Abstract 5708: Anti-proliferative and pro-apoptotic effects of (−)-epigallocatechin-3-gallate encapsulated in chitosan nanoparticles on human melanoma Mel 928 cells (2010)

Abstract We recently employed the use of nanotechnology to improve the outcome of bioactive food components for cancer chemoprevention and termed the concept as nanochemoprevention (Cancer Res. 2009; 69(5): 1712-6.). To demonstrate the proof-of-principle we encapsulated green tea polyphenol epigallocatechin-3-gallate (EGCG) in polylactic acid-polyethylene glycol (PLA-PEG) nanoparticles and showed that this formulation demonstrates greater than ten-fold dose advantage over non-encapsulated EGCG in human prostate cancer PC3 cells both in vitro and in vivo settings. Here we extended the work to human melanoma cells where we previously showed that EGCG exerts anti-proliferative and pro-apoptotic effects. Further, one disadvantage of PLA-PEG nanoparticles is the lack of its oral bioavailability. To overcome this obstacle we encapsulated EGCG in nanoparticles employing a naturally occurring polymer chitosan (hereafter referred to as nano-EGCG), a delivery system suitable for oral consumption. We first determined the usefulness of this unique formulated nano-EGCG for its cytotoxic response against human melanoma Mel 928 cells. MTT assay showed that treatment of cells with nano-EGCG, compared to EGCG in PBS, produced remarkably superior effects with an IC50 of 2.0 μM compared to 50 μM when delivered in PBS. Our next goal was to determine if nano-EGCG retains its mechanistic identity. Nano-EGCG treatment (2 μM) to the Mel 928 cells resulted in (i) a shift in Bax/Bcl-2 ratio in favor of apoptosis, (ii) an increase in PARP cleavage, (iii) a significant inhibition in caspase 3 and 9, other key regulators of apoptosis, (iv) a dramatic increase in the levels of annexin V, and (v) marked induction of p21 and p27 cyclin dependent kinase inhibitors. Importantly, 2 μM of EGCG delivered in PBS was without any effect on any of these responses. Next we determined EGCG release kinetics of the nanoformulation using simulated gastric and intestinal fluids. We observed that the EGCG release peaked at 2 hours post-incubation with 50% released into the intestinal fluid and 30% released into the gastric juice and the percent release thereafter was maintained at constant levels suggesting a sustained release phenomenon. Furthermore, to ascertain oral bioavailability of nano-EGCG compared to free EGCG, blood samples were collected from mice treated with EGCG and nano-EGCG at 1, 8 and 24 hrs post administration and the levels of EGCG were determined by LC/MS/MS. We observed a steady and sustained release of EGCG in the plasma of mice following oral administration. Several experiments indicated that this nano formulated EGCG exhibited a significant longer half-life compared to non-encapsulated EGCG when given orally to mice. Translation of these data to appropriate animal model systems could pave the way for developing new avenues for melanoma chemoprevention. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5708.