Complex brain organoid model reproducing the glioblastoma tumor niche and its immune component for the development of personalized medicine
Glioblastoma, responsible for 3,500 annual deaths in France, is an extremely aggressive brain tumor that is resistant to current treatments. Clinical trials in immunotherapy have shown only transient effects, underscoring the importance of understanding resistance mechanisms and developing more targeted therapeutic strategies.
We have developed an innovative model of glioma stem cell invasion in immunocompetent and vascularized brain organoids derived from induced pluripotent stem cells (iPSCs) (Raguin et al., submitted). This model faithfully reproduces the glioblastoma tumor niche, including vascular co-option, reprogramming of microglia into tumor-associated macrophages, and tumor recurrence following radiotherapy.
The aim of this PhD project is to derive a universal brain organoid model for the transfer of glioma cells from patients and lymphocytes to optimize the immunotherapy approach (CAR-T cells).
The project involves creating a universal model of human brain organoids that are immunologically "silent" by suppressing the expression of the HLA class I/II system in iPSCs (CRISPR/CAS9 for the ß2M and CIITA genes). Additionally, it aims to elucidate the mechanisms of immunosuppression induced by irradiation, such as the reprogramming of microglial/macrophage cells and the involvement of senescence. Various approaches to make the tumor microenvironment more conducive to immunotherapy will be explored, including activating the type I interferon pathway through genetic modification or with cGAS/STING pathway agonists. Subsequently, the use of CAR-T cells targeting an antigen overexpressed by glioblastoma cells (CD276/B7-H3) will be studied. This model could be used in personalized medicine by co-cultivating patients' tumor cells, monocytes, and CAR-T cells.
This project offers innovative perspectives for the personalized treatment of glioblastoma via immunotherapy and could represent a major advancement in this therapeutic approach.
Dynamic interplay of Rad51 nucleoprotein filament-associated proteins - Involvement in the regulation of homologous recombination
Homologous recombination (HR) is an important repair mechanism for DNA double-strand breaks induced by ionizing radiation. A key step in HR is the formation of Rad51 nucleoprotein filaments on the single-stranded DNA that is generated from these breaks. We were the first to show, using yeast as a model, that a tight control of the formation of these filaments is essential for HR not to induce chromosomal rearrangements by itself (eLife 2018, Cells 2021). In humans, the functional homologs of the yeast control proteins are tumor suppressors. Thus, the control of HR seems to be as important as the mechanism of HR itself. Our project involves the use of new molecular tools that allow a breakthrough in the study of these controls. We will use a functional fluorescent version of the Rad51 protein, first developed by our collaborators A. Taddei (Institut Curie), R. Guérois and F. Ochsenbein (I2BC, Joliot, CEA). This major advance will allow us to observe the influence of regulatory proteins on DNA repair by microscopy in living cells. We have also developed highly accurate structural models of control protein complexes associated with Rad51 filaments. We will adopt a multidisciplinary approach based on genetics, molecular biology, biochemistry, and protein structure in collaboration with W.D. Heyer (University of California, Davis, USA), to understand the function of the regulators of Rad51 filament formation. The description of the organization of these proteins with Rad51 filaments will allow us to develop new therapeutic approaches.
Development of a dosimetry system to track alpha particles in in vitro assays for Targeted Alpha Therapy
Targeted Alpha Therapy (TAT) is a promising new method of treating cancer. It uses radioactive substances called alpha-emitting radioisotopes that are injected into the patient's body. These substances specifically target cancer cells, allowing the radiation to be concentrated where it is needed most, close to the tumors. Alpha particles are particularly effective because of their short range and ability to target and destroy cancer cells.
As with any new treatment, TAT must undergo preclinical studies to test its effectiveness and compare it to other existing treatments. Much of this research is done in laboratory, where cancer cells are exposed to these radioactive substances to observe their effects, such as cell survival. However, assessing the effects of alpha particles requires special methods because they behave differently than other types of radiation.
Recently, a method for measuring the radiation dose received by cells in laboratory experiments has been successfully tested. This method uses detecto
Effects of the combination of ionizing radiation and radio-enhancing molecules in breast cancer models
The proposed program aims to evaluate the efficacy of molecules enhancing the effects of radiotherapy, in in vitro and in vivo models of breast cancer. Two types of molecules, namely an inhibitor of mitochondrial genome maintenance and an inhibitor of the Base Excision Repair pathway, will be tested for radiopotentiation efficacy in the models.
The proposed inhibitors, whether targeting mitochondrial genome maintenance or the BER pathway, are already being investigated in vitro, both in the laboratory and by collaborators. We have shown that inhibition of the mechanisms targeted leads to an impairment in DNA damage repair following genotoxic stress. During this project, we will evaluate the effects of inhibitors on DNA damage repair induced by irradiation of different types (conventional, ultra-high dose rate, even extreme dose rate) and the associated mechanisms.
Variability in response to therapeutic combinations is frequently observed when moving from in vitro to in vivo models. We will therefore evaluate the inhibitors on cell line models well characterized in the laboratory, and corresponding to different breast cancer subtypes. On the other hand, the studies will be completed by a validation of the effects observed in vitro on a murine model of breast cancer. This xenograft model, developed in immunocompetent animals, will enable us to monitor the clinical, histological and immune response of the animals and their tumors, in order to confirm the interest of the molecules for therapeutic application in support of radiotherapy.
The proposed program will benefit from the laboratory's collaborations with physicists and chemists, and IRCM's experimental facilities and platforms (irradiation, animal experimentation, microscopy, cytometry, etc.).