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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1992 Oct 15;89(20):9749–9753. doi: 10.1073/pnas.89.20.9749

Recombinant antibody-metallothionein: design and evaluation for radioimmunoimaging.

C Das 1, P V Kulkarni 1, A Constantinescu 1, P Antich 1, F R Blattner 1, P W Tucker 1
PMCID: PMC50210  PMID: 1409693

Abstract

We have produced a chimeric antibody (Ab) in which metallothionein, a well-characterized biological chelator of metals, was genetically fused to the F(ab') domain of the S107 Ab heavy chain. Coexpression with the Ab light chain that conveys specificity for the synthetic antigen phosphocholine was achieved in plasmacytoma cells. Metal- and antigen-binding domains of the Ab-metallothionein hybrid function with normal avidity and specificity. Ab-metallothionein can be efficiently loaded with 99mTc and used to specifically bind phosphocholine-haptenated cells in vitro or to localize plasma-cell ascites tumors in mice. The approach offers potential advantages for producing radiolabeled Ab for targeted radiotherapy and diagnostic imaging.

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Selected References

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  1. Barbosa M. S., Lowy D. R., Schiller J. T. Papillomavirus polypeptides E6 and E7 are zinc-binding proteins. J Virol. 1989 Mar;63(3):1404–1407. doi: 10.1128/jvi.63.3.1404-1407.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Batra J. K., Fitzgerald D. J., Chaudhary V. K., Pastan I. Single-chain immunotoxins directed at the human transferrin receptor containing Pseudomonas exotoxin A or diphtheria toxin: anti-TFR(Fv)-PE40 and DT388-anti-TFR(Fv). Mol Cell Biol. 1991 Apr;11(4):2200–2205. doi: 10.1128/mcb.11.4.2200. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berg J., Lötscher E., Steimer K. S., Capon D. J., Baenziger J., Jäck H. M., Wabl M. Bispecific antibodies that mediate killing of cells infected with human immunodeficiency virus of any strain. Proc Natl Acad Sci U S A. 1991 Jun 1;88(11):4723–4727. doi: 10.1073/pnas.88.11.4723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brown B. A., Drozynski C. A., Dearborn C. B., Hadjian R. A., Liberatore F. A., Tulip T. H., Tolman G. L., Haber S. B. Conjugation of metallothionein to a murine monoclonal antibody. Anal Biochem. 1988 Jul;172(1):22–28. doi: 10.1016/0003-2697(88)90406-x. [DOI] [PubMed] [Google Scholar]
  5. Brzezinska M., Benveniste R., Davies J., Daniels P. J., Weinstein J. Gentamicin resistance in strains of Pseudomonas aeruginosa mediated by enzymatic N-acetylation of the deoxystreptamine moiety. Biochemistry. 1972 Feb 29;11(5):761–765. doi: 10.1021/bi00755a013. [DOI] [PubMed] [Google Scholar]
  6. Burchiel S. W., Hadjian R. A., Hladik W. B., Drozynski C. A., Tolman G. L., Haber S. B., Gallagher B. M. Pharmacokinetic evaluation of technetium-99-metallothionein-conjugated mouse monoclonal antibody B72.3 in rhesus monkeys. J Nucl Med. 1989 Aug;30(8):1351–1357. [PubMed] [Google Scholar]
  7. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  8. Chu G., Hayakawa H., Berg P. Electroporation for the efficient transfection of mammalian cells with DNA. Nucleic Acids Res. 1987 Feb 11;15(3):1311–1326. doi: 10.1093/nar/15.3.1311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Desaymard C., Giusti A. M., Scharff M. D. Rat anti-T15 monoclonal antibodies with specificity for VH- and VH-VL epitopes. Mol Immunol. 1984 Oct;21(10):961–967. doi: 10.1016/0161-5890(84)90154-8. [DOI] [PubMed] [Google Scholar]
  10. Engvall E., Perlmann P. Enzyme-linked immunosorbent assay (ELISA). Quantitative assay of immunoglobulin G. Immunochemistry. 1971 Sep;8(9):871–874. doi: 10.1016/0019-2791(71)90454-x. [DOI] [PubMed] [Google Scholar]
  11. Iscove N. N., Guilbert L. J., Weyman C. Complete replacement of serum in primary cultures of erythropoietin-dependent red cell precursors (CFU-E) by albumin, transferrin, iron, unsaturated fatty acid, lecithin and cholesterol. Exp Cell Res. 1980 Mar;126(1):121–126. doi: 10.1016/0014-4827(80)90476-0. [DOI] [PubMed] [Google Scholar]
  12. Karin M., Richards R. I. Human metallothionein genes--primary structure of the metallothionein-II gene and a related processed gene. Nature. 1982 Oct 28;299(5886):797–802. doi: 10.1038/299797a0. [DOI] [PubMed] [Google Scholar]
