Journal of Nuclear Science, Engineering and Technology (JONSAT)
In cooperation with the Iranian Nuclear Society

Production, Quality Control and Biodistribution Study of F(ab')2 Fragment of Antibody PR81 Labeled with 99mTc for Diagnosis of Breast Carcinoma Bearing Mice

Document Type : Research Paper

Authors

M Salouti
M.H Babaei
H Rajabi
H Forootan
M.J Rasaee
A Bitarafan Rajabi
F Johari Daha
M Mazidi
J Mohammad Nejad
M Shafiee
Abstract
Immunoglobulin fragments offer a number of advantages for intratumor penetration over the intact antibodies. The goal of this study was to prepare F(ab')2 fragmants from antibody PR81 and 99mTc labeling in order to produce a biological radiopharmaceutical for imaging of breast cancer in nuclear medicine. The antibody PR81 was digested with 5% (w/w) pepsin at different times to find the best time for a complete digestion. The prepared F(ab')2 fragments were purified by protein A column chromatography. The immunoreactivity of the complex assessed by radioimmunoassay was determined to be 65.2% ± 5.1. 99mTc radiolabeling of F(ab')2 fragment by using HYNIC as a chelator and tricine as a
co-ligand resulted in a radiochemical purity of 70.1%±5.2 after 1 hr with a high stability. The complex was purified by the gel chromatography method to increase the radiochemical purity. Biodistribution and imaging studies in BALB/c mice with breast tumor xenograft demonstrated as a specific localization of the compound at the site of tumors 4 h post injection with high sensitivity and minimum accumulation in non target organs. The results showed that F(ab')2 fragment of PR81 is a promising candidate for radioimmunoscintigraphy of the human breast carcinoma.

Highlights

  1. 1.    جهت دریافت کارشناسی ارشد مامایی، دانشگاه تربیت مدرس (سال 1382).                                                                              

 

  1. 2.    M. Paknejad, M.J. Rasaee, F. Karami, S. Kashanian, M.A. Mohagheghi, K. Omidfar, “Production of monoclonal antibody, PR81, recognizing the tandem repeat region of MUC1 mucin,” Hybridoma and Hybridomics, 22(3): 153-158 (2003).

 

  1. 3.    M. Salouti, H. Rajabi, H. Babaei, M.J. Rasaee, R. Najafi, M. Paknejad, Z. Mohammad Hasan, T.M. Altarihi, “99mTc direct radiolabeling of PR81, a new anti-MUC1 monoclonal antibody for radioimmunoscintigraphy,” Iran J. N. M, 23: 7-16 (2005).

 

  1. 4.    M. Salouti, H. Rajabi, M.H. Babaei, M.J. Rasaee, R. Najafi, M. Paknejad, Z. Mohammad Hasan, T.M. Altarihi, J. Mohammad Nejad, “A New Monoclonal Antibody Radiopharmaceutical for Radioimmunoscintigraphy of Breast Cancer: Direct Labeling of Antibody and Its Quality Control,” DARU; 14(1): 14-19 (2006).

 

  1. 5.    M. Salouti, H. Rajabi, M.H. Babaei, M.J. Rasaee, R. Najafi, M. Shafiee, M. Mazidi, Z.M. Hasan, A. Bitarafan Rajabi, N. Namvar, T.M. Altarihi, Mohammad Nejad, “Radioimmunoscintigraphy of Breast Tumor Xenografts in mouse model by 99mTc Direct Radiolabeling of a Monoclonal Antibody PR81,” The Iranian Journal of Medical Physics, 2(8): 27-34, 60 (2005).

 

  1. 6.    M. Salouti, H. Rajabi, M.H. Babaei, M.J. Rasaee, R. Najafi, M. Shafiee, M. Mazidi, Z.M. Hasan, A. Bitarafan Rajabi, N. Namvar, T.M. Altarihi, Mohammad Nejad,

 

  1. 7.    S. Potamianos, A.D. Varvarigou, S.C. Archimandritis, “Radioimmunoscintigraphy and radioimmunotherapy in cancer: principles and application,” Anticancer Research, 20: 925– 948 (2000).

 

 

  1. 8.    M. Mariani, M. Camagna, L. Tarditi, E. Seccamani, “A new enzymatic method to obtain high–yield F(ab')2 suitable for clinical use from mouse IgG1,” Molecular Immunology, 28(1/2): 69-77 (1991).

