Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma: Role in Tumor Growth and Immune Evasion

Sabine Heitzeneder, Elena Sotillo, Jack F. Shern, Sivasish Sindiri, Peng Xu, Robert Jones, Michael Pollak, Pernille R. Noer, Julie Lorette, Ladan Fazli, Anya Alag, Paul Meltzer, Ching Lau, Cheryl A Conover, Claus Oxvig, Poul H. Sorensen, John M. Maris, Javed Khan, Crystal L. Mackall

Research output: Contribution to journalArticle

Abstract

BACKGROUND: Ewing sarcoma (EWS) manifests one of the lowest somatic mutation rates of any cancer, leading to a scarcity of druggable mutations and neoantigens. Immunotherapeutics targeting differentially expressed cell surface antigens could provide therapeutic benefit for such tumors. Pregnancy-associated plasma protein A (PAPP-A) is a cell membrane-associated proteinase produced by the placenta that promotes fetal growth by inducing insulinlike growth factor (IGF) signaling. METHODS: By comparing RNA expression of cell surface proteins in EWS (n = 120) versus normal tissues (n = 42), we comprehensively characterized the surfaceome of EWS to identify highly differentially expressed molecules. Using CRISPR/Cas-9 and anti-PAPP-A antibodies, we investigated biological roles for PAPP-A in EWS in vitro and in vivo in NSG xenograft models and performed RNA-sequencing on PAPPA knockout clones (n = 5) and controls (n = 3). All statistical tests were two-sided. RESULTS: EWS surfaceome analysis identified 11 highly differentially overexpressed genes, with PAPPA ranking second in differential expression. In EWS cell lines, genetic knockout of PAPPA and treatment with anti-PAPP-A antibodies revealed an essential survival role by regulating local IGF-1 bioavailability. MAb-mediated PAPPA inhibition diminished EWS growth in orthotopic xenografts (leg area mm2 at day 49 IgG2a control (CTRL) [n = 14], mean = 397.0, SD = 86.1 vs anti-PAPP-A [n = 14], mean = 311.7, SD = 155.0; P = .03; median OS anti-PAPP-A = 52.5 days, 95% CI = 46.0 to 63.0 days vs IgG2a = 45.0 days, 95% CI = 42.0 to 52.0 days; P = .02) and improved the efficacy of anti-IGF-1R treatment (leg area mm2 at day 49 anti-PAPP-A + anti-IGF-1R [n = 15], mean = 217.9, SD = 148.5 vs IgG2a-CTRL; P < .001; median OS anti-PAPP-A + anti-IGF1R = 63.0 days, 95% CI = 52.0 to 67.0 days vs IgG2a-CTRL; P < .001). Unexpectedly, PAPPA knockout in EWS cell lines induced interferon (IFN)-response genes, including proteins associated with antigen processing/presentation. Consistently, gene expression profiles in PAPPA-low EWS tumors were enriched for immune response pathways. CONCLUSION: This work provides a comprehensive characterization of the surfaceome of EWS, credentials PAPP-A as a highly differentially expressed therapeutic target, and discovers a novel link between IGF-1 signaling and immune evasion in cancer, thus implicating shared mechanisms of immune evasion between EWS and the placenta.

Original languageEnglish (US)
Pages (from-to)970-982
Number of pages13
JournalJournal of the National Cancer Institute
Volume111
Issue number9
DOIs
StatePublished - Sep 1 2019

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Pregnancy-Associated Plasma Protein-A
Tumor Escape
Ewing's Sarcoma
Growth
Intercellular Signaling Peptides and Proteins
Immune Evasion
Antigen Presentation
Heterografts
Placenta
Leg
Clustered Regularly Interspaced Short Palindromic Repeats
RNA Sequence Analysis
Cell Line
Neoplasms
Antibodies
Mutation Rate
Surface Antigens
Fetal Development
Transcriptome
Interferons

