Tumor versus Stromal Cells in Culture—Survival of the Fittest?

Abstract

Two of the signature genetic events that occur in human gliomas, EGFR amplification and IDH mutation, are poorly represented in experimental models in vitro. EGFR amplification, for example, occurs in 40 to 50% of GBM, and yet, EGFR amplification is rarely preserved in cell cultures derived from human tumors. To analyze the fate of EGFR amplified and IDH mutated cells in culture, we followed the development over time of cultures derived from human xenografts in nude rats enriched for tumor cells with EGFR amplification and of cultures derived from patient samples with IDH mutations, in serum monolayer and spheroid suspension culture, under serum and serum free conditions. We observed under serum monolayer conditions, that nestin positive or nestin and SMA double positive rat stromal cells outgrew EGFR amplified tumor cells, while serum spheroid cultures preserved tumor cells with EGFR amplification. Serum free suspension culture exhibited a more variable cell composition in that the resultant cell populations were either predominantly nestin/SOX2 co-expressing rat stromal cells or human tumor cells, or a mixture of both. The selection for nestin/SMA positive stromal cells under serum monolayer conditions was also consistently observed in human oligodendrogliomas and oligoastrocytomas with IDH mutations. Our results highlight for the first time that serum monolayer conditions can select for stromal cells instead of tumor cells in certain brain tumor subtypes. This result has an important impact on the establishment of new tumor cell cultures from brain tumors and raises the question of the proper conditions for the growth of the tumor cell populations of interest.

Innovative approaches to establish and characterize primary cultures: an ex vivo 3D system and the zebrafish model

“…Conversely, our 3D system provided the necessary stimuli to enrich the liposarcoma cell fraction over the stromal counterpart. More importantly, 2D culturing of near-patient material induces an in vitro adaptation process that may end up in cancer cells losing their characteristic markers ( Fan et al, 2012 ; Talasila et al, 2013 ). In our study, 3D-cultured cancer cells preserved both MDM2 amplification and gene expression associated with liposarcoma pathogenesis and aggressiveness ( Benassi et al, 2001 ; Gogou et al, 2012 ; Guo et al, 2008 ; Pazzaglia et al, 2004 ; Stratford et al, 2011 ; Somarelli et al, 2016 ), suggesting that this in vitro model could facilitate the identification of disease-specific biomarkers.…”

The angiogenic switch leads to a metabolic shift in human glioblastoma

“…41 However, the proneural GBM from our xenograft model was a primary GBM and showed high levels of EGFR amplification as well as gain of chromosome 7 and loss of chromosome 10. 8,42 Thus, this tumor most likely belongs to a subgroup of proneural tumors associated with a worse prognosis which have been classified as non-GCIMP tumors. 40 The proneural xenograft showed a purely invasive phenotype that could be switched to an angiogenic phenotype and a mesenchymal subclass upon overexpression of EGFR-DN.…”

Engraftment of Human Glioblastoma Cells in Immunocompetent Rats through Acquired Immunosuppression

“…A considerable advantage of the spheroid model compared to cell line-based models is preservation of the patient genotype [ 5 , 9 ]. In particular, EGFR amplification, a hallmark genetic aberration within GBMs is frequently lost/selected against in standard monolayer serum culture, but preserved in biopsy spheroids and in xenografts [ 6 , 10 ]. In general, lack of communication between human and rat antigens and the immune-compromised nature of the host diminish the translational relevance of results obtained from xenograft tumors.…”