SNF5 Loss Research Articles

Inhibition of MYC by the SMARCB1 tumor suppressor

SMARCB1 encodes the SNF5 subunit of the SWI/SNF chromatin remodeler. SNF5 also interacts with the oncoprotein transcription factor MYC and is proposed to stimulate MYC activity. The concept that SNF5 is a coactivator for MYC, however, is at odds with its role as a tumor-suppressor, and with observations that loss of SNF5 leads to activation of MYC target genes. Here, we reexamine the relationship between MYC and SNF5 using biochemical and genome-wide approaches. We show that SNF5 inhibits the DNA-binding ability of MYC and impedes target gene recognition by MYC in cells. We further show that MYC regulation by SNF5 is separable from its role in chromatin remodeling, and that reintroduction of SNF5 into SMARCB1-null cells mimics the primary transcriptional effects of MYC inhibition. These observations reveal that SNF5 antagonizes MYC and provide a mechanism to explain how loss of SNF5 can drive malignancy.

Abstract LB-206: Inhibition of MYC by the SMARCB1 tumor suppressor

Abstract The SMARCB1 tumor suppressor (SNF5) is a known interaction partner of the oncoprotein c-MYC (MYC), and thought to serve as a co-activator of its transcriptional properties. This role of SNF5, however, is at odds with the tumor suppressor function of SNF5, and with findings that loss of SNF5 is associated with activation of MYC target gene signatures. We have used biochemical, genetic, and genome-wide approaches to reexamine the relationship of SNF5 with MYC within the context of malignant rhabdoid tumor (MRT), an aggressive childhood cancer in which SNF5 is lost. We find that, consistent with activation of MYC target gene signatures in tumors that lack SNF5, MYC function is important for multiple aspects of MRT biology. Surprisingly, SNF5 is capable of inhibiting MYC binding to E boxes in vitro and globally within the context of chromatin. Using ATAC-seq, RNA-seq, and PRO-Seq, we demonstrate that regulation of MYC binding by SNF5 is independent of the role of SNF5 in chromatin remodeling, but instead is responsible for controlling RNA polymerase pause release at MYC-regulated genes. These findings inform a novel model of MRT tumorigenesis in which loss of SNF5 derepresses MYC function, raising the possibility that MYC contributes to disease progression in this malignancy. Citation Format: April M. Weissmiller, Jing Wang, Shelly L. Lorey, Gregory C. Howard, Ernest Martinez, Qi Liu, William P. Tansey. Inhibition of MYC by the SMARCB1 tumor suppressor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-206.

High Frequency of Ovarian Cyst Development in Vhl2B/+;Snf5+/− Mice

The new paradigm of mutations in chromatin-modifying genes as driver events in the development of cancers has proved challenging to resolve the complex influences over disease phenotypes. In particular, impaired activities of members of the SWI/SNF chromatin remodeling complex have appeared in an increasing variety of tumors. Mutations in SNF5, a member of this ubiquitously expressed complex, arise in almost all cases of malignant rhabdoid tumor in the absence of additional genetic alterations. Therefore, we studied how activation of additional oncogenic pathways might shift the phenotype of disease driven by SNF5 loss. With the use of a genetically engineered mouse model, we examined the effects of a hypomorphic Vhl2B allele on disease phenotype, with a modest up-regulation of the hypoxia response pathway. Snf5+/-;Vhl2B/+ mice did not demonstrate a substantial difference in overall survival or a change in malignant rhabdoid tumor development. However, a high percentage of female mice showed complex hemorrhagic ovarian cysts, a phenotype rarely found in either parental mouse strain. These lesions also showed mosaic expression of SNF5 by immunohistochemistry. Therefore, our studies implicate that modest changes in angiogenic regulation interact with perturbations of SWI/SNF complex activity to modulate disease phenotypes.

