New generations of metallic nanoparticles to enhance the response to radiotherapy of radioresistant and invasive cancers

L’utilisation de nanoparticules contenant un métal à numéro atomique élevé est une approche prometteuse pour amplifier les effets de la radiothérapie, un traitement aujourd’hui utilisé chez plus de 50 % des patients atteints d’un cancer. Parmi elles, les AGuIX®, nanoparticules aux propriétés théranostiques, font aujourd’hui l’objet de multiples essais cliniques. Ce travail de thèse s’inscrit dans le cadre du développement d’une seconde génération des nanoparticules AGuIX® afin d’en augmenter l’efficacité et de cibler la formation de métastases. Dans une première partie de ce travail, les atomes de gadolinium (Z = 64) des AGuIX® ont été remplacés par des atomes d’yttrium (Z = 39) ou de bismuth (Z = 83) afin de mesurer l’impact du numéro atomique sur la radiosensibilisation. Malgré un défaut de reproductibilité de la synthèse chimique, un effet radiosensibilisant supérieur a été quantifié in vitro avec les nanoparticules contenant du bismuth. La seconde partie de ce travail était dédiée à l’évaluation des nanoparticules CuPRiX, contenant à la fois du gadolinium et chélates libres. Elles permettent la chélation in-situ du cuivre, un oligo-élément impliqué dans les mécanismes de croissance et de migration cellulaire tumorale. Dans un premier temps, les travaux menés in vitro sur trois modèles cellulaires de cancers ORL, du sein, et du poumon ont montré que CuPRiX était capable de ralentir la migration l’invasion cellulaire, d’inhiber l’activité de l’enzyme cuivre-dépendante Lysyl Oxidase (LOX) impliquée dans ces processus, et de radiosensibiliser les cellules. Dans un second temps, l’efficacité de CuPRiX a été validé in vivo dans un modèle murin de cancer du sein métastatique. Cette étude a montré une forte réduction de la croissance tumorale et du nombre de métastases grâce à la chélation du cuivre, tout en conservant un bon effet radiosensibilisant. Cette nouvelle génération de nanoparticules combinant radiosensibilisation et chélation du cuivre apparait comme une stratégie prometteuse dans le traitement des cancers radiorésistants et métastatiques.The use of nanoparticles containing a metal with a high atomic number is a promising approach to amplify the effects of radiotherapy, a treatment currently used in over 50% of cancer patients. Among them, AGuIX®, a nanoparticle with theranostic properties, is currently evaluated in multiple clinical trials. This thesis work is part of the development of a second generation of AGuIX® nanoparticles to increase their efficiency and to target metastasis formation. In a first part of this work, the gadolinium atoms (Z = 64) of AGuIX® were replaced by yttrium (Z = 39) or bismuth (Z = 83) atoms to measure the impact of the atomic number on radiosensitization. Despite a lack of reproducibility of the chemical synthesis, a superior radiosensitizing effect is observed in vitro with the nanoparticles containing bismuth. The second part of this work was dedicated to the evaluation of CuPRiX nanoparticles, containing both gadolinium and free chelates. They allow the in-situ chelation of copper, a trace element involved in the growth and migration mechanisms of tumor cells. Firstly, in vitro studies on three cell models of oral, breast and lung cancers showed that CuPRiX was able to slow down cell migration and invasion, inhibit the activity of the copper-dependent enzyme Lysyl Oxidase (LOX) involved in these processes, and radiosensitize tumor cells. Secondly, the efficacy of CuPRiX was validated in vivo in a mouse model of metastatic breast cancer. This study showed a strong reduction in tumor growth and the number of metastases thanks to copper chelation, while maintaining a good radiosensitizing effect. This new generation of nanoparticles combining radiosensitization and copper chelation appears to be a promising strategy in the treatment of radioresistant and metastatic cancers

Nouvelles générations de nanoparticules métalliques permettant d’amplifier la réponse à la radiothérapie des cancers radiorésistants et invasifs

