result, patients with this diagnosis have an average median survival time of just 13 months. Treatment challenges are further exacerbated in the 30-55% of metastatic TNBC patients that develop brain metastases (BMs), where therapeutic agent efficacy is further limited by the blood-brain barrier. Nanotherapeutics may be especially poised for overcoming such treatment barriers, particularly if specifically targeted to TNBC cells within the brain lesions. The TWEAK receptor, Fn14, is minimally expressed by healthy breast and brain tissues but overexpressed in primary TNBC tumors and TNBC BMs. We previously demonstrated that Fn14-targeted, paclitaxel-loaded polymeric nanoparticles (NPs), termed ‘DARTs’, outperform Abraxane—an FDA-approved paclitaxel nanoformulation—following intravenous delivery in mouse models of primary TNBC and TNBC BM. In Part 1 of this work, we investigated the impact of using different Fn14 targeting moieties and moiety surface densities on NP binding affinity and uptake by Fn14-expressing tumor cells. Specifically, we found that DARTs functionalized with ~13 ITEM4 monoclonal antibodies, which are specific for Fn14, exhibit significantly greater uptake by TNBC cells compared to NPs with other ITEM4 surface densities or those with ITEM4 fragment antigen binding (Fab) regions for targeting. In addition, we determined that DARTs are predominantly internalized via clathrin-mediated endocytosis and traffic via the endolysosomal pathway. In Part 2 of this project, we further examined the utility of this formulation as a drug delivery platform in complementary xenograft and syngeneic models of TNBC BM. First, we determined that tumor cells were the predominant source of Fn14 expression in the TNBC brain tumor-immune microenvironment, with minimal Fn14 expression by microglia, infiltrating macrophages, monocytes, or lymphocytes. We then showed that although DARTs, non-targeted PLGA-PEG NPs and Abraxane exhibit similar accumulation in brains harboring TNBC BMs following systemic delivery, only DART NPs specifically associated with TNBC cells. Collectively, these findings deepen our understanding of the potential use of the DART NP formulation for drug delivery to TNBC patients with BMs.