Loss of Snf5 Induces Formation of an Aberrant SWI/SNF Complex

The SWI/SNF chromatin remodeling complex is highly conserved from yeast to human, and aberrant SWI/SNF complexes contribute to human disease. The Snf5/SMARCB1/INI1 subunit of SWI/SNF is a tumor suppressor frequently lost in pediatric rhabdoid cancers. We examined the effects of Snf5 loss on the composition, nucleosome binding, recruitment, and remodeling activities of yeast SWI/SNF. The Snf5 subunit is shown by crosslinking-mass spectrometry (CX-MS) and subunit deletion analysis to interact with the ATPase domain of Snf2 and to form a submodule consisting of Snf5, Swp82, and Taf14. Snf5 promotes binding of the Snf2 ATPase domain to nucleosomal DNA and enhances the catalytic and nucleosome remodeling activities of SWI/SNF. Snf5 is also required for SWI/SNF recruitment by acidic transcription factors. RNA-seq analysis suggests that both the recruitment and remodeling functions of Snf5 are required invivo for SWI/SNF regulation of gene expression. Thus, loss of SNF5 alters the structure and function of SWI/SNF.

Abstract 1972: Changes in the architecture, interactions and chromatin remodeling of SWI/SNF due to loss of Snf5

Abstract The mammalian SWI/SNF complex is a family of an estimated ∼100 complexes, each with different combinations of related subunits. Efforts in sequencing many cancer genomes has revealed that mammalian SWI/SNF complexes is one of the most frequently mutated epigenetic factors found in a broad range of cancers. In order to understand more about the subunit organization of SWI/SNF and because yeast and mammalian SWI/SNF complexes share many conserved subunits and domains; we have studied the role of the Snf5 subunit in the yeast SWI/SNF to better understand the role of the mammalian homolog SNF5 that is a known tumor suppressor gene. The basis for the loss of mammalian SNF5 being a driver mutation in pediatric rhabdoid tumors is currently not well understood or how the loss of SNF5 effects the composition, recruitment and nucleosome remodeling activity of the SWI/SNF complex. We have assessed the effects on deletion of Snf5 on the compositional integrity of SWI/SNF and have evidence for Snf5 forming a sub-complex with two other highly conserved subunits of SWI/SNF that are lost from the complex when Snf5 is deleted. In addition we have mapped the inter-subunit and intra-subunit interactions of the yeast SWI/SNF complex with lysine specific homo bi-functional crosslinkers and mass spectrometry. These data confirm the formation of a tri-subunit Snf5 sub-complex and show that this module uniquely interacts with the ATPase domain of the catalytic subunit of SWI/SNF (Snf2). These observations lead to a further investigation into the regulation of the remodeling activity of SWI/SNF by the Snf5 subunit. We have found that Snf5 is required for the ATPase domain to make stable interactions with nucleosomal DNA. And the loss of these interactions in turn also causes a reduction in the remodeling efficiency of the SWI/SNF complex. We have done additional mapping of the interactions of Snf5 as part of the SWI/SNF complex and find that the highly conserved region of yeast Snf5 corresponding to the human SNF5 protein associates with the surface of the nucleosome near the histone H2A-H2B dimer. We will discuss the relevance of these findings in highlighting how an aberrant form of SWI/SNF could be created by loss of Snf5 and provide an example of how it alters the normal function and structure of SWI/SNF. Citation Format: Blaine Bartholomew, Jim Persinger, Payel Sen, Mekonnen Dechassa Lemma, Jie Luo, Jeff Ranish. Changes in the architecture, interactions and chromatin remodeling of SWI/SNF due to loss of Snf5. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1972.

SNF5/INI1 deficiency redefines chromatin remodeling complex composition during tumor development.

