ANTHRACYCLINE MULTI-DRUG RESISTANCE REVERSAL BY VERAPAMIL- MOLECULAR MECHANISM ELUCIDATION

1. ANTHRACYCLINE MULTI-DRUG RESISTANCE REVERSAL BY VERAPAMIL- MOLECULAR MECHANISM ELUCIDATION Klara Chrzanowski The objective of my work was to examine the molecular mechanism reversal of cell resistance to anticancer drugs to verapamil. Making the mechanism clear would create reliable ground to search for new more effecient drug reversing a drug resistance to the chemotherapeutics. My hypothesis assumes verapamil molecules react with Ca2+ ions in cytoplasm forming stable, biligand complex. Hence, the inability of reduction of oxidized glutathione (GS) to its reduced form excludes removing of anthracyclines outside cell. It results in a higher level of anthracyclines inside cell. Efficiency of this process depends on the level of reduced glutathione inside the cell. The level of reduced glutathione is dependent of the concentration of Ca2+ ions, since this form of glutathione is created in the first stage of the pentose phosphate pathway. Ca 2+ Glucose 6-phosphate + NADP+ 6-phospho-gloucono-δ- lactone The mechanism of reversing DR to anthracycline may rely on the blocking of the Ca2+ ions by verapamil. Calcium ion- Verapamil Complex Structure Verification of the hypothesis was done in two ways 1) using the quantum chemical method by which thermodynamic functions, enthalpy, and free enthalpy were calculated, 2) using MS method applying ‘soft’ ionization techniques APCI and ESI, which allow samples to be analysed in a liquid mobile phase, samples of verapamil and calcium-verapamil complex solutions in methanol were studied. The following values were obtained: ∆H = -36.27 kcal/mol and ∆G = -32.42 kcal/mol. These values prove that the reaction between verapamil and the calcium ion is spontaneous and hence thermodynamically possible. The calculated values of ∆G give information whether the reaction is spontaneous and therefore thermodynamically possible or not. The sign of ∆H gives account to whether the reaction produces energy or absorbs it. The complex compound of Ca2+ ion with verapamil was synthesized and its identity confirmed by the mass spectrum. Through the Electrospray ionization technique the molecular ions of Verapamil and the Calcium- Verapamil complex were detected. Many experiments were performed with different MS ionisations and changes in the parameters of the sources were used to choose optimal conditions for the best results.