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ESR1, ERBB2, and Ki67 mRNA expression predicts stage and grade of non-muscle-invasive bladder carcinoma (NMIBC)

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Abstract

Pathological staging and grading are crucial for risk assessment in non-muscle-invasive bladder cancer (NMIBC). Molecular grading might support pathological evaluation and minimize interobserver variability. In this study, the well-established breast cancer markers ESR1, PGR, ERBB2, and MKI67 were evaluated as potential molecular markers to support grading and staging in NMIBC. We retrospectively analyzed clinical data and formalin-fixed paraffin-embedded tissues (FFPE) of patients with NMIBC. Messenger RNA (mRNA) expression of the aforementioned markers was measured by single-step reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) using RNA-specific TaqMan assays. Relative gene expression was determined by normalization to two reference genes (CALM2 and B2M) using the 40−ΔΔCT method and correlated to histopathological stage and grade. Pathological assessment was performed by an experienced uropathologist. Statistical analysis was performed using the SAS software JMP 9.0.0 version and GraphPad Prism 5.04. Of 381 cases of NMIBC, samples of 100 pTa and 255 pT1 cases were included in the final study. Spearman rank correlation revealed significant correlations between grade and expression of MKI67 (r = 0.52, p < 0.0001), ESR1 (r = 0.25, p < 0.0001), and ERBB2 (r = 0.18, p = 0.0008). In Mann-Whitney tests, MKI67 was significantly different between all grades (p < 0.0001), while ESR1 (p = 0.0006) and ERBB2 (p = 0.027) were significantly different between G2 and G3. Higher expression of MKI67 (r = 0.49; p < 0.0001), ERBB2 (r = 0.22; p < 0.0001), and ESR1 (r = 0.18; p = 0.0009) mRNA was positively correlated with higher stage. MKI67 (p < 0.0001), ERBB2 (p = 0.0058), and PGR (p = 0.0007) were significantly different between pTa and pT1. In NMIBC expression of ESR1, ERBB2 and MKI67 are significantly different between stage and grade. This potentially provides objective parameters for pathological evaluation.

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Abbreviations

ESR1/ER:

Estrogen receptor alpha

ERBB2/HER2:

erb-b2 receptor tyrosine kinase 2/human epidermal growth factor receptor 2

Cq:

Quantification cycle

FFPE:

Formalin-fixed paraffin-embedded

GOI:

Gene of interest

HR:

Hazard ratio

mRNA:

Messenger ribonucleic acid

MKI67/Ki67:

Marker of proliferation Ki-67

PGR/PgR:

Progesterone receptor

REF:

Reference gene

RT-qPCR:

Reverse transcription quantitative real-time polymerase chain reaction

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Acknowledgments

The authors thank Sefanie Herlein for excellent technical support.

Author information

Authors and Affiliations

  1. Department of Urology, University of Regensburg, Caritas Krankenhaus St. Josef, Landshuter Str. 65, 93053, Regensburg, Germany

    Johannes Breyer, Stefan Denzinger, Maximilian Burger & Wolfgang Otto

  2. STRATIFYER Molecular Pathology GmbH, Cologne, Germany

    Ralph M. Wirtz

  3. Institute of Pathology at the St Elisabeth Hospital Köln-Hohenlind, Cologne, Germany

    Ralph M. Wirtz & Sebastian Eidt

  4. BioNTech Diagnostics GmbH, Mainz, Germany

    Mark Laible & Kornelia Schlombs

  5. Department of Urology, University of Mannheim, Mannheim, Germany

    Philipp Erben & Maximilian Christian Kriegmair

  6. Institute of Pathology, University of Erlangen-Nuremberg, Erlangen, Germany

    Robert Stoehr & Arndt Hartmann

Authors
  1. Johannes Breyer
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  2. Ralph M. Wirtz
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  3. Mark Laible
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  4. Kornelia Schlombs
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  5. Philipp Erben
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  6. Maximilian Christian Kriegmair
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  7. Robert Stoehr
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  8. Sebastian Eidt
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  9. Stefan Denzinger
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  10. Maximilian Burger
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  11. Arndt Hartmann
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  12. Wolfgang Otto
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Corresponding author

Correspondence to Johannes Breyer.

Ethics declarations

Informed consent

Informed consent was obtained from all the individual participants included in the study.

Funding

The work on this topic was funded by the German Cancer Aid (Deutsche Krebshilfe), grant number 110,541.

Conflict of interest

RMW and SE are founders of STRATIFYER Molecular Pathology GmbH. RMW is an employee of STRATIFYER Molecular Pathology GmbH. ML and KS are employees of BioNTech Diagnostics GmbH.

Additional information

Johannes Breyer, Ralph M. Wirtz, Philipp Erben, Maximilian Christian Kriegmair, Maximilian Burger Arndt Hartmann, and Wolfgang Otto - on behalf of the BRIDGE Consortium

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Breyer, J., Wirtz, R.M., Laible, M. et al. ESR1, ERBB2, and Ki67 mRNA expression predicts stage and grade of non-muscle-invasive bladder carcinoma (NMIBC). Virchows Arch 469, 547–552 (2016). https://doi.org/10.1007/s00428-016-2002-1

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  • DOI: https://doi.org/10.1007/s00428-016-2002-1

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