Malignant rhabdoid tumors (MRT), a pediatric cancer that most frequently appears in the kidney and brain, generally lack SNF5 (SMARCB1/INI1), a subunit of the SWI/SNF chromatin-remodeling complex. Recent studies have established that multiple SWI/SNF complexes exist due to the presence or absence of different complex members. Therefore, the effect of SNF5 loss upon SWI/SNF complex formation was investigated in human MRT cells. MRT cells and primary human tumors exhibited reduced levels of many complex proteins. Furthermore, reexpression of SNF5 increased SWI/SNF complex protein levels without concomitant increases in mRNA. Proteomic analysis, using mass spectrometry, of MRT cells before and after SNF5 reexpression indicated the recruitment of different components into the complex along with the expulsion of others. IP-Western blotting confirmed these results and demonstrated similar changes in other MRT cell lines. Finally, reduced expression of SNF5 in normal human fibroblasts led to altered levels of these same complex members. These data establish that SNF5 loss during MRT development alters the repertoire of available SWI/SNF complexes, generally disrupting those associated with cellular differentiation. These findings support a model where SNF5 inactivation blocks the conversion of growth-promoting SWI/SNF complexes to differentiation-inducing ones. Therefore, restoration of these complexes in tumors cells provides an attractive approach for the treatment of MRTs. SNF5 loss dramatically alters SWI/SNF complex composition and prevents formation of complexes required for cellular differentiation.

Highlights of This Issue

Despite promising preclinical evidence, therapies targeting the insulin-like growth factor (IGF) fail for many Ewing sarcoma patients, suggesting the influence of additional cellular factors. Patel and colleagues performed a genomic analysis using FAIRE-seq, which unexpectedly revealed PTEN deletion and loss of PTEN expression in a large fraction of primary human tumors. PTEN deficiency was associated with hyperactivation of AKT signaling and a more aggressive cellular phenotype. Strikingly, PTEN loss decreased sensitivity to IGF1 receptor inhibitors but increased responsiveness to temsirolimus, an mTOR inhibitor, marked by increased autophagy and an associated decrease in viability. These data support PTEN status as a clinically relevant biomarker to guide patient treatment.Pediatric malignant rhabdoid tumors (MRT) generally arise from inactivation of SNF5 (SMARCB1/INI1), a subunit of the macromolecular SWI/SNF chromatin-remodeling complex. However, the mechanism by which SNF5 loss initiates MRT development remains unclear. Wei and colleagues used mass spectrometry-based proteomics to demonstrate that SNF5 loss in MRT alters the subunit composition of the SWI/SNF complex. Importantly, SNF5 restoration rescues SWI/SNF complex integrity. Therefore, SNF5 inactivation drives the appearance of MRTs by dramatically modifying the composition of SWI/SNF complexes and preventing formation of complexes required for cellular differentiation.As a synthetic lethality-based cancer therapeutic strategy, PARP inhibition selective for BRCA1/2 deficiency has shown promising effect; therefore, defining human tumors with wild-type BRCA1/2 genes that are also sensitive to PARP inhibitors is of great clinical interest. Liu and colleagues show that TGFβ, a multitasking cytokine elevated in tumor microenvironments, regulates DNA repair by simultaneously suppressing the gene expression of ATM, MSH2, and BRCA1, resulting in a “BRCAness” phenotype that includes impaired DNA repair efficiency and reduced genomic stability, as well as a synthetic lethality to PARP inhibition. This approach improves selection of breast cancer patients based on TGFβ pathway biomarkers for PARP-targeting therapy.Estrogen and its receptors, predominantly ERα, are important mediators and therapeutic targets in hormone-dependent breast cancer. Cellular and epidemiologic evidence indicates that the G protein-coupled estrogen receptor (GPR30/GPER) has tumorigenic potential. To directly determine the oncogenic role of GPER in breast tumorigenesis, Marjon and colleagues intercrossed GPER-null mice with mammary tumor–prone MMTV-PyMT transgenic mice. Mice lacking GPER displayed reduced tumors with lower mitotic index and histologic grade. Moreover, distant metastasis was greatly diminished in GPER-deficient mice. These findings reveal that GPER contributes to tumorigenesis and, importantly, to metastasis, suggesting GPER as a new diagnostic and therapeutic target in breast cancer.

