Conjugation of a Ru(II) Arene Complex to Neomycin or to Guanidinoneomycin Leads to Compounds with Differential Cytotoxicities and Accumulation between Cancer and Normal Cells

A straightforward methodology for the synthesis of conjugates between a cytotoxic organometallic ruthenium(II) complex and amino- and guanidinoglycosides, as potential RNA-targeted anticancer compounds, is described. Under microwave irradiation, the imidazole ligand incorporated on the aminoglycoside moiety (neamine or neomycin) was found to replace one triphenylphosphine ligand from the ruthenium precursor [(η6-p-cym)RuCl(PPh3)2]+, allowing the assembly of the target conjugates. The guanidinylated analogue was easily prepared from the neomycin-ruthenium conjugate by reaction with N,N′-di-Boc-N″-triflylguanidine, a powerful guanidinylating reagent that was compatible with the integrity of the metal complex. All conjugates were purified by semipreparative high-performance liquid chromatography (HPLC) and characterized by electrospray ionization (ESI) and matrix-assisted laser desorption-ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) and NMR spectroscopy. The cytotoxicity of the compounds was tested in MCF-7 (breast) and DU-145 (prostate) human cancer cells, as well as in the normal HEK293 (Human Embryonic Kidney) cell line, revealing a dependence on the nature of the glycoside moiety and the type of cell (cancer or healthy). Indeed, the neomycin-ruthenium conjugate (2) displayed moderate antiproliferative activity in both cancer cell lines (IC50 ≈ 80 μM), whereas the neamine conjugate (4) was inactive (IC50 ≈ 200 μM). However, the guanidinylated analogue of the neomycin-ruthenium conjugate (3) required much lower concentrations than the parent conjugate for equal effect (IC50 = 7.17 μM in DU-145 and IC50 = 11.33 μM in MCF-7). Although the same ranking in antiproliferative activity was found in the nontumorigenic cell line (3 2 > 4), IC50 values indicate that aminoglycoside-containing conjugates are about 2-fold more cytotoxic in normal cells (e.g., IC50 = 49.4 μM for 2) than in cancer cells, whereas an opposite tendency was found with the guanidinylated conjugate, since its cytotoxicity in the normal cell line (IC50 = 12.75 μM for 3) was similar or even lower than that found in MCF-7 and DU-145 cancer cell lines, respectively. Cell uptake studies performed by ICP-MS with conjugates 2 and 3 revealed that guanidinylation of the neomycin moiety had a positive effect on accumulation (about 3-fold higher in DU-145 and 4-fold higher in HEK293), which correlates well with the higher antiproliferative activity of 3. Interestingly, despite the slightly higher accumulation in the normal cell than in the cancer cell line (about 1.4-fold), guanidinoneomycin-ruthenium conjugate (3) was more cytotoxic to cancer cells (about 1.8-fold), whereas the opposite tendency applied for neomycin-ruthenium conjugate (2). Such differences in cytotoxic activity and cellular accumulation between cancer and normal cells open the way to the creation of more selective, less toxic anticancer metallodrugs by conjugating cytotoxic metal-based complexes such as ruthenium(II) arene derivatives to guanidinoglycosides

Understanding Parahydrogen Hyperpolarized Urine Spectra: The Case of Adenosine Derivatives

Abstract from publication:Parahydrogen hyperpolarization has emerged as a promising tool for sensitivity-enhanced NMR metabolomics. It allows for the resolution and quantification of NMR signals of certain classes of low-abundance metabolites that would otherwise be undetectable. Applications have been implemented in pharmacokinetics and doping drug detection, demonstrating the versatility of the technique. Yet, in order for the method to be adopted by the analytical community, certain limitations have to be understood and overcameovercome. One such question is NMR signal as-signment. At present, the only reliable way to establish the identity of an analyte that gives rise to certain parahydrogen hyperpolarized NMR signals is internal standard addition, which can be laborious. Herein we show that analogously to regular NMR metabolomics, generating libraries of hyperpolarized analyte signals is a viable way to address this limitation. We present hyperpolarized spectral data of adenosines and give an early example of identifying them from a urine sample with the small library. Doing so, we verify the detectability of a class of diagnos-tically valuable metabolites: adenosine and its derivatives, some of which are cancer biomarkers, and some are central to cellular energy management (e.g., ATP).Data from publication:Embedded data files contain raw NMR data acquired during preparation of this publication and that is presented in the publication. Files are named as follows:Fig2/21122303 - 2D dataset for adenosine 1aFig2/21122323 - 2D dataset for 1-methyladenosine 1bFig2/21122313 - 2D dataset for 6N-methyladenosine 1cFig2/21122123 - 2D dataset for 2'-O-methyladenosine 1dFig2/21122333 - 2D dataset for AMP 1eFig2/21122204 - 2D dataset for ADP 1fFig2/21122213 - 2D dataset for ATP 1g---Fig3/19021921 - 2D dataset of a urine SPE extract. Can be compared to datasets from Fig2.Some spectra may display signals from impurities at intensity levels below what is displayed in the article.THIS DATASET IS ARCHIVED AT DANS/EASY, BUT NOT ACCESSIBLE HERE. TO VIEW A LIST OF FILES AND ACCESS THE FILES IN THIS DATASET CLICK ON THE DOI-LINK ABOV

NMR detection in biofluid extracts at sub-[small mu ]M concentrations via para-H2 induced hyperpolarization

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Resolving dosy spectra of isomers by methanol-d(4) solvent effects

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Dosy analysis of micromolar analytes: Resolving dilute mixtures by SABRE hyperpolarization

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A New Ir-NHC Catalyst for Signal Amplification by Reversible Exchange in D2O

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