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Sensitive enzyme immunoassay for the quantification of aclacinomycin A usingβ-D-galactosidase as a label

  • Original Articles
  • Immunoassay Aclacinomycin A
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Summary

A sensitive enzyme immunoassay method (EIA) for an anticancer drug, aclacinomycin A (ACM), has been developed.With a double-antibody technique, ACM at a concentration as low as 100 pg/tube can be detected. An antibody to ACM was obtained by immunizing rabbits with an antigen prepared by coupling ACM with mercaptosuccinylated bovine serum albumin via N-maleoyl aminobutyric acid (MABA) as a coupling agent. Enzyme labeling of ACM was performed withβ-D-galactosidae (β-Gal; EC 3.2.1.23) via m-maleoyl benzoic acid (MBA). The standard curve of the assay was linear on a logit-log plot over a concentrationrange of 30 pg to 10 ng. The antibody detected ACM and its metabolites, MA144 M1 (M1), MA144 N1 (N1), MA144 S1 (S1), and aklavin (T1) equally well, but was only minimally reactive with aklavinone (D1) and 7-deoxyaklavinone (C1), thus suggesting that this EIA can detect the total amounts of ACM and its biologically active glycosides among metabolites of ACM. This EIA is practically free from interference by any other anticancer drugs. Using this assay, serum levels of ACM equivalents can be determined accurately after administration of the drug to rats at a single dose of 10 mg/kg. Since ACM is now undergoing clinical trial, the EIA of the drug will be a valuable tool in clinical pharmacological studies.

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References

  1. Anderson GW, Zimmerman JE, Callahan FM (1967) A reinvestigation of the mixed carbonic anhydride method of peptide synthesis. J Am Chem Soc 89:5012

    Google Scholar 

  2. Craven GR, Steers E, Anfinsen CB (1965) Purification, comparison, and molecular weight ofβ-D-galactosidase ofEscherichia coli K 12. J Biol Chem 240:2468

    Google Scholar 

  3. Egorin MJ, Van Echo DA, Whitacre MY, Fox BM, Aisner J, Wiernik PH, Bachur NR (1981) A phase 1 trial of aclacinomycin A. Proc AACR/ASCO 22:353

    Google Scholar 

  4. Egorin MJ, Echo DV, Fox BM, Whitacre M, Bachur NR (1982) Plasma kinetics of aclacinomycin A and its major metabolites in man. Cancer Chemother Pharmacol 8:41

    Google Scholar 

  5. Ellman GL (1962) Tissue sulfhydryl groups. Arch Biochem Biophys 82:70

    Google Scholar 

  6. Fujiwara K, Saikusa H, Yasuno M, Kitagawa T (1982) Enzyme immunoassay for the quantification of mitomycin C usingβ-galactosidase as a label. Cancer Res 42:1487

    Google Scholar 

  7. Fujiwara K, Asada H, Kitagawa T, Yamamoto K, Ito T, Tsuchiya R, Sohda M, Nakamura N, Hara K, Tomonaga Y, Ichimaru M, Takahashi S (1983) Preparation of polyamine antibody and its use in enzyme immunoassay of spermine and spermidine withβ-D-galactosidase as a label. J Immunol Methods 61:217

    Google Scholar 

  8. Furue H, Komita T, Nakao I, Furukawa I, Kanko T, Yokoyama T (1978) Clinical experiences with aclacinomycin-A. Recent Results Cancer Res 63:241

    Google Scholar 

  9. Hori S, Shirai M, Hirano S, Oki T, Inui T, Tsukagoshi S, Ishizuka M, Takeuchi T, Umezawa H (1977) Antitumor activity of new anthracycline antibiotics, aclacinomycin A and its analogs, and their toxicity. Gann 68:685

    Google Scholar 

  10. Iguchi H, Matsushita Y, Ohmori K, Hirano S, Kiyosaki T, Hori S, Tone H, Oki T (1980a) Studies on the absorption, excretion and distribution of aclacinomycin A: Absorption, excretion and distribution of aclacinomycin A in mice, rabbits, and dogs by photometric assay. Jpn J Antibiot [Suppl] 33:179

    Google Scholar 

  11. Iguchi H, Seryu Y, Kiyosaki T, Hori S, Tone T, Oki T (1980b) Studies on the absorption, excretion and distribution of14C- or3H-aclacinomycin A in mice and rabbits. Jpn J Antibiot [Suppl] 33:169

    Google Scholar 

  12. Kitagawa T, Aikawa T (1976) Enzyme coupled immunoassay of insulin using a novel coupling reagent. J Biochem (Tokyo) 79:233

    Google Scholar 

  13. Kitamura I, Oki T, Inui T (1978) A sensitive analytical method for aclacinomycin A and its analogs by thin-layer chromatography and fluorescence scanning. J Antibiot (Tokyo) 31:912

    Google Scholar 

  14. Klotz IM, Heiney RE (1959) Introduction of sulfhydryl groups into proteins using acetylmercaptosuccinic anhydride. Arch Biochem Biophys 82:70

