Isolation and characterization of tumorspheres from a recurrent pineoblastoma patient: Feasibility of a patient-derived xenograft

Abstract

The existence of tumorspheres (TSs) might confer treatment resistance to pineoblastoma (PB). The existence of PB TSs with cellular immortalization potential has not yet been reported. We developed a procedure for isolating TSs from recurrent PB (rPB) and tested whether their properties made them suitable for use as a patient-derived xenograft (PDX). Immunocytochemical staining, RT-PCR and quantitative real-time PCR showed that, among stemness proteins, CD133, musashi and podoplanin were expressed at elevated levels in rPB TSs, but nestin was not. rPB TSs cultured under neuro-glial differentiation conditions expressed TUBB3, but not GFAP, MBP or NeuN. Unlike glioblastoma TSs, rPB TSs showed no clear evidence of invasion in 3D invasion assay or increased expression of genes associated with epithelial-mesenchymal transition. An orthotopic xenograft showed that tumor xenografts replicated the histopathological features of the patient tumor and expressed similar genome profiles, as determined by short tandem repeat genotyping. These data demonstrate the isolation and the characterization of rPB TSs for the first time. Using an orthotopic xenograft, we showed that rPB TSs could replicate the patient tumor, demonstrating their potential as a PDX for precision medicine.

The mode and dynamics of glioblastoma cell invasion into a decellularized tissue-derived extracellular matrix-based three-dimensional tumor model

“…Tumorspheres (TS) were directly established from fresh GBM tissues as approved by the institutional review board of Severance Hospital, Yonsei University College of Medicine (4-2012-0212). For isolation of TS from GBM specimens, we followed previously published methods for TS isolation from human brain 17 , 34 . The patient-derived GBM cells (pdGCs; TS) were isolated and cultured in 2D, collagen hydrogel, and pdECM environments, respectively.…”

Effect of combined anti-PD-1 and temozolomide therapy in glioblastoma

“…GL261-luc cells were constructed using lentivirus-mediated transduction and puromycin-based selection, and cultured in tumorsphere complete media as previously described. 25,26 U87MG and T98G cells were purchased from Korean Cell Line Bank and grown in media according to American Type Culture Collection recommendations. U87MG cells and T98G cells revealed methylated and unmethylated MGMT promoter region, respectively.…”

Modeling germline mutations in pineoblastoma uncovers lysosome disruption-based therapy

“…Finally, very few human PB cell lines have been successfully established in culture. One line, TS13-19, isolated from sporadic PB and propagated as non-adherent tumorspheres in serum-free media 56 , was also highly sensitive to NOR (Fig. 8g).…”

“…Our in silico drug prediction analysis identified multiple tricyclic, antidepressant drugs such as NOR, which ranked at the top, as potential therapeutics for Rb/p53-deleted and Rb/p53mutated PBs as well as for Dicer/p53-and Rb/Dicer1/p53-deficient lesions. NOR inhibited growth of primary Rb/p53-and Rb/ Dicer1/p53-deficient PB cells as well as a human PB cell line 56 . NOR suppressed autophagy flux not by blocking assembly of the autolysosome but by disrupting the lysosome.…”