High 4E-BP1 expression associates with chromosome 8 gain and CDK4/6 sensitivity in Ewing sarcoma
Cornelius M. Funk, Anna C. Ehlers, Martin F. Orth, Karim Aljakouch, Jing Li, Tilman L.B. Hölting, Rainer Will, Florian H. Geyer, A. Katharina Ceranski, Franziska Willis, Endrit Vinca, Shunya Ohmura, Roland Imle, Jana Siebenlist, Angelina Yershova, Maximilian M.L. Knott, Felina Zahnow, Ana Sastre, Javier Alonso, Felix Sahm, Heike Peterziel, Anna Loboda, Martin Schneider, Ana Banito, Gabriel Leprivier, Wolfgang Hartmann, Uta Dirksen, Olaf Witt, Ina Oehme, Stefan M. Pfister, Laura Romero-Pérez, Jeroen Krijgsveld, Florencia Cidre-Aranaz, Thomas G.P. Grünewald, Julian Musa
Cornelius M. Funk, Anna C. Ehlers, Martin F. Orth, Karim Aljakouch, Jing Li, Tilman L.B. Hölting, Rainer Will, Florian H. Geyer, A. Katharina Ceranski, Franziska Willis, Endrit Vinca, Shunya Ohmura, Roland Imle, Jana Siebenlist, Angelina Yershova, Maximilian M.L. Knott, Felina Zahnow, Ana Sastre, Javier Alonso, Felix Sahm, Heike Peterziel, Anna Loboda, Martin Schneider, Ana Banito, Gabriel Leprivier, Wolfgang Hartmann, Uta Dirksen, Olaf Witt, Ina Oehme, Stefan M. Pfister, Laura Romero-Pérez, Jeroen Krijgsveld, Florencia Cidre-Aranaz, Thomas G.P. Grünewald, Julian Musa
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Research Article Genetics Oncology

High 4E-BP1 expression associates with chromosome 8 gain and CDK4/6 sensitivity in Ewing sarcoma

Abstract

Chromosome 8 (chr8) gains are common in cancer, but their contribution to tumor heterogeneity is largely unexplored. Ewing sarcoma (EwS) is defined by FET::ETS fusions with few other recurrent mutations to explain clinical diversity. In EwS, chr8 gains are the second most frequent alteration, making it an ideal model to study the relevance of chr8 gains in an otherwise silent genomic context. We report that chr8 gain–driven expression patterns correlate with poor overall survival of patients with EwS. This effect is mainly mediated by increased expression of the translation initiation factor binding protein 4E-BP1, encoded by EIF4EBP1 on chr8. Among all chr8-encoded genes, EIF4EBP1 expression showed the strongest association with poor survival and correlated with chr8 gains in EwS tumors. Similar findings emerged across multiple cancer entities in The Cancer Genome Atlas. Multiomics profiling revealed that 4E-BP1 orchestrates a pro-proliferative proteomic network. Silencing 4E-BP1 reduced proliferation, clonogenicity, spheroidal growth in vitro, and tumor growth in vivo. Drug screens demonstrated that high 4E-BP1 expression sensitizes EwS to pharmacological CDK4/6-inhibition. Chr8 gains and elevated 4E-BP1 emerge as prognostic biomarkers in EwS, with poor outcomes driven by 4E-BP1–mediated pro-proliferative networks that sensitize tumors to CDK4/6 inhibitors. Testing for chr8 gains may enhance risk stratification and therapy in EwS and other cancers.

Authors

Cornelius M. Funk, Anna C. Ehlers, Martin F. Orth, Karim Aljakouch, Jing Li, Tilman L.B. Hölting, Rainer Will, Florian H. Geyer, A. Katharina Ceranski, Franziska Willis, Endrit Vinca, Shunya Ohmura, Roland Imle, Jana Siebenlist, Angelina Yershova, Maximilian M.L. Knott, Felina Zahnow, Ana Sastre, Javier Alonso, Felix Sahm, Heike Peterziel, Anna Loboda, Martin Schneider, Ana Banito, Gabriel Leprivier, Wolfgang Hartmann, Uta Dirksen, Olaf Witt, Ina Oehme, Stefan M. Pfister, Laura Romero-Pérez, Jeroen Krijgsveld, Florencia Cidre-Aranaz, Thomas G.P. Grünewald, Julian Musa

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Figure 1

Chr8 gain drives overexpression of the clinically relevant translation initiation factor 4E-BP1 in EwS.

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Chr8 gain drives overexpression of the clinically relevant translation i...
(A) Flowchart illustrating patient stratification to either a high or low chr8 gene-expression-signature group based on hierarchical clustering (hclust) of sample-specific ssGSEA enrichment scores for the chr8 gene set (left). Heatmap of ssGSEA scores for chr8 genes in 196 EwS tumors (cohort 1) (right). Color intensity indicates the degree of gene set enrichment. (B) Kaplan-Meier overall survival analysis of cohort 1 stratified into either a high or low chr8-signature-enrichment group as described in (A). P values were determined by the Mantel-Haenszel test. (C) Kaplan-Meier overall survival analysis of 117 patients with EwS (cohort 1 with Chr8 focus) stratified into either a high or low chr8-signature-enrichment group as described in (A) but excluding samples with other inferred, recurrent CNVs. P values were determined by the Mantel-Haenszel test. (D) Overall survival batch analysis as assessed for every gene or exclusively chr8 genes covered in transcriptomic profiling of cohort 1 using the Mantel-Haenszel test. (E) Kaplan-Meier overall survival analysis of cohort 1 stratified by quartile of EIF4EBP1 expression. Percentages given for each expression quartile refer to the percentage of patients with predicted chr8 gain in the respective quartile. P values were determined by the Mantel-Haenszel test. (F) EIF4EBP1 expression as measured by microarray profiling in 196 EwS tumors (cohort 1) stratified into either a high or low chr8-signature-enrichment group as described in (A). P values were determined by 2-tailed Mann-Whitney test; horizontal bars represent means, and whiskers represent the SEM. ***P < 0.001. (G) EIF4EBP1 expression as measured by RNA-Seq in cohort 2 depending on the presence of chr8 gain as determined by methylation array. P values were determined by 2-tailed Mann-Whitney test; horizontal bars represent means, and whiskers represent the SEM. ***P < 0.001. (H) Kaplan-Meier overall survival analysis of 117 patients with EwS (cohort 1 with Chr8 focus, as in C) stratified by quartile of EIF4EBP1 expression. Percentages given for each expression quartile refer to the percentage of patients with predicted chr8 gain in the respective quartile. P values were determined by the Mantel-Haenszel test.