Abstract IA13: SWI/SNF complexes are frequently mutated in cancer: Epigenetic mechanisms and therapeutic targeting

Abstract My laboratory began studying the SWI/SNF complex when the SNF5 subunit was first identified as a tumor suppressor over a decade ago when it was found to be recurrently and specifically inactivated in the highly aggressive pediatric cancers AT/RT and rhabdoid tumors. Data emerging over the last two years implicate the SWI/SNF (BAF) chromatin remodeling complex as a major tumor suppressor as frequent inactivating mutations in at least eight SWI/SNF subunits have been identified in a broad variety of cancers and overall in 20% of all human cancers. These include inactivating mutations of SWI/SNF subunits in some cases of pediatric neuroblastoma, medulloblastoma and Burkitt's lymphoma as well as in adult ovarian, endometrioid, bladder, breast, stomach, colorectal, pancreatic, renal, hepatocellular, and lung cancers, and melanomas. The SWI/SNF complex includes both core and lineage-specific subunits and utilizes the energy of ATP to modulate chromatin structure and regulate transcription. We study the complex using mouse models, cell lines and primary human tumor samples. Our goals are to elucidate the normal function of the complex, identify the mechanisms by which subunit mutations drive cancer formation, and utilize this insight to identify and develop novel therapeutic approaches. Using mouse models, we have shown that biallelic inactivation of murine Snf5 leads to the rapid onset of cancer formation in 100% of mice with a median latency of only 11 weeks. Heterozygous mice develop rhabdoid tumors that are histologically indistinguishable from their human counterpart and Mx-Cre-mediated conditional inactivation results in 85% of mice developing aggressive mature T cell lymphomas and 15% developing rhabdoid tumors. Intriguingly, the rapid cancer onset arises neither due to defective DNA repair nor due to genome instability, as we have found that the genomes of both the murine and human SNF5-deficient cancers are diploid and indistinguishable from normal cells via high-density SNP arrays. Most recently, by sequencing the exomes of 35 human pediatric rhabdoid tumors we have shown that these cancer genomes contain an extremely low rate of mutations, with loss of SNF5 being essentially the sole recurrent event. Indeed, in two of the cancers there were no other identified mutations. Our results suggest that high mutation rates are dispensable for the genesis of cancers driven by mutation of this chromatin remodeling complex and further that epigenetic dysfunction caused by mutation of the SWI/SNF complex may underlie the broad spectrum of cancers caused by mutation of this complex. Insights into the normal function of SWI/SNF complexes, the mechanisms by which mutation of the complexes drive cancer formation, and opportunities for therapeutic intervention will be presented. Citation Format: Charles W. M. Roberts. SWI/SNF complexes are frequently mutated in cancer: Epigenetic mechanisms and therapeutic targeting. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr IA13.

Abstract 2245: Mechanism of replicative senescence induced by SNF5 loss

Abstract Cellular senescence was initially described as the phenomenon by which normal diploid cells cease to divide after reaching the Hayflick limit. However, with the identification of replicative senescence, other inducers of this process have appeared in the literature including oncogene induced senescence (OIS). Of interest, different oncogenes can initiate OIS through alternative pathways e.g. HRAS (G12V) and BRAF (V600E) that effect senescence through p16 dependent and p16 independent pathways, respectively. Alternatively, loss of tumor suppressor gene function can also induce senescence. In malignant rhabdoid tumors (MRTs), SNF5 behaves as a tumor suppressor and its re-expression in MRT cell lines results in growth arrest via the RB pathway. Paradoxically, when we knocked out SNF5 in normal human fibroblasts (NHF-1), we also observed senescence. We therefore hypothesize that perturbation of the SWI/SNF complex (e.g. knocking down SNF5) results in cellular senescence in most normal tissues. This suggests an epigenetic mechanism of senescence occurring through a p16 independent pathway, as previous publications have demonstrated a critical role for SNF5 in p16 expression. Additionally, in BRAF induced senescence, changes in levels of CD44, phospho-MEK1/2, and p16 are observed. However, we do not find these changes during SNF5 loss. Our findings suggest a relationship between SWI/SNF's chromatin remodeling activity and the regulation of cellular senescence. Importantly, our studies will identify the key signaling pathways that allow MRTs to escape cellular senescence after SNF5 loss. These results will further elucidate the role of SNF5 loss in MRT development and provide new targets for clinical intervention. Acknowledgement: We thank the members of the Weissman lab for their help, advice, and support and R01CA91048 (B.E.W.) , T32ES007126 (D.W.), UNC Dissertation Completion Fellowship (D.W.) for financial support. Citation Format: Darmood Wei, HoYoon Chung, Dennis A. Simpson, William K. Kaufmann, Bernard E. Weissman. Mechanism of replicative senescence induced by SNF5 loss. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2245. doi:10.1158/1538-7445.AM2014-2245