The use of nanoparticles containing a metal with a high atomic number is a promising approach to amplify the effects of radiotherapy, a treatment currently used in over 50% of cancer patients. Among them, AGuIX®, a nanoparticle with theranostic properties, is currently evaluated in multiple clinical trials. This thesis work is part of the development of a second generation of AGuIX® nanoparticles to increase their efficiency and to target metastasis formation. In a first part of this work, the gadolinium atoms (Z = 64) of AGuIX® were replaced by yttrium (Z = 39) or bismuth (Z = 83) atoms to measure the impact of the atomic number on radiosensitization. Despite a lack of reproducibility of the chemical synthesis, a superior radiosensitizing effect is observed in vitro with the nanoparticles containing bismuth. The second part of this work was dedicated to the evaluation of CuPRiX nanoparticles, containing both gadolinium and free chelates. They allow the in-situ chelation of copper, a trace element involved in the growth and migration mechanisms of tumor cells. Firstly, in vitro studies on three cell models of oral, breast and lung cancers showed that CuPRiX was able to slow down cell migration and invasion, inhibit the activity of the copper-dependent enzyme Lysyl Oxidase (LOX) involved in these processes, and radiosensitize tumor cells. Secondly, the efficacy of CuPRiX was validated in vivo in a mouse model of metastatic breast cancer. This study showed a strong reduction in tumor growth and the number of metastases thanks to copper chelation, while maintaining a good radiosensitizing effect. This new generation of nanoparticles combining radiosensitization and copper chelation appears to be a promising strategy in the treatment of radioresistant and metastatic cancers.L’utilisation de nanoparticules contenant un métal à numéro atomique élevé est une approche prometteuse pour amplifier les effets de la radiothérapie, un traitement aujourd’hui utilisé chez plus de 50 % des patients atteints d’un cancer. Parmi elles, les AGuIX®, nanoparticules aux propriétés théranostiques, font aujourd’hui l’objet de multiples essais cliniques. Ce travail de thèse s’inscrit dans le cadre du développement d’une seconde génération des nanoparticules AGuIX® afin d’en augmenter l’efficacité et de cibler la formation de métastases. Dans une première partie de ce travail, les atomes de gadolinium (Z = 64) des AGuIX® ont été remplacés par des atomes d’yttrium (Z = 39) ou de bismuth (Z = 83) afin de mesurer l’impact du numéro atomique sur la radiosensibilisation. Malgré un défaut de reproductibilité de la synthèse chimique, un effet radiosensibilisant supérieur a été quantifié in vitro avec les nanoparticules contenant du bismuth. La seconde partie de ce travail était dédiée à l’évaluation des nanoparticules CuPRiX, contenant à la fois du gadolinium et chélates libres. Elles permettent la chélation in-situ du cuivre, un oligo-élément impliqué dans les mécanismes de croissance et de migration cellulaire tumorale. Dans un premier temps, les travaux menés in vitro sur trois modèles cellulaires de cancers ORL, du sein, et du poumon ont montré que CuPRiX était capable de ralentir la migration l’invasion cellulaire, d’inhiber l’activité de l’enzyme cuivre-dépendante Lysyl Oxidase (LOX) impliquée dans ces processus, et de radiosensibiliser les cellules. Dans un second temps, l’efficacité de CuPRiX a été validé in vivo dans un modèle murin de cancer du sein métastatique. Cette étude a montré une forte réduction de la croissance tumorale et du nombre de métastases grâce à la chélation du cuivre, tout en conservant un bon effet radiosensibilisant. Cette nouvelle génération de nanoparticules combinant radiosensibilisation et chélation du cuivre apparait comme une stratégie prometteuse dans le traitement des cancers radiorésistants et métastatiques

Nouvelles générations de nanoparticules métalliques permettant d’amplifier la réponse à la radiothérapie des cancers radiorésistants et invasifs

