Jin Wook Hwang1, Julien Loisel-Duwattez2, Christophe Desterke1, Theodoros Latsis1, Sarah Pagliaro1, Frank Griscelli1, Annelise Bennaceur-Griscelli3, Ali G Turhan4. 1. INSERM UMR-S 935, Université Paris Sud, 94800 Villejuif, France; ESTeam Paris Sud, Université Paris Sud, 94800 Villejuif, France. 2. INSERM U1195, Université Paris Sud, Faculté de Médecine, APHP, Service de Neurologie, Bicêtre Hospital, 94276 le Kremlin Bicêtre, France. 3. INSERM UMR-S 935, Université Paris Sud, 94800 Villejuif, France. 4. INSERM UMR-S 935, Université Paris Sud, 94800 Villejuif, France; INGESTEM National IPSC Infrastructure, 94800 Villejuif, France; Division of Hematology, Paris Sud University Hospitals, Le Kremlin Bicêtre 75006, Villejuif 94800, France; ESTeam Paris Sud, Université Paris Sud, 94800 Villejuif, France. Electronic address: turviv33@gmail.com.
Abstract
BACKGROUND: Current experimental models using either human or mouse cell lines, are not representative of the complex features of GBM. In particular, there is no model to study patient-derived iPSCs to generate a GBM model. Overexpression of c-met gene is one of the molecular features of GBM leading to increased signaling via STAT3 phosphorylation. We generated an iPSC line from a patient with c-met mutation and we asked whether we could use it to generate neuronal-like organoids mimicking features of GBM. METHODS: We have generated iPSC-aggregates differentiating towards organoids. We analyzed them by gene expression profiling, immunostaining and transmission electronic microscopy analyses (TEM). RESULTS: Herein we describe that c-met-mutated iPSC aggregates spontaneously differentiate into dopaminergic neurons more rapidly than control iPSC aggregates in culture. Gene expression profiling of c-met-mutated iPSC aggregates at day +90 showed neuronal- and GBM-related genes, reproducing a genomic network described in primary human GBM. Comparative TEM analyses confirmed the enrichment of these structures in intermediate filaments and abnormal cilia, a feature described in human GBM. The c-met-mutated iPSC-derived organoids, as compared to controls expressed high levels of glial fibrillary acidic protein (GFAP), which is a typical marker of human GBM, as well as high levels of phospho-MET and phospho-STAT3. The use of temozolomide (TMZ) showed a preferential cytotoxicity of this drug in c-met-mutated neuronal-like organoids. GENERAL SIGNIFICANCE: This study shows the feasibility of generating "off-the shelf" neuronal-like organoid model mimicking GBM using c-met-mutated iPSC aggregates and its potential future use in research.
BACKGROUND: Current experimental models using either human or mouse cell lines, are not representative of the complex features of GBM. In particular, there is no model to study patient-derived iPSCs to generate a GBM model. Overexpression of c-met gene is one of the molecular features of GBM leading to increased signaling via STAT3 phosphorylation. We generated an iPSC line from a patient with c-met mutation and we asked whether we could use it to generate neuronal-like organoids mimicking features of GBM. METHODS: We have generated iPSC-aggregates differentiating towards organoids. We analyzed them by gene expression profiling, immunostaining and transmission electronic microscopy analyses (TEM). RESULTS: Herein we describe that c-met-mutated iPSC aggregates spontaneously differentiate into dopaminergic neurons more rapidly than control iPSC aggregates in culture. Gene expression profiling of c-met-mutated iPSC aggregates at day +90 showed neuronal- and GBM-related genes, reproducing a genomic network described in primary humanGBM. Comparative TEM analyses confirmed the enrichment of these structures in intermediate filaments and abnormal cilia, a feature described in humanGBM. The c-met-mutated iPSC-derived organoids, as compared to controls expressed high levels of glial fibrillary acidic protein (GFAP), which is a typical marker of humanGBM, as well as high levels of phospho-MET and phospho-STAT3. The use of temozolomide (TMZ) showed a preferential cytotoxicity of this drug in c-met-mutated neuronal-like organoids. GENERAL SIGNIFICANCE: This study shows the feasibility of generating "off-the shelf" neuronal-like organoid model mimicking GBM using c-met-mutated iPSC aggregates and its potential future use in research.