Isabelle Forfar

The antimalarial ferroquine: role of the metal and intramolecular hydrogen bond in activity and resistance.

Inhibition of hemozoin biocrystallization is considered the main mechanism of action of 4-aminoquinoline antimalarials including chloroquine (CQ) but cannot fully explain the activity of ferroquine (FQ) which has been related to redox properties and intramolecular hydrogen bonding. Analogues of FQ, methylferroquine (Me-FQ), ruthenoquine (RQ), and methylruthenoquine (Me-RQ), were prepared. Combination of physicochemical and molecular modeling methods showed that FQ and RQ favor intramolecular hydrogen bonding between the 4-aminoquinoline NH group and the terminal amino group in the absence of water, suggesting that this structure may enhance its passage through the membrane. This was further supported by the use of Me-FQ and Me-RQ where the intramolecular hydrogen bond cannot be formed. Docking studies suggest that FQ can interact specifically with the {0,0,1} and {1,0,0} faces of hemozoin, blocking crystal growth. With respect to the structure-activity relationship, the antimalarial activity on 15 different P. falciparum strains showed that the activity of FQ and RQ were correlated with each other but not with CQ, confirming lack of cross resistance. Conversely, Me-FQ and Me-RQ showed significant cross-resistance with CQ. Mutations or copy number of pfcrt, pfmrp, pfmdr1, pfmdr2, or pfnhe-1 did not exhibit significant correlations with the IC(50) of FQ or RQ. We next showed that FQ and Me-FQ were able to generate hydroxyl radicals, whereas RQ and me-RQ did not. Ultrastructural studies revealed that FQ and Me-FQ but not RQ or Me-RQ break down the parasite digestive vacuole membrane, which could be related to the ability of the former to generate hydroxyl radicals.

New ferrocenic pyrrolo[1,2-a]quinoxaline derivatives: synthesis, and in vitro antimalarial activity.

Following our search for antimalarial compounds, novel series of ferrocenic pyrrolo[1,2-a]quinoxaline derivatives 1-2 were synthesized from various substituted nitroanilines and tested for in vitro activity upon the erythrocytic development of Plasmodiumfalciparum strains with different chloroquine-resistance status. The pyrrolo[1,2-a]quinoxalines 1 were prepared in 6-8 steps through a regioselective palladium-catalyzed monoamination by coupling 4-chloropyrrolo[1,2-a]quinoxalines with 1,3-bis(aminopropyl)piperazine or -methylamine using Xantphos as the ligand. The ferrocenic bispyrrolo[1,2-a]quinoxalines 2 were prepared by reductive amination of previously described bispyrrolo[1,2-a]quinoxalines 9 with ferrocene-carboxaldehyde, by treatment with NaHB(OAc)(3). The best results were observed with ferrocenic pyrrolo[1,2-a]quinoxalines linked by a bis(3-aminopropyl)piperazine. Moreover, it was observed that a methoxy group on the pyrrolo[1,2-a]quinoxaline nucleus and no substitution on the terminal N-ferrocenylmethylamine function enhanced the pharmacological activity. Selected compounds 1b, 1f-h, 1l and 2a were tested for their ability to inhibit beta-haematin formation, the synthetic equivalent of hemozoin, by using the HPIA (heme polymerization inhibitory activity) assay. Of the tested compounds, only 2a showed a beta-haematin formation inhibition, but no inhibition of haem polymerization was observed with the other selected ferrocenic monopyrrolo[1,2-a]quinoxaline derivatives 1b, 1f-h and 1l, as the IC(50) values were superior to 10 equivalents.

Synthesis and antimalarial activities of rhenium bioorganometallics based on the 4-aminoquinoline structure

A bioorganometallic approach to malaria therapy led to the discovery of ferroquine (FQ, SSR97193). To assess the importance of the electronic properties of the ferrocenyl group, cyclopentadienyltricarbonylrhenium analogues related to FQ, were synthesized. The reaction of [N-(7-chloro-4-quinolinyl)-1,2-ethanodiamine] with the cyrhetrenylaldehyde complexes (η(5)-C(5)H(4)CHO)Re(CO)(3) and [η(5)-1,2-C(5)H(3)(CH(2)OH)(CHO)]Re(CO)(3) produces the corresponding imine derivatives [η(5)-1,2-C(5)H(3)(R)(CHN-CH(2)CH(2)NH-QN)]Re(CO)(3) R=H 3a; R=CH(2)OH 3b; QN=N-(7-Cl-4-quinolinyl). Reduction of 3a and 3b with sodium borohydride in methanol yields quantitatively the amine complexes [η(5)-1,2-C(5)H(3)(R)(CH(2)-NH-CH(2)CH(2)NH-QN)]Re(CO)(3) R=H 4a; R=CH(2)OH 4b. To establish the role of the cyrethrenyl moiety in the antimalarial activity of this series, purely organic parent compounds were also synthesized and tested. Evaluation of antimalarial activity measured in vitro against the CQ-resistant strains (W2) and the CQ-susceptible strain (3D7) of Plasmodium falciparum indicates that these cyrhetrene conjugates are less active compared to their ferrocene and organic analogues. These data suggest an original mode-of-action of FQ and ferrocenyl analogues in relationship with the redox pharmacophore.

