Abstract 3235: Improved in vitro efficacy study of RGD grafted docetaxel encapsulated solid lipid nanoparticles on breast cancer cells
Authors: Patel Deepa H. , Naik Sachin , Misra Ambikanandan 
Published: June 2012

DC- encapsulated solid lipid nanoparticles (SLN) were prepared using Supercritical fluid technology using CO2 as an anti-solvent. RGD-conjugation was done by carbodiimide coupling method. The SLN were characterized for particle size and zeta potential by zeta sizer (Nano-ZS), entrapment efficiency and in-vitro drug release profile using HPLC assay, and RGD-conjugation efficiency by amino acid analysis method. The possibility of drug-lipid -RGD interaction was ascertained using a differential scanning colorimetry and X-ray diffraction techniques. To establish antiproliferative effect, the RGD-conjugated SLN of DC(RGD-SLN-DC), unconjugated SLN(DC-SLN) and equal amounts of the free drug were studied on MDA-MB-231 cells by CCK-8 assay method. DNA content analysis was done on flowcytometry by propadium iodide staining. The mode of cell death at different time and concentration was determined by FITC-Annexine V assay.

Results: The DC encapsulated RGD conjugated and unconjugated SLN were found to be nanosized with optimum drug entrapment. The in-vitro release profiles of unconjugated and conjugated SLN were found to be similar except the fast drug release was observed in case of RGD conjugated pegylated SLN due to the fast hydration process of PEG molecules on the surface of the particles. The cytotoxicity indicated by IC50 values suggests that RGD conjugated SLN at 72hrs are 2.1 and 6.3 times more cytotoxic than unconjugated SLN and drug solution for for MDA-MB-231 cells. The results shows that the RGD conjugated SLN at the concentration of 4nM showed 97% G2 phase arrest as compared to 65% G2 phase arrest with unconjugated SLN at the same concentration at 48hrs. Two types of mode of cell death were found during the FITC-Annexin V assay. At 48 hrs, the treatment of 4nM of drug solution with cells resulted in 23.5% & 2.8 % necrotic & apoptotic cell fragments respectively. While at similar drug equivalent concentration, the RGD-SLN-DC and DC-SLN showed 2.9% & 76.3% and 3.4% & 42.3% necrotic & apoptotic cell fragments respectively. With increase in time and concentration the mode of cell death by apoptosis was found to be increasing.

Conclusions: To conclude, RGD conjugation to SLN improved antiproliferative activity when assessed in vitro in breast cancer cells compared to free drug and unconjugated SLN. The DNA content analysis depicted the cell cycle arrest in G2 phase was more even at lower drug equivalent concentration of RGD-SLN-DC. The mode of cell death was found to be mainly by necrosis at low drug equivalent concentration (1nM) and by apoptosis at high drug equivalent concentration (10nM) of RGD conjugated drug encapsulated SLN for breast cancer cells. Hence, it may be concluded that drug nanocarriers once conjugated with RGD can provide prolonged drug release to cytoplasm and can affectively target breast cancer and can probably reduce the limitations associated with breast cancer chemotherapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3235. doi:10.1158/1538-7445.AM2011-3235