SNF5/INI1 Deficiency Destabilizes the SWI/SNF Complex During Malignant Rhabdoid Tumor Development

Malignant rhabdoid tumors (MRTs), a pediatric cancer that frequently arises in the kidney, lack SNF5, a subunit of the SWI/SNF chromatin-remodeling complex. Recent studies have also shown that BAF180/PBRM1, another SWI/SNF subunit, was mutated in adult renal carcinoma. So now BAF180 is recognized as tumor suppressor gene. Intriguingly, both SNF5 and BAF180 regulate the cell cycle through control of expression of p21. Therefore, we hypothesized that SNF5 loss may affect BAF180 function(s) during MRT development.

Abstract IA12: SWI/SNF (BAF) complexes in tumor suppression

Abstract Data emerging over the last two years implicate the SWI/SNF (BAF) chromatin remodeling complex as a major tumor suppressor as frequent inactivating mutations in at least seven SWI/SNF subunits have been identified in a variety of cancers. These include inactivating mutations of the gene encoding the ARID1A (BAF250a) subunit in ovarian, endometrioid, bladder, stomach, colorectal and pancreatic cancers; of the PBRM1 (BAF180) subunit in renal carcinomas; of the ARID2 subunit in hepatocellular, lung, and pancreas carcinomas as well as melanomas; of the BRD7 subunit in breast cancers; and of the BRG1 (SMARCA4) subunit in non-small cell lung cancers and medulloblastomas. The SWI/SNF complex includes both core and lineage-specific subunits and utilizes the energy of ATP to modulate chromatin structure and regulate transcription. My laboratory began studying the SWI/SNF complex when the SNF5 subunit was first identified as a tumor suppressor over a decade ago when it was found to be recurrently and specifically inactivated in a highly aggressive type of pediatric cancer called malignant rhabdoid tumor. We now study the complex using mouse models, cell lines and primary human tumor samples. Our goals are to elucidate the normal function of the complex, identify the mechanisms by which subunit mutations drive cancer formation, and utilize this insight to identify and develop novel therapeutic approaches. Using mouse models, we have shown that biallelic inactivation of murine Snf5 leads to the rapid onset of cancer formation in 100% of mice with a median latency of only 11 weeks. Heterozygous mice develop rhabdoid tumors that are histologically indistinguishable from their human counterpart and Mx-Cre-mediated conditional inactivation results in 85% of mice developing aggressive mature T cell lymphomas and 15% developing rhabdoid tumors. Intriguingly, the rapid cancer onset arises neither due to defective DNA repair nor due to genome instability, as we have found that the genomes of both the murine and human SNF5-deficient cancers are diploid and indistinguishable from normal cells via high-density SNP arrays. Most recently, by sequencing the exomes of 35 human pediatric rhabdoid tumors we have shown that these cancer genomes contain an extremely low rate of mutations, with loss of SNF5 being essentially the sole recurrent event. Indeed, in two of the cancers there were no other identified mutations. Our results suggest that high mutation rates are dispensable for the genesis of cancers driven by mutation of this chromatin remodeling complex and further that epigenetic dysfunction caused by mutation of the SWI/SNF complex may underlie the broad spectrum of cancers caused by mutation of this complex. Insights into the normal function of SWI/SNF complexes, the mechanisms by which mutation of the complexes drive cancer formation, and opportunities for therapeutic intervention will be presented. Citation Format: Charles W. M. Roberts. SWI/SNF (BAF) complexes in tumor suppression. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr IA12.

Abstract SY07-01: The SWI/SNF chromatin remodeling complex is frequently mutated in cancer: Mechanisms and potential therapeutic insights.

