The CoREST complex is a therapeutic vulnerability in malignant peripheral nerve sheath tumors

Abstract

Malignant peripheral nerve sheath tumor (MPNST) is a highly aggressive sarcoma that may be seen in patients with neurofibromatosis type 1 (NF1) or occur sporadically. While surgery is the primary treatment for localized MPNST with a 61.9% overall survival rate, metastatic disease is often fatal due to resistance to systemic therapies which underscores the urgent need for effective treatments. MPNSTs frequently harbor inactivating driver mutations in the PRC2 epigenetic repressor complex suggesting epigenetic therapies may represent a specific vulnerability in these tumors. Here, we investigate the role of the LSD1-HDAC1-CoREST (LHC) repressor complex in mediating MPNST tumor growth and progression. Our findings demonstrate that the LHC small molecule inhibitor, corin, induces apoptosis and significantly inhibits proliferation in MPNST cells. Transcriptomic analysis of corin-treated MPNST cells demonstrates specific increases in genes associated with axonogenesis and neuronal differentiation as well as altered extracellular matrix; additionally, corin treatment is shown to inhibit MPNST invasion in vitro. These results underscore the critical role of the LHC complex in facilitating MPNST growth and progression and suggest that targeting the LHC complex represents a promising therapeutic approach for this aggressive malignancy.

Keywords: CoREST; Corin; Epigenetics; HDAC; LSD1; MPNST.

Fig. 1

The LHC inhibitor, corin, blocks tumor cell growth and promotes apoptosis in MPNST. (A) Dose–response curves for MPNST cells treated with DMSO, corin, HDACi (entinostat), or LSD inhibitor (GSK-LSD1). Treatment was for 72 h except for the four JHU cell lines which were treated for 120 h. Y axis represents proliferation rate normalized to DMSO. N = 4 (B) Flow cytometry analysis of MPNST cells treated with DMSO or 1 μM Corin for 72 h and stained with Annexin V. N = 3 (C) Quantification of apoptosis data in (B) for early apoptotic, apoptotic/necrotic and late apoptotic/necrotic cells treated with DMSO vs. corin. Cells in red font are PRC2 mutants and cells in green font are PRC2 wildtype. (D) Phase-contrast images of cells treated with DMSO or 1 μM Corin or HDACi for 72 h taken at 10X magnification. Paired t test was performed for apoptosis analysis of DMSO vs. corin treatment. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 2

Corin treatment of MPNST cells alters expression of axonogenesis and invasion genes independent of NF1 and PRC2 mutation status. (A) Heatmap and hierarchical clustering of genes differentially regulated by Corin in four MPNST cell lines. N = 2 (B) PCA plot for MPNST cells depicted in (A) following corin treatment. (C) Volcano plot representing differentially expressed genes (DEGs) in all four MPNST cell lines treated with corin. Red circles indicate genes upregulated by corin, while blue circles indicate gene downregulated by corin. Genes of interests are labeled. (D) Venn diagram illustrating the overlap of differentially upregulated genes among the four MPNST cell lines treated with corin. (E) Gene ontology analysis of the 1435 genes upregulated by corin in all four cell lines. BP: Biological process, and CC: Cellular component. (F) Heatmap of the five genes of interests labeled based on the GO analysis. (G) Kaplan Meier survival curves of the five genes. Kaplan Meier survival curves of the five genes (n = 59; “high” and “low” expression defined by median expression across patients). (H) Homer Motif analysis of the 1435 genes upregulated by Corin in all four cell lines.

Fig. 3

Inhibition of the LHC complex promotes histone acetylation without a corresponding increase in chromatin accessibility. (A) Western blot of histone marks in PRC2 + (wildtype) and PRC2- (mutant) MPNST cells treated with DMSO vs corin. N = 2 (B) Bar graph quantifying the western blot in (A). (C) Plot profile and heatmap of chromatin accessibility in S462 MPNST cells treat with DMSO, corin or HDACi (entinostat). X axis represents the center of peak spanning -3 k to 3 k bp. Y axis represents the accessibility signal. (D) Genome annotation illustrating alteration of accessibility peaks at different genome annotations in S462 cells following treatment with DMSO, corin, or HDACi (entinostat). (E) Individual chromatin accessibility gene tracks for MAPT, PDGFB, NGFR, SPP1 and GDF15 in S462 cells following treatment with DMSO, corin or HDACi (entinostat) N = 2.

Fig. 4

Inhibition of the LHC complex leads to decreased tumor cell invasion in MPNST. (A) qPCR analysis of expression of genes of interest in S462, T265, MPNST724 and STS26T MPNST cells following treatment with DMSO or 1 μM corin for 24 h. N = 3 (B) Boyden chamber invasion assay of S462, T265, MPNST724 and STS26T MPNST cells treated with DMSO or 1 μM corin. N = 3 (C) Quantification of Boyden chamber invasion assay in S462, T265, MPNST724 and STS26T MPNST cells treated with DMSO or 1 μM corin. (D) Working model depicting the repressive function of the CoREST complex on the expression of differentiation and anti-invasion genes in MPNST cells by blocking the recruitment of differentiation and anti-invasion-associated transcription factors, leading to a a stem-like cell phenotype that is invasive. In MPNST, particularly in PRC2-null tumors, chromatin is in a highly open state. Treatment with corin inhibits the binding of the CoREST complex to chromatin leading to recruitment of differentiation and anti-invasion-associated transcription factors and the de-repression of differentiation and anti-invasion genes, resulting in a more differentiated and less invasive cell phenotype.