Overcoming extractability hurdles of a 14C labeled taxane analogue milataxel and its metabolite from xenograft mouse tumor and brain tissues

Taxane analogue milataxel have been shown to bound to proteins/tissues irreversibly. Extraction of the aforementioned bound drug and metabolite was proven to be difficult task. Nonetheless, an extraction method had to be developed to accurately determine drug concentration in tissues over time. This method would enable Taxolog, Inc. (Fairfield, NJ, USA) to accurately map the fate of drug in mice and it would also enable to better design drug dosing scheme for its maximum efficiency. A productive extraction technique for milataxel (MAC-321, TL-139) in nude mice with various xenograft human tumors was developed by extracting analytes from tumors using a novel extraction procedure and analyzing samples by LC-MS. This extraction technique entails disrupting tissue cells with hexane followed by acidic methanol (MeOH), with the aid of a tissuemizer and sonic cell disrupter. An average extractability of 75% was achieved as confirmed by the recovery of 14C labeled milataxel, as compared to 4-48.5% extraction efficiency using solvents and/or combination of solvents such as acetonitrile (ACN), ethanol, ethyl acetate, MeOH/acetic acid in water, and chloroform/MeOH. This extraction technique allowed for quantitation of milataxel and its major metabolite s-lactate (M-10) from tumors and brain tissue samples using HPLC coupled with electro-spray ionization mass spectrometry (HPLC-ESI-MS). Ratios of M-10 metabolite to milataxel were determined to be approximately 3:1 and 2:1 in SKMES human lung carcinoma tumors and A-375 melanoma tumors, respectively, and declined in concentration over 20 days. However, levels of milataxel and M-10 were determined to be equal at 8 h in HCT-15 human colon carcinoma tumors with M-10 levels dropping sharply over a 10-day period.