Serum-free culture success of glial tumors is related to specific molecular profiles and expression of extracellular matrix–associated gene modules

Glial tumors consist of a heterogeneous group of primary CNS neoplasms. While the clinical outcome of these tumors varies substantially between different grades (World Health Organization [WHO] grades I–IV), only grade I tumors can be treated curatively.1 In the last decade, substantial effort has been put into characterizing the pathogenic mechanisms underlying this complex group of glial cell–derived tumors. These efforts have led to the establishment of several molecular subclasses of glioma, which differentiate types of gliomas based on their intrinsic molecular distinctions and similarities, rather than the conventional characteristics used for histology-based grading.2–5 Indeed, gene expression–based clustering of glioma from several grades within the WHO grading system proved more predictive of survival than the histological classification.4, 6 Thus far, however, this gain of insight has not led to improved clinical outcome for patients, since there are no specific treatment strategies designed to target specific subtypes of glioma as of yet. Consequently, the development of preclinical models that accurately reflect the molecular heterogeneity of glioma is imperative for both translationally relevant drug screening programs and the advancement of patient-tailored treatment options. Commercially available cell cultures of glioma have been demonstrated to poorly mimic the molecular aberrations found in patient samples.7,8 Furthermore, the xenograft models derived from these cultures do not sufficiently recapitulate the histological hallmarks of glioma.9 With the advent of the cancer stem cell hypothesis, several groups have implemented serum-free (SF) cell culture regimens, originally developed for neural stem cell propagation, to establish glioma stem-like cell (GSC) cultures from fresh tumor tissue.10,11 By analyzing gene expression profiles of GSCs and serum supplemented (SS) cultures, a distinct separation between the aforementioned and the parental tumors was revealed by unsupervised clustering.7 Several groups have reported GSC cultures to be superior with regard to retaining the original patient gene expression signature and histological phenotype in xenografts.7,11,12 This has led to a wide variety of applications for these cell culture assays, ranging from inquiries into fundamental hypotheses13,14 to preclinical testing of novel agents.15–17 Several contradicting publications have since been published on the optimal cell culture methodology,18,19 mandatory molecular aberrations for successful propagation,20–22 and positive selection for gene expression signatures related to specific molecular subtypes in vitro.14, 23 Most of these previous publications have focused on the characterization of successfully propagated specimens of glioblastoma multiforme (GBM; WHO grade IV) from adults. Reports on the culture success of grade II and grade III gliomas are sparse.20,24 Therefore, we undertook a characterization study of a large cohort (N = 261), which addresses the distribution of glioma from all histological entities for the outcome of GSC culture attempt. Within equal WHO grades, correlations between cell culture outcome and patient overall survival were assessed. Tumor samples of both successful and unsuccessful cultures (n = 46 in total) were also subjected to molecular analysis, and a number of molecular traits that influence cell culture success rate were identified, as well as genes that may play a role in this process. These results emphasize the need for, and provide leads to, the development of improved culture protocols supporting growth of all subtypes of glioma. This is essential for implementation of this model in drug screening programs for personalized treatment strategies.