Metformin-encapsulating immunoliposomes conjugated with anti-TROP 2 antibody fragments for the active targeting of triple-negative breast cancer

Abstract

Trophoblast cell-surface antigen 2 (TROP 2) has re-emerged as a promising biomarker in triple-negative breast cancer (TNBC), with high overexpression in many TNBC cases. However, despite its potential and approval as an antibody–drug-conjugate for TNBC treatment, TROP 2-targeted delivery systems are currently underexplored. Therefore, this study was aimed at exploiting the potential of TROP 2 targeting by encapsulating metformin (Met), an antidiabetic drug associated with tumor growth inhibitory properties, inside liposomes decorated with TROP 2-targeting single-chain variable fragments (scFvs). The optimization of scFv grafting resulted in Met-immunoliposomes with an average diameter of less than 200 nm, low polydispersity index (∼0.1), negative surface charge (<−10 mV), high Met drug loading (>150 mg g⁻¹), and high affinity towards TROP 2 binding. Furthermore, Met-immunoliposomes were reproducible, and the scFv conjugation was stable in the presence of serum for five days. Their cellular uptake increased 4 folds in two-dimensional and 9 folds in three-dimensional TNBC models owing to the high affinity towards TROP 2 binding. Finally, it was observed that the therapeutic effect of Met in suppressing cancer cell growth and proliferation was superior when using anti-TROP 2 scFv-grafted Met-immunoliposomes, which completely stopped the spheroid growth and inhibited the expression of adenosine triphosphate. This study is one of the first reports to explore the combination of nanoparticle-based drug delivery systems to target the TROP 2 protein in TNBC, and to the best of our knowledge, this is the first report to specifically combine the use of scFvs with TROP 2 targeting to deliver therapeutics for TNBC treatment.