title: Improved Synthesis and In Vitro Evaluation of an Aptamer Ribosomal Toxin Conjugate.
abstract: Delivery of toxins, such as the ricin A chain, Pseudomonas exotoxin, and gelonin, using antibodies has had some success in inducing specific toxicity in cancer treatments. However, these antibody-toxin conjugates, called immunotoxins, can be bulky, difficult to express, and may induce an immune response upon in vivo administration. We previously reported delivery of a recombinant variant of gelonin (rGel) by the full-length prostate-specific membrane antigen (PSMA) binding aptamer, A9, to potentially circumvent some of these problems. Here, we report a streamlined approach to generating aptamer-rGel conjugates utilizing a chemically synthesized minimized form of the A9 aptamer. Unlike the full-length A9 aptamer, this minimized variant can be chemically synthesized with a 5' terminal thiol. This facilitates the large scale synthesis and generation of aptamer toxin conjugates linked by a reducible disulfide linkage. Using this approach, we generated aptamer-toxin conjugates and evaluated their binding specificity and toxicity. On PSMA(+) LNCaP prostate cancer cells, the A9.min-rGel conjugate demonstrated an IC50 of ∼60 nM. Additionally, we performed a stability analysis of this conjugate in mouse serum where the conjugate displayed a t1/2 of ∼4 h, paving the way for future in vivo experiments.
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine , Bronx, New York City, New York.
authors.forename: Linsley
authors.initials: L
authors.lastname: Kelly
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine , Bronx, New York City, New York.
authors.forename: Christina
authors.initials: C
authors.lastname: Kratschmer
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine , Bronx, New York City, New York.
authors.forename: Keith E
authors.initials: KE
authors.lastname: Maier
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine , Bronx, New York City, New York.
authors.forename: Amy C
authors.initials: AC
authors.lastname: Yan
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine , Bronx, New York City, New York.
authors.forename: Matthew
authors.initials: M
authors.lastname: Levy
mesh_terms: D000954:Antigens, Surface / Q000008:administration & dosage* / Q000737:chemistry / Q000235:genetics; D052157:Aptamers, Nucleotide / Q000008:administration & dosage / Q000138:chemical synthesis / Q000737:chemistry; D045744:Cell Line, Tumor; D043425:Glutamate Carboxypeptidase II / Q000008:administration & dosage* / Q000737:chemistry / Q000235:genetics; D006801:Humans; D000922:Immunotoxins / Q000737:chemistry* / Q000235:genetics; D039161:Integrin alpha6beta4 / Q000008:administration & dosage* / Q000737:chemistry / Q000235:genetics; D009369:Neoplasms / Q000188:drug therapy* / Q000235:genetics; D054789:Ribosome Inactivating Proteins, Type 1 / Q000737:chemistry* / Q000235:genetics / Q000627:therapeutic use; D012276:Ricin / Q000627:therapeutic use
chemical_list: D000954:Antigens, Surface; D052157:Aptamers, Nucleotide; D000922:Immunotoxins; D039161:Integrin alpha6beta4; D054789:Ribosome Inactivating Proteins, Type 1; C025674:GEL protein, Gelonium multiflorum; D012276:Ricin; C473848:FOLH1 protein, human; D043425:Glutamate Carboxypeptidase II