Tumour Stem Cells in Major Types of Tumours of the Central Nervous System in Children
According to the new edition of the WHO Classification of Tumours of the Central Nervous System, tumours of the CNS in children are graded and classified according to a combination of histological and molecular characteristics, the clinical features of which and pathways of tumourigenesis are significantly different from adult tumours
[14]. Here reviews the origins and markers of CSCs in the following major types of tumours of the CNS in children (
Table 1).
Table 1. Tumour stem cell markers and markers for their differentiated forms from reports according to the WHO Classification of Tumours of the Central Nervous System 2021 edition.
2.1. Gliomas, Glioneuronal Tumours, and Neuronal Tumours
The catalogue of gliomas, glioneuronal tumours, and neuronal tumours in the 2021 WHO classification contained unaltered types from the 2017 edition, such as ependymal tumours, and glioneuronal and neuronal tumours, while their subtypes expanded by divergent molecular signatures. Paediatric-type diffuse low-grade gliomas and paediatric-type diffuse high-grade gliomas were new types of tumours to clearly separate these prognostically and biologically distinct groups of tumours from adult ones, as well as circumscribed astrocytic gliomas that substituted other astrocytic tumours in the 2017 WHO classification
[5][62].
The most common low-grade glioma in children was pilocytic astrocytoma, belonging to the tumour type of circumscribed astrocytic gliomas, caused by extracellular-signal-regulated kinase (ERK) constitutive activation from its upstream BRAF mutations (BRAF-KIAA1549 fusion or BRAF-V600E) or NF1 alterations
[63], and the less common active mutations of FGFR1 and PTPN11, and NTRK2 fusion
[64]. The CSCs from pilocytic astrocytoma were identified in the highly pioneering work in insolating CSCs from CNS tumours by primary tumour sphere culture
[11]. CD133 and Nestin were first used as CSC markers for pilocytic astrocytoma, while pluripotent markers SOX2, Nanog, Oct4, CD44, Integrin α6 (CD49f), SSEA-1, or CD15, ATP Binding ABCG1, and neuro-stem cell markers such as Podoplanin, and markers for different precursor/progenitor cells such as BLBP and A2B5, were later introduced by different research groups
[11][23][24][25][26]. Specific markers to confirm the differentiation status of pilocytic astrocytoma CSCs were reported as Beta-III Tubulin, GFAP, Oligodendrocyte Marker O4, S100B, NF, and EAAT1/2
[11][24][25][26]. There were also controversial markers such as Olig2 and PDGFRα, which were regarded as not only CSC markers, but also differentiation markers, in different reports
[24][26][27][28]. This situation was ascribed to the definition of CSCs in CNS, since neuro-stem cells, neural precursor cells, and even neuron/oligodendrocyte/glial progenitor cells were all transformable to the CSCs in generating the tumours in different studies
[13]. The latest hypothesis was that pilocytic astrocytoma CSCs were more differentiated radial glia/oligodendrocyte precursor cell-like cells than those immature neuro stem cell-like CSCs in high-grade gliomas
[28]. In pleomorphic xanthoastrocytoma, another low-grade circumscribed astrocytic glioma, CSCs were preliminarily studied and it was revealed that CD133 was low, while a high level of CD15 was detected, with gradually decreased GFAP-positive cells by passages, in a xenograft model
[29]. In other low-grade gliomas, CSCs were expectedly discovered by the recognition of CSC markers of CD133, SOX2, Nestin, and the confirmation of their differentiation by oligodendrocyte lineage markers of CNPase, glial markers of GFAP, and NFP, in oligodendroglioma (defined as diffuse low-grade glioma, MAPK pathway-altered in the 2021 WHO classification)
[15]; by CSC markers of SOX2, AGR2, and differentiation markers of GFAP, Beta-III Tubulin, in dysembryoplastic neuroepithelial tumours
[30]; by CSC markers of CD133, Nestin, SOX2, Nanog, CD44, EGFR, hTERT, BLBP, GFAP-delta, Olig2, and ASCL1; and by differentiation markers of Beta-III Tubulin, GFAP, Synaptophysin, MAP2, NeuN, BMP2, BMPR1B, and PSANACM in central neurocytoma
[31][32].
