Manganese Antioxidants for Delivery to Gut and Inflammation Control: design and evaluation – MAGIC

The MAGIC project involved developments in synthetic chemistry of SOD mimics:
-a series of purely artificial molecules, on which we worked to improve activity, sub-cellular targeting and stability (inertia) in a cellular context.
-a series of peptide mimics, for which a combinatorial approach with an activity-based screening has been implemented.
Cellular biology approaches were used to test the activity of these compounds directly in biological environments. These compounds are intended to restore the normal life of a cell under oxidative stress. Most of the metallic compounds used nowadays in inorganic medicinal chemistry are exploited for their toxicity properties: anti-cancer intended to kill cancer cells, anti-bacterial intended to kill bacteria, anti-fungal intended to kill fungi, anti-parasite to kill parasites. In contrast, with the series of SOD mimics we want to have non-toxic compounds that restore the redox balance of the cell. Therefore, we had to develop specific assays that were quite distinct from those in the literature. Since biological activity depends on (a) the amount that has entered the cells, (b) the intracellular localization, we set up methods for quantification of complexes in cells and intracellular imaging. The study of speciation (nature of the complexes) in a cellular context could be initiated at the end of the project and is still in progress (two papers, one in revision and one in preparation).
A last frontier is the in vivo passage. We approached it with a mice model of colitis. We had to think about delivery methods in the intestine avoiding acid destruction during the passage through the stomach, which cannot be explained here (patent considered). Promising results were recorded and will be the subject of a new ANR funding from April 2022, led by one of us (P. Seksik) and with a similar consortium, project this time more focused on bio-medical applications. In parallel, a project of the ENS team very focused on analytical developments (speciation and imaging of metal complexes in cellular conditions; functional studies of pathways; screening) was also initiated and was funded by the ANR with a start in April 2021.
Overall, the MAGIC project has been the occasion of multidisciplinary developments as anticipated. It has opened up new avenues of research, and has seen the perpetuation of the collaboration between the three teams.

In addition to the development of SOD-active metal complexes with improved activity and stability in biological context, the project allowed the establishment and optimization of methods for the evaluation of these SOD mimics in biological context: non-toxicity, anti-inflammatory properties related to anti-oxidant properties, possibility to correlate to cellular distribution and intracellular amount, first approaches to the study of intracellular speciation, promising studies on mouse model and first approaches to delivery issues.
Interesting results have been obtained, both from the point of view of the rational or combinatorial design of SOD mimes and their studies in biological context, with important advances in imaging and on the speciation issue (publications in progress).

Outstanding features:
In the wake of recent studies of chemistry in the cell, the MAGIC project aimed at monitoring and characterizing small metal complexes in cells. This is an emerging field in biological inorganic chemistry for which one must transpose to cells the knowledge acquired in the chemist's round flask. Improving cellular penetration, controlling the subcellular targeting of SODm are real challenges, as is the determination of the speciation of these SODm in cells. Physicochemical techniques for the quantification and imaging of metal cations at the subcellular level are in full expansion: in addition to conventional fluorescence imaging with labeled complexes, we have used innovative techniques such as X-ray microfluorescence for direct subcellular mapping of Mn.
Future prospects: For the development of peptide mimetics, the project was the occasion for the consolidation of a partnership with the team of Prof. V. Pecoraro (Michigan Univ.) with student exchanges (see publication). It also allowed to explore other properties of SOD mimics: as an adjuvant limiting the secondary effects of oxaliPt, as an inhibitor of the effects of Cu(II) in the process of generation of Aß plaques by metal exchange (Cu-Mn) with French partners (CNRS Toulouse, LCC and Cochin Hospital).
Promising results have been recorded and will be the subject of a new ANR funding from April 2022, led by one of us (P. Seksik) and with a similar consortium, this time more focused on bio-medical applications. In parallel, a project of the ENS team very focused on analytical developments (speciation and imaging of metal complexes in cellular conditions; functional studies of pathways; screening) was also born and was funded by the ANR with a start in April 2021.

Scientific articles
SOD mimics: from the tool box of the chemists to cellular studies, C. Policar*, J. Bouvet, H. C. Bertrand, N. Delsuc, Curr. Op. Chem. Biol., April. 2022, ASAP (IF 2021: 7.64)

Evaluation of the compounds commonly known as superoxide dismutase and catalase mimics in cellular models, A. Vincent, M. Thauvin, E. Quévrain, E.Mathieu, S.Layani, P. Seksik, I. Batinic-Haberle, S. Vriz, C. Policar,* N. Delsuc*, Journal of Inorganic Biochemistry, 219, 2021, 111431, IF(2020) 3.94 doi.org/10.1016/j.jinorgbio.2021.111431

Intracellular location matters: rationalization of the anti-inflammatory activity of a manganese(II) superoxide dismutase mimic complex, E. Mathieu […] P. Seksik, N. Delsuc, Clotilde Policar*, Chem.Commun., 2020, 56, 7885 - 7888, IF(2020) 5.996, doi.org/10.1039/D0CC03398G