Abstract Data emerging over the last two years implicate the SWI/SNF (BAF) chromatin remodeling complex as a major tumor suppressor as frequent inactivating mutations in at least six SWI/SNF subunits have been identified in a variety of cancers. These include inactivating mutations of the gene encoding the ARID1A (BAF250a) subunit in ovarian, endometrioid, bladder, stomach, colorectal and pancreatic cancers; of the PBRM1 (BAF180) subunit in renal carcinomas; of the ARID2 subunit in hepatocellular, lung, and pancreas carcinomas as well as melanomas; of the BRD7 subunit in breast cancers; and of the BRG1 (SMARCA4) subunit in non-small cell lung cancers and medulloblastomas. The SWI/SNF complex includes both core and lineage-specific subunits and utilizes the energy of ATP to modulate chromatin structure and regulate transcription. My laboratory began studying the SWI/SNF complex when the SNF5 subunit was first identified as a tumor suppressor over a decade ago when it was found to be recurrently and specifically inactivated in a highly aggressive type of pediatric cancer called malignant rhabdoid tumor. We now study the complex using mouse models, cell lines and primary human tumor samples. Our goals are to elucidate the normal function of the complex, identify the mechanisms by which subunit mutations drive cancer formation, and utilize this insight to identify and develop novel therapeutic approaches. Using mouse models, we have shown that biallelic inactivation of murine Snf5 leads to the rapid onset of cancer formation in 100% of mice with a median latency of only 11 weeks. Heterozygous mice develop rhabdoid tumors that are histologically indistinguishable from their human counterpart and Mx-Cre-mediated conditional inactivation results in 85% of mice developing aggressive mature T cell lymphomas and 15% developing rhabdoid tumors. Intriguingly, the rapid cancer onset arises neither due to defective DNA repair nor due to genome instability, as we have found that the genomes of both the murine and human SNF5-deficient cancers are diploid and indistinguishable from normal cells via high-density SNP arrays. Most recently, by sequencing the exomes of 35 human pediatric rhabdoid tumors we have shown that these cancer genomes contain an extremely low rate of mutations, with loss of SNF5 being essentially the sole recurrent event. Indeed, in two of the cancers there were no other identified mutations. Our results suggest that high mutation rates are dispensable for the genesis of cancers driven by mutation of this chromatin remodeling complex and further that epigenetic dysfunction caused by mutation of the SWI/SNF complex may underlie the broad spectrum of cancers caused by mutation of this complex. Insights into the normal function of SWI/SNF complexes, the mechanisms by which mutation of the complexes drive cancer formation, and opportunities for therapeutic intervention will be presented. Citation Format: Charles W. M. Roberts. The SWI/SNF chromatin remodeling complex is frequently mutated in cancer: Mechanisms and potential therapeutic insights. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr SY07-01. doi:10.1158/1538-7445.AM2013-SY07-01

Activation of β-catenin/TCF targets following loss of the tumor suppressor SNF5

The SWI/SNF chromatin remodeling complex is a master regulator of developmental cell-fate decisions, although the key target pathways are poorly characterized. Here, we interrogated the contribution of the SWI/SNF subunit and tumor suppressor SNF5 to the regulation of developmental pathways using conditional mouse and cell culture models. We find that loss of SNF5 phenocopies β-catenin hyperactivation and that SNF5 is essential for regulating Wnt/β-catenin pathway target expression. These data provide insight into chromatin-based mechanisms that underlie developmental regulation and elucidate the emerging theme that mutation of this tumor suppressor complex can activate developmental pathways by uncoupling them from upstream control.