High-grade gliomas in children are relatively less common, but their prognosis is extremely poor. Since the WHO 2021 classification withdrew glioblastoma in children and established paediatric-type diffuse high-grade gliomas as a new catalogue, old entities of glioblastoma, anaplastic astrocytoma, diffuse intrinsic pontin gliomas (DIPG), and other high-grade gliomas were reclassified to the new tumour type. The CSCs of paediatric high-grade gliomas were isolated as early as 2003, with stemness markers of Nestin and Musashi, along with the differentiation markers of Beta-III Tubulin, GFAP, and Oligodendrocyte Marker O4
[12]. Later, CD133, SOX2, CD44, SEEA-1, ALDH, L1CAM, Olig2, Nanog (partial, case-by-case), Podoplanin, and Bmi-1 were recognized as additional CSC markers, and MAP2, NFP, CNPase, NeuN, and MBP were introduced as differentiation markers
[15][20][21][22]. Additionally, in H3 K27-altered Diffuse Midline Gliomas (DMG), a group of devastating highly malignant tumours raised in children, the driver mutation itself could have interfered with differentiation and promoted stem cell proliferation with maintained stemness
[65]. Studies on DIPGs by human-derived primary cell lines or iPSC harbouring H3K27M not only revealed consistent CSC markers of CD133, SOX2, Nestin, PAX6, Vimentin, and differentiation markers of MAP2, GFAP, NeuN, NOTCH 1, and CSPG4 in previous studies on high-grade gliomas, but also introduced new CSC markers of ALDH and controversial markers of Olig2 and PDGFRα; due to the origins of DIPG, CSCs differed in different research groups
[16][17][18][19].
Ependymal tumours are groups of tumours derived from the glial cell lining of the ventricular system, the subgroups of which have been carefully sorted by molecular signatures and anatomic locations. These tumours can develop in both children and adults, and treatments are challenging due to the locations and chemo-radiation resistances of tumours in children. Although the driven mutations of the supratentorial ependymoma (ZFTA fusion-positive or YAP1 fusion-positive) and posterior fossa ependymoma (either group PFA or PFB) are fully elucidated, according to the WHO 2021 classification
[5][66], there is still no efficient treatment to control them. In the ground-breaking work from 2005, CSCs of ependymoma were supposed to be a transformation of radial glia cells (RGCs), since ependymomas recapitulated the gene expression profiles of regionally specified RGCs. The CSC markers of Nestin, CD133, RC2, BLBP, and differentiation markers of β-III Tubulin, MAP2, GFAP, S100, CNPase, and NG2 were accepted
[33]. In later studies, the driver mutations of ZFTA (RELA) fusion and YAP1 fusion were both confirmed to contribute to the oncogenic signalling by inducing neural progenitor cells or neural stem cells to form ependymomas
[67][68], and single-cell RNA sequencing revealed subgroups of tumour cells called undifferentiated ependymal cells (UECs) that might act as CSCs in ependymomas, since their RNA profile overlapped with most of progenitors in different lineages, with spatiotemporally specific signatures in separating CSCs of supratentorial and anteroposterior ependymoma
[35][36]. In total, the CSC markers for ependymoma were expanded (there were numerous markers from single-cell sequencing and, here, researchers list important ones) to include Vimentin, HES1, PBX1, SOX9, PAX3, Protein c-Fos, ZFP36, LGR5, EGR1, JUN, and ATF3, and differentiation markers of Synaptophysin, O4, Olig1/2, APC, CSPG2, DNAAF1, RSPH1, and CAPS were added
[34][35][36].
2.2. Choroid Plexus Tumours
Choroid plexus tumours originate from choroid plexus epithelial cells, which derive from neuroepithelial progenitors with MYC overexpression and a loss of p53, and are extremely rare in adults but are commonly seen in young children under 1 year of age
[37][38]. According to the 2021 WHO classification, three subtypes of choroid plexus tumours were grouped as choroid plexus papilloma, atypical choroid plexus papilloma, and choroid plexus carcinoma
[5]. Although recent molecular subgroups based on epigenetic profiles have been matched with subtypes in view of pathological characteristics, patients with choroid plexus papilloma still have poor prognoses, with a 5-year overall survival of 65%
[69][70]. Since choroid plexus papilloma is regarded as a fully differentiated benign papillary neoplasm closely resembling non-neoplastic choroid plexus tissue, most studies on CSCs of choroid plexus tumours have focused on choroid plexus carcinoma
[38]. The markers for CSCs in choroid plexus carcinoma are MYC, Nestin, ATOH1, BLBP, GFAP, Geminin, GDF7, and differentiation markers are mainly for choroid plexus epithelial cells, such as TTR, AQP1, OTX2, GMNC, MCIDAS, FOXJ1, CCNO, TAp73, and MYB
[37][38][39].
