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Place Amphi Dorothy Hodgkin
Title
Abstract
Photoacoustics (PA) is an emerging technology that turns light into sound waves, with several applications, one of which is the non-invasive, high-resolution imaging of biological tissues. PA contrast agents for biological applications should be photostable, NIR-absorbing molecules or objects with a high extinction coefficient, and lowlight emission. PA imaging is a relatively inexpensive, non-invasive imaging technique, and currently there is a growing demand on the market for new biocompatible PA contrast agents.
This thesis aims to develop a pH-responsive PA probe for cellular imaging, capable of sensing acidic environments like lysosomes through shifts in its optical properties. The overarching goal of this study is to develop a stimulus-responsive PA agent by synthetizing and characterizing a pH-sensitive molecule, and formulate it into biocompatible, PA active, pH-sensitive polymeric nanoparticles (NPs).
Two push-pull BODIPY derivatives were synthesized: one symmetrically 3,5-disubstituted with 3-nitro-4-phenol units, and an analog asymmetrically functionalized with one nitrophenol and one julolidine moiety to induce a stronger red-shift. The first one exhibits attractive properties for a PA dye candidate, absorbing at 666 nm in dichloromethane (DCM) with a relatively high extinction coefficient (εDCM =6.6 105 M-1cm-1), weak emission of fluorescence (ΦF=0.7), at 685 nm (Δν=319 cm-1) and short lifetime (τ~0.4 ns). Deprotonation of the molecule by addition of trimethylamine in DCM induces a change in the absorption profile, characterized by band broadening, a red-shift toward 745 nm (1686 cm-1), and quenched fluorescence. Titration carried out in aqueous media on the PEGyilated derivative of the symmetrical BODIPY yielded an apparent pKa of 7.6 by absorption and 4.9 by emission of fluorescence. The asymmetrically functionalized BODIPY absorbs at 684 nm in DCM and emits at 701 nm. Compared to the other dye, its absorption is 396 cm-1 redshifted and exhibits a lower extinction coefficient (εDCM = 3.2 105 M-1cm-1). Therefore, the symmetrical BODIPY was selected as the best dye candidate and clicked to PLA-PEG (Mw~104 Da) via CuAAC. Subsequently, the grafted dye was formulated into NPs by the solvent-evaporation method with variable dye loadings. The NPs were ~70 nm to ~150 nm in size (increasing with dye-loading), negatively charged (ζ-potential of ~7 mV), and showed no sign of aggregation when refrigerated (+4°C) for over two months. J-aggregate formation was observed between 734 nm and 752 nm. NPs displayed high PA activity, with extinction coefficients up to 109 M-1 cm-1, performing comparably with best-in-class PA contrast agents. NPs were shown to respond to pH changes in the PA excitation spectra, with an average pKa of ~7.8. No cytotoxicity was observed in preliminary HeLa or E. coli viability assays, which makes them suitable for future in-vivo testing.
PhD supervision
- Director: Rachel Méallet, Professor, ISMO, University Paris-Saclay
- Co-director: Gilles Clavier, Research Director, PPSM, CNRS - ENS Paris-Saclay
Members of the jury
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Mireille Blachard, Research Director, ISM, Université de Bordeaux, Rapportrice
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Olivier Galangau, Associate Researcher, ISCR, Université de Rennes, Rapporteur
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Andrea Pasc, Professor, L2CM, Université de Lorraine, Examinatrice
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Jérôme Hannedouche, Directeur de Recherches, Softmat, Université de Toulouse, Examinateur