Thérapie laser assistée par nanoparticules plasmoniques du rétinoblastome

Le rétinoblastome (RB), qui touche environ 1 enfant sur 18 000 est le cancer infantile le plus fréquent. Il existe sous forme héréditaire, avec un risque accru de bilatéralité, ou sporadique et est lié à des mutations du gène Rb. Bien que les chances de survie soient très élevées dans les pays dits développés, il n’est pas rare que la solution finale pour l’éradication du cancer soit l’énucléation, autrement dit le retrait de l’oeil. En effet, parmi les stades de développement de la tumeur, qui nait initialement à la surface de la rétine, on compte l’essaimage de la tumeur vers la partie centrale de l’oeil, le corps vitré. Les cellules tumorales prennent alors une nature flottante et leur éradication en devient d’autant plus compliquée. Si initialement des traitements de cryothérapie et thermothérapie peuvent être utilisés, le seul traitement admissible à ce stade est la chimiothérapie intravitréenne. L’humeur vitrée, principalement composée de collagène et d’acide hyaluronique, présente une forte viscosité et rend plus difficile le ciblage des tumeurs par les médicaments. C’est ainsi que, pour éviter l’envahissement du cerveau par le nerf optique, enlever l’oeil parait une solution tout à fait préférable. Comme alternative aux traitements chimiothérapeutiques usuels, souvent lourds en effets secondaires, nous proposons ici une preuve de concept pour le traitement laser assisté par nanoparticules plasmoniques du rétinoblastome. Cette méthode présente l’avantage de produire une très forte augmentation de température hautement localisée, permettant de tuer les cellules cancéreuses sans affecter les tissus alentour. Bien que l’interaction laser-nanoparticules ait fait l’objet de beaucoup de recherche pour des applications dans le domaine médical, on ne retrouve qu’étonnamment peu de cas appliqués au traitement du cancer dans l’oeil. Les cellules Y79 utilisées pour l’étude, de par leur nature flottante, apportent de nouveaux enjeux à la démonstration in vitro. Ayant une forte tendance à former des amas cellulaires elles apportent en effet une structure tridimensionnelle à l’étude. Dans un premier temps, afin de simplifier le système étudiées les cellules seront artificiellement attachées au fond des pétris à l’aide d’un polymère. Les utilisations de deux types de lasers seront comparées, ainsi que différents paramètres tels que la fluence laser, la concentration en nanoparticules, le temps d’irradiation et le taux de re-prolifération cellulaire 18h après irradiation. Dans un deuxième temps, afin de se rapprocher des conditions réelles, un simili-vitré constitué d’acide hyaluronique et de milieu de culture cellulaire est réalisé et le traitement est mené directement sur les amas cellulaires. Nous avons ainsi pu montrer que, malgré la viscosité, les nanoparticules se rendaient jusqu’au coeur des amas. En utilisant un laser pulsé nanoseconde et des nanoparticules d’or de 100nm, nous sommes parvenus à un taux de mort cellulaire d’environ 80% pour une fluence d’irradiation de 0,2μJ/μm2 dans le simili-corps vitré.----------Abstract The retinoblastoma (RB), an infantile eye cancer, concerns 1 of 18 000 births and is usually diagnosed before 6 years old. It exists under two forms, hereditary or sporadic, and is linked with the Rb gene mutation. Two stages are described in the literature. At the beginning, the tumor grows on the retina. At this stage, the usual treatments, depending on the tumor size, are cryotherapy or surgery complemented with localized chemotherapy. Then the tumor detaches from the retina and starts floating in the vitreous. The viscosity of the vitreal chamber prevents any surgery and the only possible treatment is chemotherapy. Moreover, at this step, the risk of migration towards the brain is highly increased. Unfortunately, to avoid dramatic complications, enucleation is often the best option. As an alternative and efficient localized therapy, we have investigated laser induced plasmon-mediated hyperthermia which offers the advantage of producing a highly localized heat increase of the cancer cells, thus killing these cells by avoiding any side effects on healthy tissues. This work aims to establish a proof of concept for in vitro treatment of RB cells with nanosecond laser and 100nm spherical gold nanoparticles (SNG) and extend this new therapy to phantom reproducing vitreous of the eye. Nanosecond laser surgery, assisted by plasmonic effect from gold nanoparticles have been investigated in numerous cases of cancers. However, the Y79 cells, which are floating cells, bring new issues for in vitro studies: it adds a new dimension as they naturally grow in 3D clusters. The first study is performed by artificially attaching the cells to the petridishes with poly-L-lysine and the dead cells are counted with PI fluorescence. Furthermore, dead cells 18h and 24h hours after irradiation were evaluated in order to estimate the reproliferation after treatment. In vitro studies were performed by varying different parameters such as the laser power, the exposure time to laser irradiation per cell and the concentration of nanoparticles. To mimic the eye, we are artificially fabricating a phantom of the vitreous in which RB cells are cultured. The vitreous is mainly composed of water (98%), hyaluronan, collagen and proteins. The hyaluronan, a polymer naturally present under different sizes in the eye, has been chosen to mimic the vitreous. The obtained gel was characterized by measuring its viscosity to validate the model. Cellular growth rate and cells distribution were obtained and compare to those from conventional culture medium. We have verified that the nanoparticles can reach their target RB cells in this phantom and that we can kill up to 80% of Y79 cells with a 0,2μJ/μm2 irradiation. Furthermore, we noticed that viscosity of the surrounding medium has increased the efficiency of the treatment

