Non-Specific Binding of the Experimental Anti-Cancer Drug 5,6-Dimethylxanthenone-4-Acetic Acid (DMXAA) in Liver Microsomes from Various Species

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Total (added) drug concentrations other than unbound concentrations have been used to estimate the in-vitro enzyme kinetic parameters for 5,6-dimethylxanthenone-4-acetic acid (DMXAA), an experimental anti-cancer drug. This study aimed to investigate the non-specific binding of DMXAA to liver microsomes from various species and to microsomes from human lymphoblastoid cells expressing drug-metabolising enzymes, and to examine the effect of the binding on the estimation of enzyme kinetic parameters for DMXAA in-vitro. The separation of unbound DMXAA was conducted by ultrafiltration and DMXAA concentrations were determined by validated HPLC. The results indicated that DMXAA was bound to liver microsomes and lymphoblastoid cell microsomes to a small extent (free fraction in microsomes, fu(mic) mostly > 0.85). Correction for the unbound DMXAA concentration resulted in slightly lower apparent Michaelis-Menten constant (Km) values, but with the maximal velocity of reaction (Vmax) unchanged, leading to slightly higher unbound Vmax/Km values. These results indicate that the non-specific binding of DMXAA to microsomes is insignificant and has little impact on the enzyme kinetic estimation in-vitro.

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Journal of Pharmacy and Pharmacology, v. 54, issue 7, p. 997-1004