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Neuronatin is a modifier of estrogen receptor-positive breast cancer incidence and outcome

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Abstract

Purpose

Understanding the molecular mediators of breast cancer survival is critical for accurate disease prognosis and improving therapies. Here, we identified Neuronatin (NNAT) as a novel antiproliferative modifier of estrogen receptor-alpha (ER+) breast cancer.

Experimental design

Genomic regions harboring breast cancer modifiers were identified by congenic mapping in a rat model of carcinogen-induced mammary cancer. Tumors from susceptible and resistant congenics were analyzed by RNAseq to identify candidate genes. Candidates were prioritized by correlation with outcome, using a consensus of three breast cancer patient cohorts. NNAT was transgenically expressed in ER+ breast cancer lines (T47D and ZR75), followed by transcriptomic and phenotypic characterization.

Results

We identified a region on rat chromosome 3 (142–178 Mb) that modified mammary tumor incidence. RNAseq of the mammary tumors narrowed the candidate list to three differentially expressed genes: NNAT, SLC35C2, and FAM210B. NNAT mRNA and protein also correlated with survival in human breast cancer patients. Quantitative immunohistochemistry of NNAT protein revealed an inverse correlation with survival in a univariate analysis of patients with invasive ER+ breast cancer (training cohort: n = 444, HR = 0.62, p = 0.031; validation cohort: n = 430, HR = 0.48, p = 0.004). NNAT also held up as an independent predictor of survival after multivariable adjustment (HR = 0.64, p = 0.038). NNAT significantly reduced proliferation and migration of ER+ breast cancer cells, which coincided with altered expression of multiple related pathways.

Conclusions

Collectively, these data implicate NNAT as a novel mediator of cell proliferation and migration, which correlates with decreased tumorigenic potential and prolonged patient survival.

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Acknowledgements

We thank Jozef Lazar for his assistance in generating the SS-3BN congenic strains.

Funding

This work was supported by a seed grant from the Wisconsin Breast Cancer Showhouse, the MCW Cancer Center, the Advancing a Healthier Wisconsin Endowment, and the Dr. Nancy Laning Sobczak Fund for Breast Cancer (M.J.F, H.R., C.B.). Support was also received from the NCI (R01CA193343 (M.J.F), R01CA188575 (H.R)); the Mary Kay Foundation (Grant No. 024-16 (M.J.F) and 017-29 (C.B.)); the METAvivor Foundation (M.J.F and H.R.); Susan G. Komen Grants KG091116 (H.R., E.P.M, I.C., A.J.K., C.D.S., J.A.H., and H.H.) and CCR17483233 (C.B), an American Cancer Society Institutional Research Grant (#86-004-26 (C.B.)); and the National Center for Research Resources, the National Center for Advancing Translational Sciences, and the Office of the Director of the NIH via the Clinical & Translational Science Institute (#8KL2TR000056 (C.B.)), NIH Office of the Director and NIEHS (K01ES025435 (J.W.P.)). The study was also supported by W81XWH-12-2-0050, HU0001-16-2-0004 from the U.S. Department of Defense through the Henry M. Jackson Foundation for the Advancement of Military Medicine (C.D.S., A.J.K., J.A.H., and H.H.). The views expressed in this article are those of the authors and do not reflect the official policy of the Department of the Army/Navy/Air Force, Department of Defense (DOD), or US Government.

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Authors and Affiliations

  1. Genomic Sciences and Precision Medicine Center, Medical College of Wisconsin, 8701 Watertown Plank Rd, Milwaukee, WI, 53226, USA

    Cody Plasterer, Shirng-Wern Tsaih, Caitlin O’Meara, Angela Lemke, Dana Murphy, Jennifer Smith & Michael J. Flister

  2. Department of Physiology, Medical College of Wisconsin, Milwaukee, WI, USA

    Cody Plasterer, Shirng-Wern Tsaih, Caitlin O’Meara, Angela Lemke, Dana Murphy & Michael J. Flister

  3. Department of Pathology, Medical College of Wisconsin, Milwaukee, WI, USA

    Amy R. Peck, Yunguang Sun & Hallgeir Rui

  4. Division of Biostatistics, Thomas Jefferson University, Philadelphia, PA, USA

    Inna Chervoneva

  5. Simmons Cancer Institute, Southern Illinois University School of Medicine, Springfield, IL, USA

    Sophia Ran

  6. Department of Medical Microbiology, Immunology, and Cell Biology, Southern Illinois University School of Medicine, Springfield, IL, USA

    Sophia Ran

  7. John P. Murtha Cancer Center, Uniformed Services University and Walter Reed National Military Medical Center, Bethesda, MD, USA

    Albert J. Kovatich, Jeffrey A. Hooke & Craig D. Shriver

  8. Chan Soon-Shiong Institute of Molecular Medicine at Windber, Windber, PA, USA

    Hai Hu

  9. Medical Oncology, Thomas Jefferson University, Philadelphia, PA, USA

    Edith P. Mitchell

  10. Department of Radiation Oncology, Medical College of Wisconsin, Milwaukee, WI, USA

    Carmen Bergom

  11. Department of Radiology, Medical College of Wisconsin, Milwaukee, WI, USA

    Amit Joshi

  12. Joseph J. Zilber School of Public Health, University of Wisconsin-Milwaukee, Milwaukee, WI, USA

    Paul Auer

  13. Department of Pediatrics and Human Development, College of Human Medicine, Michigan State University, Lansing, MI, USA

    Jeremy Prokop

  14. Department of Pharmacology and Toxicology, Michigan State University, East Lansing, MI, USA

    Jeremy Prokop

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  1. Cody Plasterer
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  21. Michael J. Flister
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Corresponding author

Correspondence to Michael J. Flister.

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Conflict of interest

The authors have no conflicts of interest to declare.

Ethical approval

All applicable international, national, and institutional guidelines for the care and use of animals were followed. The Institutional Animal Care and Use Committee (IACUC) of the Medical College of Wisconsin approved all animal studies. All procedures involving animals were conducted in accordance with the National Institutes of Health guidelines concerning the use and care of experimental animals. All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. All histological breast cancer tissues were archival, de-identified specimens approved for use under waiver of consent by MCW IRB protocol PRO00028590.

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Plasterer, C., Tsaih, SW., Peck, A.R. et al. Neuronatin is a modifier of estrogen receptor-positive breast cancer incidence and outcome. Breast Cancer Res Treat 177, 77–91 (2019). https://doi.org/10.1007/s10549-019-05307-8

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