Systematic multi-omics cell line profiling uncovers principles of Ewing sarcoma fusion oncogene-mediated gene regulation

Orth, Martin F.; Surdez, Didier; Faehling, Tobias; Ehlers, Anna C; Marchetto, Aruna; Grossetête, Sandrine; Volckmann, Richard; Zwijnenburg, Danny; Gerke, Julia S.; Zaïdi, Sakina; Alonso, Javier; Sastre, Ana; Baulande, Sylvain; Sill, Martin; Cidre‐Aranaz, Florencia; Ohmura, Shunya; Kirchner, Thomas; Hauck, Stefanie M.; Reischl, Eva; Gymrek, Melissa; Pfister, Stefan M.; Strauch, Konstantin; Koster, Jan; Delattre, Olivier; Grünewald, Thomas G.P.

doi:10.1016/j.celrep.2022.111761

Cited by 31 publications

(32 citation statements)

References 153 publications

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“…Strikingly, even when only considering patients with localized disease (i.e., without evidence for metastasis at diagnosis), this association remained significant (P=0.0309, Figure 1b), indicating that chr8 gain is functionally involved in mediating an unfavorable disease phenotype. In support of this hypothesis, it is intriguing that while chr8 gain is found only in approximately 50% of primary tumors, around 80% of EwS cell lines, which are expected to be derived from highly aggressive tumor clones, exhibit chr8 gains (mostly trisomies) [16][17][18][19]21,[24][25][26][27][28] . Together, these findings suggest that genes located on chr8 contribute to aggressive cellular behavior and disease progression in EwS.…”

Section: Chromosome 8 Gain Drives Overexpression Of the Clinically Re...mentioning

confidence: 91%

“…Strikingly, this association remained significant (P=0.0309, Figure 1b) even when only considering patients with localized disease (i.e., without evidence for metastasis at diagnosis), indicating that chr8 gain is functionally involved in mediating an unfavorable disease phenotype. In support of this hypothesis, it is intriguing that while chr8 gain is only found in approximately 50% of primary tumors, around 80% of EwS cell lines, which are expected to be derived from highly aggressive tumor clones, exhibit chr8 gains (mostly trisomies) [16][17][18][19]21,[24][25][26][27][28] . Since previous studies have reported that chr8 gains can co-occur with other recurrent chromosomal gains and losses that may have an effect on patient overall survival [16][17][18]27 , such as chr1q gains, chr12 gains, and 16q loss, we reassessed our Cohort 1 now only focusing on those patients that show a predicted exclusive chr8 gain or none of the abovementioned CNVs.…”

Section: Chromosome 8 Gain Drives Overexpression Of the Clinically Re...mentioning

confidence: 91%

“…Chr8 gain is present in approximately 50% of EwS cases, often in form of chr8 trisomy, making it the second most frequently observed recurrent somatic alteration in EwS following FET::ETS fusions [16][17][18][19][20][21][22] . Previous studies focused solely on specific correlations regarding the role of (partial) chr8 gains in EwS [16][17][18][19]21,[23][24][25][26][27][28] , and suggested that chr8 gains may be an early event in EwS tumorigenesis 29 . However, the precise functional and clinical impact of whole chr8 gains in EwS remains unclear.…”

Section: Introductionmentioning

confidence: 99%

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Chromosome 8 gain drives cancer progression by hijacking the translation factor 4E-BP1 sensitizing for targeted CDK4/6 inhibition

Funk

Orth²,

Aljakouch

et al. 2022

Preprint

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To refine patient care in the context of precision oncology, it is increasingly important to understand mechanisms underlying inter-individual tumor heterogeneity, especially in oligomutated cancers. Ewing sarcoma (EwS) is genetically characterized by pathognomonic FET::ETS gene fusions (in most cases EWSR1::FLI1) while featuring a general paucity of other recurrent somatic alterations that might account for observed diversity in clinical patient outcome. Following FET::ETS fusions, chromosome (chr) 8 gain is the second most common recurrent somatic alteration in EwS. However, its pathophysiological role and potential clinical implications remain unclear. Here, we report that gene expression signatures indicative for chr8 gain are significantly associated with poor overall survival of EwS patients, and that this effect is predominantly mediated by expression of the translation initiation factor binding protein EIF4EBP1 (4E-BP1). High EIF4EBP1 expression shows the strongest association with poor overall survival of EwS patients among all genes located on chr8, which as well holds true in a subgroup analysis of patients with localized disease, emphasizing the prognostic role of 4E-BP1 in EwS. Consistently, chr8 gain was associated with higher intra-tumoral EIF4EBP1 expression in EwS patient samples. RNA interference-mediated knockdown of 4E-BP1 significantly decreased proliferation, clonogenicity, and spheroidal growth of EwS cells in vitro as well as tumor growth of EwS xenografts in vivo. To identify potential mechanisms via which 4E-BP1 mediates its phenotypic effect, we performed integrated proteomic and transcriptomic profiling of EwS cell lines with and without silencing of 4E-BP1 revealing that 4E-BP1 guides a multifunctional proteomic network including hubs associated with RNA processing, translational regulation, and chromatin modification. Collectively, we establish a significant association between chr8 gain and unfavorable prognosis in EwS. We show that this association primarily depends on the expression of the translation initiation factor binding protein 4E-BP1 regulating a multifunctional proteomic network. Our data suggest that cytogenetic testing for chr8 gain in EwS tumors may help to improve risk-stratification of patients and adaptation of the therapeutic management.

