title: Synthesis, characterization, and labeling with 99mTc/188Re of peptide conjugates containing a dithia-bisphosphine chelating agent.
abstract: Radiolabeling of small receptor-avid peptides at specific predetermined chelation sites with radioactive metals has been an effective approach for production of target-specific radiopharmaceuticals for diagnosis and therapy of diseases. Among various electron-donating groups found on chelator frameworks, phosphines are unique because they display versatile coordination chemistry with a wide range of transition metals. We have recently reported the utility of a dithia-bis(hydroxymethyl)phosphine-based (P2S2) bifunctional chelating agent (BFCA) containing air-stable primary phosphine groups to form 99mTc-labeled receptor-avid peptides by the preconjugation approach. Here we report a novel strategy for labeling small peptides with both 99mTc and 188Re using the P2S2-COOH (6,8-bis[3-(bis(hydroxymethyl)phosphanyl)propylsulfanyl]octanoic acid) BFCA by a postconjugation radiolabeling approach. The first step in this approach involves the coupling of the corresponding (PH2)2S2-COOH intermediate to the N-terminus of the peptide(s). Formylation of P-H bonds with aqueous formaldehyde in the presence of HCl in ethanol affords the corresponding (hydroxymethyl)phosphine-P2S2-peptide conjugates in the form of an oxidatively stable phosphonium salt. The P2S2-peptide conjugates are generated (where the PH2 groups are converted to P(CH2OH)2 groups) by treatment of the P2S2-peptide phosphonium salt(s) with 1 M sodium bicarbonate solution at pH 8.5. Complexation of BFCA conjugates with 99mTc is achieved by direct reduction with Sn(II) tartarate to yield the 99mTc-P2S2-peptide conjugate in near quantitative yields. Complexation of the BFCA conjugates with 188Re is achieved by transchelation with 188Re citrate in yields of >/=90%. In this study, (PH2)2S2-COOH BFCA was conjugated to model peptides. The glycineglycine ethyl ester (GlyGlyOEt)-(PH2)2S2-COOH BFCA conjugate was converted to the hydroxymethylene phosphine form and complexed with 99mTc to produce the 99mTcO2-P2S2-GlyGlyOEt conjugate 8 in RCPs of >/=95%. This singular 99mTc product is stable over 24 h in aqueous solution as confirmed by HPLC. Identical retention times of the 99mTcO2-P2S2-GlyGlyOEt complex and its cold rhenium analogue (ReO2-P2S2-GlyGlyOEt) on HPLC indicates similarity in structures at the macroscopic and the tracer levels. The utility of this postconjugation strategy was further demonstrated by synthesizing a P2S2-D-Lys6-LHRH conjugate and producing its corresponding 99mTc complex in RCPs of >/=88%. Finally, the P2S2-5-Ava-BBN[7-14]NH2 bombesin (BBN) analogue was synthesized, the PH2 groups converted to P(CH2OH)2 groups and subsequently labeled with 188Re to yield a 188Re-labeled bombesin analogue with a RCP of >/=90%. The biological integrity of this conjugate was demonstrated in both in vitro and in vivo. The results of this investigation demonstrate that the (PH2)2S2-COOH BFCA can be conveniently used as a precursor for labeling small receptor-avid peptides with diagnostic (99mTc) and therapeutic (188Re) radionuclides via the postconjugation approach in high yields.
authors.affiliation: Department of Radiology, University of Missouri-Columbia, Columbia, Missouri 65211, USA.
authors.forename: H
authors.initials: H
authors.lastname: Gali
authors.forename: T J
authors.initials: TJ
authors.lastname: Hoffman
authors.forename: G L
authors.initials: GL
authors.lastname: Sieckman
authors.forename: N K
authors.initials: NK
authors.lastname: Owen
authors.forename: K V
authors.initials: KV
authors.lastname: Katti
authors.forename: W A
authors.initials: WA
authors.lastname: Volkert
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001682:Biological Availability; D002614:Chelating Agents / Q000138:chemical synthesis / Q000378:metabolism / Q000493:pharmacokinetics; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D009363:Neoplasm Proteins / Q000378:metabolism; D009368:Neoplasm Transplantation; D015609:Organotechnetium Compounds / Q000138:chemical synthesis / Q000378:metabolism / Q000493:pharmacokinetics; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism / Q000493:pharmacokinetics*; D011868:Radioisotopes; D011869:Radioligand Assay; D019275:Radiopharmaceuticals / Q000138:chemical synthesis* / Q000378:metabolism / Q000493:pharmacokinetics*; D018004:Receptors, Bombesin / Q000378:metabolism; D012211:Rhenium; D013667:Technetium; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured
chemical_list: D002614:Chelating Agents; D009363:Neoplasm Proteins; D015609:Organotechnetium Compounds; D010455:Peptides; D011868:Radioisotopes; D019275:Radiopharmaceuticals; D018004:Receptors, Bombesin; C109881:dithio-bisphosphine tc99m; D012211:Rhenium; D013667:Technetium