Synthesis of new 4-(E)-alkenylpyrrolo[1,2-a]quinoxalines as antileishmanial agents by Suzuki-Miyaura cross-coupling reactions

A series of new 4-(E)-alkenylpyrrolo[1,2-a]quinoxaline derivatives, structural analogues of alkaloid chimanine B, was synthesized in good yields using efficient palladium(0)-catalyzed Suzuki-Miyaura cross-coupling reactions. These new compounds were tested for in vitro antiparasitic activity upon Leishmania amazonensis and Leishmania infantum strains. Biological results showed activity against the promastigote forms of L. amazonensis and L. infantum with IC50 ranging from 0.5 to 7 μM. From a Structure-Activity Relationships point of view, these pharmacological results mainly enlightened the importance of the 4-lateral C6, C7 or C8 α-unsaturated trans-alkenyl chain of unsubstituted pyrrolo[1,2-a]quinoxaline moiety.

Reaction between 5-(phenoxymethyl)-2-amino-2-oxazoline and N-benzyl-3-(ethoxycarbonyl)-4-piperidinone hydrochloride: A structural investigation

Abstract The two isomeric 7-benzyl-2-(phenoxymethyl)-2,3,6,7,8,9-hexahydro-5H-[1,3]oxazolo[3,2-a] pyrido[4,3-d]pyrimidin-5-one 3 and 7-benzyl-2-(phenoxymethyl)-1,2,6,7,8,9-hexahydro-5H-[1,3]oxazolo[3,2-a]pyrido[3,4-e]pyrimidin-5-one 4 were synthesized by a one-step cyclocondensation from 5-(phenoxymethyl)-2-amino-2-oxazoline 1 with N-benzyl-3-(ethoxycarbonyl)-4-piperidinone hydrochloride 2. Their structures were assigned by comparison of two dimensional NMR spectra (HMBC, NOESY) with the results obtained from theoretical calculations. The structure of one related hydrolysis compound was established using X-ray crystallography, allowing to further confirm the structure assignment.

Potential of FTIR spectroscopy for analysis of tears for diagnosis purposes

AbstractIt has been widely reported that the tear film, which is crucially important as a protective barrier of the eye, undergoes biochemical changes as a result of a wide range of ocular pathology. This tends to suggest the possibility of early detection of ocular diseases on the basis of biochemical analysis of tears. However, studies of tears by conventional methods of biomolecular and biochemical analysis are often limited by methodological difficulties. Moreover, such analysis could not be applied in the clinic, where structural and morphological analyses by, mainly, slit-lamp biomicroscopy remains the recommended method. In this study, we assessed, for the first time, the potential of FTIR spectroscopy combined with advanced chemometric processing of spectral data for analysis of raw tears for diagnosis purposes. We first optimized sampling and spectral acquisition (tears collection method, tear sample volume, and preservation of the samples) for accurate spectral measurement. On the basis of the results, we focused our study on the possibility of discriminating tears from normal individuals from those of patients with different ocular pathologies, and showed that the most discriminating spectral range is that corresponding to variations of CH2 and CH3 of lipid aliphatic chains. We also report more subtle discrimination of tears from patients with keratoconus and those from patients with non-specific inflammatory ocular diseases, on the basis of variations in spectral ranges attributed notably to lipid and carbohydrate vibrations. Finally, we also succeeded in distinguishing tears from patients with early-stage and late-stage keratoconus on the basis of spectral features attributed to protein structure. Therefore, this study strongly suggests that FTIR spectral analysis of tears could be developed as a valuable and cost-saving tool for biochemical-based detection of ocular diseases, potentially before the appearance of the first morphological signs of diseases. Combined with supervised modelling methods and with use of a spectral data base acquired for representative patients, such a spectral approach could be a useful addition to current methods of clinical analysis for improvement of patient care. FigurePCA-based discrimination between tears from keratoconus patients and patients with others ocular pathology. Scatter plot of spectra depending on PC1 and PC2 (percentage of total variance) scores (a) and statistical significance of PC-dis mean scores differences (b). ****p < 0.0001. Spectra from patients with keratoconus (K), allergies (A), rosacea (R), dry syndrome (S), conjunctivitis (Co), and lachrymal hypersensitivity (Hy). Arrows, misclassified keratoconus spectra. Ellipse overlaid on the data points serves as visual guide to the eye

Investigation of blood vessels in glioblastoma at a micrometric scale: a comparative study by synchrotron and conventional micro-FTIR