 

  1. 9.    D.W. Rea, M.E. Ultee, “A novel method for controlling the pepsin digestion of antibodies,” JIM, 157: 165-173 (1993).

 

10. S. Demignot, M.C. Garnett, R.W. Baldwin, “Mouse IgG2b monoclonal antibody fragmentation. Preparation and purification of Fab, Fc and Fab/c fragments,” JIM, 121: 209-217 (1989).

 

11. D.E. Milenic, J.M. Esteban, D. Colcher, “Comparison of methods for the generation of immunoreactive fragments of a monoclonal antibody (B72.3) reactive with human carcinomas,” JIM, 120: 71-83 (1989).

 

12. R. Kurkela, L. Vuolas, P. Vihko, “Preparation of F(ab')2 fragments from monoclonal mouse IgG1 suitable for use in radioimmunoimaging,” JIM,  110: 229-236 (1988).

 

13. T. Ghazanfari, Z. Mohammad, R. Yaraie, “Recognition and determination of immunogenicity of a spontaneous tumor in a BALB/c mouse,” Daneshvar, 24: 65-72 (1998).

 

14. B.A. Rhodes, D.A. Torvestad, K. Breslow, “99mTc labeling and acceptance testing of radiolabeled antibodies and antibody fragments. In: Burcheil SW, Rhodes BA, editors. Tumor Imaging. New York: Mason, 111 (1982).

 

15. M.H. Babaei, R. Farshidfar, R. Najafi, “Evaluation of tumor targeting with radiolabeled F(AB')2 fragment of a humanized monoclonal antibody,” DARU, 10(2): 49-53 (2002).

 

16. T.M. Behr, S. Memtsoudis, R.M. Sharkey, R.D. Blumenthal, R.M. Dunn, S. Gratz, “Experimental studies on the role of antibody fragments in cancer radio-immunotherapy: influence of radiation dose and dose rate on toxicity and anti-tumor efficacy,” Int. J. Cancer, 77: 787-795 (1998).

17. R. Kurkela, L. Vuolas, P. Vihko, “Preparation of F(ab')2 fragments from monoclonal mouse IgG1 suitable for use in radioimaging,” JIM, 110: 229-236 (1988).

 

18. H.J. Haisma, J. Hilgers, V.R. Zurawski, “Iodination of monoclonal antibodies for diagnosis and radiotherapy using a convenient one vial method,” J. Nucl. Med. 27: 1890-1895 (1986).

 

19. T. Lindmo, E. Boven, F. Cuttitta, J. Fedorko, P.J. Bunn, “Determination of the immunoreactive fraction of radiolabeled monoclonal antibodies by linear extrapolation of binding at infinite antigen excess,” J. Immunol Methods, 65: 55-63 (1983).

 

20. M.J. Abrams, M. Juweid, C.I. TenKate, “Technetium-99m human polyclonal IgG radiolabeled via the hydrazino nicotinamide derivative for imaging focal sites of infection in rats,” J. Nucl. Med. 31: 2022-2028 (1990).

 

21. A. Johnstone, R. Thorpe, “Basic techniques. In: Johnstone A, Thorpe R, editors. Immunochemistry in practice. Blackwell Science Ltd, 1-34 (1996).

 

22. H.J. Lee, W. Pardridge, “Monoclonal antibody radiopharmaceuticals: cationization, pegylation, radiometal chelation, pharmacokinetics, and tumor imaging,” Bioconjugate Chem, 14: 546-553 (2003).

 

23. M.J. Verhaar, B.A. Zonnenberg, J.M.H. Klerk, G.H. Blijham, “Radioimmunodiagnosis and therapy,” Cancer Treatment Reviews, 26: 3-10 (2000).

 

24. Y. Yamaguchi, H. Kim, K. Kato, K. Masuda, I. Shimada, Y. Arata, “Proteolytic fragmentation with high specificity of mouse immunoglobulin,” G. JIM, 181: 259-267 (1995).

 

25. L.M. Lamki, “Radioimmuniscintigraphy of cancer, problems, pitfalls and prospects. In: Freeman, L.M. (ed) Nuclear Medicine Annual 1990, Raven Press, New York, 113-150 (1990).

