Simplified method for conjugating macrocyclic bifunctional chelating agents to antibodies via 2-iminothiolane.

A one-step method for conjugating macrocyclic chelators to antibodies using the protein modification reagent 2-iminothiolane controls aggregation, maintains immunoreactivity, and produces consistent chelate/antibody ratios. Conjugation conditions have been investigated with the macrocyclic chelates 6-[p-(bromoacetamido)benzyl]-1,4,8,11-tetraazacyclotetradecane-N,N ',N",N"'-tetraacetic acid and 2-[p-(bromoacetamido)benzyl]-1,4,7,10-tetraazacyclododecane-N,N',N ",N"'-tetraacetic acid, with three different monoclonal antibodies. The bifunctional chelating agents are prepared by bromoacetylation of their amine precursors using a two-phase H2O/CHCl3 system, which improves product purity.

[1]  S. Carroll,et al.  Substituted 2-iminothiolanes: reagents for the preparation of disulfide cross-linked conjugates with increased stability. , 1990, Bioconjugate chemistry.

[2]  H. Mikola,et al.  The synthesis and use of activated N-benzyl derivatives of diethylenetriaminetetraacetic acids: alternative reagents for labeling of antibodies with metal ions. , 1989, Analytical biochemistry.

[3]  M. Eaton,et al.  Towards tumour targeting with copper-radiolabelled macrocycle–antibody conjugates , 1989 .

[4]  S. Denardo,et al.  The Peptide Way to Macrocyclic Bifunctional Chelating Agents: Synthesis of 2‐(p‐Nitrobenzyl)‐1,4,7,10‐tetraazacyclododecane‐N,N′,N′′,N′′′‐tetraacetic Acid (VI) and Study of Its Yttrium(III) Complex. , 1988 .

[5]  G. Denardo,et al.  Copper-67-labeled monoclonal antibody Lym-1, a potential radiopharmaceutical for cancer therapy: labeling and biodistribution in RAJI tumored mice. , 1988, Journal of nuclear medicine : official publication, Society of Nuclear Medicine.

[6]  B. Longenecker,et al.  Comparative radiolabeling and distribution of a tumour-directed monoclonal antibody. , 1988, International journal of radiation applications and instrumentation. Part B, Nuclear medicine and biology.

[7]  R. R. Robinson,et al.  Chimeric mouse-human IgG1 antibody that can mediate lysis of cancer cells. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[8]  D. Blakey,et al.  Effect of chemical deglycosylation of ricin A chain on the in vivo fate and cytotoxic activity of an immunotoxin composed of ricin A chain and anti-Thy 1.1 antibody. , 1987, Cancer research.

[9]  G. Denardo,et al.  Immunochemical aspects of monoclonal antibodies important for radiopharmaceutical development. , 1986, International journal of radiation applications and instrumentation. Part B, Nuclear medicine and biology.

[10]  S. Denardo,et al.  Copper chelates as probes of biological systems: stable copper complexes with a macrocyclic bifunctional chelating agent. , 1985, Analytical biochemistry.

[11]  C. Meares,et al.  Conjugation of antibodies with bifunctional chelating agents: isothiocyanate and bromoacetamide reagents, methods of analysis, and subsequent addition of metal ions. , 1984, Analytical Biochemistry.

[12]  H. Penefsky.  A centrifuged-column procedure for the measurement of ligand binding by beef heart F1. , 1979, Methods in enzymology.

[13]  C. Meares,et al.  The attachment of metal-chelating groups to protein: tagging of albumin by diazonium coupling and use of the products as radiopharmaceuticals. , 1978, The International journal of applied radiation and isotopes.

[14]  H. Stetter,et al.  Complex Formation with Tetraazacycloalkane‐N,N′,N″,N‴‐tetraacetic Acids as a Function of Ring Size , 1976 .

[15]  J W Hershey,et al.  Methyl 4-mercaptobutyrimidate as a cleavable cross-linking reagent and its application to the Escherichia coli 30S ribosome. , 1973, Biochemistry.

[16]  Sidney Udenfriend,et al.  Fluorescamine: A Reagent for Assay of Amino Acids, Peptides, Proteins, and Primary Amines in the Picomole Range , 1972, Science.

[17]  A. Seligman,et al.  ENZYME ALTERABLE ALKYLATING AGENTSVI. SYNTHESIS, CHEMICAL PROPERTIES, TOXICITIES, AND CLINICAL TRIAL OF HALOACETATES AND HALOACETAMIDES CONTAINING ENZYME-SUSCEPTIBLE BONDS. , 1963, Journal of the National Cancer Institute.

[18]  R. Thiers,et al.  Contamination in trace element analysis and its control. , 2006, Methods of biochemical analysis.