MICROTUBULAR DISRUPTION PROLONGS THE EXPRESSION OF HUMAN DINEDIPHOSPHOGLUCURONATE-GLUCURONOSYLTRANSFERASE-1 GENE TRANSFERRED INTO GUNN RAT LIVERS

DNA delivered to the liver by asialoglycoprotein receptor-mediated end ocytosis is degraded in lysosomes within 48 h, To test the hypothesis that microtubular disruption should promote transgene persistence by i nterrupting endosomal translocation to lysosomes, plasmids containing bacterial chloramphenicol acetyltransferase (pSV2-CAT) or human biliru bin-UDP-glucuronosyltransferase-1 (pSVK3-hBUGT(1)) genes were complexe d with asialoglycoprotein-polylysine conjugates, and 1 mg of the compl exed DNA was injected intravenously into bilirubin-UDP-glucuronosyltra nsferase-deficient Gunn rats, 30 min before DNA injection, one group r eceived 0.75 mg of colchicine/kg of body weight intraperitoneally, whi ch was shown by immunofluorescent confocal microscopy to disrupt the m icrotubular network, Control rats received normal saline, In colchicin e-pretreated rats receiving pSV2-CAT, hepatic chloramphenicol acetyltr ansferase activity persisted for 9-14 weeks, whereas in the saline pre treated group the activity was detectable for 48 h only, In colchicine -pretreated Gunn rats receiving pSVK3-hBUGT(1), the DNA persisted in l iver for 10 weeks, bilirubin glucuronides were excreted in bile, and s erum bilirubin levels declined by 25-35% in 2-4 weeks and remained red uced for 8 weeks, Without colchicine pretreatment, the DNA was detecta ble in liver for 2 days only, and serum bilirubin levels were not redu ced, Thus, microtubular disruption provides a noninvasive method for p rolonging the effect of liver-targeted gene therapy.