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

Heitzeneder, S., Sotillo, E., Shern, J. F., Sindiri, S., Xu, P., Jones, R., ... Mackall, C. L. (2019). Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma: Role in Tumor Growth and Immune Evasion. Journal of the National Cancer Institute, 111(9), 970-982. https://doi.org/10.1093/jnci/djy209

Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma : Role in Tumor Growth and Immune Evasion. / Heitzeneder, Sabine; Sotillo, Elena; Shern, Jack F.; Sindiri, Sivasish; Xu, Peng; Jones, Robert; Pollak, Michael; Noer, Pernille R.; Lorette, Julie; Fazli, Ladan; Alag, Anya; Meltzer, Paul; Lau, Ching; Conover, Cheryl A; Oxvig, Claus; Sorensen, Poul H.; Maris, John M.; Khan, Javed; Mackall, Crystal L.

In: Journal of the National Cancer Institute, Vol. 111, No. 9, 01.09.2019, p. 970-982.

Research output: Contribution to journalArticle

Heitzeneder, S, Sotillo, E, Shern, JF, Sindiri, S, Xu, P, Jones, R, Pollak, M, Noer, PR, Lorette, J, Fazli, L, Alag, A, Meltzer, P, Lau, C, Conover, CA, Oxvig, C, Sorensen, PH, Maris, JM, Khan, J & Mackall, CL 2019, 'Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma: Role in Tumor Growth and Immune Evasion', Journal of the National Cancer Institute, vol. 111, no. 9, pp. 970-982. https://doi.org/10.1093/jnci/djy209
Heitzeneder, Sabine ; Sotillo, Elena ; Shern, Jack F. ; Sindiri, Sivasish ; Xu, Peng ; Jones, Robert ; Pollak, Michael ; Noer, Pernille R. ; Lorette, Julie ; Fazli, Ladan ; Alag, Anya ; Meltzer, Paul ; Lau, Ching ; Conover, Cheryl A ; Oxvig, Claus ; Sorensen, Poul H. ; Maris, John M. ; Khan, Javed ; Mackall, Crystal L. / Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma : Role in Tumor Growth and Immune Evasion. In: Journal of the National Cancer Institute. 2019 ; Vol. 111, No. 9. pp. 970-982.
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title = "Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma: Role in Tumor Growth and Immune Evasion",
abstract = "BACKGROUND: Ewing sarcoma (EWS) manifests one of the lowest somatic mutation rates of any cancer, leading to a scarcity of druggable mutations and neoantigens. Immunotherapeutics targeting differentially expressed cell surface antigens could provide therapeutic benefit for such tumors. Pregnancy-associated plasma protein A (PAPP-A) is a cell membrane-associated proteinase produced by the placenta that promotes fetal growth by inducing insulinlike growth factor (IGF) signaling. METHODS: By comparing RNA expression of cell surface proteins in EWS (n = 120) versus normal tissues (n = 42), we comprehensively characterized the surfaceome of EWS to identify highly differentially expressed molecules. Using CRISPR/Cas-9 and anti-PAPP-A antibodies, we investigated biological roles for PAPP-A in EWS in vitro and in vivo in NSG xenograft models and performed RNA-sequencing on PAPPA knockout clones (n = 5) and controls (n = 3). All statistical tests were two-sided. RESULTS: EWS surfaceome analysis identified 11 highly differentially overexpressed genes, with PAPPA ranking second in differential expression. In EWS cell lines, genetic knockout of PAPPA and treatment with anti-PAPP-A antibodies revealed an essential survival role by regulating local IGF-1 bioavailability. MAb-mediated PAPPA inhibition diminished EWS growth in orthotopic xenografts (leg area mm2 at day 49 IgG2a control (CTRL) [n = 14], mean = 397.0, SD = 86.1 vs anti-PAPP-A [n = 14], mean = 311.7, SD = 155.0; P = .03; median OS anti-PAPP-A = 52.5 days, 95{\%} CI = 46.0 to 63.0 days vs IgG2a = 45.0 days, 95{\%} CI = 42.0 to 52.0 days; P = .02) and improved the efficacy of anti-IGF-1R treatment (leg area mm2 at day 49 anti-PAPP-A + anti-IGF-1R [n = 15], mean = 217.9, SD = 148.5 vs IgG2a-CTRL; P < .001; median OS anti-PAPP-A + anti-IGF1R = 63.0 days, 95{\%} CI = 52.0 to 67.0 days vs IgG2a-CTRL; P < .001). Unexpectedly, PAPPA knockout in EWS cell lines induced interferon (IFN)-response genes, including proteins associated with antigen processing/presentation. Consistently, gene expression profiles in PAPPA-low EWS tumors were enriched for immune response pathways. CONCLUSION: This work provides a comprehensive characterization of the surfaceome of EWS, credentials PAPP-A as a highly differentially expressed therapeutic target, and discovers a novel link between IGF-1 signaling and immune evasion in cancer, thus implicating shared mechanisms of immune evasion between EWS and the placenta.",
author = "Sabine Heitzeneder and Elena Sotillo and Shern, {Jack F.} and Sivasish Sindiri and Peng Xu and Robert Jones and Michael Pollak and Noer, {Pernille R.} and Julie Lorette and Ladan Fazli and Anya Alag and Paul Meltzer and Ching Lau and Conover, {Cheryl A} and Claus Oxvig and Sorensen, {Poul H.} and Maris, {John M.} and Javed Khan and Mackall, {Crystal L.}",
year = "2019",
month = "9",
day = "1",
doi = "10.1093/jnci/djy209",
language = "English (US)",
volume = "111",
pages = "970--982",
journal = "Journal of the National Cancer Institute",
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publisher = "Oxford University Press",
number = "9",

