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PRC2 loss amplifies Ras-driven transcription and confers sensitivity to BRD4-based therapies


The polycomb repressive complex 2 (PRC2) exerts oncogenic effects in many tumour types1. However, loss-of-function mutations in PRC2 components occur in a subset of haematopoietic malignancies, suggesting that this complex plays a dichotomous and poorly understood role in cancer2,3. Here we provide genomic, cellular, and mouse modelling data demonstrating that the polycomb group gene SUZ12 functions as tumour suppressor in PNS tumours, high-grade gliomas and melanomas by cooperating with mutations in NF1. NF1 encodes a Ras GTPase-activating protein (RasGAP) and its loss drives cancer by activating Ras4. We show that SUZ12 loss potentiates the effects of NF1 mutations by amplifying Ras-driven transcription through effects on chromatin. Importantly, however, SUZ12 inactivation also triggers an epigenetic switch that sensitizes these cancers to bromodomain inhibitors. Collectively, these studies not only reveal an unexpected connection between the PRC2 complex, NF1 and Ras, but also identify a promising epigenetic-based therapeutic strategy that may be exploited for a variety of cancers.

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Figure 1: PRC2 components are deleted or mutated in human MPNSTs and exert tumour-suppressive activity.
Figure 2: Suz12 and Nf1 mutations cooperate to promote widespread tumour development in mice.
Figure 3: SUZ12 and JQ1 regulate PRC2 targets and the Ras transcriptional signature.
Figure 4: JQ1 and MEK inhibitors cooperate to promote cell death, suppress Ras transcriptional output and promote tumour regression.

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Primary accessions

Gene Expression Omnibus

Data deposits

All microarray data have been deposited in the Gene Expression Omnibus database under accession number GSE52777.


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This work was supported by the following organizations: The US Department of Defense (W81XWH-11-1-0138), the Ludwig Center at DF/HCC and the Children’s Tumor Foundation (K.C.); T.D. was a recipient of the Young Investigator Award of the Children’s Tumor Foundation; FWO-Flanders G.0784.10N (E.L.). E.B. was a recipient of an Emmanuel Vanderschueren Fellowship from the Vlaamse Liga tegen Kanker. Association Neurofibromatoses et Recklinghausen, Ligue Française Contre les Neurofibromatoses, Association pour la Recherche sur le Cancer, Comité de Paris de la Ligue Contre le Cancer, the French Clinical Research program (PHRC 2002, P. Wolkenstein) and INSERM (Nf1GeneModif project) (M.V. and E.P.). We thank the Platform of Biological Resources, Assistance Publique Hôpitaux de Paris, Hôpital Henri Mondor, Créteil, France, for providing tissue samples.

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Authors and Affiliations



T.D. and K.C. conceived and designed the functional studies and mouse experiments. E.L. E.B., M.V., E.P. and A.L. conceived, designed and performed the genomic studies. T.D. performed the cellular, mouse, microarray and ChiP-seq experiements. H.B. coordinated human sample acquisition and analysis. E.L., M.U., V.M., H.K. and M.V. provided human MPNST samples. N.O. performed immunohistological staining. K.H. provided the SUZ12 mice and advice. J.L.H. performed pathological analysis. W.C. assisted in evaluating mouse neurofibromas. J.B. provided compounds and advice. T.D. and K.C. wrote the manuscript. All authors discussed the results and commented on the manuscript.

Corresponding authors

Correspondence to Eric Legius or Karen Cichowski.

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Competing interests

The authors declare no competing financial interests.

Extended data figures and tables

Extended Data Figure 1 Mutational data in human MPNSTs and further biological analysis of SUZ12 loss.

a, Schematic overview of the deletions in the NF1, SUZ12 and EED regions observed in human MPNSTs (green, germline deletion; red, somatic deletion; yellow, duplication). b, List of the amino-acid changes or deletions found in SUZ12 and EED in human MPNSTs. c, Schematic representation of the location of SUZ12 and EED mutations (red, truncating mutation; green, missense mutation with amino-acid change noted). d, Immunoblots of lysates from primary human MPNSTs. Tumours with homozygous inactivating mutations in one of the PRC2 components (SUZ12 or EED) show complete loss of H3K27Me3. *Homozygous inactivation of EED. e, Immunoblots comparing NF1, SUZ12 and H3K27me3 expression of four human MPNST cell lines. *Cell line derived from an MPNST of a patient with an NF1 microdeletion. The human GBM cell lines A172 and U251 were used as a control. Corresponding NF1 mutations are reported in Extended Data Table 1 (S462, L2; 90-8TL, L3) and elsewhere in these NF1-deficient lines. p53 mutations, when known, are denoted and reported elsewhere20. f, Proliferation curves used to derive bar graphs shown in Fig. 1d (red, LacZ control; green, SUZ12 reconstituted). g, Relative proliferation of several SUZ12 WT cell lines: colon (RKO, colo741, HCT-116) or GBM (T98G), after introduction of a control or SUZ12 lentivirus. None of these cell lines exhibited a significant decrease in proliferation under normal growth conditions or cell death in limiting growth factors, in contrast to SUZ12-deficient cells shown in Fig. 1d. h, Effects of shSUZ12 (S1) on colony formation (top) and SUZ12 expression (bottom) in A172 GBM cells, which are NF1 WT (see Extended Data Fig. 1e). i, Effects of shSUZ12 (S1) on SUZ12 expression and colony formation in WM3526 melanoma cells, which are NF1 WT. Error bars, s.d. (n = 3, biological replicates).