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Section: Chromosome 8 Gain Drives Overexpression Of the Clinically Re...mentioning

confidence: 91%

Section: Chromosome 8 Gain Drives Overexpression Of the Clinically Re...mentioning

confidence: 91%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Chromosome 8 gain drives cancer progression by hijacking the translation factor 4E-BP1 sensitizing for targeted CDK4/6 inhibition

Funk

Orth²,

Aljakouch

et al. 2022

Preprint

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“…First, we compared pseudobulk chromatin accessibility data from our experiment to context-specific data of the well-characterized fusion EWSR1-FLI1 in Ewing sarcoma. A high-confidence set of loci bound by EWSR1-FLI1 across 15 Ewing sarcoma cancer cell lines was previously published 33 . We examined the normalized ATAC signal at these 1,879 EWSR1-FLI1 bound sites after combining all nuclei for each variant in our library (Fig 4a).…”

Section: Resultsmentioning

confidence: 99%

Discovering chromatin dysregulation induced by protein-coding perturbations at scale

Frenkel,

Hujoel,

Morris

et al. 2023

Preprint

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Although population-scale databases have expanded to millions of protein-coding variants, insight into variant mechanisms has not kept pace. We present PROD-ATAC, a high-throughput method for discovering the effects of protein-coding variants on chromatin. A pooled library of variants is expressed in a disease-agnostic cell line, and single-cell ATAC resolves each variant's effect on chromatin. Using PROD-ATAC, we characterized the effects of >100 oncofusions (a class of cancer-causing chimeric proteins) and controls and revealed that pioneer activity is a common feature of fusions spanning an enormous range of fusion frequencies. Further, fusion-induced dysregulation can be context-agnostic as observed mechanisms often overlapped with cancer and cell-type specific prior knowledge. We also showed that gain-of-function pioneering is common among oncofusions. This work provides a global view of fusion-induced chromatin. We uncovered convergent mechanisms among disparate oncofusions and shared modes of dysregulation across different cancers. PROD-ATAC is generalizable to any set of protein-coding variants.

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“…Analysis of DRIP signal in non-EWS::FLI1 transcriptional regulated genes 39 (ACTB, ACTG1, CFL1 and TPT1) showed that EwS cells accumulated higher levels of R-loops than IMR-90 (Fig. 5E; Supplementary Fig.…”

Section: Ews::fli1 Promotes R-loops Accumulation Independently Of Its...mentioning

confidence: 99%

EWS::FLI1-DHX9 interaction promotes Ewing sarcoma sensitivity to DNA topoisomerase 1 poisons by altering R-loop metabolism

Olmedo-Pelayo,

Granado-Calle,

Delgado-Bellido

et al. 2023

Preprint

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Drug resistance is one of the major factors associated with poor outcome of cancer patients. Treatment of Ewing sarcoma (EwS), an aggressive neoplasm mainly affecting children, adolescents and young adults, is associated with therapy failure and tumor relapse in 30-80% of the cases. Thus, it supports the need to explore the mechanisms modulating drug activity. Here, we describe a novel mechanism of drug sensitivity based on the role of EWS::FLI1 in R-loop metabolism. Our results demonstrate that EWS::FLI1 promotes R-loop formation favoring the interaction between DHX9 and elongating RNA polymerase II. In addition, we discovered that EWS::FLI1 kidnaps DHX9 preventing the resolution of TOP1 poisoning-associated R-loops. Our findings indicate that R-loops accumulation promotes replicative stress, genome instability and cell sensitivity to SN-38. Collectively, these results uncover a novel mechanism behind EwS sensitivity to genotoxic agents, with relevant implications for EwS treatment.

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Systematic multi-omics cell line profiling uncovers principles of Ewing sarcoma fusion oncogene-mediated gene regulation

Cited by 31 publications

References 153 publications

Chromosome 8 gain drives cancer progression by hijacking the translation factor 4E-BP1 sensitizing for targeted CDK4/6 inhibition

Chromosome 8 gain drives cancer progression by hijacking the translation factor 4E-BP1 sensitizing for targeted CDK4/6 inhibition

Discovering chromatin dysregulation induced by protein-coding perturbations at scale

EWS::FLI1-DHX9 interaction promotes Ewing sarcoma sensitivity to DNA topoisomerase 1 poisons by altering R-loop metabolism

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