The use of nanoparticles containing a metal with a high atomic number is a promising approach to amplify the effects of radiotherapy, a treatment currently used in over 50% of cancer patients. Among them, AGuIX®, a nanoparticle with theranostic properties, is currently evaluated in multiple clinical trials. This thesis work is part of the development of a second generation of AGuIX® nanoparticles to increase their efficiency and to target metastasis formation. In a first part of this work, the gadolinium atoms (Z = 64) of AGuIX® were replaced by yttrium (Z = 39) or bismuth (Z = 83) atoms to measure the impact of the atomic number on radiosensitization. Despite a lack of reproducibility of the chemical synthesis, a superior radiosensitizing effect is observed in vitro with the nanoparticles containing bismuth. The second part of this work was dedicated to the evaluation of CuPRiX nanoparticles, containing both gadolinium and free chelates. They allow the in-situ chelation of copper, a trace element involved in the growth and migration mechanisms of tumor cells. Firstly, in vitro studies on three cell models of oral, breast and lung cancers showed that CuPRiX was able to slow down cell migration and invasion, inhibit the activity of the copper-dependent enzyme Lysyl Oxidase (LOX) involved in these processes, and radiosensitize tumor cells. Secondly, the efficacy of CuPRiX was validated in vivo in a mouse model of metastatic breast cancer. This study showed a strong reduction in tumor growth and the number of metastases thanks to copper chelation, while maintaining a good radiosensitizing effect. This new generation of nanoparticles combining radiosensitization and copper chelation appears to be a promising strategy in the treatment of radioresistant and metastatic cancers.L’utilisation de nanoparticules contenant un métal à numéro atomique élevé est une approche prometteuse pour amplifier les effets de la radiothérapie, un traitement aujourd’hui utilisé chez plus de 50 % des patients atteints d’un cancer. Parmi elles, les AGuIX®, nanoparticules aux propriétés théranostiques, font aujourd’hui l’objet de multiples essais cliniques. Ce travail de thèse s’inscrit dans le cadre du développement d’une seconde génération des nanoparticules AGuIX® afin d’en augmenter l’efficacité et de cibler la formation de métastases. Dans une première partie de ce travail, les atomes de gadolinium (Z = 64) des AGuIX® ont été remplacés par des atomes d’yttrium (Z = 39) ou de bismuth (Z = 83) afin de mesurer l’impact du numéro atomique sur la radiosensibilisation. Malgré un défaut de reproductibilité de la synthèse chimique, un effet radiosensibilisant supérieur a été quantifié in vitro avec les nanoparticules contenant du bismuth. La seconde partie de ce travail était dédiée à l’évaluation des nanoparticules CuPRiX, contenant à la fois du gadolinium et chélates libres. Elles permettent la chélation in-situ du cuivre, un oligo-élément impliqué dans les mécanismes de croissance et de migration cellulaire tumorale. Dans un premier temps, les travaux menés in vitro sur trois modèles cellulaires de cancers ORL, du sein, et du poumon ont montré que CuPRiX était capable de ralentir la migration l’invasion cellulaire, d’inhiber l’activité de l’enzyme cuivre-dépendante Lysyl Oxidase (LOX) impliquée dans ces processus, et de radiosensibiliser les cellules. Dans un second temps, l’efficacité de CuPRiX a été validé in vivo dans un modèle murin de cancer du sein métastatique. Cette étude a montré une forte réduction de la croissance tumorale et du nombre de métastases grâce à la chélation du cuivre, tout en conservant un bon effet radiosensibilisant. Cette nouvelle génération de nanoparticules combinant radiosensibilisation et chélation du cuivre apparait comme une stratégie prometteuse dans le traitement des cancers radiorésistants et métastatiques

Novel generation of ultrasmall high Z tailored nanoparticles for tumor cell radiosensitization

International audienc

A New Generation of Ultrasmall Nanoparticles Inducing Sensitization to Irradiation and Copper Depletion to Overcome Radioresistant and Invasive Cancers

