Studies of cisplatin and hemoglobin interactions using nanospray mass spectrometry and liquid chromatography with inductively-coupled plasma mass spectrometry

Abstract

Interactions of cisplatin with hemoglobin (Hb) were studied using both nanoelectrospray mass spectrometry (nanoESI-MS) and a combination of size exclusion high performance liquid chromatography with inductively coupled plasma mass spectrometry (HPLC-ICPMS). Size exclusion HPLC separation of free and protein-bound cisplatin followed by simultaneous monitoring of 195Pt and 57Fe demonstrated the presence of Hb-bound Pt complexes. Nanospray quadrupole time-of-flight mass spectrometry studies of the Hb-cisplatin complexes further demonstrated the specific binding of cisplatin to the alpha-chain, heme-alpha, beta-chain, and heme-beta units of hemoglobin. Accurate mass measurements and tandem mass spectrometry information confirmed the Hb-cisplatin complexes. The formation of Hb-cisplatin complexes was observed at the sub-microM to microM concentration levels of cisplatin, which are relevant to clinical levels. These findings and the techniques developed for cisplatin-Hb interaction studies are useful for understanding of drug-protein interactions.

Characterization of Intact Hemoglobin and Oxaliplatin Interaction by Nanoelectrospray Ionization Tandem Mass Spectrometry

“…Human Hb A 0 and horse heart myoglobin were purchased from Sigma. The purity of the Hb was determined by gel electrophoresis and nanoelectrospray MS, as described previously (32,33 ). The myoglobin was used as received without further purification.…”

“…Several MS studies have reported binding of platinum drugs to glutathione (26 ) and proteins, including albumin (27,28 ), ubiquitin (29,30 ), transferrin (31 ), and Hb (32,33 ). We previously reported nanoelectrospray ionization MS (nanoESI-MS) characterization of cisplatin, carboplatin, and oxaliplatin adducts with subunits of Hb and the release of a heme group as a result of drug binding (33 ).…”

Investigation of interaction between human hemoglobin A0 and platinum anticancer drugs by capillary isoelectric focusing with whole column imaging detection

“…A recent review summarizes the progress made in the applications of modern analytical methodologies for antitumor metallodrug-protein studies [20]. ESI-MS was recently shown as an extremely useful tool for studying the cisplatin-protein interactions [19,[21][22][23][24][25]. Also, HPLC [19,24,[26][27][28] and a complementary high-resolution technique with an orthogonal separation mechanism, CE [29][30][31], are growing in importance for characterizing the protein binding for different metal complexes.…”

“…Also, HPLC [19,24,[26][27][28] and a complementary high-resolution technique with an orthogonal separation mechanism, CE [29][30][31], are growing in importance for characterizing the protein binding for different metal complexes. The potential of resolving intact and proteinbound drug forms appears to be prominent when combined with a metal-specific mode of detection by using inductively coupled plasma MS (ICP-MS) [19,25,[27][28][29].…”

The application of inductively coupled plasma mass spectrometry in clinical pharmacological oncology research

“…No compatibility problems were described. The interaction of cisplatin with hemoglobin was also extensively studied with SEC using a mobile phase of 10 mM ammonium bicarbonate (Mandal, Teixeira, & Li, 2003;Mandal, Kalke, & Li, 2004). A similar procedure was utilized to separate oxaliplatin and oxaliplatin bound to holo-transferrin (Zhao, Mandal, & Li, 2005).…”