Cell surface retention of PrP^C by anti-PrP antibody prevents protease-resistant PrP formation

Abstract

Because of the emergence of variant CJD and accumulation of iatrogenic CJD cases, the establishment of therapeutics for prion disease is urgently needed. One possible strategy for therapeutics is an inhibition of an abnormal isoform of prion protein (PrP^Sc) formation in the infected host. The C-terminal portion of prion protein (PrP), corresponding to a protease-resistant core fragment of PrP^Sc, is essential for prion propagation. Antibodies to the C-terminal portion of PrP are known to inhibit PrP^Sc accumulation in prion-persistently-infected cells. Here we show that, in addition to monoclonal antibodies (mAbs) to the C-terminal portion of PrP, a mAb recognizing the octapeptide repeat region in the N-terminal portion of PrP that is dispensable for PrP^Sc formation, reduced PrP^Sc accumulation in cells persistently infected with prion; mAb 110 and 31C6, recognizing linear epitope aa 59–90 and aa 143–149, respectively, and mAbs 44B1 and 72, recognizing discontinuous epitope located within aa 155–231 and aa 89–231, respectively, inhibited the PrP^Sc formation when the cells were cultured with medium containing these mAbs. The 50% effective doses were as low as ∼1 nM, and, regardless of their epitope specificity, the inhibitory mAbs shared the ability to bind cellular prion protein (PrP^C) expressed on the cell surface. Flow cytometric analysis revealed that mAbs which bound to the cell surface during cell culture were not internalized even after their withdrawal from the growth medium. Retention of the mAb-PrP^C complex on the cell surface was also confirmed by the fact that internalization was enhanced by treatment of cells with dextran sulfate. These results suggest that anti-PrP mAb antagonized PrP^Sc formation by interfering with the regular PrP^C degradation pathway.