}

TY - JOUR

T1 - Pregnancy-Associated Plasma Protein-A (PAPP-A) in Ewing Sarcoma

T2 - Role in Tumor Growth and Immune Evasion

AU - Heitzeneder, Sabine

AU - Sotillo, Elena

AU - Shern, Jack F.

AU - Sindiri, Sivasish

AU - Xu, Peng

AU - Jones, Robert

AU - Pollak, Michael

AU - Noer, Pernille R.

AU - Lorette, Julie

AU - Fazli, Ladan

AU - Alag, Anya

AU - Meltzer, Paul

AU - Lau, Ching

AU - Conover, Cheryl A

AU - Oxvig, Claus

AU - Sorensen, Poul H.

AU - Maris, John M.

AU - Khan, Javed

AU - Mackall, Crystal L.

PY - 2019/9/1

Y1 - 2019/9/1

N2 - BACKGROUND: Ewing sarcoma (EWS) manifests one of the lowest somatic mutation rates of any cancer, leading to a scarcity of druggable mutations and neoantigens. Immunotherapeutics targeting differentially expressed cell surface antigens could provide therapeutic benefit for such tumors. Pregnancy-associated plasma protein A (PAPP-A) is a cell membrane-associated proteinase produced by the placenta that promotes fetal growth by inducing insulinlike growth factor (IGF) signaling. METHODS: By comparing RNA expression of cell surface proteins in EWS (n = 120) versus normal tissues (n = 42), we comprehensively characterized the surfaceome of EWS to identify highly differentially expressed molecules. Using CRISPR/Cas-9 and anti-PAPP-A antibodies, we investigated biological roles for PAPP-A in EWS in vitro and in vivo in NSG xenograft models and performed RNA-sequencing on PAPPA knockout clones (n = 5) and controls (n = 3). All statistical tests were two-sided. RESULTS: EWS surfaceome analysis identified 11 highly differentially overexpressed genes, with PAPPA ranking second in differential expression. In EWS cell lines, genetic knockout of PAPPA and treatment with anti-PAPP-A antibodies revealed an essential survival role by regulating local IGF-1 bioavailability. MAb-mediated PAPPA inhibition diminished EWS growth in orthotopic xenografts (leg area mm2 at day 49 IgG2a control (CTRL) [n = 14], mean = 397.0, SD = 86.1 vs anti-PAPP-A [n = 14], mean = 311.7, SD = 155.0; P = .03; median OS anti-PAPP-A = 52.5 days, 95% CI = 46.0 to 63.0 days vs IgG2a = 45.0 days, 95% CI = 42.0 to 52.0 days; P = .02) and improved the efficacy of anti-IGF-1R treatment (leg area mm2 at day 49 anti-PAPP-A + anti-IGF-1R [n = 15], mean = 217.9, SD = 148.5 vs IgG2a-CTRL; P < .001; median OS anti-PAPP-A + anti-IGF1R = 63.0 days, 95% CI = 52.0 to 67.0 days vs IgG2a-CTRL; P < .001). Unexpectedly, PAPPA knockout in EWS cell lines induced interferon (IFN)-response genes, including proteins associated with antigen processing/presentation. Consistently, gene expression profiles in PAPPA-low EWS tumors were enriched for immune response pathways. CONCLUSION: This work provides a comprehensive characterization of the surfaceome of EWS, credentials PAPP-A as a highly differentially expressed therapeutic target, and discovers a novel link between IGF-1 signaling and immune evasion in cancer, thus implicating shared mechanisms of immune evasion between EWS and the placenta.

