1H HR-MAS NMR Based Metabolic Profiling of Cells in Response to Treatment with Chlorin-Polymer Systems

Gjuroski, Ilche; Hädener, Marianne; Diserens, Gaëlle; Vermathen, Peter; Furrer, Julien; Vermathen, Martina (7 July 2016). 1H HR-MAS NMR Based Metabolic Profiling of Cells in Response to Treatment with Chlorin-Polymer Systems (Unpublished). In: International Conference of Porphyrins and Phthalocyanines ICPP-9. Nanjing, China. 03.-08.07.2016.

The application of polymer based nanoparticles has found great recognition as delivery systems for porphyrinic photosensitizers in photodynamic therapy (PDT). Polymer-based nanoparticles have the capability to enhance porphyrin solubility and stability under physiological conditions and ideally can improve their uptake and phototoxic efficiency in the tumour target tissue. Recently, we have shown by 1H NMR spectroscopy that amino acid derivatives of chlorin e6 (xCE) are monomerized in aqueous solution by encapsulation into PVP [1] or Kolliphor P188 [2], a biodegradable triblock copolymer. While the cellular uptake of the porphyrinic compounds could be readily proved by fluorescence detection after cell incubation little is known about the metabolic response of the cells towards the porphyrin polymer systems and their single components in the dark. Therefore, the aim of this study was to directly monitor the metabolic response of cancer cells following incubation with the chlorin-polymer systems applying 1H High Resolution Magic Angle Spinning (HR-MAS) NMR spectroscopy. HR-MAS NMR is a technique, which provides well resolved 1H NMR spectra directly from semi-solid materials like living cells. Combined with multivariate statistical analysis it allows to monitor alterations in small metabolites following interventions like drug treatment. [3, 4]
In the present study, cultured HeLa cells were incubated with (i) serine-chlorin e6 (SerCE) encapsulated into a PVP-matrix (SerCE-PVP), as well as with (ii) SerCE and (iii) PVP alone and (iv) with pure buffer (PBS) serving as control. For each group, 8 samples were prepared leading to a total of 32 samples. After incubation the cells were washed, the pellets were taken up in D2O-based PBS and the cell suspensions were directly submitted to HR-MAS NMR spectroscopy. Partial least squared discriminant analysis (PLS-DA) of the cell metabolite spectra lead to a complete separation between the SerCE-PVP versus PVP-treatment groups but also between SerCE and PVP each versus control. The load values revealed main alterations deriving from lipid components suggesting for higher lipid mobility in response to either SerCE-PVP or the single components. Specific to SerCE treatment in the presence or absence of PVP were alterations in choline containing compounds indicating interactions with phospholipids. Further studies are currently in progress to compare the effect of PVP and Kolliphor P188 as delivery system for SerCE on the metabolic response of HeLa cells.

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