Abstract
To identify cellular and molecular changes that driver pediatric low grade glioma (PLGG) progression, we analyzed putative cancer stem cells (CSCs) and evaluated key biological changes in a novel and progressive patient-derived orthotopic xenograft (PDOX) mouse model. Flow cytometric analysis of 22 PLGGs detected CD133+ (<1.5%) and CD15+ (20.7 ± 28.9%) cells, and direct intra-cranial implantation of 25 PLGGs led to the development of 1 PDOX model from a grade II pleomorphic xanthoastrocytoma (PXA). While CSC levels did not correlate with patient tumor progression, neurosphere formation and in vivo tumorigenicity, the PDOX model, IC-3635PXA, reproduced key histological features of the original tumor. Similar to the patient tumor that progressed and recurred, IC-3635PXA also progressed during serial in vivo subtransplantations (4 passages), exhibiting increased tumor take rate, elevated proliferation, loss of mature glial marker (GFAP), accumulation of GFAP−/Vimentin+ cells, enhanced local invasion, distant perivascular migration, and prominent reactive gliosis in normal mouse brains. Molecularly, xenograft cells with homozygous deletion of CDKN2A shifted from disomy chromosome 9 to trisomy chromosome 9; and BRAF V600E mutation allele frequency increased (from 28% in patient tumor to 67% in passage III xenografts). In vitro drug screening identified 2/7 BRAF V600E inhibitors and 2/9 BRAF inhibitors that suppressed cell proliferation. In summary, we showed that PLGG tumorigenicity was low despite the presence of putative CSCs, and our data supported GFAP−/Vimentin+ cells, CDKN2A homozygous deletion in trisomy chromosome 9 cells, and BRAF V600E mutation as candidate drivers of tumor progression in the PXA xenografts.
Highlights
Pediatric low grade gliomas (PLGGs) are slow growing tumors accounting for 1/3 of all childhood brain tumors [1]
To gain insight into in vivo tumor evolution and progression, we examined if the histopathological features and, more importantly, the progression nature of the original patient tumor were replicated in the patient-derived orthotopic xenograft (PDOX) tumors during longterm serial subtransplantations in mouse brains, followed by the analysis of the underlying cellular and molecular (e.g. BRAF V600E mutation and CDKN2A deletion) changes in tumor cells and in the host normal brain cells that drove or accompanied the PDOX tumor progression to identify new therapeutic targets
The overall yields of tumor cells from childhood LGG were low Despite extensive collaborative effort, the tumor tissues obtained for PLGGs were still limited, frequently less than 3 x 3 x 3 mm3 (Table 1)
Summary
Pediatric low grade gliomas (PLGGs) are slow growing tumors accounting for 1/3 of all childhood brain tumors [1]. Mouse models that replicate key biological features of PLGG are highly desired to identify mechanism of recurrence/malignant degeneration and enable preclinical studies of PLGG. We have shown that direct injection of fresh surgical specimens into anatomicallymatched locations in the brains of immunodeficient mice facilitates establishment of clinically-relevant orthotopic xenograft mouse models that replicate the histology, invasive growth, and key genetic features of primary patient tumors [4,5,6,7,8]. The added advantage of patientderived orthotopic xenograft (PDOX) mouse model is that the normal brain responses toward xenograft growth, which is difficult to obtain from patient surgical samples, can be analyzed simultaneously together with brain tumor cells. PDOX mouse models of PLGGs, have not been previously established
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