Extended Data Figure 2 Suz12 and Nf1 mutations cooperate to promote widespread tumour development in mice.

a, Semi-quantitative PCR showing loss of the WT Suz12 and Nf1 allele in Nf1/Suz12 mouse tumours. b, Table listing the tumours observed in Nf1+/−; Suz12+/− and Suz12+/− mice. Tumour types denoted with a red asterisk occur in patients with NF1 and an increased frequency in patients with NF1 microdeletions. Tumour types denoted with a blue asterisk represent spontaneous tumour types/lesions that have been shown to harbour NF1 mutations humans. c, Semi-quantitative PCR showing loss of the WT Suz12, Nf1 and p53 allele in Nf1/p53/Suz12 mouse MPNST. d, Haematoxylin and eosin staining of a GBM from an Nf1/p53/Suz12 mouse.

Extended Data Figure 3 Microarray data analysis (GSEA) and pERK levels in response to SUZ12 modulation and JQ1 treatment.

a, Table showing that PRC2 (EZH2) and Ras signatures were suppressed in MPNST cells reconstituted with SUZ12 or treated with JQ1. Also shown are normalized enrichment score, P value and false discovery rate. b, EZH2 signature enrichment plots (using the KONDO_EZH2 data set obtained from the Molecular Signatures Database) in MPNSTs reconstituted with SUZ12, versus vector control, or treated with JQ1, versus vehicle control (100 nM, 24 h, triplicate samples). Plots show a significant downregulation of the EZH2 signature after SUZ12 reconstitution and JQ1 treatment. c, Microarray expression analysis in human MPNST cells shows that expression of MYC and MYCN is unaltered after SUZ12 reconstitution or treatment with JQ1. Error bars, s.d. (n = 3, biological replicates). d, Phospho-ERK levels did not appreciably change in response to SUZ12 loss (U251), reconstitution (90-8TL) or JQ1 treatment (90-8TL).

Extended Data Figure 4 Consequences of SUZ12 loss on drug sensitivity and H3K27me3 levels.

a, Graph depicting log2 fold change in MPNST cell number (90-8TL) after 3 days in response to drugs and siRNA sequences indicated. PD-0325901 also cooperates with genetic BRD4 ablation to kill cells. b, Relative expression of BRD4 as determined by real-time PCR after exposure to BRD4-specific siRNAs (90-8TL). c, SUZ12 ablation using shSUZ12 S1 conferred increased sensitivity to combined PD-901/JQ1 in NF1-deficient but not NF1 WT colon cancer and GBM cell lines. Log2 fold change in cell number was determined. RKO, DLD1 and Caco cell lines were treated with 1 μM JQ1 and 1 μM PD901 each time alone or in combination. The Lovo cell line was treated with 500 nM JQ1 and 250 M PD901; SW480 was treated with 250 nM PD901 and 1 μM JQ1; LN229 and A172 cell lines were treated with 500 nM JQ1 and 1 μM PD901. d, Immunohistochemistry comparing H3K27me3 expression in an additional human MPNST that was WT for SUZ12 and other PRC2 genes, and one that was null for SUZ12. Arrow depicts positive staining of blood vessel. Staining of additional tumours is shown in Extended Data Table 1. Error bars, s.d. (n = 3, biological replicates).

Extended Data Table 1 Combined deletion and mutational data for tumours listed in Fig. 1

Supplementary information

Supplementary Discussion

This file contains an extended discussion about the mutation analysis, mouse models, microarray analysis and PRC2 loss and the Ras transcriptional signatures. (PDF 151 kb)

Supplementary Table 1

This file contains SUZ12 signature. Genes significantly downregulated (2 fold) after reconstitution of SUZ12 in the 90-8TL cell line. (XLSX 120 kb)

Supplementary Table 2

This file contains Ras signature gene set. List of upregulated Ras signature genes (starting from Bild_HRAS_Oncogenic_signature, KRAS.300_UP.V1_UP, KRAS.600_UP.V1_UP, KRAS.600.LUNG.BREAST_UP.V1_UP and KRAS.BREAST_UP.V1_UP) that were significantly repressed by JQ1, PD-901 or combined JQ1/PD901 in MPNSTs (Class comparison of all treatments at p-value <0.001). (XLSX 26 kb)

Supplementary Table 3

This file shows genes significantly changing under different drug treatments. List of genes that significantly change (p<0.001) after 24h of JQ1, PD901 or PD901/JQ1 treatment (as compared to DMSO). For each gene the parametric p-value, False discovery rate and fold change compared to DMSO is indicated. (XLSX 508 kb)

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De Raedt, T., Beert, E., Pasmant, E. et al. PRC2 loss amplifies Ras-driven transcription and confers sensitivity to BRD4-based therapies. Nature 514, 247–251 (2014).

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