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Platinum metallodrug‐protein binding studies by capillary electrophoresis‐inductively coupled plasma‐mass spectrometry: Characterization of interactions between Pt(II) complexes and human serum albumin
Author(s) -
Timerbaev Andrei R.,
Aleksenko Svetlana S.,
PolecPawlak Kasia,
Ruzik Rafal,
Semenova Olga,
Hartinger Christian G.,
Oszwaldowski Slawomir,
Galanski Mathea Sophia,
Jarosz Maciej,
Keppler Bernhard K.
Publication year - 2004
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200305984
Subject(s) - chemistry , human serum albumin , capillary electrophoresis , platinum , cisplatin , chromatography , mass spectrometry , serum albumin , stoichiometry , reaction rate constant , bovine serum albumin , kinetics , blood proteins , biochemistry , organic chemistry , medicine , physics , surgery , chemotherapy , quantum mechanics , catalysis
Characterizing how platinum metallocomplexes bind to human serum albumin (HSA) is essential in evaluating anticancer drug candidates. Using cisplatin as a reference complex, the application of capillary electrophoresis (CE) to reliably assess drug/HSA interactions was validated. Since this complex is small compared to the size of the protein, the binding response could only be recognized when applying CE coupled to a (platinum) metal‐specific mode of detection, namely inductively coupled plasma‐mass spectrometry (ICP‐MS). This coupling allowed for confirmation of a specific affinity of cisplatin and novel Pt complexes to HSA, measurement of the kinetics of binding reactions, and determination of the number of drug molecules attached to the protein. As the cisplatin/HSA molar ratio increased, the reaction rate became faster with a maximum on the kinetic curve appearing at about 50 h of incubation at 20 times excess of cisplatin. The reaction was characterized as a pseudo‐first order reaction with the rate constant k = 0.003 min −1 at 37°C. When incubated with a 20‐fold excess of cisplatin, HSA bound up to 10 mol of Pt per mol of the protein. This is indicative for a strong metal‐protein coordination occurring at several HSA sites other than the only protein cysteine residue. Structural analogs of cisplatin, bearing aminoalcohol ligands, showed comparable protein binding reactivity and stoichiometry but a common equilibrium was not reached even after one week of incubation. Also apparent was a two‐step mechanism of the binding reaction. Results demonstrated the suitability of CE‐ICP‐MS as a rapid assay for high‐throughput studying of drug/HSA interactions.