Novel chelating agents for potential clinical applications of copper

https://doi.org/10.1016/S0969-8051(01)00287-6Get rights and content

Abstract

Copper offers a unique selection of radioisotopes (60Cu, 61Cu, 62Cu, 64Cu, and 67Cu) with half-lives ranging from 9.8 min to 61.9 h suitable for imaging and/or radiotherapy. In peptide/antibody targeted radiotherapy one of the most studied chelating agents for copper, 1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid (TETA), has been employed in clinical trials, but transchelation to ceruloplasmin and/or superoxide dismutase in vivo has been noted. In this study, a series of novel hexadentate chelating agents based on N,N′,N″-tris(2-pyridylmethyl)-1,3,5-cis,cis,-triaminocyclohexane (tachpyr) have been synthesized and the serum stability of their copper complexes was evaluated as compared to TETA. Copper complexes of tachpyr modified at the 3, 4, or 5 position or with replacement of pyridine by imidazole have serum stability comparable to Cu[TETA]. When the complexes were cross-challenged, Cu[TETA] versus tachpyr or 1,3,5-cis,cis,-triaminocyclohexane- N,N′,N″-tris-(2-methyl-(N-methylimidazole)) (IM), tachpyr and IM appear to have superior copper chelation ability to TETA. When challenged by a large excess of non-radioactive copper, copper exchange with the tachpyr radio-copper complex was observed. However, tachpyr clearly exhibited a significant preference for Cu(II) over Zn(II) or Fe(III). Therefore, tachpyr, 1,3,5-cis,cis,-triaminocyclohexane-N,N′,N″-tri-(3-methyl-2-methylpyridineimine) (tachpyr(3-Me)), 1,3,5-cis,cis,-triaminocyclohexane-N,N′,N″-tri-(4-methyl-2-methylpyridineimine) (tachpyr(4-Me)), 1,3,5-cis,cis,-triaminocyclohexane-N,N′,N″-tri-(5-methyl-2-methylpyridineimine) (tachpyr(5-Me)) and IM easily form copper complexes with high stability. These novel chelating agents provide an attractive lead for creation of new copper radiopharmaceuticals for diagnosis and therapy applications.

Introduction

Copper radioisotopes offer an attractive basis for the development of radiopharmaceuticals. Potentially useful copper radionuclides include 60Cu, 61Cu, 62Cu, 64Cu, and 67Cu, which provide a wide selection of nuclear properties for imaging and/or therapy [6]. Availability of these isotopes for pre-clinical and clinical research has greatly improved in recent years [18], [24], [25]. Numerous developments have led to various applications for copper radioisotopes such as myocardial perfusion agents [15] and hypoxia imaging agents [13], [39], detection of multi-drug resistance [19], peptide targeted imaging and therapy [3], [21], hypoxia targeted therapy [20], radioimmunoimaging [1], [42], and radioimmunotherapy [14], [28], [29].

For peptide and antibody-targeted imaging and therapy, it is highly desirable that the complex of radioisotopes such as 64Cu and 67Cu be stable in vivo in order to minimize release of the isotope to normal tissue thus reducing the background and minimizing radiation exposure to normal tissue. Among chelating agents, 1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid (TETA) has been reported to form a stable copper complex in an in vitro serum stability study [9] and its use as a chelating agent for copper radiopharmaceuticals has advanced to clinical trials [2], [28]. However, evidence of transchelation of radio-copper to ceruloplasmin in the plasma of lymphoma patients from the TETA antibody conjugate (2IT-BAT-Lym-1) has been presented [26]. It has also been reported that Cu(II) dissociates from TETA-Octreotide, and binds to the protein superoxide dismutase in rat liver [5]. Additionally, the work of Pippin [37] demonstrated Cu[TETA] to be relatively labile under mild acid conditions. Thus, development of novel, more stable, chelating agents for use of 64Cu and 67Cu for imaging or therapy is an ongoing effort [4], [12], [17].

Novel hexadentate chelating agents based on cis,cis-1,3,5-triaminocyclohexane (tach) have been reported for their chelation properties with different transition metal ions [7], [16], [22], [31], [32], [34]. Recently, we have reported on Cu(II) complexation with N,N′,N″-tris(2-pyridylmethyl)-1,3,5-cis,cis,-triaminocyclohexane (tachpyr, 1), and the stability in serum for its potential use as a radiopharmaceutical [30]. Herein, we report the synthesis of a series of novel hexadentate chelating agents (1–10) illustrated in Fig. 1 based on tachpyr and its analog, 1,3,5-cis,cis,-triaminocyclohexane- N,N′,N″-tris-(2-methyl-(N-methylimidazole)) (IM, 10), the evaluation of this series of new chelates, their complexation characteristics with 64Cu and/or 67Cu and the stability of complexes in human serum in comparison to macrocyclic polyaminocarboxylate TETA (11) and 1,4,7,10,13,16-hexaazacyclooctadecane-1,4,7,10,13,16-hexaacetic acid (HEHA, 12).

