title: Radiometal-labeled peptide-PNA conjugates for targeting bcl-2 expression: preparation, characterization, and in vitro mRNA binding.
abstract: A new antisense peptide-peptide nucleic acid (peptide-PNA) conjugate, designed for targeting bcl-2 expression, has been radiolabeled, characterized, and evaluated for bcl-2 mRNA binding in a cell-free system. A PNA complementary to the first six codons of the bcl-2 gene was synthesized by standard solid-phase Fmoc chemistry and conjugated to a new derivative of 1,4,7,10-tetraazacyclododecane-N,N',N",N'"-tetraacetic acid (DOTA) that allows macrocyclic radiometal chelates to be incorporated into any sequence position of a peptide-PNA conjugate. The DOTA-PNA conjugate was then coupled to a membrane-permeating transduction peptide, PTD-4, designed for intracellular delivery of the radiolabeled PNA. The conjugate was characterized by HPLC and ESI-MS and labeled with (111)In and (90)Y to high specific activities (>1000 Ci/mmol) with high radiochemical purity. Northern blot analysis showed that (90)Y-PTD-4-K(DOTA)-anti-bcl-2-PNA bound specifically to as little as 50 fmol of bcl-2 mRNA, a result equivalent to that obtained with the analogous (32)P-labeled DNA antisense oligonucleotide. Thus, the mRNA targeting properties of (111)In- and (90)Y-PTD-4-K(DOTA)-anti-bcl-2-PNA demonstrate potential for diagnostic imaging and targeted radiotherapy applications in bcl-2-positive cancers.
authors.affiliation: Department of Veterinary Medicine and Surgery, University of Missouri Research Reactor, Columbia, Missouri 65211, USA. LewisMic@missouri.edu
authors.forename: Michael R
authors.initials: MR
authors.lastname: Lewis
authors.forename: Fang
authors.initials: F
authors.lastname: Jia
authors.forename: Fabio
authors.initials: F
authors.lastname: Gallazzi
authors.forename: Yi
authors.initials: Y
authors.lastname: Wang
authors.forename: Jiuli
authors.initials: J
authors.lastname: Zhang
authors.forename: Nalini
authors.initials: N
authors.lastname: Shenoy
authors.forename: Susan Z
authors.initials: SZ
authors.lastname: Lever
authors.forename: Mark
authors.initials: M
authors.lastname: Hannink
mesh_terms: D000970:Antineoplastic Agents / Q000138:chemical synthesis* / Q000494:pharmacology*; D001483:Base Sequence; D016503:Drug Delivery Systems; D015195:Drug Design; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D019254:Genes, bcl-2 / Q000235:genetics*; D007205:Indium Radioisotopes; D020135:Peptide Nucleic Acids / Q000138:chemical synthesis* / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D010455:Peptides / Q000138:chemical synthesis* / Q000378:metabolism / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D015021:Yttrium Radioisotopes
chemical_list: D000970:Antineoplastic Agents; D007205:Indium Radioisotopes; D020135:Peptide Nucleic Acids; D010455:Peptides; D012333:RNA, Messenger; D015021:Yttrium Radioisotopes