Synthesis of aminobenzyltriethylenetetraamino-hexaacetic acid: Conjugation of the chelator to protein by an alkylamine linkage

Kuldeep K. Bhargava, Z. Y. Zhang, Christopher J. Palestro, Seetharama A. Acharya

Research output: Contribution to journalArticle

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Abstract

The conjugation of a chelating agent to an antibody as an anchoring site for a radionuclide is the first step in the successful preparation of a radiolabeled antibody for a diagnostic and therapeutic application. The high affinity of the protein bound chelator towards radionuclide ensures a higher selectivity in the delivery of the radionuclide to the targeted tissue. 4-Aminobenzylderivativetriethlenetetraaminohexaacetic acid (TTHA), a hexadentate chelating agent has been now prepared for conjugation with proteins in view of the higher affinity of TTHA metal ions as compared to DTPA. The latent crosslinking potential of α-hydroxy aldehydes has been used to conjugate the new chelating agent to proteins through an alkylamine linkage. On incubation of amino benzyl TTHA with glycoladehyde at neutral pH and room temperature, the reagent is converted to oxoethyl amino benzyl TTHA. On addition of albumin to this reaction mixture, the oxo ethylamino benzyl TTHA generates reversible schiff base adducts with the amino groups of albumin. The reduction of the Schiff base adducts of the chelator with the protein by sodium cyanoborohydride stabilizes the schiff base adducts as stable alkylamine linkages. 4-Thiocyanatobenzyl TTHA has also been prepared and conjugated to albumin through a thiocarbamoyl linkage. Both preparations of TTHA conjugated albumin complexed with 99mTc and 111In, with high affinity and no decomposition of the complex was noticed for at least up to 6 hrs after the preparation. The radiolabels complexed with these TTHA -albumin conjugates could not be 'chased' out by free DTPA. A comparison of the biodistribution of 111In, bound to the TTHA conjugated through an alkylamine and a thiocarbamoyl linkage showed that 111In complexed with alkylamine linked TTHA was retained in blood to a level nearly 17% higher compared to that seen with thicarbamoyl linked TTHA, one hr after the injection into mice. Thus, the alkylamine linkage appears to be more stable under the in vivo conditions. The glycolaldehyde mediated alkylation procedure offers a mild, simple and rapid method for preparation of drug-protein (antibody) conjugates.

Original languageEnglish (US)
Pages (from-to)761-770
Number of pages10
JournalProtein Journal
Volume18
Issue number7
StatePublished - 1999

Fingerprint

Chelating Agents
Proteins
Chelation
Acids
Radioisotopes
Antibodies
Albumins
Schiff Bases
Pentetic Acid
Alkylation
Aldehydes
Crosslinking
Metal ions
Blood
Sodium
triethylenetetraminehexaacetic acid
Tissue
Decomposition
Drug Compounding
Metals

Keywords

  • α-hydroxy aldehydes
  • Amadori rearrangement
  • Chelation
  • Conjugation

ASJC Scopus subject areas

  • Biochemistry
  • Analytical Chemistry
  • Organic Chemistry
  • Bioengineering

Cite this

Synthesis of aminobenzyltriethylenetetraamino-hexaacetic acid : Conjugation of the chelator to protein by an alkylamine linkage. / Bhargava, Kuldeep K.; Zhang, Z. Y.; Palestro, Christopher J.; Acharya, Seetharama A.

In: Protein Journal, Vol. 18, No. 7, 1999, p. 761-770.

Research output: Contribution to journalArticle

Bhargava, Kuldeep K. ; Zhang, Z. Y. ; Palestro, Christopher J. ; Acharya, Seetharama A. / Synthesis of aminobenzyltriethylenetetraamino-hexaacetic acid : Conjugation of the chelator to protein by an alkylamine linkage. In: Protein Journal. 1999 ; Vol. 18, No. 7. pp. 761-770.
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AU - Acharya, Seetharama A.

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KW - α-hydroxy aldehydes

KW - Amadori rearrangement

KW - Chelation

KW - Conjugation

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