183. Hemochromatosis: AAV Coding an Antisense Gene Against DMT-1 Iron Transporter in Intestinal Epithelial Cells Acts as RNAi. And Inhibits Iron Transport

Abstract

Hemochromatosis, the most prevalent inborn genetic disease (1:200 in Caucasians) is characterized by an increased iron absorption by enterocytes. Iron accumulates in the liver promoting oxidative stress, DNA damage and cirrhosis. Long-term reduction of intestinal iron transport by inhibition of the transporter DMT-1 could conceivably be achieved in vivo by oral administration of AAV vectors carrying a gene that can transcribe a short antisense RNA. Studies presented show that an AAV carrying a short antisense coding gene preceded by a CMV promoter and a SV40 poly-A signal strongly reduces apical uptake of iron in intestinal Caco-2 cells. The anti DMT-1 antisense gene led to a marked reduction (−80%) in DMT-1 mRNA and to very low DMT-1 antisense mRNA levels, suggesting that after hybridizing both RNA molecules are hydrolyzed into short ds RNA by the enzyme Dicer.DMT-1 levels are controlled by intracellular iron, such that sustained reductions in enterocytes are likely to lead to compensatory enhanced traslation by increased stability of DMT-1 mRNA. Present studies address whether the same RNAi-like mechanism inhibits the basolateral transport of iron by ferroportin a basolateral transporter that releases iron from enterocytes into the circulation. The latter does not present the feedback that DMT-1 has. Overall, studies presented suggest that enhanced the iron transport seen in hemochromatosis could be blunted by AAV-mediated gene therapy.