Nuclear Magnetic Resonance (NMR) Study of the Effect of Cisplatin on the Metabolic Profile of MG-63 Osteosarcoma Cells

Abstract

In the present study, 1 H HRMAS NMR spectroscopy was used to assess the changes in the intracellular metabolic profile of MG-63 human osteosarcoma (OS) cells induced by the chemotherapy agent cisplatin (CDDP) at different times of exposure. Multivariate analysis was applied to the cells spectra, enabling consistent variation patterns to be detected and drug-specific metabolic effects to be identified. Statistical recoupling of variables (SRV) analysis and spectral integration enabled the most relevant spectral changes to be evaluated, revealing significant time-dependent alterations in lipids, choline-containing compounds, some amino acids, polyalcohols, and nitrogenated bases. The metabolic relevance of these compounds in the response of MG-63 cells to CDDP treatment is discussed.

Metabolic Markers of MG-63 Osteosarcoma Cell Line Response to Doxorubicin and Methotrexate Treatment: Comparison to Cisplatin

“…Cho levels are possibly also affected by an involvement in enhanced phospholipid biosynthesis through PC formation or PTC-cycle regulation. 19 The GPC peak correlated positively with the PUFA resonance, through STOCSY (data not shown), which suggests a close metabolic link to phosphatidylcholine (PTC) hydrolysis, which possibly releases PUFAs into the cytosol and thus contributes to the NMR spectrum. It is important to note that STOCSY correlations between different compounds may either reflect metabolic links or be simply casual, ultimate confirmation of metabolic relationships requiring additional demonstration, for example, through specific pathway analysis.…”

“…For NMR analysis, the duplicates were mixed when necessary to provide samples with a large enough number of cells (ideally between 5 and 10 × 10 6 cells) for good signal-to-noise spectra to be obtained. For cDDP assays, procedures were those described elsewhere 19 for an IC 50 value (at 24 h) of 30 μM.…”

“…The spectra obtained for cDDP exposure studies have been recorded at a field strength of 700 MHz in similar conditions. 19 A standard 1D spectrum (pulse program "noesypr1d", Bruker library) was acquired for each sample, with spectral width of 9803.92 Hz, 32 K data points, 100 ms mixing time, 4 s relaxation delay, and 256 scans. All 1D spectra were processed with a line broadening of 0.3 Hz and zero filling to 64 K, manually phased, and baseline corrected.…”

“…One of the MTX-treated samples revealed an abnormally strong profile of lipid resonances, and for this reason it was not considered for multivariate analysis. In addition, the spectra obtained in an earlier study 19 of MG-63 cells (same cell line) exposed to cDDP were reprocessed, as described next. Data matrices were built from standard 1D spectra (δ 0.25−9.80), excluding the water region (δ 4.9−5.1), and spectra were aligned using recursive segment wise peak alignment 35 to minimize chemical shift variations and were normalized to probabilistic quotient normalization (PQN) 36 to account for dilutionindependent effects on spectral area (MATLAB version 7.12.0, The MathWorks Inc.).…”

A molecular view of cisplatin's mode of action: interplay with DNA bases and acquired resistance

“…The detailed knowledge of the molecular basis of the cDDP-guanine/adenine and cDDPglutathione interplay gathered in this work will pave the way for future studies on other platinum and palladium-based agents previously characterized by the authors and assessed as to their cytotoxicity towards several human cancer cell lines. [29][30][31][32][33][34][35][36][37] This will provide valuable clues for the rational design of novel cisplatin-like drugs, displaying a higher efficiency, lower toxicity, decreased acquired resistance and possible oral administration. …”