Paragangliomas arise through an autonomous vasculo-angio-neurogenic program inhibited by imatinib

Abstract

Tumours can be viewed as aberrant tissues or organs sustained by tumorigenic stem-like cells that engage into dysregulated histo/organogenetic processes. Paragangliomas, prototypical organoid tumours constituted by dysmorphic variants of the vascular and neural tissues found in normal paraganglia, provide a model to test this hypothesis. To understand the origin of paragangliomas, we built a biobank comprising 77 cases, 18 primary cultures, 4 derived cell lines, 80 patient-derived xenografts and 11 cell-derived xenografts. We comparatively investigated these unique complementary materials using morphofunctional, ultrastructural and flow cytometric assays accompanied by microRNA studies. We found that paragangliomas contain stem-like cells with hybrid mesenchymal/vasculoneural phenotype, stabilized and expanded in the derived cultures. The viability and growth of such cultures depended on the downregulation of the miR-200 and miR-34 families, which allowed high PDGFRA and ZEB1 protein expression levels. Both tumour tissue- and cell culture-derived xenografts recapitulated the vasculoneural paraganglioma structure and arose from mesenchymal-like cells through a fixed developmental sequence. First, vasculoangiogenesis organized the microenvironment, building a perivascular niche which in turn supported neurogenesis. Neuroepithelial differentiation was associated with severe mitochondrial dysfunction, not present in cultured paraganglioma cells, but acquired in vivo during xenograft formation. Vasculogenesis was the Achilles’ heel of xenograft development. In fact, imatinib, that targets endothelial-mural signalling, blocked paraganglioma xenograft formation (11 xenografts from 12 cell transplants in the control group versus 2 out of 10 in the treated group, P = 0.0015). Overall our key results were unaffected by the SDHx gene carrier status of the patient, characterized for 70 out of 77 cases. In conclusion, we explain the biphasic vasculoneural structure of paragangliomas and identify an early and pharmacologically actionable phase of paraganglioma organization.Electronic supplementary materialThe online version of this article (10.1007/s00401-017-1799-2) contains supplementary material, which is available to authorized users.

Cancer Stem Cells in Pheochromocytoma and Paraganglioma

“…Contrary to the findings within other adrenal tumors, OCT4, CD133, and NANOG expression was not detected in any of the samples from Oudjik et al, however expression of other stem cell markers such as DLK1/PREF1, NGFR, LIN28, SOX2, and THY1 was observed in 12-40% of cases, whereas the expression of Nestin, SOX17 and CD117 was identified less frequently in 2-3% of cases (65). In contrast, a case study of a PCC in pregnancy showed high expression of Nestin (67), which, as CD133, has also been detected in PGLs (66). As Nestin marks sustentacular cells, it raises questions regarding their role in PCCs/PGLs.…”

“…For example, the expression of OCT4 is inconsistent as a study by Looijenga et al concluded that PCCs/PGLs are negative for OCT4 (63), yet a subsequent study by Alexander et al showed strong and diffuse cytoplasmic staining of OCT4 in PCCs and metastases (64). Oudijk et al analyzed the expression of relevant CSC markers in tissue microarrays of a large number of PCCs and PGLs and showed that frequently CSC marker expression was associated with cluster 1 SDHx-mutated tumors (65), though expression of CSC markers has also been observed in PGLs from patients without SDHx mutations (66). Contrary to the findings within other adrenal tumors, OCT4, CD133, and NANOG expression was not detected in any of the samples from Oudjik et al, however expression of other stem cell markers such as DLK1/PREF1, NGFR, LIN28, SOX2, and THY1 was observed in 12-40% of cases, whereas the expression of Nestin, SOX17 and CD117 was identified less frequently in 2-3% of cases (65).…”

“…As PCCs/PGLs occasionally express CD117 (Table 1), the tyrosine kinase inhibitor imatinib has for example been tested on in vitro cultures of PGL cell cultures, where it was shown to inhibit growth of both SDHx-unrelated and related tumor cells. Furthermore, it prevented xenograft formation in mice transplanted with patient-derived cells (66). CSCs are known to be more resistant to conventional chemo-and radio-therapy compared to non-CSC populations.…”

Extracellular Vesicles in Feto–Maternal Crosstalk and Pregnancy Disorders

“…EVs are in fact characterized by the phenotype of the parental cells, and, for example, given that endothelial cells express the marker CD31, also known as platelet endothelial cell adhesion molecule (PECAM1) [40,41]; therefore, endothelial-derived EVs are identified by the surface expression of CD31 [36]. In the same way, because mesenchymal stem cells expose CD90 [42][43][44], it has been observed that their derived vesicles are recognized by the positivity to CD90, which is a glycophosphatidylinositol cell surface protein, originally discovered as a thymocyte antigen [36].…”

Advances in paraganglioma–pheochromocytoma cell lines and xenografts

“…Verginelli et al also recently described attempts to develop PDX models of paragangliomas (Verginelli et al 2018), using a total of 90 PGL fragments from 16 patients and reporting an overall take rate of 89% (80/90). Xenografts were investigated 4.5-10 months posttransplantation and found to present as 4-6 mm nodules that infiltrated adjacent murine neurovascular bundles.…”