International audienceAn emerging target to overcome cancer resistance to treatments is copper, which is upregulated in a wide variety of tumors and may be associated with cancer progression and metastases. The aim of this study was to develop a multimodal ultrasmall nanoparticle, CuPRiX, based on the clinical AGuIX nanoparticle made of the polysiloxane matrix on which gadolinium chelates are grafted. Such hybrid nanoparticles allow: (i) a localized depletion of copper in tumors to prevent tumor cell dissemination and metastasis formation and (ii) an increased sensitivity of the tumor to radiotherapy (RT) due to the presence of high Z gadolinium (Gd) atoms. CuPRiX nanoparticles are obtained by controlled acidification of AGuIX nanoparticles. They were evaluated in vitro on two cancer cell lines (lung and head and neck) using the scratch-wound assay and clonogenic cell survival assay. They were able to reduce cell migration and invasion and displayed radiosensitizing properties

Involvement of HIF-1α in the Detection, Signaling, and Repair of DNA Double-Strand Breaks after Photon and Carbon-Ion Irradiation

International audienceHypoxia-Inducible Factor 1α (HIF-1α), which promotes cancer cell survival, is the main regulator of oxygen homeostasis. Hypoxia combined with photon and carbon ion irradiation (C-ions) stabilizes HIF-1α. Silencing HIF-1α under hypoxia leads to substantial radiosensitization of Head-and-Neck Squamous Cell Carcinoma (HNSCC) cells after both photons and C-ions. Thus, this study aimed to clarify a potential involvement of HIF-1α in the detection, signaling, and repair of DNA Double-Strand-Breaks (DSBs) in response to both irradiations, in two HNSCC cell lines and their subpopulations of Cancer-Stem Cells (CSCs). After confirming the nucleoshuttling of HIF-1α in response to both exposure under hypoxia, we showed that silencing HIF-1α in non-CSCs and CSCs decreased the initiation of the DSB detection (P-ATM), and increased the residual phosphorylated H2AX (γH2AX) foci. While HIF-1α silencing did not modulate 53BP1 expression, P-DNA-PKcs (NHEJ-c) and RAD51 (HR) signals decreased. Altogether, our experiments demonstrate the involvement of HIF-1α in the detection and signaling of DSBs, but also in the main repair pathways (NHEJ-c and HR), without favoring one of them. Combining HIF-1α silencing with both types of radiation could therefore present a potential therapeutic benefit of targeting CSCs mostly present in tumor hypoxic niches

Involvement of HIF-1α in the Detection, Signaling, and Repair of DNA Double-Strand Breaks after Photon and Carbon-Ion Irradiation

Hypoxia-Inducible Factor 1α (HIF-1α), which promotes cancer cell survival, is the main regulator of oxygen homeostasis. Hypoxia combined with photon and carbon ion irradiation (C-ions) stabilizes HIF-1α. Silencing HIF-1α under hypoxia leads to substantial radiosensitization of Head-and-Neck Squamous Cell Carcinoma (HNSCC) cells after both photons and C-ions. Thus, this study aimed to clarify a potential involvement of HIF-1α in the detection, signaling, and repair of DNA Double-Strand-Breaks (DSBs) in response to both irradiations, in two HNSCC cell lines and their subpopulations of Cancer-Stem Cells (CSCs). After confirming the nucleoshuttling of HIF-1α in response to both exposure under hypoxia, we showed that silencing HIF-1α in non-CSCs and CSCs decreased the initiation of the DSB detection (P-ATM), and increased the residual phosphorylated H2AX (γH2AX) foci. While HIF-1α silencing did not modulate 53BP1 expression, P-DNA-PKcs (NHEJ-c) and RAD51 (HR) signals decreased. Altogether, our experiments demonstrate the involvement of HIF-1α in the detection and signaling of DSBs, but also in the main repair pathways (NHEJ-c and HR), without favoring one of them. Combining HIF-1α silencing with both types of radiation could therefore present a potential therapeutic benefit of targeting CSCs mostly present in tumor hypoxic niches

Proof of Concept of the Radiosensitizing Effect of Gadolinium Oxide Nanoparticles in Cell Spheroids and a Tumor-Implanted Murine Model of Chondrosarcoma

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Transfer of the ultrasmall theranostic gadolinium-based nanoparticle AGuIX® to clinical medicine

