Calixto-Hope G. Lucas

Background: Tumor invasion and malignant transformation are spatiotemporally coordinated processes that shape clinical behavior, diagnostic classification, and treatment response. Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive tumors that can arise sporadically or in people with neurofibromatosis type 1 (NF-1), yet how distinct spatial niches underlie MPNST intratumor heterogeneity, local invasion, or malignant transformation from benign precursors is not fully understood. Methods: We integrate spatial analysis of 50,896 transcriptomes from 15 human MPNSTs with deep learning image analysis to define the interplay between genetic alterations, the immune microenvironment, and epigenetic cell fate regulation in MPNST invasion and transformation. Results: Spatial transcriptomics reveal distinct infiltrating immune cells and immunomodulatory spatial clusters in sporadic compared to NF-1–associated MPNSTs. Local invasion is marked by spatially defined immunomodulatory and dedifferentiation programs. Analysis of 2 NF-1–associated MPNSTs with contiguous morphologically defined neurofibroma and MPNST components demonstrates that NF-1–associated tumors exhibit a similar malignant transformation process associated with dynamic immune infiltration, chromosome 9p loss, and dedifferentiation concomitant with increased cell proliferation. Finally, deep learning morphology encoders were trained on spatial transcriptomic data to predict spatial transcriptomic cluster identity and chromosome 9p loss from hematoxylin and eosin–stained images although the generalizability of such encoders remains a challenge. Conclusions: Our data provide a spatial framework for peripheral nerve sheath tumor transformation although the limited sample size of the present cohort will require additional validation of the proposed model.

Background: High-grade astrocytoma with piloid features (HGAP) is a recently defined tumor type that is not well understood. Prognostic factors of clinical outcomes are not well established. Methods: Methylation profiling was performed on tumor samples, many at the National Cancer Institute (NCI) Laboratory of Pathology, and others from publicly available sources. Methylation classifier scores of ≥0.90 to the HGAP class on the NCI-Bethesda classifier version 3 were included. Clinical features were collected from the medical record. Survival analyses were performed using the Kaplan-Meier and Cox-proportional hazards methods. Results: The cohort comprised 421 patients. There were high rates of ATRX alteration (62%), CDKN2A/B homozygous loss (78%) and MGMT promoter methylation (53%). MAPK alterations were identified in 74% of evaluable samples. The median age was 46 years, and posterior fossa location was predominant (52%). The median overall survival was 88 months. Older age (P = .01) and the presence of an ATRX alteration (P = .04) were found to be negative prognostic factors. The presence of cystic features on magnetic resonance imaging (MRI) was found to be favorably prognostic (P = .01). Factors that were not significantly associated with prognosis included histologic high-grade features, CDKN2A/B homozygous deletion, MGMT promoter methylation, extent of resection, and presence of NF1 syndrome. Conclusions: This large cohort establishes relative frequencies of several important markers. Additionally, older age, the presence of an ATRX alteration, and cystic features on MRI were found to be prognostic. Our work may aid in optimizing treatment regimens for patients with this tumor type.

Background: Atypical teratoid rhabdoid tumor (ATRT) is the most common malignant brain tumor in infants. ATRT is associated with inactivation/deletion of SMARCB1, a member of the SWI/SNF chromatin remodeling complex. SMARCB1 loss contributes to tumorigenicity by compromising SWI/SNF activity at specific loci associated with the CoREST repressor complex, which regulates transcription at critical gene promoters and enhancers. We therefore explored the role of the CoREST repressor complex in ATRT. Methods: We evaluated the effects of the bifunctional LSD1/HDAC1/2 small molecule CoREST inhibitor, corin, on ATRT tumor cell growth, apoptosis, differentiation, gene expression, and chromatin accessibility. Results: Corin inhibited the growth of ATRT cells regardless of their epigenetic subgroup. Corin caused increased tumor cell apoptosis and differentiation. ATAC-seq showed increased chromatin accessibility in corin-treated ATRT cells, with changes seen at genes associated with neuronal differentiation and synaptic function. RNA-seq confirmed increased expression of neuronal differentiation genes in ATRT cells treated with corin. Knockdown of RCOR2 phenocopied the effects of corin and desensitized the cells to the drug, confirming corin specificity to the CoREST complex. Corin suppressed orthotopic ATRT tumor growth, leading to significant extension of lifespan in ATRT mouse models. Corin caused increased histone acetylation (H3K9ac) and methylation (H3K4Me1) in ATRT orthotopic xenografts, consistent with on-target pharmacodynamics. Conclusions: The CoREST inhibitor, corin, suppressed tumor growth, induced differentiation, and promoted apoptosis in ATRT, leading to significantly increased survival of mice bearing ATRT orthotopic xenografts. Our results suggest a potential application of CoREST complex inhibitors in patients with ATRT.

Supplementary Fig. 13. UMAP markers for CyTOF gating of T cells from intracranial mouse glioblastoma allografts after radiotherapy, IL6 blockade, and immune checkpoint inhibition. a, UMAP cell density plots showing protein expression used for assigning cell types in Fig. 8c.

Supplementary Fig. 5. UMAP markers for CyTOF gating of myeloid cells from intracranial mouse glioblastoma allografts. a, UMAP cell density plots showing protein expression used for assigning cell types in Fig. 4e.

Supplementary Fig. 7. CCL2 and CXCL10 from the tumor microenvironment do not contribute to glioblastoma overall survival. a, Body weights for C57J/B6 WT mice (n=8) vs C57J/B6 Il6-/- mice (n=8) harboring intracranial SB28 glioblastoma allografts. See also Fig. 6g. b, Kaplan–Meier curves for overall survival from SB28 intracranial glioblastoma allografts in C57J/B6 WT (n=8 mice) vs C57J/B6 Ccl2-/- mice (n=8 mice) vs Cxcl10-/- mice (n=8 mice) showing no change in overall survival for mice lacking CCL2 or CXCL10 in the tumor microenvironment. Log rank test. c, Body weights for C57J/B6 WT (n=8 mice) vs C57J/B6 Ccl2-/- mice (n=8 mice) vs Cxcl10-/- mice (n=8 mice) harboring intracranial SB28 glioblastoma allografts. d, Body weights for C57J/B6 WT mice harboring SB28 intracranial glioblastoma allografts with doxycycline-induced IL6 (n=10 mice) or EV (n=10 mice) overexpression. See also Fig. 6i.

Supplementary Fig. 1. Swimmer plot. a, Clinical course and treatment history for patients included as ICI-responders and non-responders.

Supplementary Fig. 10. UMAP markers for CyTOF gating of myeloid cells from intracranial mouse glioblastoma allografts after IL6 blockade and immune checkpoint inhibition. a, UMAP cell density plots showing protein expression used for assigning cell types in Fig. 7d.

Supplementary Fig. 3. Gene circuit network changes between ICI responders and non-responders before and after ICI treatment. a, Network of gene circuits mapping differentially expressed genes between ICI responders (far left) and non-responders (middle left) post-ICI and pre-ICI specimens, and differentially expressed genes between pre-ICI (middle right) and post-ICI (far right) specimens from ICI responders and non-responders. Nodes represent pathways and edges represent shared genes between pathways (p≤0.05, FDR0.05). n = 14 match-paired glioblastoma samples from 7 patients.