title: Preparation and properties of clickable amino analogues of the duocarmycins: factors that affect the efficiency of their fluorescent labelling of DNA.
abstract: Herein we report the synthesis of three DNA-alkylating amino analogues of the duocarmycins that carry an alkyne functional group suitable for copper-catalysed click chemistry. The alkyne-containing substituents are connected via a side chain position which projects away from the minor groove, and have only a small effect on DNA alkylation and cytotoxicity. The efficiency of click reactions with fluorophore azides was studied using alkylated ctDNA by analysing the adenine adducts produced after thermal depurination. Click reactions "on DNA" were sensitive to steric effects (tether length to the alkyne) and, surprisingly, to the nature of the fluorophore azide. With the best combination of click partners and reagents, adducts could be detected in the nuclei of treated cells by microscopy or flow cytometry, provided that an appropriate detergent (Triton X-100 and not Tween 20) was used for permeabilisation. The method is sensitive enough to detect adducts at physiologically relevant concentrations, and could have application in the development of nitro analogues of the duocarmycins as hypoxia-activated anticancer prodrugs.
authors.affiliation: Auckland Cancer Society Research Centre, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag 92019, Auckland 1142 (New Zealand), Fax: (+64) 9373-7502. m.tercel@auckland.ac.nz.
authors.forename: Moana
authors.initials: M
authors.lastname: Tercel
authors.forename: Sarah P
authors.initials: SP
authors.lastname: McManaway
authors.forename: H D Sarath
authors.initials: HD
authors.lastname: Liyanage
authors.forename: Frederik B
authors.initials: FB
authors.lastname: Pruijn
mesh_terms: D000478:Alkylation; D000480:Alkynes / Q000737:chemistry; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000138:chemical synthesis* / Q000494:pharmacology; D045744:Cell Line, Tumor; D057930:Click Chemistry / Q000379:methods*; D004247:DNA / Q000737:chemistry*; D004285:Dogs; D004354:Drug Screening Assays, Antitumor; D000080890:Duocarmycins; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D007211:Indoles / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D011758:Pyrroles / Q000138:chemical synthesis / Q000494:pharmacology
chemical_list: D000480:Alkynes; D000903:Antibiotics, Antineoplastic; D000080890:Duocarmycins; D005456:Fluorescent Dyes; D007211:Indoles; D011758:Pyrroles; C066098:duocarmycin SA; D004247:DNA
keywords: DNA; antitumor agents; click chemistry; duocarmycins; fluorescent probes