A method to study drug concentration-depth profiles in tissues: mitomycin C in dog bladder wall

Abstract

Determination of the depth of penetration of locally applied drug therapy and evaluation of possible mechanisms of drug transport require knowledge of drug concentration-versus-tissues depth profiles. A method to determine the drug concentration-depth profile is needed. We have devised such a method and used it to determine the penetration of mitomycin C (MMC) in the dog bladder wall after intravesical drug instillation. This method is based on sectioning of frozen tissue into 40-microns segments, followed by drug extraction and high-pressure liquid chromatography analysis. Tissue concentrations could be detected with a sensitivity of 1 ng/sample, or 20 ng/g for tissue samples of approximately 2 x 2 cm. This sensitivity was sufficient to describe the penetration of MMC in the bladder wall of dogs, using an identical instillation technique, dwell time, and MMC concentration as in human patients. Tissue concentrations were expressed relative to tissue weight or tissue protein contents. For MMC, standardization to tissue weight yielded a better mathematical fit of the concentration-versus-depth profiles than standardization to protein content. The time interval between tissue harvesting and freezing was critical. The MMC concentration at the urothelial side of dog bladders was 2- to 10-fold higher in samples processed immediately after harvesting, compared to samples processed after 1 hr or longer. This significant decrease was not due to drug metabolism in situ. In separate in vitro experiments, we found that the degradation of MMC in 8% tissue homogenate was relatively slow, with only a 30% decline in concentration over 24 hr.(ABSTRACT TRUNCATED AT 250 WORDS)