Epigenetic inactivation of the tumor suppressor BIN1 drives proliferation of SNF5-deficient tumors

Emerging evidence demonstrates that subunits of the SWI/SNF chromatin remodeling complex are specifically mutated at high frequency in a variety of human cancer types. SNF5 (SMARCB1/INI1/BAF47), a core subunit of the SWI/SNF complex, is inactivated in the vast majority of rhabdoid tumors (RT), an aggressive type of pediatric cancer. SNF5-deficient cancers are diploid and genomically stable, suggesting that epigenetically based changes in transcription are key drivers of tumor formation caused by SNF5 loss. However, there is limited understanding of the target genes that drive cancer formation following SNF5 loss. Here we performed comparative expression analyses upon three independent SNF5-deficient cancer data sets from both human and mouse and identify downregulation of the BIN1 tumor suppressor as a conserved event in primary SNF5-deficient cancers. We show that SNF5 recruits the SWI/SNF complex to the BIN1 promoter, and that the marked reduction of BIN1 expression in RT correlates with decreased SWI/SNF occupancy. Functionally, we demonstrate that re-expression of BIN1 specifically compromises the proliferation of SNF5-deficient RT cell lines. Identification of BIN1 as a SNF5 target gene reveals a novel tumor suppressive regulatory mechanism whose disruption can drive cancer formation.

TCR-dependent transformation of mature memory phenotype T cells in mice.

A fundamental goal in cancer research is the identification of the cell types and signaling pathways capable of initiating and sustaining tumor growth, as this has the potential to reveal therapeutic targets. Stem and progenitor cells have been implicated in the genesis of select lymphoid malignancies. However, the identity of the cells in which mature lymphoid neoplasms are initiated remains unclear. Here, we investigate the origin of peripheral T cell lymphomas using mice in which Snf5, a chromatin remodelling-complex subunit with tumor suppressor activity, could be conditionally inactivated in developing T cells. In this model of mature peripheral T cell lymphomas, the cell of origin was a mature CD44hiCD122loCD8⁺ T cell that resembled a subset of memory cells that has capacity for self-renewal and robust expansion, features shared with stem cells. Further analysis showed that Snf5 loss led to activation of a Myc-driven signaling network and stem cell transcriptional program. Finally, lymphomagenesis and lymphoma proliferation depended upon TCR signaling, establishing what we believe to be a new paradigm for lymphoid malignancy growth. These findings suggest that the self-renewal and robust proliferative capacities of memory T cells are associated with vulnerability to oncogenic transformation. Our findings further suggest that agents that impinge upon TCR signaling may represent an effective therapeutic modality for this class of lethal human cancers.

Abstract LB-237: Epigenetic antagonism between Polycomb and SWI/SNF complexes duringoncogenic transformation

Abstract While epigenetic alterations are widely present in cancers, in the context of genomic instability it has been difficult to determine the degree to which epigenetic alterations serve as primary drivers of tumorigenesis. Cancers initiated by mutation of the SNF5 chromatin remodeling subunit, despite being highly aggressive and lethal, are diploid and genomically stable, making them an ideal model for this purpose. SNF5 is a core subunit of the SWI/SNF chromatin complex and is mutated in the large majority of malignant rhabdoid tumors (MRT), highly lethal cancers that occur in kidney, brain, and soft tissues of young children. Inactivating mutations in SNF5 have also recently been found to occur in a variety of other cancers and mutations in other SWI/SNF subunits occur frequently in ovarian, endometrial, renal, and lung cancers. Given our findings of genomic stability, we hypothesize that the consequences of SNF5 loss are fundamentally epigenetic in nature; that characterization of the epigenetic changes present in our model systems, rather than in cancers with genome instability, will identify those epigenetic alterations capable of driving oncogenic transformation; and lastly that cancers carrying mutations in the SWI/SNF complex can be effectively treated with epigenetically targeted therapies. Analysis of Drosophila mutants suggests that Polycomb and SWI/SNF complexes can serve antagonistic developmental roles. However, the relevance of this relationship to human disease is unclear. Here we have investigated functional relationships between these epigenetic regulators in oncogenic transformation. Mechanistically, we show that loss of the SNF5 tumor suppressor leads to elevated expression of the Polycomb gene EZH2 and that Polycomb targets are broadly H3K27-trimethylated and repressed in SNF5-deficient fibroblasts and cancers. Further, we show antagonism between SNF5 and EZH2 in the regulation of stem cell-associated programs and that Snf5 loss activates those programs. Finally, using conditional mouse models, we show that inactivation of Ezh2 blocks tumor formation driven by Snf5 loss. Collectively, we demonstrate that the rapid onset of tumor formation following SNF5 loss arises due to imbalanced epigenetic antagonism between the SWI/SNF complex and the Polycomb complex PRC2. Our work reveals essential roles for epigenetic modifications during tumor formation and demonstrates that inactivation of EZH2 can have therapeutic efficacy against cancer in vivo. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-237. doi:10.1158/1538-7445.AM2011-LB-237

