Inter and intra-tumor heterogeneity of paediatric type diffuse high-grade gliomas revealed by single-cell mass cytometry

Abstract

Paediatric-type diffuse high-grade gliomas (PDHGG) are aggressive tumors affecting children and young adults, with no effective treatment. These highly heterogeneous malignancies arise in different sites of the Central Nervous System (CNS), carrying distinctive molecular alterations and clinical outcomes (inter-tumor heterogeneity). Moreover, deep cellular and molecular profiling studies highlighted the coexistence of genetically and phenotypically different subpopulations within the same tumor mass (intra-tumor heterogeneity). Despite the recent advances made in the field, the marked heterogeneity of PDHGGs still impedes the development of effective targeted therapies and the identification of suitable biomarkers. In order to fill the existing gap, we used mass cytometry to dissect PDHGG inter- and intra-heterogeneity. This is one of the most advanced technologies of the “-omics” era that, using antibodies conjugated to heavy metals, allows the simultaneous measurement of more than 40 markers at single-cell level. To this end, we analyzed eight PDHGG patient-derived cell lines from different locational and molecular subgroups. By using a panel of 15 antibodies, directly conjugated to metals or specifically customized to detect important histone variants, significant differences were highlighted in the expression of the considered antigens. The single-cell multiparametric approach realized has deepened our understanding of PDHGG, confirming a high degree of intra- and inter-tumoral heterogeneity and identifying some antigens that could represent useful biomarkers for the specific PDHGG locational or molecular subgroups.

Inhibition of exosome biogenesis affects cell motility in heterogeneous sub-populations of paediatric-type diffuse high-grade gliomas

“…The genetic and phenotypic inter- and intra-tumoral heterogeneity may be one of the most challenging obstacles in the development of effective therapies for cancer [ 47 – 49 ]. Bulk DNA sequencing, single-cell RNAseq and more recently single-cell proteomic analyses have provided evidence of the intra-tumour heterogeneity in PDHGG [ 10 , 50 , 51 ]. Moreover, diffusely infiltrating gliomas are heterogeneous tumours organized into functional cellular networks as demonstrated in vitro and in vivo using patient-derived cell lines and orthotopic xenograft models [ 10 , 52 , 53 ].…”

Glial fibrillary acidic protein, neurofilament light, matrix metalloprotease 3 and fatty acid binding protein 4 as non-invasive brain tumor biomarkers

“…In addition, only a small proportion of resected tumors are used for histological testing, which is complicated with inter-observer variability, with as many as 12% of tumors being misdiagnosed [ 2 ]. Due to significant intra-tumor heterogeneity [ 3 ], these surgical samples also reflect poorly the entire molecular landscape of the tumor, potentially missing detection of currently established prognostic markers ( e.g., IDH mutation and MGMT promoter hypermethylation) [ 4 – 6 ] and limiting our ability to identify other viable targetable mutations.…”

“…Tumor heterogeneity [ 3 ] influences initial diagnosis and management and remains relevant even during adjuvant therapy, when serial imaging is used to monitor for true tumor recurrence (TTR) versus pseudo-progression (PP; ~15% of cases) or radiation necrosis (RN; ~10% of cases) [ 8 ]. Despite advances in imaging modalities, pathological tissue assessment remains the gold standard for distinguishing among these entities, implying that a notable proportion of patients may undergo unnecessary surgery for RN or PP.…”