ABSTRACT
Chemoselective methionine bioconjugation with alkyne-bearing oxaziridine and alkyne-bearing iodonium salts was investigated as a new platform for site-selective radiolabeling of proteins and peptides with fluorine-18. Alkyne-bearing sulfimide conjugates, resulting from oxaziridine modification, underwent copper-assisted alkyne-azide cycloaddition (CuAAC) with an 18F-labeled PEGylated azide to afford 18F-labeled triazoles in excellent radiochemical yields. Diazoester sulfonium salt bioconjugates, formed from alkyne-bearing 2-diazoiodonium salts, gave low yields of 18F-labeled triazoles and were shown to be unstable to CuAAC conditions. Photolytic removal of the diazo group, however, afforded the trialkylsulfonium salt which smoothly underwent CuAAC with the 18F-labeled PEGylated azide to afford high radiochemical yields of the desired 18F-labeled click product. Overall, the results establish the viability of chemoselective methionine bioconjugation as a method for preparing site-selective 18F-labeled PET radioligands.
Subject(s)
Fluorine Radioisotopes , Methionine/chemistry , Peptides/chemistry , Proteins/chemistry , Click Chemistry/methods , Radiopharmaceuticals , Serum Albumin, Bovine/chemistryABSTRACT
A two-step degradation-reconstruction approach to the carbon-14 radiolabeling of alkyl carboxylic acids is presented. Simple activation via redox-active ester formation was followed by nickel-mediated decarboxylative carboxylation to afford a range of complex compounds with ample isotopic incorporations for drug metabolism and pharmacokinetic studies. The practicality and operational simplicity of the protocol were demonstrated by its use in an industrial carbon-14 radiolabeling setting.
Subject(s)
Carboxylic Acids/chemistry , Radiopharmaceuticals/chemistry , Carbon Isotopes/chemistry , Carbon Radioisotopes/chemistry , Carboxylic Acids/chemical synthesis , Catalysis , Decarboxylation , Isotope Labeling/methods , Nickel/chemistry , Radiopharmaceuticals/chemical synthesisABSTRACT
2-Iminothiolane has found utility in the growing area of antibody-drug conjugates by serving as a lysine-thiolating agent and the junction between the antibody and the cytotoxic payload during random conjugation of a monoclonal antibody. 2-(14)C-Iminothiolane was prepared from commercially available [(14)C]KCN using a four-step sequence in an overall 10% radiochemical yield. Stable-labeled 2-(13)C,(15)N-iminothiolane was also prepared from [(13)C(15)N]KCN in a similar manner. The Ë labeled Traut's reagent produced by this sequence showed comparable reactivity as the commercially available unlabeled reagent with a representative monoclonal antibody and could serve as highly informative analytical tools to investigate antibody-drug conjugate formation via the random conjugation process.
Subject(s)
Carbon Radioisotopes/chemistry , Imidoesters/chemical synthesis , Nitrogen Isotopes/chemistry , Imidoesters/chemistryABSTRACT
The synthesis of a 16-residue, stable isotopically labeled peptide is described for use as a LC-MS/MS (Liquid chromatography-mass spectrometry/mass spectrometry) internal standard in bioanalytical studies. This peptide serves as a single universal surrogate peptide capable of quantifying a wide variety of immunoglobulin G and Fc-fusion protein drug candidates in animal species used in pre-clinical drug development studies. An efficient synthesis approach for this peptide was developed using microwave-assisted solid phase peptide synthesis (SPPS) techniques, which included the use of a pseudoproline dipeptide derivative. The corresponding conventional room temperature SPPS was unsuccessful and gave only mixtures of truncated products. Stable-labeled leucine was incorporated as a single residue via manual coupling of commercially available Fmoc-[(13) C6 , (15) N]-l-leucine onto an 11-unit segment followed by automated microwave-assisted elaboration of the final four residues. Using this approach, the desired labeled peptide was prepared in high purity and in sufficient quantities for long-term supplies as a bioanalytical internal standard. The results strongly demonstrate the importance of utilizing both microwave-assisted peptide synthesis and pseudoproline dipeptide techniques to allow the preparation of labeled peptides with highly lipophilic and sterically hindered side-chains.
Subject(s)
Chromatography, Liquid/standards , Mass Spectrometry/standards , Peptide Fragments/chemical synthesis , Solid-Phase Synthesis Techniques/methods , Amino Acid Sequence , Carbon Radioisotopes/chemistry , Chromatography, Liquid/methods , Humans , Immunoglobulin Fc Fragments/chemistry , Immunoglobulin G/chemistry , Mass Spectrometry/methods , Microwaves , Molecular Sequence Data , Nitrogen Isotopes/chemistry , Reference StandardsABSTRACT
For the first time, [3α-(3) H] 17α-hydroxy pregnenolone (1) was synthesized through a multiple step sequence. The presence of [3ß-(3) H] isomer in RP-HPLC purified product was identified by tritium NMR. The [3ß-(3) H] isomer was then separated from [3α-(3) H] 17α-hydroxy pregnenolone with chiralPAK AD-H column. [3α-(3) H] pregnenolone (2) was synthesized from commercial available 5-pregnen-3,20-dione in one step with an improved procedure.
