RESUMO
Drug resistance is one of the main problems of cancer treatment. For this reason, combination therapy is commonly used for years. The combination of a chemotherapeutic, carboplatin, and the epigenetic drug decitabine is a new approach to modulate drug resistance. Nanoparticulate systems can overcome the drawbacks associated with the drug combinations. An analytical method that can detect and quantify carboplatin and decitabine which is encapsulated into the nanoparticles is necessary for nanoparticle development. In the literature, there is no analytical method in which carboplatin and decitabine are determined simultaneously. The primary purpose of this study is to develop and validate a novel, and stability-indicating high-performance liquid chromatography method for simultaneous determination of carboplatin and decitabine in pharmaceutical preparations in addition to developing the first nanoformulation for this drug combination. Therefore, various experimental parameters were optimized. The chromatographic separation was achieved using an XSelect® CSH C18 (250 × 4.6 mm I.D., 5 µm) column and a mobile phase consisting of methanol:water (containing 0.1% phosphoric acid) (3:97, v/v). The mobile phase pH was adjusted to 7.0 with 5 M NaOH. The developed method was successfully applied for the simultaneous determination and quantification of carboplatin and decitabine co-encapsulated in nanoparticles and released into in vitro dissolution medium.
Assuntos
Carboplatina/análise , Decitabina/análise , Nanopartículas/química , Química Farmacêutica , Cromatografia Líquida de Alta Pressão , Estabilidade de MedicamentosRESUMO
DNA hypermethylation is an epigenetic event that is commonly found in malignant cells and is used as a therapeutic target for ß-decitabine (ß-DEC) containing hypomethylating agents (eg Dacogen® and guadecitabine). ß-DEC requires cellular uptake and intracellular metabolic activation to ß-DEC triphosphate before it can get incorporated into the DNA. Once incorporated in the DNA, ß-DEC can exert its hypomethylating effect by trapping DNA methyltransferases (DNMTs), resulting in reduced 5-methyl-2'-deoxycytidine (5mdC) DNA content. ß-DEC DNA incorporation and its effect on DNA methylation, however, have not yet been investigated in patients treated with ß-DEC containing therapies. For this reason, we developed and validated a sensitive and selective LC-MS/MS method to determine total intracellular ß-DEC nucleotide (ß-DEC-XP) concentrations, as well as to quantify ß-DEC and 5mdC DNA incorporation relative to 2'-deoxycytidine (2dC) DNA content. The assay was successfully validated according to FDA and EMA guidelines in a linear range from 0.5 to 100 ng/mL (ß-DEC), 50 to 10,000 ng/mL (2dC), and 5 to 1,000 ng/mL (5mdC) in peripheral blood mononuclear cell (PBMC) lysate. An additional calibrator at a concentration of 0.1 ng/mL was added for ß-DEC to serve as a limit of detection (LOD). Clinical applicability of the method was demonstrated in patients treated with guadecitabine. Our data support the use of the validated LC-MS/MS method to further explore the intracellular pharmacokinetics in patients treated with ß-DEC containing hypomethylating agents.