Les glioblastomes sont des tumeurs cérébrales qui sont extrêmement agressives, et qui, en dépit de l'arsenal thérapeutique (chirurgie, radiothérapie ou chimiothérapie), ne laissent pas plus de 16 mois d'espérance de vie aux patients. Dans le cadre de cette thèse, nous proposons d'utiliser certaines toxines en tant que vecteurs pour l'administration de médicaments anticancéreux, et notamment pour le traitement du gliome. Les travaux présentés ici se concentrent sur l’utilisation de variants de la maurocalcine (MCa) et des analogues de la chlorotoxine (CTX). La MCa est une toxine issue du venin du scorpion Scorpio maurus palmatus, qui est capable de pénétrer dans les cellules facilement et rapidement. Il a été prouvé que la MCa peut entrer dans la cellule avec une cargaison. C’est en exploitant cette capacité présente chez deux de ces variants que nous avons synthétisé avec succès deux nouveaux composés à base de cette toxine avec de la doxorubicine et un dérivé du platine. Les études de toxicité et de caractérisation de ces composés qui ont été réalisé on permit de mettre en évidence l’intérêt et le potentiel de la MCa. La seconde partie de ces travaux de thèse portée sur la CTX et des peptides semblables, également extrait de venin de scorpion. Ils ont la particularité de fixer / interagir uniquement avec les cellules cancéreuses d'origine gliale. Après une rapide caractérisation de ces analogues de la CTX, l’un d’eux la Lqh-8/6 a été utilisé avec succès pour l'administration ciblée de doxorubicine. L’ensemble des travaux menés durant cette thèse constitue une base de départ solide pour une amélioration des systèmes de vectorisation, surtout en cancérologie de molécules actives. De plus ces résultats mettent aussi en avant l’avantage de l’utilisation d’un système de couplage « universel » basé sur la chimie click. / Glioblastoma are cerebral tumors that are extremely aggressive, and that, in spite of a battery of therapeutic interventions (surgery, radiotherapy or chemotherapy), leave no more than 16 months life expectancy to the patients. As part of this thesis, we propose to use some selected toxins as vectors for the delivery of anticancer drugs, and namely for the treatment of glioma. The works presented here concentrate on the use of variants of maurocalcine (MCa) and the analogues of chlorotoxine (CTX). MCa is a toxin from of the scorpion Maurus palmatus that has cell penetrating propriety. It has been proved that MCa can enter the cell with cargoes. While exploiting this present capacity to two of these variants we synthesized successfully two new compounds with this toxin with the doxorubicine and a by-product of the platinum. Toxicity studies and characterization of these compounds that have been made were permitted to highlight the interest and potential of the MCa. The second part of the thesis work focused on the CTX and similar peptides, also extracted from scorpion venom. They have the particularity to fix/ interact only with cancer cell from neuroectodermal origin. After a fast characterization of these analogues of CTX, one of them (Lqh-8/6) was successfully used for the targeted administration of doxorubicin. All work conducted during this thesis constitutes a solid starting point for an improvement of the systems of vectorization of active molecules, especially in cancer research Moreover, these results also emphasize the advantage of the use of a system of "universal" coupling based on the click chemistry.