Abstract 66: Epigenetic inactivation of the tumor suppressor BIN1 drives proliferation of SNF5-deficient tumors

Abstract SNF5 (SMARCB1/INI1/BAF47), a core subunit of SWI/SNF chromatin remodeling complexes, is a potent tumor suppressor that is specifically inactivated in a variety of aggressive pediatric cancers. We have recently demonstrated that SNF5 acts as a tumor suppressor not genetically via maintenance of genome integrity, but epigenetically via transcriptional regulation. However, few of the target genes essential for oncogenesis following SNF5 loss have been identified. Here, we demonstrate that aberrant epigenetic silencing of the tumor suppressor gene BIN1 is a conserved event in SNF5-deficient cancers and contributes to tumorigenesis. Comparative genomewide expression analysis reveals that BIN1 expression is strongly downregulated in SNF5-deficient malignant rhabdoid tumors (MRT) relative to other human tumors, and we also observe decreased Bin1 expression in murine Snf5-deficient lymphomas. The underlying mechanism is epigenetic silencing as reintroduction of SNF5 into deficient tumor cell lines results in increased SWI/SNF recruitment to the BIN1 promoter and increased BIN1 expression. Furthermore, expression of BIN1 specifically compromises the proliferation of SNF5-deficient MRT cell lines. BIN1 encodes a family of alternatively spliced adaptor proteins that have been implicated in cellular differentiation, tumor suppression, and immune escape of transformed cells, but its regulation has not been well characterized. Identification of BIN1 as a key SNF5 target reveals a novel mechanism by which epigenetically based changes can drive cancer formation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 66. doi:10.1158/1538-7445.AM2011-66

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Loss of the tumor suppressor Snf5 leads to aberrant activation of the Hedgehog-Gli pathway

Aberrant activation of the Hedgehog (Hh) pathway can drive tumorigenesis. To investigate the mechanism by which glioma-associated oncogene family zinc finger-1 (GLI1), a crucial effector of Hh signaling, regulates Hh pathway activation, we searched for GLI1-interacting proteins. We report that the chromatin remodeling protein SNF5 (encoded by SMARCB1, hereafter called SNF5), which is inactivated in human malignant rhabdoid tumors (MRTs), interacts with GLI1. We show that Snf5 localizes to Gli1-regulated promoters and that loss of Snf5 leads to activation of the Hh-Gli pathway. Conversely, re-expression of SNF5 in MRT cells represses GLI1. Consistent with this, we show the presence of a Hh-Gli-activated gene expression profile in primary MRTs and show that GLI1 drives the growth of SNF5-deficient MRT cells in vitro and in vivo. Therefore, our studies reveal that SNF5 is a key mediator of Hh signaling and that aberrant activation of GLI1 is a previously undescribed targetable mechanism contributing to the growth of MRT cells.

Epigenetic Antagonism between Polycomb and SWI/SNF Complexes during Oncogenic Transformation

Epigenetic alterations have been increasingly implicated in oncogenesis. Analysis of Drosophila mutants suggests that Polycomb and SWI/SNF complexes can serve antagonistic developmental roles. However, the relevance of this relationship to human disease is unclear. Here, we have investigated functional relationships between these epigenetic regulators in oncogenic transformation. Mechanistically, we show that loss of the SNF5 tumor suppressor leads to elevated expression of the Polycomb gene EZH2 and that Polycomb targets are broadly H3K27-trimethylated and repressed in SNF5-deficient fibroblasts and cancers. Further, we show antagonism between SNF5 and EZH2 in the regulation of stem cell-associated programs and that Snf5 loss activates those programs. Finally, using conditional mouse models, we show that inactivation of Ezh2 blocks tumor formation driven by Snf5 loss.