2.3. Embryonal Tumours
CNS embryonal tumours are different groups of highly malignant tumours mainly affecting young children, with gradually increased incidence over a long time period
[6][71]. In the new WHO classification, the two types of embryonal tumours are medulloblastomas and other CNS embryonal tumours. Molecular subtypes of medulloblastomas are extraordinarily famous and practical due to their perfect correlation with clinical features
[5]. The first report of CSCs in medulloblastoma was the pioneering work on isolating multiple types of paediatric brain tumours in 2003, with anaplastic astrocytoma and glioblastoma, by the same CSC markers as Nestin and Musashi
[12]. The mouse model confirmed that the SHH subgroup medulloblastoma arose from both granule neuron precursors (GNPs) and multipotent neural stem cells, by CSC markers of p75NTR, TrkC, Zic1, MATH1, SOX1, SOX2, PLZF, DACH1, Multimerin 1, PAX6, ATOH1, MycN, and differentiation markers of NeuN, GABRA6, Synaptophysin, PAX2, BLBP, and O4
[40][41]. Then, in medulloblastoma cell lines of SHH, group3/4 subtypes, CSCs were successfully discriminated by markers of CD133, Nestin, SOX1, and SOX2, and markers of GFAP, CD44, CD24, and Beta-III Tubulin were applied to indicate differentiation
[42]. Furthermore, an interesting phenomenon had been noticed that, in all four subgroups of medulloblastoma, a small group of Wnt-active cells existed and could impair the stemness of CSCs by reducing Bmi-1 and SOX2 levels
[72].
Embryonal tumour with multilayered rosettes (ETMR) is another type of common aggressive embryonal tumour in young children, with C19MC-altered or DICER1-altered molecular signatures. Although ETMRs consist of populations resembling neural stem cells, radial glial cells, and more differentiated cells, the neuronal and glial cell markers can be detected in limited parts of tumour tissues
[48]. CSC components have been found in different histological subtypes of ETMRs by markers of LIN28A/B, HMGA2, Nestin, Vimentin, Oct4, SOX2, Nanog, CRABP1, DNMT3B, SOX3, SOX11, PAX6, SALL4, POU3F2, MEIS1/2, MYCN, Wee1, and CHEK2, and their induced differentiation cells have been marked with AQP4, GFAP, Synaptophysin, NeuN, and NFP
[47][48][49].
Atypical teratoid/rhabdoid tumours (AT/RT) have been divided into three molecular subtypes, TYR, SHH, and MYC, which predominantly affect infants or young children, with a remarkably simple alteration of SWItch/sucrose nonfermentable related, Matrix-associated, Actin-dependent regulator of Chromatin, Subfamily B1 (SMARCB1) or SMARCA4, but with devastating outcomes
[73]. After the first report of isolating CSCs of AT/RT from patients by a marker of CD133 through sphere culture and a xenograft model
[43], SMARCB1-deficient pluripotent stem cells-based experiments revealed that the origin of CSCs in AT/RT, SMARCB1-deficient neural progenitor-like cells efficiently gave rise to AT/RT-like tumours and their stemness signatures worsened the prognosis
[46]. The markers of AT/RT CSCs are general markers of CD133, Nestin, Musashi, Nanog, Oct4, and SOX2, with ALDH, SALL4, MYC, LIN28A/B, NCAM, PAX6, and KLF4, and the induced differentiation markers are MAP2, Vimentin, GFAP, synaptophysin, CD99, S-100, EMA, and SMA
[43][44][45][46].