 

26. A.M. Verbruggen, “Radiopharmaceuticals: state of the art,” Eur. J. Nucl. Med. 17: 346-364 (1990).

 

27. S.W. Schwarz, J.M. Connett, C.J. Anderson, et al, “Evaluation of a direct method for technetium labeling intact and F(ab')2 1A3, an anticolorectal monoclonal antibody,” Nucl. Med. Biol. 21: 619-626 (1994).

 

28. S.K. Larsen, G. Caldwell, J.D. Higgins, M.S. Abram, H.F. Solomon, “Technetium complex of tricine: useful precursor for the 99mTc labelling of hydrazino nicotinamide modified proteins,” J. Label Compd Radiopharm, 35: 1-2 (1994).

 

29. B.W. Wessels, “Current status of animal radioimmunotherapy,” Cancer Res, 50, 970-973 (1990).

Keywords

Breast Carcinoma
Radioimmunoscintigraphy
Antibody PR81
F(ab′)2
Fragment
99mTc Labeling
  1. 1.    جهت دریافت کارشناسی ارشد مامایی، دانشگاه تربیت مدرس (سال 1382).                                                                              

 

  1. 2.    M. Paknejad, M.J. Rasaee, F. Karami, S. Kashanian, M.A. Mohagheghi, K. Omidfar, “Production of monoclonal antibody, PR81, recognizing the tandem repeat region of MUC1 mucin,” Hybridoma and Hybridomics, 22(3): 153-158 (2003).

 

  1. 3.    M. Salouti, H. Rajabi, H. Babaei, M.J. Rasaee, R. Najafi, M. Paknejad, Z. Mohammad Hasan, T.M. Altarihi, “99mTc direct radiolabeling of PR81, a new anti-MUC1 monoclonal antibody for radioimmunoscintigraphy,” Iran J. N. M, 23: 7-16 (2005).

 

  1. 4.    M. Salouti, H. Rajabi, M.H. Babaei, M.J. Rasaee, R. Najafi, M. Paknejad, Z. Mohammad Hasan, T.M. Altarihi, J. Mohammad Nejad, “A New Monoclonal Antibody Radiopharmaceutical for Radioimmunoscintigraphy of Breast Cancer: Direct Labeling of Antibody and Its Quality Control,” DARU; 14(1): 14-19 (2006).

 

  1. 5.    M. Salouti, H. Rajabi, M.H. Babaei, M.J. Rasaee, R. Najafi, M. Shafiee, M. Mazidi, Z.M. Hasan, A. Bitarafan Rajabi, N. Namvar, T.M. Altarihi, Mohammad Nejad, “Radioimmunoscintigraphy of Breast Tumor Xenografts in mouse model by 99mTc Direct Radiolabeling of a Monoclonal Antibody PR81,” The Iranian Journal of Medical Physics, 2(8): 27-34, 60 (2005).

 

  1. 6.    M. Salouti, H. Rajabi, M.H. Babaei, M.J. Rasaee, R. Najafi, M. Shafiee, M. Mazidi, Z.M. Hasan, A. Bitarafan Rajabi, N. Namvar, T.M. Altarihi, Mohammad Nejad,

 

  1. 7.    S. Potamianos, A.D. Varvarigou, S.C. Archimandritis, “Radioimmunoscintigraphy and radioimmunotherapy in cancer: principles and application,” Anticancer Research, 20: 925– 948 (2000).

 

 

  1. 8.    M. Mariani, M. Camagna, L. Tarditi, E. Seccamani, “A new enzymatic method to obtain high–yield F(ab')2 suitable for clinical use from mouse IgG1,” Molecular Immunology, 28(1/2): 69-77 (1991).

 

  1. 9.    D.W. Rea, M.E. Ultee, “A novel method for controlling the pepsin digestion of antibodies,” JIM, 157: 165-173 (1993).

 

10. S. Demignot, M.C. Garnett, R.W. Baldwin, “Mouse IgG2b monoclonal antibody fragmentation. Preparation and purification of Fab, Fc and Fab/c fragments,” JIM, 121: 209-217 (1989).

 

11. D.E. Milenic, J.M. Esteban, D. Colcher, “Comparison of methods for the generation of immunoreactive fragments of a monoclonal antibody (B72.3) reactive with human carcinomas,” JIM, 120: 71-83 (1989).

