Phenotypic and Genotypic Characterization of Orthotopic Human Glioma Models and Its Relevance for the Study of Anti‐glioma Therapy

Abstract

Most human gliomas are characterized by diffuse infiltrative growth in the brain parenchyma. Partly because of this characteristic growth pattern, gliomas are notorious for their poor response to current therapies. Many animal models for human gliomas, however, do not display this diffuse infiltrative growth pattern. Furthermore, there is a need for glioma models that represent adequate genocopies of different subsets of human gliomas (e.g., oligodendrogliomas). Here, we assessed the intracerebral growth patterns and copy number changes [using multiplex ligation-dependent probe amplification (MLPA)/comparative genomic hybridization (CGH)] of 15 human glioma lines in nude mice. Most xenografts present with compact growing lesions intracerebrally. Only the E98 and, to a lesser degree, E106 xenograft lines (propagated through subcutaneous growth) consistently produced intracerebral tumors, displaying diffuse infiltrative growth in the brain parenchyma. In contrast, four xenograft lines (E434, E468, E473 and E478), established by direct intracerebral inoculation of human glioma cells and serially propagated intracerebrally, consistently showed extensive diffuse infiltration throughout the brain. After several passages, the neoplastic cells still carry typical chromosomal aberrations [(-1p/-19q in oligodendroglioma, +7/-10 in glioblastoma multiforme (GBM)]. Especially these latter four models and the E98 line thus represent adequate geno- and phenocopies of human gliomas and form an attractive platform to investigate different therapeutic approaches in a preclinical setting.

Isolation of targeting nanobodies against co-opted tumor vasculature

“…[5][6][7][8][9][10][11]29,30 Although gliomas are certainly at the extreme end of a spectrum with respect to infiltrative growth, it was recently shown that targeting of the angiogenic pathway also in subcutaneous xenografts leads to enhanced invasive growth and metastases. 31,32 Such adverse effects are not expected to occur when directing therapy against the existing tumor vasculature.…”

“…28 In vivo Selection of Tumor Vessel Binding Nanobodies via Biopanning of Phage Display Libraries To grow intracerebral E98 and E434 tumors, Balb/c nude mice were transcranially injected with the glioblastoma xenograft line E98 (n ¼ 2) or the oligodendroglioma line E434 (n ¼ 2) essentially as described earlier. 11 In short, a mouse carrying a subcutaneous E98 tumor and one carrying a cerebral E434 xenograft were killed. The tumors were sterile removed, minced to small pieces and filtered through a 70-mm mesh nylon filter.…”

“…17 In vivo biopannings with peptide phage display libraries have been successfully used to obtain organ-specific vascular targeting peptides in animal models. 18,19 In addition, we previously provided proof of concept that via in vivo biopanning of phage antibody-display libraries in orthotopic E98 glioma xenografts, 11,20 antibodies can be isolated that recognize subsets of tumor vessels (submitted). In that work, we used a nanobody phage display library, cloned from lymphocytes of an immune-naive Llama.…”

“…We performed in vivo biopanning of this phage library in the orthotopic glioma xenograft model E98 and the oligodendroglioma xenograft line E434. E98 was used because it displays angiogenic as well as cooptive growth, 11 whereas E434 is a hallmark example of a xenograft, which exclusively displays diffuse infiltrative growth with no apparent sprouting angiogenesis. 11 This approach resulted in the isolation of a number of tumor vessel recognizing nanobodies, also against non-angiogenic blood vessels in diffuse infiltrative tumor areas.…”

“…However, a number of phase II studies have now revealed that, despite inducing a radiological response, these inhibitors do not prolong overall survival and allow disease progression, 5,6 likely because diffuse infiltrative growth is angiogenesis independent and provides a mechanism of escape from anti-angiogenic therapies. [4][5][6][7][8][9][10][11] The phenotypic differences that exist in GBM are reflected in the vasculature, which is highly heterogeneous. 12 Vessels may be angiogenic as well as more mature or co-opted, and the latter ones may be unsusceptible to anti-angiogenic therapy.…”