Thioester Linked Cationic Lipopolymers for co-delivery of TRAIL plasmid and its complementary siRNA targets: One Stone Two Bird Approach for Cancer Therapy

Thapa, Bindu (Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta)
KC, Remant (Department of Chemical and Material Engineering, University of Alberta)
Uludag, Hasan (Department of Chemical and Material Engineering, University of Alberta)

Introduction

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in variety of cancer cells without affecting normal cells, making it as a potential agent for cancer therapy. In spite of promising preclinical results, TRAIL protein therapy tested so far failed to exert robust anticancer activity in patients due to resistance induction in malignant cells and its rapid renal clearance. Hence, we aim (i) to use TRAIL plasmid as promising alternative since it produces TRAIL protein at the site of action in higher concentration, (ii) to identify novel small interfering RNA (siRNA) targets, which sensitize breast cancer cells against TRAIL, and (iii) to co-deliver TRAIL plasmid and identified complementary siRNA targets.  Both plasmid DNA and siRNA require effective delivery vehicles. Moreover, co-delivery is more complicated to achieve sufficient outcome for a robust effect. We, herein, proposed thioester linked cationic lipopolymers, which display sufficient binding and delivery capacity of individual as well as cocktails of these therapeutics agents.

Materials and Methods

To identify novel siRNA targets that sensitize breast cancer cells against TRAIL, a siRNA library against 446 human apoptosis-related proteins were screened in triple negative breast cancer MDA-MB-231 cells in presence or absence of TRAIL protein using cationic lipopolymers as siRNA delivery agent. A library of cationic lipopolymers (PEI-L) was prepared by rational grafting of aliphatic lipids (L) onto small molecular weight (MW: 0.6, 1.2 and 1.8 kDa) polyethyleneimine (PEI) with amide or thioester bond.  Efficiency of this library to deliver   individual and cocktail nucleic acid therapeutics was explored and potential polymer was identified.

Results

Based on growth inhibition of MDA-MB-231 cells, sixteen siRNAs were found to sensitize TRAIL-induced cell death. Among them, novel and the most promising targets BCL2L12 and SOD1 were further evaluated. Silencing of both these targets significantly sensitized TRAIL-induced cell death in MDA-MB-231 cells and TRAIL-resistant MCF-7 cells. Among the polymers, thioester linked polymer resulted higher DNA transfection efficiency while employed in co-delivery of DNA/siRNA cocktail. A range of other polymers was found effective in delivery of individual molecules, but not in co-delivery simultaneously. We were able to optimize the proper composition of plasmid DNA:siRNA as well as polymer/nucleic ratio to generate maximum effect using these particular polymers. We further compare the efficacy between co-delivery and separate delivery, and the outcome of co-delivery was significantly higher than the delivery of separate complexes.

Discussion and Conclusion

Co-delivery of plasmid DNA and siRNA was feasible to include complementary therapeutic functionalities into a single carrier designed using thioester composed cationic lipopolymers. It maximized the therapeutic benefit via enhancing each other’s transfection efficiency and generating synergistic effects. The therapeutic benefit by dual delivery of TRAIL plasmid and its complementary siRNA targets with single carrier provided a more effective way to treat breast cancer in vitro.

Acknowledgements

Bindu Thapa was supported by Alberta Innovates Graduate Studentship

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