ESR1, ERBB2, and Ki67 mRNA expression predicts stage and grade of non-muscle-invasive bladder carcinoma (NMIBC)
- 548 Downloads
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.
KeywordsNMIBC ERBB2 ESR1 MKI67 Stage Grade
Estrogen receptor alpha
erb-b2 receptor tyrosine kinase 2/human epidermal growth factor receptor 2
Gene of interest
Messenger ribonucleic acid
Marker of proliferation Ki-67
Reverse transcription quantitative real-time polymerase chain reaction
The authors thank Sefanie Herlein for excellent technical support.
Compliance with ethical standards
Informed consent was obtained from all the individual participants included in the study.
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.
- 4.Otto W, Denzinger S, Fritsche HM, Burger M, Rößler W, Bertz S, May M, Hartmann A, Hofstädter F, Wieland WF, Eder F (2013) Introduction and first clinical application of a simplified immunohistochemical validation system confirms prognostic impact of KI-67 and CK20 for stage T1 urothelial bladder carcinoma: single-center analysis of eight biomarkers in a series of three hundred six patients. Clin Genitourin Cancer 11:537–544CrossRefPubMedGoogle Scholar
- 5.Sylvester RJ, van der Meijden A, Oosterlinck W, Witjes JA, Bouffioux C, Denis L, Newling DW, Kurth K (2006) Predicting recurrence and progression in individual patients with stage Ta, T1 bladder cancer using EORTC risk tables: a combined analysis of 2596 patients from seven EORTC trials. Eur Urol 49:466–465CrossRefPubMedGoogle Scholar
- 9.Choi W, Porter S, Kim S, Willis D, Plimack ER, Hoffman-Censits J, Roth B, Cheng T, Tran M, Lee IL, Melquist J, Bondaruk J, Majewski T, Zhang S, Pretzsch S, Baggerly K, Siefker-Radtke A, Czerniak B, Dinney CP, McConkey DJ (2014) Identification of distinct basal and luminal subtypes of muscle-invasive bladder cancer with different sensitivities to frontline chemotherapy. Cancer Cell 25:152–165CrossRefPubMedPubMedCentralGoogle Scholar
- 13.Lopez-Beltran A, Luque RJ, Alvarez-Kindelan J, Quintero A, Merlo F, Carrasco JC, Requena MJ, Montironi R (2004) Prognostic factors in stage T1 grade 3 bladder cancer survival: the role of G1-S modulators (p53, p21Waf1, p27kip1, Cyclin D1, and Cyclin D3) and proliferation index (ki67-MIB1). Eur Urol 45:606–612CrossRefPubMedGoogle Scholar
- 14.Gontero P, Gillo A, Fiorito C, Oderda M, Pacchioni D, Casetta G, Peraldo F, Zitella A, Tizzani A, Ricceri F (2014) Prognostic factors of ‘high-grade’ Ta bladder cancers according to the WHO 2004 classification: are these equivalent to ‘high-risk’ non-muscle-invasive bladder cancer? Urol Int 92:136–142CrossRefPubMedGoogle Scholar
- 18.Amsellem-Ouazana D, Bièche I, Molinié V, Elie C, Vieillefond A, Tozlu S, Botto H, Debré B, Lidereau R (2006) Is quantitative real-time RT-PCR an adjunct to immunohistochemistry for the evaluation of ErbB2 status in transitional carcinoma of the bladder? Eur Urol 49:1035–1042CrossRefPubMedGoogle Scholar
- 20.Iyer G, Al-Ahmadie H, Schultz N, Hanrahan AJ, Ostrovnaya I, Balar AV, Kim PH, Lin O, Weinhold N, Sander C, Zabor EC, Janakiraman M, Garcia-Grossman IR, Heguy A, Viale A, Bochner BH, Reuter VE, Bajorin DF, Milowsky MI, Taylor BS, Solit DB (2013) Prevalence and co-occurrence of actionable genomic alterations in high-grade bladder cancer. J Clin Oncol 31:3133–3140CrossRefPubMedPubMedCentralGoogle Scholar
- 22.Tan W, Boorjian S, Advani P, Farmer S, Lohse C, Cheville J, Kwon E, Leibovich B (2015) The estrogen pathway: estrogen receptor-α, progesterone receptor, and estrogen receptor-β expression in radical cystectomy urothelial cell carcinoma specimens. Clin Genitourin Cancer 13:476–484CrossRefPubMedGoogle Scholar