Effective and cancer-specific siRNA delivery to human hepatocellular carcinoma cells mediated by poly(beta-amino ester) nanoparticles

摘要

Introduction: Liver cancer is the second leading cause of cancer mortality worldwide. Hepatocellular carcinoma (HCC) accounts for approximately 90% of primary liver cancers and has a five year post-diagnosis survival rate of only 12%. Limited efficacy of the current treatment options is one of the most challenging problems existing in the field of liver oncology. Gene therapy offers promising advantages that might overcome these limitations. Unfortunately, the poor selectivity in vector targeting still represents a main obstacle in gene therapy. Here we demonstrate that bioreducible poly(beta-amino ester)s (PBAEs) nanoparticles suggest an effective and cancer-specific siRNA delivery strategy to target human HCC cells. Materials and Methods: A human liver epithelial cell line (THLE-3) and a human hepatocellular carcinoma cell line (HepG2) were separately cultured and treated with nanoparticles formed by PBAEs and either AllStars Hs Cell Death siRNA (death siRNA) or AllStars Negative Control siRNA (Qiagen, Netherlands). While the death siRNA consists of a blend of siRNAs that silence important genes related to cell survival, the negative control has no target within mammalian cells. Effective delivery of death siRNA leads to cell death and the negative control allows evaluation of non-specific cell toxicity. Two bioreducible PBAEs, that we synthesized and evaluated at five different concentrations, were used as vectors to carry either death siRNA or negative control siRNA. Three days post-treatment, DAPI/PI staining was applied to differentiate live from dead cells. Live cells were counted using the Image J software. Results and Discussion: Our bioreducible PBAE nanoparticles lead to effective siRNA delivery and concomitant siRNA-mediated cell death to the human HCC cell line. The bioreducible polymer R646 (Figure 1), at 270ng/ul was considered to form the favorite formulation. A siRNA-mediated cell death of 89.1 ± 5% for HepG2 and 30.2 ± 16% for THLE-3 cells were observed, with a statistically significant difference between the HCC cells when compared to the healthy hepatocytes (p＜0.05). Non-specific toxicity to healthy hepatocytes remained below 26.1 ± 6%. Conclusion: Bioreducible poly(beta-amino ester)s nanoparticles not only seem to provide high siRNA-mediated death to HCC cells but also enable cancer-target in vitro. Even though further validation in vitro and in vivo is required, these findings suggest that biomaterial mediated siRNA delivery using bioreducible poly(beta-amino ester)s is a potentially viable strategy for liver cancer treatment.