Probing the action of a novel anti-leukaemic drug therapy at the single cell level using modern vibrational spectroscopy techniques

Acute myeloid leukaemia (AML) is a life threatening cancer for which there is an urgent clinical need for novel therapeutic approaches. A redeployed drug combination of bezafibrate and medroxyprogesterone acetate (BaP) has shown anti-leukaemic activity in vitro and in vivo. Elucidation of the BaP mechanism of action is required in order to understand how to maximise the clinical benefit. Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, Synchrotron radiation FTIR (S-FTIR) and Raman microspectroscopy are powerful complementary techniques which were employed to probe the biochemical composition of two AML cell lines in the presence and absence of BaP. Analysis was performed on single living cells along with dehydrated and fixed cells to provide a large and detailed data set. A consideration of the main spectral differences in conjunction with multivariate statistical analysis reveals a significant change to the cellular lipid composition with drug treatment; furthermore, this response is not caused by cell apoptosis. No change to the DNA of either cell line was observed suggesting this combination therapy primarily targets lipid biosynthesis or effects bioactive lipids that activate specific signalling pathways

StearoylCoA Desaturase-5: A Novel Regulator of Neuronal Cell Proliferation and Differentiation

Recent studies have demonstrated that human stearoylCoA desaturase-1 (SCD1), a Δ9-desaturase that converts saturated fatty acids (SFA) into monounsaturated fatty acids, controls the rate of lipogenesis, cell proliferation and tumorigenic capacity in cancer cells. However, the biological function of stearoylCoA desaturase-5 (SCD5), a second isoform of human SCD that is highly expressed in brain, as well as its potential role in human disease, remains unknown. In this study we report that the constitutive overexpression of human SCD5 in mouse Neuro2a cells, a widely used cell model of neuronal growth and differentiation, displayed a greater n-7 MUFA-to-SFA ratio in cell lipids compared to empty-vector transfected cells (controls). De novo synthesis of phosphatidylcholine and cholesterolesters was increased whereas phosphatidylethanolamine and triacylglycerol formation was reduced in SCD5-expressing cells with respect to their controls, suggesting a differential use of SCD5 products for lipogenic reactions. We also observed that SCD5 expression markedly accelerated the rate of cell proliferation and suppressed the induction of neurite outgrowth, a typical marker of neuronal differentiation, by retinoic acid indicating that the desaturase plays a key role in the mechanisms of cell division and differentiation. Critical signal transduction pathways that are known to modulate these processes, such epidermal growth factor receptor (EGFR)Akt/ERK and Wnt, were affected by SCD5 expression. Epidermal growth factor-induced phosphorylation of EGFR, Akt and ERK was markedly blunted in SCD5-expressing cells. Furthermore, the activity of canonical Wnt was reduced whereas the non-canonical Wnt was increased by the presence of SCD5 activity. Finally, SCD5 expression increased the secretion of recombinant Wnt5a, a non-canonical Wnt, whereas it reduced the cellular and secreted levels of canonical Wnt7b. Our data suggest that, by a coordinated modulation of key lipogenic pathways and transduction signaling cascades, SCD5 participates in the regulation of neuronal cell growth and differentiation

Inhibition of fatty acid desaturation is detrimental to cancer cell survival in metabolically compromised environments.

BACKGROUND: Enhanced macromolecule biosynthesis is integral to growth and proliferation of cancer cells. Lipid biosynthesis has been predicted to be an essential process in cancer cells. However, it is unclear which enzymes within this pathway offer the best selectivity for cancer cells and could be suitable therapeutic targets. RESULTS: Using functional genomics, we identified stearoyl-CoA desaturase (SCD), an enzyme that controls synthesis of unsaturated fatty acids, as essential in breast and prostate cancer cells. SCD inhibition altered cellular lipid composition and impeded cell viability in the absence of exogenous lipids. SCD inhibition also altered cardiolipin composition, leading to the release of cytochrome C and induction of apoptosis. Furthermore, SCD was required for the generation of poly-unsaturated lipids in cancer cells grown in spheroid cultures, which resemble those found in tumour tissue. We also found that SCD mRNA and protein expression is elevated in human breast cancers and predicts poor survival in high-grade tumours. Finally, silencing of SCD in prostate orthografts efficiently blocked tumour growth and significantly increased animal survival. CONCLUSIONS: Our data implicate lipid desaturation as an essential process for cancer cell survival and suggest that targeting SCD could efficiently limit tumour expansion, especially under the metabolically compromised conditions of the tumour microenvironment

Multi-dimensional TOF-SIMS analysis for effective profiling of disease-related ions from the tissue surface

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) emerges as a promising tool to identify the ions (small molecules) indicative of disease states from the surface of patient tissues. In TOF-SIMS analysis, an enhanced ionization of surface molecules is critical to increase the number of detected ions. Several methods have been developed to enhance ionization capability. However, how these methods improve identification of disease-related ions has not been systematically explored. Here, we present a multi-dimensional SIMS (MD-SIMS) that combines conventional TOF-SIMS and metal-assisted SIMS (MetA-SIMS). Using this approach, we analyzed cancer and adjacent normal tissues first by TOF-SIMS and subsequently by MetA-SIMS. In total, TOF- and MetA-SIMS detected 632 and 959 ions, respectively. Among them, 426 were commonly detected by both methods, while 206 and 533 were detected uniquely by TOF- and MetA-SIMS, respectively. Of the 426 commonly detected ions, 250 increased in their intensities by MetA-SIMS, whereas 176 decreased. The integrated analysis of the ions detected by the two methods resulted in an increased number of discriminatory ions leading to an enhanced separation between cancer and normal tissues. Therefore, the results show that MD-SIMS can be a useful approach to provide a comprehensive list of discriminatory ions indicative of disease states