Purpose: The water channel, aquaporin (AQP)1, is highly specific to the choroid plexus (CP) epithelium within the brain. It is therefore a potential target through which therapeutic agents could be selectively directed to the CP. Here we describe the conjugation of a monoclonal antibody (mAb), raised against an extra-cellular domain of AQP1, to the A chain of ricin (RCA). This reagent should allow study of a highly specific chemical lesion of the CP. Methods: A published method was used to couple the anti-AQP1 mAb to the A chain of ricin using an SPDP cross-linker. The conjugate was purified by Superdex S-200 chromatography. Column fractions were analysed by SDS-PAGE and Western blotting. Fractions containing conjugate were assayed for cytotoxicity on rat CP cells in culture, using a WST-1 viability assay to assess cell death. Results: SDS-PAGE and Western blotting demonstrated separation of conjugated anti-AQP1 mAb from unconjugated ricin by the Superdex column. Cultured rat CP cells were killed with an IC50 of 1.81 μg/ml when exposed to the anti-AQP1 mAb-RCA conjugation reaction products. Selectivity of this reagent was demonstrated by the higher IC50 exhibited by non-AQP1-expressing primary fibroblast cultures and cultures exposed to the reaction products from the non-specific IgG-RCA conjugation attempt. Conclusion: We have been able to demonstrate selective cytotoxicity in vitro of an anti-AQP1 mAb-RCA conjugate. We hope that this work will generate further interest in the use of this site-specific conjugate to target the CP in conditions such as hydrocephalus, tumours that arise from here, and other AQP1-expressing tumours.
- Choroid plexus
ASJC Scopus subject areas
- Pediatrics, Perinatology, and Child Health
- Clinical Neurology