    Google Scholar 

  15. Komiyama T, Oki T, Inui T (1979) A proposed reaction mechanism for the enzymatic reductive cleavage of glycosidic bond in anthracycline antibiotics. J Antibiot (Tokyo) 32:1219

    Google Scholar 

  16. Malspeis L, Neidhart J, Staubus A, Kear T, Booth J (1981) HPLC determination of aclacinomycin A (NSC 208734. ACM) in plasma and application to preliminary clinical pharmacokinetic studies. Proc Am Assoc Cancer Res 22:242

    Google Scholar 

  17. Matsuzawa Y, Kiyosaki T, Oki T, Takeuchi T, Umezawa H (1982) Radioimmunoassay for aclacinomycin A. Gann 73:229

    Google Scholar 

  18. Ogasawara T, Masuda Y, Goto S, Mori S, Oki T (1981) High performance liquid chromatographic determination of aclacinomycin A and its related compounds. II. Reverse phase HPLC determination of aclacinomycin A and its metabolities in biological fluids using fluorescence detection. J Antibiot (Tokyo) 34:52

    Google Scholar 

  19. Oki T (1977) New anthracycline antibiotics. J Antibiot (Tokyo) 30:570

    Google Scholar 

  20. Oki T (1980) New anthracycline antibiotics. Jpn J Antibiot [Suppl] 33:179

    Google Scholar 

  21. Oki T, Matsuzawa Y, Yoshimoto A, Numata K, Kitamura I, Hori S, Takamatsu A, Umezawa H, Ishizuka M, Naganawa H, Suda H, Hamada M, Takeuchi T (1975) New antitumor antibiotics, aclacinomycin A and B. J Antibiot (Tokyo) 28:830

    Google Scholar 

  22. Oki T, Takeuchi T, Oka S, Umezawa H (1980) Current status of Japanese studies with the new anthracycline antibiotic, aclacinomycin A. Recent Results Cancer Res 74:207

    Google Scholar 

  23. Oki T, Takeuchi T, Oka S, Umezawa H (1981) New anthracycline antibiotic aclacinomycin A: Experimental studies and corrections with clinical trials. Recent Results Cancer Res 76:21

    Google Scholar 

  24. Pal SB (ed) (1978) Enzyme-labelled immunoassay of hormones and drugs. Ulm, West Germany. de Gruyter, Berlin New York

    Google Scholar 

  25. Rich DH, Gesellchen PD, Tong A, Cheung A, Buckner CK (1975) Alkylating derivatives of amino acids and peptides. Synthesis ofN-maleoyl amino acids, [1-(N-maleoylglycyl)cysteinyl]oxytosin, and [1-(N-maleoyl-11-aminodecanoyl) cysteinyl] oxytosin. Effects on vasopressin-stimulated water loss from isolated toad bladder. J Med Chem 18:1004

    Google Scholar 

  26. Tanaka H, Yoshioka T, Shimauchi Y, Matsuzawa Y, Oki T, Inui T (1980) Chemical modification of anthracycline antibiotics. I. Demethoxycarbonylation, 10-epimerization and 4-0-methylation of aclacinomycin A. J Antibiot (Tokyo) 33:1323

    Google Scholar 

  27. Van Vunakis H, Levine L (1974) Use of the double-antibody and nitrocellulose membrane filtration technique to separate free antigen from antibody bound antigen in radioimmunoassay. In: Mule SJ, Sunshine I, Braude M (eds) Immunological assay of drugs of abuse. CRC, Cleveland, p 23

    Google Scholar 

  28. Wisdom GB (1976) Enzyme-immunoassay. Clin Chem 22:1243

    Google Scholar 

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Authors and Affiliations

  1. Second Departments of Internal Medicine, School of Medicine, Nagasaki University, Sakamoto-machi 7, 852, Nagasaki, Japan

    Masanori Sohda, Noriaki Nakamura & Kohei Hara

  2. Faculty of Pharmaceutical Sciences, Nagasaki University, Bunkyo-machi 1-14, 852, Nagasaki, Japan

    Kunio Fujiwara, Hitoshi Saikusa & Tsunehiro Kitagawa

  3. Central Research Laboratories, Sanraku-Ocean Co., Ltd, 9-1, Jonan 4-chome, 251, Fujisawa, Japan

    Hiroshi Tone

Authors
  1. Masanori Sohda
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  2. Kunio Fujiwara
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  3. Hitoshi Saikusa
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  4. Tsunehiro Kitagawa
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  5. Noriaki Nakamura
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  6. Kohei Hara
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  7. Hiroshi Tone
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Sohda, M., Fujiwara, K., Saikusa, H. et al. Sensitive enzyme immunoassay for the quantification of aclacinomycin A usingβ-D-galactosidase as a label. Cancer Chemother. Pharmacol. 14, 53–58 (1985). https://doi.org/10.1007/BF00552726

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  • DOI: https://doi.org/10.1007/BF00552726

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