Specific adducts recognised by a monoclonal antibody against cisplatin-modified DNA

• Published in: Biochemical pharmacology Vol. 70; no. 12; pp. 1717 - 1725
• Main Authors: Meczes, Emma L., Azim-Araghi, Ali, Ottley, Christopher J., Pearson, D. Graham, Tilby, Michael J.
• Format: Journal Article
• Language: English
• Published: New York, NY Elsevier Inc 05.12.2005; Elsevier Science
• Subjects: Antibodies, Monoclonal - immunology; Antineoplastic agents; Biological and medical sciences; Chromatography, Ion Exchange; Cisplatin; Cisplatin - metabolism; DNA adduct; DNA Adducts - analysis; DNA Adducts - immunology; Enzyme-Linked Immunosorbent Assay; General aspects; Humans; ICP-MS; Immunoassay; Ion-exchange chromatography; Medical sciences; Oligodeoxyribonucleotides - immunology; Oligonucleotide; Pharmacology. Drug treatments; DNA adduct; Immunoassay; Oligonucleotide; ICP-MS; Ion-exchange chromatography; Cisplatin; Antineoplastic agent; Monoclonal antibody; Inductive coupling plasma spectrometry; Ion exchange chromatography; Immunological method; Alkylating agent; Analysis method; DNA; Mechanism of action; Molecular adduct; Platinum II Complexes
• DOI: 10.1016/j.bcp.2005.09.025
• ISSN: 0006-2952

Numerous clinical or experimental studies have employed monoclonal antibody CP9/19 for quantification of cisplatin DNA adducts. The nature of adducts recognised by CP9/19 on polymeric DNA were defined using synthetic deoxynucleotides reacted with cisplatin. Total adduct levels were determined by atomic absorption spectrometry. The nature of adducts formed were confirmed by analysis of enzymatic hydrolysates using an established ion-exchange chromatography method combined with inductively coupled plasma mass spectrometry. Of the Pt bound to oligonucleotide A (TTTTTGGTTTTTGGTTTTTGGTTTTTGGTTTTT), 77% was recovered in a product consistent with the expected 1,2 intra-strand cross-link between GG. For oligonucleotide B (TTTTTAGTTTTTAGTTTTTAGTTTTTAGTTTTT), 62% of the bound Pt was recovered in a product consistent with the 1,2 intra-strand cross-link between AG. Of Pt bound to oligothymydylic acid, 65% was recovered in a product not previously described, small quantities of which were also formed on oligonucleotides A and B. The concentrations of adducts required to cause 50% reduction of signal in a competitive enzyme-linked immunosorbant assay (ELISA) ( K-values) were determined. Adducts on sequences containing no guanine or only non-adjacent guanine residues, including sequences containing adenines adjacent to guanines, exhibited low or undetectable immunoreactivities ( K-values = from 1 to >100 pmoles Pt per assay well). Adducts formed on oligodeoxynucleotides containing guanine doublets interspersed amongst thymine residues were the most immunoreactive ( K-values: 2–7 fmoles adduct per assay well), comparable to adducts on calf-thymus DNA. The only cisplatin–DNA adducts recognised with high sensitivity by antibody CP9/19 were those involving adjacent guanine residues but immunorecognition of these was influenced by the surrounding DNA sequence.