Enhanced hepatocyte uptake and liver targeting of methotrexate using galactosylated albumin as a carrier.

Liver targeting of drugs has wide therapeutic implications due to numerous liver-related diseases. Using conjugates of methotrexate (MTX) to variously galactosylated bovine serum albumin (BSA), we studied whether we could enhance the liver targeting of MTX, a model drug, via galactose receptors selectively abundant on the hepatocytes. Here, we report that the galactosylation of the carrier protein BSA significantly enhanced the hepatocyte uptake and liver targetability of MTX. In vitro, the amount of MTX taken up by rat hepatocytes was positively correlated with the galactose content in BSA. MTX conjugates were relatively stable in plasma, but released MTX with time in liver homogenates. These results imply that the conjugates would exert low toxicity in the blood, but have therapeutic activity in the liver by liberating MTX. In vivo, MTX-galactosylated BSA conjugates (MTX-L(24)BSA) showed significantly different pharmacokinetics from free MTX or MTX-BSA conjugates. The plasma level of free MTX rapidly declined in a biexponential fashion with an apparent terminal half-life of 0.35 h. MTX-BSA conjugates showed the slowest decline with an apparent terminal half-life of 6 h, whereas MTX-L(24)BSA showed a biphasic pattern; a rapid distributive phase with a half-life of 0.567 h and a slow terminal phase. MTX-L(24)BSA showed the highest liver targetability, when evaluated in terms of two indices based on the area under the total amount of radioactivity-time curve (AUQ); Te*(liver), % AUQ(liver) to total AUQ, and te*, the ratio of AUQ(liver) to AUQ(kidney). Compared with free MTX and MTX-BSA, MTX-L(24)BSA showed about twofold higher Te*(liver) of 87.5%. The te* of MTX-L(24)BSA was 25- and fourfold higher than those of free MTX and MTX-BSA, respectively. Moreover, MTX-L(24)BSA showed a gradual increase in the therapeutically active intact form of MTX in the liver while showing the lowest level of intact MTX in the kidney. These results suggest that galactosylated BSA has a great potential as an hepatocyte-directed and more effective liver targeting carrier of drugs for liver diseases.