Enzo Cadoni

Coumarins comprise a large class of natural-based and synthetic compounds containing a 2Hchromen-2-one ring. Derivatives with hydroxy groups are able to donate a H˙, exhibiting antioxidant activities similar to phenols and quinones. In this work, 3-(pyridin-2-yl)coumarins with and without hydroxy group/s were prepared and their antioxidant ability and inhibition activity towards lipoxygenase enzymes were studied. The studied derivatives have shown potent lipoxygenase activities with IC₅₀ in the μM range, with the radical scavenging properties tuneable according to the number and position of the hydroxy groups. The study of the protonation constants of the 3-(pyridin-2-yl)coumarins, carried out in aqueous solutions, helped in evaluating the absorption and the potential biological properties of the molecules, strongly affected by their protonation state and charge. The antioxidant properties and the way of enzymatic inhibition of the studied 3-(pyridin-2-yl)coumarins are discussed on the basis of the calculated specific thermochemical descriptors and molecular docking.

The novel copper complex [Cu(phen)2(salubrinal)](ClO4)2 (C0SAL) has been synthesised and characterised. Copper(ii) is coordinated by salubrinal through the thionic group, as shown by the UV-Vis, IR, ESI-MS and tandem mass results, together with the theoretical calculations. The formed complex showed a DPPH radical scavenging ability higher than that of salubrinal alone. Studies on lipid oxidation inhibition showed that the C0SAL concentration, required to inhibit the enzyme, was lower than that of salubrinal. The inhibition of the enzyme could take place via allosteric modulation, as suggested by docking calculations. C0SAL showed a good cytotoxic activity on A2780 cells, 82 fold higher than that of the precursor salubrinal and 1.4 fold higher than that of [Cu(phen)2(H2O)](ClO4)2. Treatment with C0SAL in SKOV3 ovarian cancer cells induced expression of GRP-78 and DDIT3 regulators of ER-stress response. The cytotoxic effect of C0SAL was reverted in the presence of TUDCA, suggesting that C0SAL induces cell death through ER-stress. In A2780 cells treated with C0SAL γ-H2AX was accumulated, suggesting that DNA damage was also involved.

Five coumarin derivatives, able to coordinate Fe(III) and exhibit a ratiometric and turn-on fluorescence response, were prepared and tested. The derivative with the –OCH₃ substituent exhibited a high fluorescence enhancement in the presence of Fe(III), Ga(III), Au(III) and Y(III). Thanks to the higher stability of the Fe(III) complexes, this molecule could be proposed as a selective fluorescent sensor for the detection of Fe(III) in CH₃CN and CH₃CN : H₂O (1 : 1, v/v) solutions, showing a linear response up to 35.0 μM Fe(III) concentration and a limit of detection of 2.4 μM Fe(III) concentration. The fluorescence response towards this ion was also tested on a solid surface with a simple cellulose paper-strip test. The peculiarity of the proposed sensor lies in its easy and low-cost synthesis, relevant ratiometric response and, since it is derived from a natural product, low impact on the environment and its organisms.

Rationale: Development of therapy‐resistant cancer is a major problem in clinical oncology, and there is an urgent need for novel markers identifying development of the resistant phenotype. Lipidomics represents a promising approach to discriminate lipid profiles of malignant phenotype cells. Alterations in phospholipid distribution or chemical composition have been reported in various pathologies including cancer. Here we were curious whether quantitative differences in phospholipid composition between cisplatin‐resistant and ‐sensitive model cancer cell lines could be revealed by mass spectrometric means. Methods: The phospholipid contents of cell membranes of the cancer cell lines CCRF‐CEM and A2780, both responsive and resistant to cisplatin, were analyzed by solid‐phase extraction (SPE) and electrospray ionization mass spectrometry (ESI‐MS and tandem mass spectrometry (MS/MS)). Extracts were obtained by disruption of cells with a dounce tissue grinder set followed by centrifugation. To minimize the enzymatic activity, phospholipids were extracted from cell extracts by SPE immediately after the cell lysis and analyzed by MS. Both supernatant and pellet fractions of cell extracts were analyzed. Results: A phospholipid profile specific for cell lines and their phenotypes was revealed. We have documented by quantitative analysis that phosphocholines PC P‐34:0, PC 34:1, PC 20:2_16:0, LPC 18:1 and LPC 16:0 PLs were present in the 200–400 μM concentration range in CCRF‐CEM cisplatin‐responsive cells, but absent in their cisplatin‐resistant cells. Similarly, PC 34:1, LPC 18:1 and LPC 16:0 were increased in cisplatin‐responsive A2780 cells, and PC 20:2_16:0 was downregulated in cisplatin‐resistant A2780 cells. Conclusions: In this work we showed that the ESI‐MS analysis of the lipid content of the therapy‐resistant and ‐sensitive cells can clearly distinguish the phenotypic pattern and determine the potential tumor response to cytotoxic therapy. Lipid entities revealed by mass spectrometry and associated with development of therapy resistance can thus support molecular diagnosis and provide a potential complementary cancer biomarker.