International audienceAGuIX® (Activation-and-Guidance by Irradiation X) are sub-5nm theranostic nanoparticles composed of a polysiloxane matrix and gadolinium chelates. Developed by the Lyon University, they underwent extensive pre-clinical evaluation and were recently translated to clinical evaluation in the treatment of brain metastases and advanced cervical cancer. This presentation will summarize the principal in cellulo and pre-clinical results that led to their first-in-man administration.In collaboration with other teams, we did the proof of concept of their radiosensitizing efficacy in cellular (2D/3D cultures) and preclinical models of three radioresistant tumors: head and neck squamous cell carcinoma, metastatic melanoma and chondrosarcoma tumors (Miladi et al, Nanomedicine 2015; Kotb et al, Theranostics 2016; Ollier et al, in preparation). AGuIX® enter cancer cells by passive diffusion and macropinocytosis, localize in cytoplasm as free particle or entrapped in lysosomes, in close vicinity to the mitochondria (Rima et al, Biomaterials 2013). The interaction of radiation with gadolinium atoms produce a variety of secondary emissions, such as Auger shower, leading to the production of high reactive oxygen species levels that can trigger an intra‐mitochondrial stress and nuclear DNA damage leading to cell death (Simonet al., submitted). Relative biological efficiency (RBE) in cancer cells is quite comparable to that observed in response to carbon ions, suggesting the existence of common mechanisms through the amplification of the local dose (Wozny et al, Nanomedicine 2017). AGuIX®nanoparticles have already shown their efficiency as a radiosensitizer in at least 12 preclinical models of cancer (Lux et al, BJR 2018). Biodistribution studies after intravenous administration have shown passive uptake of the nanoparticles in tumors due to enhanced permeability and retention (EPR) effect combined with rapid renal elimination. We are now testing the next generation of AGuIX® nanoparticles containing other metals or functionalized with therapeutic or targeting moieties (D. Vernos’ poster).Supported by LabEx PRIME

Transfer of the ultrasmall theranostic gadolinium-based nanoparticle AGuIX® to clinical medicine

International audienceAGuIX® (Activation-and-Guidance by Irradiation X) are sub-5nm theranostic nanoparticles composed of a polysiloxane matrix and gadolinium chelates. Developed by the Lyon University, they underwent extensive pre-clinical evaluation and were recently translated to clinical evaluation in the treatment of brain metastases and advanced cervical cancer. This presentation will summarize the principal in cellulo and pre-clinical results that led to their first-in-man administration.In collaboration with other teams, we did the proof of concept of their radiosensitizing efficacy in cellular (2D/3D cultures) and preclinical models of three radioresistant tumors: head and neck squamous cell carcinoma, metastatic melanoma and chondrosarcoma tumors (Miladi et al, Nanomedicine 2015; Kotb et al, Theranostics 2016; Ollier et al, in preparation). AGuIX® enter cancer cells by passive diffusion and macropinocytosis, localize in cytoplasm as free particle or entrapped in lysosomes, in close vicinity to the mitochondria (Rima et al, Biomaterials 2013). The interaction of radiation with gadolinium atoms produce a variety of secondary emissions, such as Auger shower, leading to the production of high reactive oxygen species levels that can trigger an intra‐mitochondrial stress and nuclear DNA damage leading to cell death (Simonet al., submitted). Relative biological efficiency (RBE) in cancer cells is quite comparable to that observed in response to carbon ions, suggesting the existence of common mechanisms through the amplification of the local dose (Wozny et al, Nanomedicine 2017). AGuIX®nanoparticles have already shown their efficiency as a radiosensitizer in at least 12 preclinical models of cancer (Lux et al, BJR 2018). Biodistribution studies after intravenous administration have shown passive uptake of the nanoparticles in tumors due to enhanced permeability and retention (EPR) effect combined with rapid renal elimination. We are now testing the next generation of AGuIX® nanoparticles containing other metals or functionalized with therapeutic or targeting moieties (D. Vernos’ poster).Supported by LabEx PRIME