DS10 - Défi des autres savoirs

Development of new fluorescent phenanthridine-based probes for the site-specific detection and quantification of superoxide radical anion in biological systems – Vivo2

Submission summary

During the last decade, the development of new techniques for the detection of superoxide radical anion (O2•–) in biological systems has continued to receive increasing attention. Superoxide is positioned upstream in most radical cascades leading to Reactive Oxygen and Nitrogen Species (RONS) and irreversible oxidative damages to biomolecules. Therefore, the ability of rigorous detection of superoxide is of major importance.
However, currently, no method fulfills the requirements for the rigorous characterization of the role of O2•– in biological systems and the characterization of RONS in vitro and in vivo remains a challenge. Many probes have been criticized and discarded due to unreliable results, detection artefacts and/or poor reproducibility.
For the last decade hydroethidine (HE) has been shown to be a promising probe for the fluorescent detection of O2•–. Unfortunately, the use of HE in biological systems is limited by four main problems:

(i) limited understanding of the mechanism of HE reaction with O2•–,
(ii) lack of site-specific analogs of HE,
(iii) degradation of HE by heme proteins leading to misleading signals,
and (iv) requirement of HPLC-based products separation coupled with fluorescence, electrochemical or MS detection.

Our teams have been actively involved in developing techniques for the detection and the characterization of RONS (e. g. spin trapping, SOD mimics, fluorescence-based RONS probes) for the last 20 years and their significant contributions have been recognized in the field of free radical/redox biology. The major objectives of this proposal can be summarized as follows:

(i) determination of the reaction mechanism of HE with O2•– and other biologically-relevant oxidants, including experimental (kinetics, products characterization) and theoretical studies (DFT calculations), using HE analogs as model systems,
(ii) targeting HE type probes to intracellular and extracellular compartments for site-specific detection of O2•–,
(iii) supramolecular and other approaches to protect HE from non-specific oxidation by heme proteins,
(iv) the development of innovative probes for the direct and specific detection of O2•– by fluorescence,
(v) application of optimized probes for detection of O2•– in biological systems.

The molecular structure of the probe plays a key role in its performance, including selectivity and specificity. As the development of HE probe was not based on rational design, there is an opportunity to develop new more efficient HE-analogs for the detection and quantification of O2•–. This project involves one French partner (ICR) as well as two foreign partners, led by Dr. Kalyanaraman (Medical College of Wisconsin, Milwaukee, USA) and by Dr. Sikora (Lodz University of Technology, Poland). These partners have already funds to perform the work described in the proposal. Thus, no financial support is requested from ANR for these two groups. All collaborating groups have a strong experience and expertise and are among leaders in their field. Also, the project will capitalize and benefit from well-established and long-term collaborations between the three groups.

This project illustrates the need for joint and interdisciplinary effort to push forward the frontiers in the free radical and biomedical fields.

Project coordinator

Institut de Chimie Radicalaire (ICR), UMR 7273, Equipe SREP (Laboratoire public)

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.

Partner

Lodz University of Technology
Medical College of Wisconsin
Institut de Chimie Radicalaire (ICR), UMR 7273, Equipe SREP

Help of the ANR 204,327 euros
Beginning and duration of the scientific project: September 2016 - 48 Months

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