Targeting PSMA on tumor cells using novel aptamer-gelonin conjugates.

Andrew D. Ellington, John W. Marks III, Ted Chitai Chu, Laura Lavery, Sarah Faulkner, Matthew Levy, Michael G. Rosenblum. The University of Texas at Austin, Austin, TX, UT M.D. Anderson Cancer Center, Houston, TX

PSMA is a transmembrane receptor whose expression increases on prostatic tumor cells during disease progression. This protein is also expressed on the endotheluim of tumor vasculature but not normal vasculature. Aptamers are small nucleic acids selected to bind proteins such as cell-surface tumor antigens with high affinity and specificity. Aptamers have the potential to serve as replacements for cell-targeting antibodies or other cell-targeting ligands. We utilized a modified RNA aptamer (designated PSMAA) known to bind the external domain of PSMA with high affinity (previously reported Ki = 2 nM). The 22 kDa PSMAA molecule was conjugated to recombinant gelonin(rGel) toxin using the heterobifunctional cross-linking agent SPDP. The PSMAA/rGel conjugate was then purified by ion exchange and gel permeation chromatography. The final product was uncontaminated by free aptamer or free rGel and migrated as a single species (~50 kDa) by SDS-PAGE. Analysis of the construct demonstrated that the rGel component was enzymatically active compared to free rGel. In addition, the conjugate was found to bind specifically to PSMA-expressing LNCAP cells. Cytotoxicity studies of the PSMAA/rGel conjugate demonstrated an I.C.50 of 32 nM on antigen-positive LNCAP cells compared to an I.C.50 of 350,000 nM on PC3 cells, which contain much less antigen; a targeting index of approximately 10,000 fold. Internalization studies should reveal the details of toxin conjugate entry, and animal model studies are ongoing. These data suggest that aptamers can be used to successfully deliver protein molecules such as toxins to tumor cells and may provide a novel approach to development of targeted therapeutic agents. The fact that aptamers can be chemically synthesized and thereby site-specifically conjugated makes them especially interesting as targeting ligands. The PSMAA-rGel construct may prove useful to target PSMA on prostate cells and on tumor vasculature. Research conducted, in part, by the Clayton Foundation for Research.

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