Anti-inflammatory activity of superoxide dismutase mimics functionalized with cell-penetrating peptides, E. Mathieu […] P. Seksik, N. Delsuc, C. Policar*, Dalton trans, 2020, 2323-2330, IF(2020) 4.174, doi: 10.1039/c9dt04619d

An easy-to-implement combinatorial approach involving an activity-based assay for the discovery of a peptidyl copper complex mimicking superoxide dismutase, A. Vincent, J. Rodon-Fores, E. Tauziet […] P. Seksik, […] C. Policar and N. Delsuc*, Chem. Commun., 2020, 56, 399-402, IF(2018-19) 6.164, doi : 10.1039/C9CC07920C

Rational De Novo Design of a Cu-Metalloenzyme for Superoxide Dismutation; E.Mathieu, C. Policar*, and V. Pecoraro*, Chem. Eur. J., 2020, 26, 249-258, IF(2018-19) 5.16, doi.org/10.1002/chem.201903808

Oxaliplatin-induced neuropathy: The preventive effect of a new Super-Oxide Dismutase modulator ; M.-A. Guillaumont, […] H. C. Bertrand, […] C. Policar, R. Coriat, Oncotarget, 2019, 10, 6418-6431, IF(2016) 5.06, doi.org/10.18632/oncotarget.27248

A metallo pro-drug to target Cu(II) in the context of Alzheimer’s disease A. Conte-Daban, […] N. Delsuc, C. Policar* and C. Hureau*, Chem. Eur. J., 2018, 24, 5095 –5099 ; IF(2018) 5.16 ; doi : 10.1002/chem.201706049

A cell-penetrant manganese SOD-mimic is able to complement MnSOD and exerts an anti-inflammatory effect on cellular and animal models of inflammatory bowel diseases, E. Mathieu, […], N.Delsuc, […], F. Chain, P. Langella, […], P. Seksik, and C.Policar*, Inorg Chem. 2017, 56, 2545-2555 ; IF(2017) 4.86 ; doi: dx.doi.org/10.1021/acs.inorgchem.6b02695

Popularization

Theatrical conference «Metals, life and the chemist«, C. Policar, la Reine Blanche cycle «Des savants sur les planches »

Publication of a highlight in the newsletter of the French Chemical Society and of a «highlight« on the CNRS website, «live from the laboratories»

Interview on France Culture, Emission divers aspects de la pensée contemporaine 22 mai 2016

Television show: E=M6, September 23, 2020 «the mysteries of the belly«.
Pilège conference March 30, 2021: general public conference on 'inflammation and IBD'.

Superoxide dismutases (SOD) are very efficient redox metalloproteins, which protect the cell from oxidative stress. Their catalytic activity of superoxide dismutation can be reproduced by low-molecular weight Mn-complexes, called SOD-mimics (SODm) and the overall characteristics of SODs (tuned redox potential, electrostatic guidance of superoxide, compartmentation in organelles) can serve as a guideline in the design of efficient SODm. Oxidative stress, mainly produced in the mitochondria, is involved in inflammation, including Inflammatory Bowel Diseases (IBD), chosen as the biological target. We wish to develop manganese SODm directly inspired from the mitochondrial Mn-SOD that could exert an anti-inflammatory effect through an intracellular antioxidant activity. These will be studied in cellular models of oxidative stress relevant to IBD.
This project will involve several steps. First, using a modular approach, we will conjugate a SODm, already developed by the consortium and known to be active in cells, to various vectors and probes to obtain a series of SOD-mimics with tuned cell-penetration properties or organelle targeting, which could be detected inside cells. We will also develop a new series of peptide-based Mn SODm. We will then determine their intrinsic anti-superoxide activity —kinetics of the reaction with superoxide. Their anti-inflammatory effects on several cell models relevant for IBD, intestinal epithelial cells and monocytes/macrophages, will be evaluated by measuring markers of oxidative stress and inflammation and reactive oxygen species (ROS). We will determine the intracellular content in complexes and their sub-cellular location by innovative imaging techniques.
What are the main challenges in this project? This project aims at performing inorganic chemistry inside cells and is thus in line with emergent studies dealing with the control and characterization of small metal complexes in cells. This is a very active new field in inorganic chemical biology for which we need to translate the chemical knowledge we have acquired in the chemist’s round-bottom flasks into cells. Enhancing cell penetration and controlling the targeting of SODm to specific organelles is a real challenge, as is the determination of their speciation (or nature) in cells. Physico-chemical techniques to quantify and map metal cations at the sub-cellular level are now emerging: we will apply conventional fluorescence with tagged complexes but also the most recent techniques, such as X-fluorescence for direct sub-cellular mapping of Mn. Success here will certainly lead to a breakthrough in bio-inorganic chemistry as, at present, little information is available on the subcellular distribution of Mn-complexes SODm. This approach will provide guidelines for the rational improvement of antioxidant SODm with an intracellular activity.
The project in inorganic biological chemistry dealing with bio-inspired catalytic SOD-mimics design, evaluation and characterization in cells, and sub-cellular imaging will be developed by a consortium with multidisciplinary expertise.