 

12. R. Kurkela, L. Vuolas, P. Vihko, “Preparation of F(ab')2 fragments from monoclonal mouse IgG1 suitable for use in radioimmunoimaging,” JIM,  110: 229-236 (1988).

 

13. T. Ghazanfari, Z. Mohammad, R. Yaraie, “Recognition and determination of immunogenicity of a spontaneous tumor in a BALB/c mouse,” Daneshvar, 24: 65-72 (1998).

 

14. B.A. Rhodes, D.A. Torvestad, K. Breslow, “99mTc labeling and acceptance testing of radiolabeled antibodies and antibody fragments. In: Burcheil SW, Rhodes BA, editors. Tumor Imaging. New York: Mason, 111 (1982).

 

15. M.H. Babaei, R. Farshidfar, R. Najafi, “Evaluation of tumor targeting with radiolabeled F(AB')2 fragment of a humanized monoclonal antibody,” DARU, 10(2): 49-53 (2002).

 

16. T.M. Behr, S. Memtsoudis, R.M. Sharkey, R.D. Blumenthal, R.M. Dunn, S. Gratz, “Experimental studies on the role of antibody fragments in cancer radio-immunotherapy: influence of radiation dose and dose rate on toxicity and anti-tumor efficacy,” Int. J. Cancer, 77: 787-795 (1998).

17. R. Kurkela, L. Vuolas, P. Vihko, “Preparation of F(ab')2 fragments from monoclonal mouse IgG1 suitable for use in radioimaging,” JIM, 110: 229-236 (1988).

 

18. H.J. Haisma, J. Hilgers, V.R. Zurawski, “Iodination of monoclonal antibodies for diagnosis and radiotherapy using a convenient one vial method,” J. Nucl. Med. 27: 1890-1895 (1986).

 

19. T. Lindmo, E. Boven, F. Cuttitta, J. Fedorko, P.J. Bunn, “Determination of the immunoreactive fraction of radiolabeled monoclonal antibodies by linear extrapolation of binding at infinite antigen excess,” J. Immunol Methods, 65: 55-63 (1983).

 

20. M.J. Abrams, M. Juweid, C.I. TenKate, “Technetium-99m human polyclonal IgG radiolabeled via the hydrazino nicotinamide derivative for imaging focal sites of infection in rats,” J. Nucl. Med. 31: 2022-2028 (1990).

 

21. A. Johnstone, R. Thorpe, “Basic techniques. In: Johnstone A, Thorpe R, editors. Immunochemistry in practice. Blackwell Science Ltd, 1-34 (1996).

 

22. H.J. Lee, W. Pardridge, “Monoclonal antibody radiopharmaceuticals: cationization, pegylation, radiometal chelation, pharmacokinetics, and tumor imaging,” Bioconjugate Chem, 14: 546-553 (2003).

 

23. M.J. Verhaar, B.A. Zonnenberg, J.M.H. Klerk, G.H. Blijham, “Radioimmunodiagnosis and therapy,” Cancer Treatment Reviews, 26: 3-10 (2000).

 

24. Y. Yamaguchi, H. Kim, K. Kato, K. Masuda, I. Shimada, Y. Arata, “Proteolytic fragmentation with high specificity of mouse immunoglobulin,” G. JIM, 181: 259-267 (1995).

 

25. L.M. Lamki, “Radioimmuniscintigraphy of cancer, problems, pitfalls and prospects. In: Freeman, L.M. (ed) Nuclear Medicine Annual 1990, Raven Press, New York, 113-150 (1990).

 

26. A.M. Verbruggen, “Radiopharmaceuticals: state of the art,” Eur. J. Nucl. Med. 17: 346-364 (1990).

 

27. S.W. Schwarz, J.M. Connett, C.J. Anderson, et al, “Evaluation of a direct method for technetium labeling intact and F(ab')2 1A3, an anticolorectal monoclonal antibody,” Nucl. Med. Biol. 21: 619-626 (1994).

 

28. S.K. Larsen, G. Caldwell, J.D. Higgins, M.S. Abram, H.F. Solomon, “Technetium complex of tricine: useful precursor for the 99mTc labelling of hydrazino nicotinamide modified proteins,” J. Label Compd Radiopharm, 35: 1-2 (1994).

 

29. B.W. Wessels, “Current status of animal radioimmunotherapy,” Cancer Res, 50, 970-973 (1990).

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