L'accueil des urgences pédiatriques au CHU d'Angers (enquête auprès des médecins généralistes)

ANGERS-BU Médecine-Pharmacie (490072105) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Multiplexed immunolabelling of cancer using bioconjugated plasmonic gold-silver alloy nanoparticles

Reliable protein detection methods are vital for advancing biological research and medical diagnostics. While immunohistochemistry and immunofluorescence are commonly employed, their limitations underscore the necessity for alternative approaches. This study introduces immunoplasmonic labelling, utilizing plasmonic nanoparticles (NPs), specifically designed gold and gold–silver alloy NPs (Au:Ag NPs), for multiplexed and quantitative protein detection. These NPs, when coupled with antibodies targeting proteins of interest, enable accurate counting and evaluation of protein expression levels while overcoming issues such as autofluorescence. In this study, we compare two nanoparticle functionalization strategies—one coating based on thiolated PEG and one coating based on calix[4]arenes—on gold and gold–silver alloy nanoparticles of varying sizes. Overall results tend to demonstrate a greater versatility for the calix[4]arene-based coating. With this coating and using the classical EDC/sulfo-NHS cross-linking procedure, we also demonstrate the successful multiplexed immunolabelling of Her2, CD44, and EpCAM in breast cancer cell lines (SK-BR-3 and MDA-MB-231). Furthermore, we introduce a user-friendly software for automatic NP detection and classification by colour, providing a promising proof-of-concept for the practical application of immunoplasmonic techniques in the quantitative analysis of biopsies in the clinical setting.info:eu-repo/semantics/publishe

Multiplexed Plasmonic Nano-Labeling for Bioimaging of Cytological Stained Samples

Reliable cytopathological diagnosis requires new methods and approaches for the rapid and accurate determination of all cell types. This is especially important when the number of cells is limited, such as in the cytological samples of fine-needle biopsy. Immunoplasmonic-multiplexed- labeling may be one of the emerging solutions to such problems. However, to be accepted and used by the practicing pathologists, new methods must be compatible and complementary with existing cytopathology approaches where counterstaining is central to the correct interpretation of immunolabeling. In addition, the optical detection and imaging setup for immunoplasmonic-multiplexed-labeling must be implemented on the same cytopathological microscope, not interfere with standard H&E imaging, and operate as a second easy-to-use imaging method. In this article, we present multiplex imaging of four types of nanoplasmonic markers on two types of H&E-stained cytological specimens (formalin-fixed paraffin embedded and non-embedded adherent cancer cells) using a specially designed adapter for SI dark-field microscopy. The obtained results confirm the effectiveness of the proposed optical method for quantitative and multiplex identification of various plasmonic NPs, and the possibility of using immunoplasmonic-multiplexed-labeling for cytopathological diagnostics

Rapid Improvement after Starting Elexacaftor–Tezacaftor–Ivacaftor in Patients with Cystic Fibrosis and Advanced Pulmonary Disease

International audienceRationale: Elexacaftor-tezacaftor-ivacaftor is a CFTR (cystic fibrosis [CF] transmembrane conductance regulator) modulator combination, developed for patients with CF with at least one Phe508del mutation. Objectives: To evaluate the effects of elexacaftor-tezacaftor- ivacaftor in patients with CF and advanced respiratory disease. Methods: A prospective observational study, including all patients aged ⩾12 years and with a percent-predicted FEV1 (ppFEV1) <40 who initiated elexacaftor-tezacaftor-ivacaftor from December 2019 to August 2020 in France was conducted. Clinical characteristics were collected at initiation and at 1 and 3 months. Safety and effectiveness were evaluated by September 2020. National-level transplantation and mortality figures for 2020 were obtained from the French CF and transplant centers and registries. Measurements and Main Results: Elexacaftor-tezacaftor- ivacaftor was initiated in 245 patients with a median (interquartile range) ppFEV1 = 29 (24-34). The mean (95% confidence interval) absolute increase in the ppFEV1 was +15.1 (+13.8 to +16.4; P < 0.0001), and the mean (95% confidence interval) in weight was +4.2 kg (+3.9 to +4.6; P < 0.0001). The number of patients requiring long-term oxygen, noninvasive ventilation, and/or enteral tube feeding decreased by 50%, 30%, and 50%, respectively (P < 0.01). Although 16 patients were on the transplant waiting list and 37 were undergoing transplantation evaluation at treatment initiation, only 2 received a transplant, and 1 died. By September 2020, only five patients were still on the transplantation path. Compared with the previous 2 years, a twofold decrease in the number of lung transplantations in patients with CF was observed in 2020, whereas the number of deaths without transplantation remained stable. Conclusions: In patients with advanced disease, elexacaftor-tezacaftor-ivacaftor is associated with rapid clinical improvement, often leading to the indication for lung transplantation being suspended