Controlled gene/drug delivery via triggerable liposomes for triple negative breast cancer treatment

Abstract

Triple negative breast cancer (TNBC) is a highly aggressive breast cancers with highest mortality rate and poorest outcomes. The challenges of TNBC treatment are rooted in lack of estrogen receptors (ER), progesterone receptors (PR), and human epidermal growth factor 2 (HER2), resulting endocrine therapy or receptors targeted therapies are not effective against TNBC. Clustered regularly interspaced short palindromic repeat-associated protein 9 (CRISPR/Cas9)-based gene therapy is a revolutionary strategy for TNBC treatment, while its clinical applications are limited by lack of safe and efficient non-viral delivery system. In this thesis, we developed light triggerable liposomes mediated CRISPR/Cas9 delivery for TNBC treatment. CRISPR/Cas9 was loaded inside liposomes with encapsulation efficiency higher than 70%. FDA-approved photosensitiser (PS) verteporfin was used to produce reactive oxygen species (ROS) when it was activated by external light, leading to liposome membrane destruction and thus release of CRISPR/Cas9 in cells. The knockdown of A20 gene-Exon3 by CRISPR/Cas9 in TNBC cells significantly supressed the cell proliferation and migration in vitro. CRISPR/Cas9-based gene editing via light triggerable liposomes with spatial and temporal control provides a promising option for TNBC treatment.

The application of triggerable liposomes can be extended to deep tissue therapy by using X-ray as external stimuli. PS in liposomes not only works for liposome rupture, but also serves as photodynamic therapy (PDT) reagent to induce cell death through extensive ROS generation. X‑ray induced photodynamic therapy (X-PDT) via protoporphyrin IX (PPIX) & perfluorooctyl bromide (PFOB) co-loaded liposomes for in vitro TNBC treatment was developed. Relief of hypoxia in cancer cells by addition of PFOB improved ROS production with 4.9-fold of PPIX under 2Gy radiation and it turned out to induce 17% increase in cell death through either apoptosis or necrosis pathway.

These findings in this thesis suggest that developed intelligent triggerable liposomes are superior to deliver CRISPR/Cas9 or photosensitisers efficiently for TNBC treatment. Future works were prospected to combine CRISPR/Cas9 gene editing with X-PDT via triggerable liposomes to achieve precise synthetic therapy.