[3H]DU41165: A high affinity ligand and novel photoaffinity labeling reagent for the progesterone receptor

Kevin G. Pinney, Kathryn E. Carlson, John A. Katzenellenbogen

Research output: Contribution to journalArticlepeer-review

Abstract

17α-Acetoxy-6-fluoro-16-methylene-(9β, 10α )pregna-4,6-dien-3,20-dione (DU41165), a retroprogestin (9β,10α) embodying a fluorine-substituted dienone system, has been prepared in high specific activity tritium-labeled form (4 Ci/mmol) and shown to be a high affinity ligand for the progesterone receptor (PgR) and a highly selective photoaffinity labeling reagent for PgR. The radiosynthesis involved conversion of DU41231 (the 17α-hydroxy analog of DU41165) to DU41165 by treatment with tritum-labeled acetic anhydride. The binding affinity of DU41165 for PgR was determined by both a competitive binding assay and a direct binding assay (Scatchard analysis) to be 1.6-2.2-times higher than that of the high affinity synthetic progestin promegestone (R5020). In unlabeled form, DU41165 demonstrates photoinactivation of PgR to the extent of 60% at 60 min. In radiolabeled form [3H]DU41165 demonstrates specific covalent attachment with an efficiency of 5-7%. SDS-polyacrylamide gel electrophoresis of photoattached [3H]DU41165 confirms that there is covalent labeling of both the B subunit (Mr = 118,000), and the A subunit (Mr = 88,000) of PgR in a molar ratio of approximately 1:3.

Original languageEnglish (US)
Pages (from-to)179-189
Number of pages11
JournalJournal of Steroid Biochemistry
Volume35
Issue number2
DOIs
StatePublished - Feb 1990

ASJC Scopus subject areas

  • Biochemistry
  • Endocrinology

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