Localized drug delivery systems have been widely studied as potential replacements for conventional chemotherapy with the capability of providing sustained and controlled drug release in specific targeted sites. They offer numerous benefits over conventional chemotherapy such as enhancing the stability of embedded drugs and preserving their anticancer activity, providing sustained and controlled drug release in the tumor site, reducing toxicity and diminishing subsequent side effects, minimizing the drug loss, averting the need for frequent administrations, and minimizing the cost of therapy. The aim of this study is to develop a localized drug delivery system with niosomes embedded in a chitosan hydrogel with targeting capabilities. The incorporation of niosomes into a chitosan hydrogel has several advantages over each individually being used. First, embedding niosomes in a chitosan hydrogel can yield control over drug release especially for small molecule drugs. Second, chitosan hydrogel may improve the release time and dosage of drugs from niosomes by protecting them with an extra barrier, resulting in tunable release rates. Third, as a localized delivery system, chitosan hydrogels can prevent the migration of niosomes away from the targeted tumor sites. Finally, chitosan has mucoadhesive property which can be used in the targeting of the tumor cells with the mucin over expression. To enhance the specific targeting, the capacity of chitosan to target MUC1 overexpression in cancer cells will be analyzed. Similarly, the incorporation of chlorotoxin in this system will be achieved and evaluated. Chlorotoxin, a 36-amino acid peptide, is purified from Leiurus quinquestriatus scorpion venom with a distinct characteristic of binding preferentially to neuroectoderma tumors such as glioma, but not to normal tissue. The overexpression of MUC1, a mucin antigen, in certain cancer cells has been used as an attractive therapeutic target in the design of a drug delivery system consisting of chitosan with a distinct mucoadhesive property. To determine the level of MUC1expression in different cell lines, Cell based Enzyme Linked Immunosorbent Assay (Cell ELISA) was developed for the first time. Attenuated Total Reflectance- Fourier Transform Infra-Red (ATR-FTIR) Spectroscopy is used to investigate the possible molecular interaction between chlorotoxin and glioma cells. This study presents a new approach in monitoring the biochemical and biophysical changes in glioma cells after being exposed to CTX. In addition to characterizing the signature spectra of CTX and glioma cells, we evaluated the differences in biochemical compositions of the spectra of the glioma cells treated with and without CTX over different incubation time periods. The results indicate that the proposed localized drug delivery system with the distinct tumor targeting features and extended release profiles would tune and control the specific delivery of chemotherapeutics in tumor sites.

abstract: Glioblastoma (GBM) is a highly invasive and deadly late stage tumor that develops from abnormal astrocytes in the brain. With few improvements in treatment over many decades, median patient survival is only 15 months and the 5-year survival rate hovers at 6%. Numerous challenges are encountered in the development of treatments for GBM. The blood-brain barrier (BBB) serves as a primary obstacle due to its innate ability to prevent unwanted molecules, such as most chemotherapeutics, from entering the brain tissue and reaching malignant cells. The GBM cells themselves serve as a second obstacle, having a high level of genetic and phenotypic heterogeneity. This characteristic improves the probability of a population of cells to have resistance to treatment, which ensures the survival of the tumor. Here, the development and testing of two different modes of therapy for treating GBM is described. These therapeutics were enhanced by pathogenic peptides known to improve entry into brain tissue or to bind GBM cells to overcome the BBB and/or tumor cell heterogeneity. The first therapeutic utilizes a small peptide, RVG-29, derived from the rabies virus glycoprotein to improve brain-specific delivery of nanoparticles encapsulated with a small molecule payload. RVG-29-targeted nanoparticles were observed to reach the brain of healthy mice in higher concentrations 2 hours following intravenous injection compared to control particles. However, targeted camptothecin-loaded nanoparticles were not capable of producing significant treatment benefits compared to non-targeted particles in an orthotopic mouse model of GBM. Peptide degradation following injection was shown to be a likely cause for reduced treatment benefit. The second therapeutic utilizes chlorotoxin, a non-toxic 36-amino acid peptide found in the venom of the deathstalker scorpion, expressed as a fusion to antibody fragments to enhance T cell recognition and killing of GBM. This candidate biologic, known as anti-CD3/chlorotoxin (ACDClx) is expressed as an insoluble protein in Nicotiana benthamiana and Escherichia coli and must be purified in denaturing and reducing conditions prior to being refolded. ACDClx was shown to selectively activate T cells only in the presence of GBM cells, providing evidence that further preclinical development of ACDClx as a GBM immunotherapy is warranted. / Dissertation/Thesis / Doctoral Dissertation Biological Design 2019