Method for radiolabeling antibody fragments
AssigneeImmunomedics, Inc.
InventorsDan Shochat, Hans J. Hansen, Robert S. Wu
Life of this patent
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Abstract
A protein containing at least one pendant sulfhydryl group is directly radiolabeled with a radiometal which binds tightly to sulfhydryl groups, using one or more pendant sulfhydryl groups on the protein as endogenous ligands and optionally using an exogenous ligand which binds tightly to the radiometal ion to further stabilize the chelate.
Claims20
A method for direct radio labeling of a monovalent antibody fragment, comprising the step of contacting, in solution, a mixture of (a) a monovalent Fab-SH, Fab'-SH or Fabc-SH monovalent antibody fragment produced by reductive cleavage, using a thiol reducing agent, of a divalent F(ab).sub.2 or F(ab').sub.2 precursor fragment or whole immunoglobulin, respectively, and (b) a stannous ion reducing agent for pertechnetate or perrhenate, wherein said mixture is substantially free of low molecular weight thiol compounds, with 99m-pertechnetate, 186-perrhenate or 188-perrhenate ions, and recovering the resultant solution of radiolabeled monovalent antibody fragment without purification.
A kit suitable for radio-labeling a monovalent antibody fragment with Tc-99m, Re-186 or Re-188, comprising, in a single container, a mixture of
- (a) a monovalent Fab-SH, Fab'-SH Or Fabc-SH antibody fragment produced by reductive cleavage, using a thiol reducing agent, of a divalent F(ab).sub.2 or F(ab').sub.2 precursor fragment or whole immunoglobulin, respectively, and
- (b) a source of stannous ions for reduction of pertechnetate or perrhenate;
- wherein said mixture is substantially free of low molecular weight thiol compounds.
A method of radio-labeling a protein consisting essentially of the steps of:
- a) incubating a protein containing disulfide bonds, selected from the group consisting of F(ab).sub.2, F(ab').sub.2 and F(abc).sub.2 immunoglobulins, with a thiol disulfide bond-reducing agent, the period of incubation being sufficient to cleave interchain heavy-heavy chain disulfide bonds to sulfhydryl groups and produce Fab, Fab' or Fabc fragments;
- b) purifying the reduced protein to substantially remove the thiol disulfide bond-reducing agent and impurities;
- c) adding SnCl.sub.2 to the reduced protein in a sufficient amount to reduce a radionuclide anion selected from the group consisting of 99m-TcO.sub.4.sup.-, 186-REO.sub.4.sup.- and 188-REO.sub.4.sup.-, the radionuclide to be added in a subsequent step;
- and
- d) radio labeling the purified reduced protein by adding said radionuclide anion, whereby the SnCl.sub.2 reduces the radionuclide, and the reduced radionuclide and the sulfhydryl-containing immunoglobulin fragments form radionuclide-containing and sulfur-containing radio-labeled complexes,
- and wherein said method does not require conjugation to the protein of a chelator for ions of the radionuclide and does not require purification after addition of SnCl.sub.2.
Dependencies are inferred from the claim text.