AB - BACKGROUND: Ewing sarcoma (EWS) manifests one of the lowest somatic mutation rates of any cancer, leading to a scarcity of druggable mutations and neoantigens. Immunotherapeutics targeting differentially expressed cell surface antigens could provide therapeutic benefit for such tumors. Pregnancy-associated plasma protein A (PAPP-A) is a cell membrane-associated proteinase produced by the placenta that promotes fetal growth by inducing insulinlike growth factor (IGF) signaling. METHODS: By comparing RNA expression of cell surface proteins in EWS (n = 120) versus normal tissues (n = 42), we comprehensively characterized the surfaceome of EWS to identify highly differentially expressed molecules. Using CRISPR/Cas-9 and anti-PAPP-A antibodies, we investigated biological roles for PAPP-A in EWS in vitro and in vivo in NSG xenograft models and performed RNA-sequencing on PAPPA knockout clones (n = 5) and controls (n = 3). All statistical tests were two-sided. RESULTS: EWS surfaceome analysis identified 11 highly differentially overexpressed genes, with PAPPA ranking second in differential expression. In EWS cell lines, genetic knockout of PAPPA and treatment with anti-PAPP-A antibodies revealed an essential survival role by regulating local IGF-1 bioavailability. MAb-mediated PAPPA inhibition diminished EWS growth in orthotopic xenografts (leg area mm2 at day 49 IgG2a control (CTRL) [n = 14], mean = 397.0, SD = 86.1 vs anti-PAPP-A [n = 14], mean = 311.7, SD = 155.0; P = .03; median OS anti-PAPP-A = 52.5 days, 95% CI = 46.0 to 63.0 days vs IgG2a = 45.0 days, 95% CI = 42.0 to 52.0 days; P = .02) and improved the efficacy of anti-IGF-1R treatment (leg area mm2 at day 49 anti-PAPP-A + anti-IGF-1R [n = 15], mean = 217.9, SD = 148.5 vs IgG2a-CTRL; P < .001; median OS anti-PAPP-A + anti-IGF1R = 63.0 days, 95% CI = 52.0 to 67.0 days vs IgG2a-CTRL; P < .001). Unexpectedly, PAPPA knockout in EWS cell lines induced interferon (IFN)-response genes, including proteins associated with antigen processing/presentation. Consistently, gene expression profiles in PAPPA-low EWS tumors were enriched for immune response pathways. CONCLUSION: This work provides a comprehensive characterization of the surfaceome of EWS, credentials PAPP-A as a highly differentially expressed therapeutic target, and discovers a novel link between IGF-1 signaling and immune evasion in cancer, thus implicating shared mechanisms of immune evasion between EWS and the placenta.

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