Glioblastoma, the most malignant brain tumor in humans, is characterized by being severely angiogenic and an increase in vascularization generally worsens the prognosis of patients. Finding the best approach to characterize glioma blood vessels (BVs) is very important in view of helping to determine any specific biomolecular markers of these tumors. In previous work by conventional FTIR spectroscopy we were able to discriminate some molecular markers in order to differentiate between normal and tumor BVs in glioma tissue sections. The aim of the present study was to assess whether FTIR microspectroscopy using a synchrotron radiation (SR) source could provide advantages over a classical globar IR source for detailed spectral analysis on such small features like micro-BVs. Using chemometric analysis such as PCA and HCA, the results show that a high brilliant SR beam provides a very satisfying quality signal compared with the globar source to study spectral images for relevant analysis of glioma big and micro-BVs and determination of subtle molecular markers characterizing them from the surrounding tissue.

Monitoring biological effects of 20 nm versus 100 nm silica nanoparticles induced on a human renal cell line using Fourier transform infrared spectroscopy

Due to their small size, nanoparticles (NPs) and particularly silica NPs (SiO2-NPs) exhibit unique properties that confer them especially great biological reactivity. Thus, intensive research studies are performed to elucidate their potential toxicity and/or their interaction with biological systems. Standard assays focus on one specific biological event which require the use of several tests to determine all potential effects induced by SiO2-NPs. Moreover, because of the interaction between the reagent and SiO2-NPs, the reliability of colorimetric or fluorometric methods is questioned when assessing nanomaterials. Therefore, Fourier transform infrared (FTIR) spectroscopy was used as a reagent-free and time-saving tool, combined with Principal Component Analysis (PCA), to reveal biochemical effects induced by 100 nm versus 20 nm SiO2-NPs on human kidney cells (HK-2) at two subtoxic concentrations (10 and 25 μg ml−1) during 24 h. This technique allows revealing dose-dependent responses and differences in biological effects between 20 nm and 100 nm SiO2-NPs with a greater impact of 20 nm SiO2-NPs. Moreover, SiO2-NP cell uptake has been highlighted. Toxic biochemical effects induced by 20 nm SiO2-NPs are detected as hallmarks of lipid peroxidation (1746 cm−1) and apoptosis (1654, 1746 and 2922 cm−1). This study demonstrates that this technique is more sensitive than cytotoxic assays and may be appropriate as a simple screening tool to quickly monitor cellular biochemical changes induced by NPs.

Discrimination between two different grades of human glioma based on blood vessel infrared spectral imaging.

AbstractGliomas are brain tumours classified into four grades with increasing malignancy from I to IV. The development and the progression of malignant glioma largely depend on the tumour vascularization. Due to their tissue heterogeneity, glioma cases can be difficult to classify into a specific grade using the gold standard of histological observation, hence the need to base classification on a quantitative and reliable analytical method for accurately grading the disease. Previous works focused specifically on vascularization study by Fourier transform infrared (FTIR) spectroscopy, proving this method to be a way forward to detect biochemical changes in the tumour tissue not detectable by visual techniques. In this project, we employed FTIR imaging using a focal plane array (FPA) detector and globar source to analyse large areas of glioma tumour tissue sections via molecular fingerprinting in view of helping to define markers of the tumour grade. Unsupervised multivariate analysis (hierarchical cluster analysis and principal component analysis) of blood vessel spectral data, retrieved from the FPA images, revealed the fine structure of the borderline between two areas identified by a pathologist as grades III and IV. Spectroscopic indicators are found capable of discriminating different areas in the tumour tissue and are proposed as biomolecular markers for potential future use of grading gliomas. Graphical AbstractInfrared imaging of glioma blood vessels provides a means to revise the pathologists’ line of demarcation separating grade III (GIII) from grade IV (GIV) parts.

FTIR spectroscopic metabolome analysis of lyophilized and fresh Saccharomyces cerevisiae yeast cells

The yeast Saccharomyces cerevisiae is widely used as a biological eukaryotic model and also serves as a production organism in biotechnology. One of the methods used to avoid degradation of the yeast cell content is lyophilization. The use of lyophilized yeast cells has several advantages over fresh ones: samples can be easily transported and/or stored and variations of their metabolomic profiles do not occur during transport or storage. Fourier transform infrared (FTIR) spectroscopy is one of the most emerging approaches in modern biology that permits operation on very small quantities of whole cells without the need for extractions or purifications. This technique is very sensitive and not only allows the discrimination between different cell genotypes but also between different growth conditions. FTIR spectra provide interesting data on the metabolic status of the whole cell. Modern multivariate data processing was applied to analyse live fresh or lyophilized S. cerevisiae cells from different growth media. This study clearly demonstrates that yeast cells coming from an identical biological medium can be used indiscriminately for FTIR analysis whether they are analysed directly as live fresh cells or after lyophilization which is a freeze-drying process. Moreover, FTIR data obtained using lyophilized cells showed less variability.