Separation and identification of platinum adducts with DNA nucleotides by capillary zone electrophoresis and capillary zone electrophoresis coupled to mass spectrometry

Abstract

Platinum adducts are supposed to be the cytotoxic lesions in DNA after platinum-containing anticancer therapy. Various adducts are formed upon interaction of platinum complexes with nucleotides, but contribution of individual adducts to antitumor activity and toxicity of platinum complexes still remains to be examined. A capillary zone electrophoresis (CZE) method is described that is suitable to separate individual platinum adducts. We investigated the formation of adducts following the reaction of cis-diamminedichloroplatinum (II) (cisplatin) with various DNA nucleotides. Baseline separation of unmodified and modified nucleotides (adducts) was achieved using uncoated fused-silica capillaries and basic separation buffers. In order to elucidate the observed peak pattern, a coupled CZE-electrospray ionization-mass spectrometry (ESI)-MS approach was applied. After incubation of mononucleotides with cisplatin, monochloro, monoaqua and bifunctional adduct species were detected. Consequently, the migration order of nucleotides and individual platinum adducts could be determined. Moreover, the time-dependent conversion from monochloro to monoaqua and subsequently to bifunctional adducts was monitored. In conclusion, individual platinum adducts were separated by CZE and identified by CZE-ESI-MS. Formation and conversion of distinct species were confirmed. Potential applications comprise studies of novel platinum complexes, investigations of platinum-adduct formation with DNA, and determination of platinum-DNA adducts in cells.

Effect of reactivity on cellular accumulation and cytotoxicity of oxaliplatin analogues

“…For the investigation of the reactivity of platinum complexes towards the nucleotides 5 0 -dAMP and 5 0 -dGMP, we modified and validated a previously described method using capillary electrophoresis [29]. Briefly, 300 lM concentration of each platinum complex were incubated simultaneously with 150 lM …”

Enhancing lipophilicity as a strategy to overcome resistance against platinum complexes?

“…For the investigation of the reactivity of platinum complexes towards the nucleotides dAMP and dGMP, we modified and validated a method previously described [24]. Briefly, 150 μM of each platinum complex was incubated with 150 μM dGMP and 150 μM dAMP under physiological conditions (4 mM NaH 2 PO 4 , 4 mM NaCl, pH 7.4, 37°C) up to 72 h. The samples were collected after 0, 24, 48 and 72 h of incubation (n = 3).…”

CE-MS in Forensic Sciences with Focus on Forensic Toxicology

“…Warnke et al investigated the formation of adducts following the reaction of cis-diamminedichloroplatinum (II) (cisplatin) with various DNA nucleotides [157]. Baseline separation of unmodified and modified nucleotides (adducts) was achieved through CZE within 11 min in one single run (Figure 9.14).…”