Section snippets

Materials and methods

64Cu and 67Cu were purchased from Washington University (St. Louis, MO) and Brookhaven National Laboratory (Upton, NY), respectively. Human serum was purchased from Gemini Bio-Products (Calabasas, CA) and stored at −20°C. The serum was centrifuged, passed through a 0.22 μm filter, stored at 4°C, and used within 24 h for in vitro serum stability studies. Instant thin layer chromatography (ITLC) silica gel impregnated glass fiber was purchased from Gelman Sciences (Ann Arbor, MI). Silica gel

Organic synthesis

The syntheses of the novel ligands in this study ultimately began with the base framework of tach which has until recently been problematic to routinely prepare in reasonable amounts and purity [7].

With this triamine in hand, the pyridyl methyl substituted analogs of tachpyr, 2–6, were reasonably prepared by first forming the corresponding imines with subsequent reduction in good yield with borohydride. Isolation generally did not require chromatography and overall yields were reasonable.

Discussion

67Cu has a 62-h physical half-life, a 185 keV gamma emission that is suitable for patient imaging, a therapeutic β emission (Emax = 577 keV), and for these reasons has been described as an ideal therapeutic isotope for radioimmunotherapy [29]. A clinical trial using 67Cu 2IT-BAT-Lym-1 for treatment of non-Hodgkin’s lymphoma has demonstrated favorable imaging, dosimetry, and therapeutic effectiveness [14]. However, transchelation of 67Cu from the TETA chelate conjugate of 2IT-BAT-Lym-1 to

Conclusion

All of the tachpyr ligands evaluated instantly formed copper complexes with 100% yield based on the uptake of radioactivity. However, modification at the 6-position of pyridine ring or at the secondary amine on tachpyr made the complex less stable. Copper complexes of tachpyr modified at the 3, 4, or 5 position or with replacement of pyridine by imidazole have serum stability comparable to Cu[TETA]. When the complexes were cross-challenged between TETA and tachpyr or IM, tachpyr and IM appear

Acknowledgements

We thank R. Planalp for his helpful discussions concerning the Cu(II) coordination chemistry of tachpyr for this study.

References (42)

  • N Petrovic et al.

    Copper incorporation into superoxide dismutase in menkes lymphoblasts

    J. Biol. Chem.

    (1996)
  • N Petrovic et al.

    Identification of an apo-superoxide dismutase (Cu,Zn) pool in human lymphoblasts

    J. Biol. Chem.

    (1996)
  • S.V Torti et al.

    Tumor cell cytotoxicity of a novel metal chelator

    Blood

    (1998)
  • C.J Anderson et al.

    Copper-64-labeled antibodies for PET imaging

    J. Nucl. Med.

    (1992)
  • C.J Anderson et al.

    64Cu-TETA-Octreotide as a PET imaging agent for patients with neuroendocrine tumors

    J. Nucl. Med.

    (2001)
  • C.J Anderson et al.

    Radiotherapy, toxicity and dosimetry of copper-64-TETA- octreotide in tumor-bearing rats

    J. Nucl. Med.

    (1998)
  • P Barbaro et al.

    Synthesis and characterization of the tetraazamacrocycle 4,10- dimethyl-1,4,7,10-tetraazacyclododecane-1,7-diacetic acid (H2Me2DO2A) and of its neutral copper(II) complex Cu(Me2DO2A). A new 64Cu-labeled macrocyclic complex for positron emission tomography imaging

    J. Chem .Soc.-Dalton. Trans.

    (2000)
  • L.A Bass et al.

    In vivo transchelation of copper-64 from TETA-octreotide to superoxide dismutase in rat liver

    Bioconjugate Chem.

    (2000)
  • L.L Chappell et al.

    Synthesis, conjugation, and radiolabeling of a novel bifunctional chelating agent for 225Ac radioimmunotherapy applications

    Bioconjugate Chem.

    (2000)
  • D.L Comins et al.

    Lithiation of methoxypyridines directed by alpha-amino alkoxides

    J. Org. Chem.

    (1990)
  • J.M Connett et al.

    Radioimmunotherapy with a 64Cu-labeled monoclonal antibodya comparison with, 67Cu

    Proc. Natl. Acad. Sci. U. S. A.

    (1996)
  • Cited by (0)

    View full text