In vivo loss of tumorigenicity in a patient-derived orthotopic xenograft mouse model of ependymoma

Abstract

IntroductionEpendymomas (EPN) are the third most common malignant brain cancer in children. Treatment strategies for pediatric EPN have remained unchanged over recent decades, with 10-year survival rates stagnating at just 67% for children aged 0-14 years. Moreover, a proportion of patients who survive treatment often suffer long-term neurological side effects as a result of therapy. It is evident that there is a need for safer, more effective treatments for pediatric EPN patients. There are ten distinct subgroups of EPN, each with their own molecular and prognostic features. To identify and facilitate the testing of new treatments for EPN, in vivo laboratory models representative of the diverse molecular subtypes are required. Here, we describe the establishment of a patient-derived orthotopic xenograft (PDOX) model of posterior fossa A (PFA) EPN, derived from a metastatic cranial lesion.MethodsPatient and PDOX tumors were analyzed using immunohistochemistry, DNA methylation profiling, whole genome sequencing (WGS) and RNA sequencing.ResultsBoth patient and PDOX tumors classified as PFA EPN by methylation profiling, and shared similar histological features consistent with this molecular subgroup. RNA sequencing revealed that gene expression patterns were maintained across the primary and metastatic tumors, as well as the PDOX. Copy number profiling revealed gains of chromosomes 7, 8 and 19, and loss of chromosomes 2q and 6q in the PDOX and matched patient tumor. No clinically significant single nucleotide variants were identified, consistent with the low mutation rates observed in PFA EPN. Overexpression of EZHIP RNA and protein, a common feature of PFA EPN, was also observed. Despite the aggressive nature of the tumor in the patient, this PDOX was unable to be maintained past two passages in vivo.DiscussionOthers who have successfully developed PDOX models report some of the lowest success rates for EPN compared to other pediatric brain cancer types attempted, with loss of tumorigenicity not uncommon, highlighting the challenges of propagating these tumors in the laboratory. Here, we discuss our collective experiences with PFA EPN PDOX model generation and propose potential approaches to improve future success in establishing preclinical EPN models.

Human Patient-Derived Brain Tumor Models to Recapitulate Ependymoma Tumor Vasculature

“…Because the NeuralBasal base media in N was formulated with hypotonic osmolarity to specifically favor neuronal growth [22] and the E media contained VEGF as an essential growth factor for endothelial cell growth, we tested whether low osmolarity or VEGF was necessary for ependymoma in vitro growth. Therefore, for EPN-4 (Figure 2B(c)), three media conditions were compared: N+E, N+E without VEGF ("noV"), and DMEM with the same supplements as N+E ("+Supp").…”

“…Cell culture approaches of the 2D monolayer or 3D neurosphere/spheroid have largely been unsuccessful or, at most, inconsistent in preserving ependymoma in vivo characteristics in the long term. Recent efforts in the patient-derived orthotopic xenograft model of ependymoma have reported the in vivo loss of tumorigenicity, highlighting challenges to this approach [22]. Monolayers were grown in tumor medium: DMEM/L-glutamine supplemented with 15% FBS.…”