  13. Kearney J. F., Barletta R., Quan Z. S., Quintáns J. Monoclonal vs. heterogeneous anti-H-8 antibodies in the analysis of the anti-phosphorylcholine response in BALB/c mice. Eur J Immunol. 1981 Nov;11(11):877–883. doi: 10.1002/eji.1830111106. [DOI] [PubMed] [Google Scholar]
  14. Kirsch I. R., Ravetch J. V., Kwan S. P., Max E. E., Ney R. L., Leder P. Multiple immunoglobulin switch region homologies outside the heavy chain constant region locus. Nature. 1981 Oct 15;293(5833):585–587. doi: 10.1038/293585a0. [DOI] [PubMed] [Google Scholar]
  15. Kreitman R. J., Chaudhary V. K., Waldmann T., Willingham M. C., FitzGerald D. J., Pastan I. The recombinant immunotoxin anti-Tac(Fv)-Pseudomonas exotoxin 40 is cytotoxic toward peripheral blood malignant cells from patients with adult T-cell leukemia. Proc Natl Acad Sci U S A. 1990 Nov;87(21):8291–8295. doi: 10.1073/pnas.87.21.8291. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kuziel W. A., Takashima A., Bonyhadi M., Bergstresser P. R., Allison J. P., Tigelaar R. E., Tucker P. W. Regulation of T-cell receptor gamma-chain RNA expression in murine Thy-1+ dendritic epidermal cells. Nature. 1987 Jul 16;328(6127):263–266. doi: 10.1038/328263a0. [DOI] [PubMed] [Google Scholar]
  17. Kwan S. P., Rudikoff S., Seidman J. G., Leder P., Scharff M. D. Nucleic acid and protein sequences of phosphocholine-binding light chains. J Exp Med. 1981 May 1;153(5):1366–1370. doi: 10.1084/jem.153.5.1366. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lees R. S., Garabedian H. D., Lees A. M., Schumacher D. J., Miller A., Isaacsohn J. L., Derksen A., Strauss H. W. Technetium-99m low density lipoproteins: preparation and biodistribution. J Nucl Med. 1985 Sep;26(9):1056–1062. [PubMed] [Google Scholar]
  19. Lieberman R., Potter M., Mushinski E. B., Humphrey W., Jr, Rudikoff S. Genetics of a new IgVH (T15 idiotype) marker in the mouse regulating natural antibody to phosphorylcholine. J Exp Med. 1974 Apr 1;139(4):983–1001. doi: 10.1084/jem.139.4.983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Morelock M. M., Tolman G. L. Metallothionein: a bifunctional chelator for the radiolabeling of biologically active molecules. Experientia Suppl. 1987;52:247–253. doi: 10.1007/978-3-0348-6784-9_19. [DOI] [PubMed] [Google Scholar]
  21. Neuberger M. S., Williams G. T., Fox R. O. Recombinant antibodies possessing novel effector functions. Nature. 1984 Dec 13;312(5995):604–608. doi: 10.1038/312604a0. [DOI] [PubMed] [Google Scholar]
  22. Poljak R. J., Amzel L. M., Avey H. P., Chen B. L., Phizackerley R. P., Saul F. Three-dimensional structure of the Fab' fragment of a human immunoglobulin at 2,8-A resolution. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3305–3310. doi: 10.1073/pnas.70.12.3305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Robertson S. M., Kettman J. R., Miller J. N., Norgard M. V. Murine monoclonal antibodies specific for virulent Treponema pallidum (Nichols). Infect Immun. 1982 Jun;36(3):1076–1085. doi: 10.1128/iai.36.3.1076-1085.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sawyer J. R., Tucker P. W., Blattner F. R. Metal-binding chimeric antibodies expressed in Escherichia coli. Proc Natl Acad Sci U S A. 1992 Oct 15;89(20):9754–9758. doi: 10.1073/pnas.89.20.9754. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Schnee J. M., Runge M. S., Matsueda G. R., Hudson N. W., Seidman J. G., Haber E., Quertermous T. Construction and expression of a recombinant antibody-targeted plasminogen activator. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6904–6908. doi: 10.1073/pnas.84.19.6904. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Tucker P. W., Marcu K. B., Newell N., Richards J., Blattner F. R. Sequence of the cloned gene for the constant region of murine gamma 2b immunoglobulin heavy chain. Science. 1979 Dec 14;206(4424):1303–1306. doi: 10.1126/science.117549. [DOI] [PubMed] [Google Scholar]
  27. Williams G. T., Neuberger M. S. Production of antibody-tagged enzymes by myeloma cells: application to DNA polymerase I Klenow fragment. Gene. 1986;43(3):319–324. doi: 10.1016/0378-1119(86)90223-4. [DOI] [PubMed] [Google Scholar]

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