Design and Optimization of Trastuzumab-Functionalized Nanolipid Carriers for Targeted Capecitabine Delivery:...

Breast cancer remains a leading cause of cancer-related mortality in women globally. The main purpose of the research to develop, optimise and characterise a trastuzumab (TZ)-functionalized nanolipid carrier (NCs) encapsulating capecitabine, as a targeted strategy to breast cancer cells, to enhance therapeutic efficacy and reduce the severe side effects associated with conventional chemotherapy.

Capecitabine encapsulated NCs (CBNCs) were prepared by thin-film hydration technique, optimized by Box-Behnken design. The optimized formulation CBNCs were subsequently conjugated with TZ by using EDC-NHS chemistry. The prepared formulations of NCs were evaluated by FTIR, DSC, XRD, FESEM, TEM, AFM, drug loading, entrapment efficiency, average particle size, PDI, zeta potential, in vitro drug release. The successful surface conjugation of TZ was tested by BCA assay and SDS-PAGE analysis. In vitro targeting efficiency and cytotoxicity initially tested in MCF-7 cells (HER2-low expressing) and subsequently validated in SKBR3 cells (HER2-overexpressing) to confirm receptor-mediated uptake and specificity.

Optimized CBNCs were found spherical, nanosized (194.6 nm), with a zeta potential -25.55 mV for CBNCs, which increased to -57.76 mV upon TZ conjugation. The formulation showed 8.5% drug loading capacity and 84.26% drug release over 72 h. FTIR and DSC showed compatibility of drug and lipid components with no major shifting in characteristic peaks. TEM and AFM confirmed formation of stable, spherical discrete nanostructures. TZ conjugation showed minor alternation in average size/surface charge/morphology/texture. Successful TZ conjugation onto CBNCs was confirmed by BCA assay and SDS-PAGE. Fluorescence microscopy confirmed successful cellular internalization. MTT assay on SKBR3 cells demonstrated significantly higher cytotoxicity for TZ-CBNCs compared to CBNCs and free drug, thereby validating the HER2-specific targeting effect beyond preliminary results obtained in MCF-7 cells.

In view of the desired physicochemical properties, controlled drug release, and in vitro anticancer effectiveness, further in vivo investigations should be prioritized to validate its clinical application in HER2-positive breast cancer treatment. Nonetheless, the use of HER2-low MCF-7 cells in early assays highlights the importance of complementary validation in HER2-overexpressing models, as addressed by SKBR3 testing in this study.