Charge-reversible lipid derivative: A novel type of pH-responsive lipid for nanoparticle-mediated siRNA delivery.

RNA interference induced by small interfering RNA (siRNA) is a promising strategy for the treatment of various intractable diseases including cancer. Lipid nanoparticles (LNP) composed of ionizable lipids and siRNA are known as a leading siRNA delivery system. However, LNPs composed of conventional ionizable lipids will be aggregated in the physiological environment because of loss of ionization. Therefore, the inclusion of hydrophilic polymer-conjugated lipids such as polyethylene glycol (PEG)-conjugated lipid is required to improve the LNP stability. Herein, we synthesized a novel charge-reversible lipid derivative, dioleoylglycerophosphate-diethylenediamine conjugate (DOP-DEDA). The surface of LNP composed of DOP-DEDA (DOP-DEDA LNP) was constantly ionized and positively charged at pH 6.0, almost neutral at pH 7.4, and negatively charged at pH 8.0. Importantly, DOP-DEDA LNP were stable in the physiological milieu without PEG-conjugated lipid. DOP-DEDA LNP disrupted the red blood cells only under the low-pH condition in a hemolysis assay, suggesting that the interaction between DOP-DEDA LNP and biological membranes is pH-dependent. DOP-DEDA LNP encapsulating siRNA against polo-like kinase 1 (siPLK1) highly suppressed the expression of PLK1 mRNA and its protein. The cellular uptake of DOP-DEDA LNP was increased in an apolipoprotein E3 (apoE3) dose-dependent manner. In addition, DOP-DEDA LNP was taken up into cancer cells via both clathrin- and caveola-mediated endocytosis pathways. These findings indicate that LNP composed of this charge-reversible lipid should be a highly stable and potent siRNA delivery vector.

Influence of the carbamate fungicide benomyl on the gene expression and activity of aromatase in the human breast carcinoma cell line MCF-7.

The carbamate fungicide benomyl reportedly inhibited the growth of the human breast cancer cell line MCF-7 by inducing apoptosis. However, influence of benomyl on the expression and activity of aromatase of MCF-7 cells remains to be examined, since benomyl was identified as an endocrine disruptor. We here confirmed through cell cycle analysis and immunofluorescence staining that benomyl damaged microtubules and caused apoptosis. We also found that benomyl inhibited histone deacetylase (HDAC) 1 and accumulated acetylated histone H3 in MCF-7 cells. Additionally, benomyl enhanced the levels of aromatase protein and mRNA, albeit at high concentrations. It is thus likely that benomyl enhanced the promoter activity of the aromatase gene via acetylation of histone H3 as does the HDAC inhibitor Vorinostat. In conclusion, benomyl remains to be a risk factor as an endocrine disruptor for breast cancer.