2.4. Germ Cell Tumours
Intracranial germ cell tumours (GCT) are less seen in children in Western countries as they account for 0.4–3.4% of all paediatric CNS tumours, while in Asia, they account for as high as 11%. Since GCTs have various types based on their cell components, the origins of GCTs have been explored by experts with long-standing controversy in “germ cell” and “pluripotent stem cell” theories
[74]. According to early reports of GCTs, stem-cell-related proteins of C-KIT, Oct3/4, transcription factor AP-2 gamma (TFAP2C, or AP-2γ), Nanog, and germ-cell-specific proteins of Melanoma-associated antigen A4 (MAGE-A4), cancer-testis antigen 1B (CTAG1B, or NY-ESO-1), expressed in tumour tissue, and primordial germ cells are regarded as tumour generators to convince “germ cell” theory
[75]. Nevertheless, the Oct4-activated neural stem cell can trigger the formation of teratoma in the brain and raise the possibility of “pluripotent stem cell” theory
[76]. However, it has been widely accepted that germinoma is a prototype of all GCTs, by CSC markers of Oct4, Nanog, SOX17 (germinoma), SOX2 (infantile GCTs), and differentiation markers of PLAP and KIT (germinoma differentiation markers from ES cells, rare in intracranial none germinoma GCTs (NGGCTs))
[50]. Due to the complexity of tumour composition, it is inaccurate to use markers or histopathology alone using small specimens for diagnosis, also in the identification of CSCs in GCTs
[77]. Recent milestone research has supposed that germinomas are raised from primordial germ cells, while NGGCTs originate from embryonic stem cells by transcriptome and methylome analysis, and CSC markers are separated into different subtypes, such as Lin28A, SOX2 KLF2/4 as general CSC markers, PIWIL1, DAZL, DDX4, NANOS3, and ERVW-1 for germinomas, and Nanog and Oct4 for NGGCTs
[51].
2.5. Tumours of the Sellar Region
The most seen tumour in the sellar region in children is the adamantinomatous craniopharyngioma (ACP), which is a distinct type of tumour other than papillary craniopharyngioma in the WHO 2021 classification, due to their different clinical demographics, radiologic features, histopathologic findings, genetic alterations, and methylation profiles
[5]. Interestingly, several studies found that CSCs of ACPs shared the same origin with paediatric rare tumour of pituitary adenoma, from pituitary stem cells (PSCs)
[53]. The CSC markers for ACPs were Nestin, SOX2, Oct4, CD133, KLF4, SOX9, β-catenin, MYC, SCA1, HESX1, and KLF2/4, and the differentiation markers were similar to pituitary hormone-secreting cells, such as PIT-1 (or POU1F1), TBX19 (or TPIT), and SF-1 (or NR5A1)
[52][53][54]
2.6. Other Tumours
In other less common assorted CNS tumours in children, CSCs differ from each other due to their different oncogenesis alterations and tumour origins.
In a catalogue of pineal tumours, pineoblastoma caused by the recurrent homozygous deletion of DROSHA and the microduplication of PDE4DIP in primitive neuroepithelia acted as an aggressive malignancy, and their CSCs were marked with CD133, Musashi, Podoplanin, and neuro-glial differentiation by Beta-III Tubulin
[55][56].
In cranial and paraspinal nerve tumours, neurofibroma and its high-grade form malignant peripheral nerve sheath tumour (MPNST), mainly presented in the department of neurology or dermatology, were more frequently seen than schwannoma in children. CSCs in these tumours were somewhat different, the markers of which were PLP, Nestin, P75, GAP43, and Sox10, with GFAP, S100, and GAP43 (as a Schwann cell marker) in differentiated cells, in neurofibroma and MPNST
[58][59]; and Oct4, SOX2, Nanog, MYC, KLF4, CD133, CD44, and CXCR4 in CSCs of schwannoma
[57].
CNS Ewing sarcoma belongs to the catalogue of mesenchymal, non-meningothelial tumours, driven by EWS gene fusions, which might originate from mesenchymal stem cells since their CSCs are confirmed by CD44, CD59, CD73, CD29, and CD54, CD90, CD105, and CD166, with differentiation markers of chondrocyte lineage-SOX9, COL10A1, PPARg2, FABP4, LPL, and osteogenic differentiation SPP1, ALPL, and RUNX2
[60].
Meningioma is rare in young children and its CSCs have been mainly reported and isolated from adult patients, with markers of Oct4, SOX2, Nanog, MYC, KLF4, CD133, and Nestin
[61].