therascreenPITX2 RGQ PCR assay for the assessment of PITX2 DNA-methylation status to investigate the role of the transcription factor PITX2 and the regulation of the Wnt/ß-catenin pathway in pathophysiological processes.

Napieralski, Rudolf; Schricker, Gabriele; Piednoir, Elodie; Manner, Olivia; Bona, Alexandre; Segalas, Samantha; Schueren, Elisabeth; Lauber, Juergen; Perkins, Jonathan A.; Noske, Aurelia; Magdolen, Viktor; Weichert, Wilko; Kiechle, Marion; Wilhelm, Olaf G.

doi:10.1038/protex.2018.022

Cited by 4 publications

(7 citation statements)

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“…The present retrospective study was conducted by applying a fully analytically validated quantitative methylation-specific real-time PCR format [16] and an optimized clinically validated cut-off to extend previously published studies. Hartmann et al [10] investigated the value of PITX2 DNA methylation in 241 lymph node-positive, ER-positive and HER2-negative breast cancer patients, a patient cohort with the same clinicopathological characteristics as the present study, and showed the significant role of PITX2 DNA methylation in the prediction of outcome after adjuvant anthracycline treatment, at that time by applying a microarray-based test.…”

Section: Discussionmentioning

confidence: 99%

“…After bisulfite conversion with the Epitect Fast DNA Bisulfite Kit (QIAGEN, Hilden, Germany), the PMR of 3 CpG motifs of the PITX2 gene P2 [11] was quantified by qMSP using the therascreen PITX2 RGQ PCR kit, containing the quantitative PCR (qPCR) reaction mix, primer and probes as well as positive and negative controls. The assay was performed on the Rotor-Gene Q MDx real-time PCR platform (QIAGEN, Hilden, Germany) and automatically assessed by the Rotor-Gene AssayManager® software V2.1 with installed Gamma Plug-in and PITX2 -specific assay profile (therascreen_PITX2_FFPE_CE Assay Profile v1.0) for data analysis and quality control [16]. …”

Section: Methodsmentioning

confidence: 99%

“…However, these assays were all research tests, not analytically validated and therefore do not qualify according to Hayes [15] as suitable for the clinical routine setting. The present study aimed at validating the PITX2 DNA methylation status by applying a fully analytically validated assay [16] in a prospective, predefined analysis to predict outcome in lymph node-positive breast cancer patients receiving adjuvant anthracycline-based chemotherapy. The results show that the PITX2 DNA methylation assay can be used to further improve the concept of individualized cancer therapy decision-making and can support physicians on a case-by-case basis to optimize the treatment plan for each single breast cancer patient.…”

Section: Introductionmentioning

confidence: 99%

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Clinical Validation of PITX2 DNA Methylation to Predict Outcome in High-Risk Breast Cancer Patients Treated with Anthracycline-Based Chemotherapy

et al. 2018

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Background: Breast cancer patients at high risk for recurrence are treated with anthracycline-based chemotherapy, but not all patients do equally benefit from such a regimen. To further improve therapy decision-making, biomarkers predicting outcome are of high unmet medical need. Methods: The percent DNA methylation ratio (PMR) of the promoter gene coding for the Paired-like homeodomain transcription factor 2 (PITX2) was determined by a validated methylation-specific real-time polymerase chain reaction (PCR) test. The multicenter study was conducted in routinely collected archived formalin-fixed paraffin-embedded (FFPE) tissue from 205 lymph node-positive breast cancer patients treated with adjuvant anthracycline-based chemotherapy. Results: The cut-off for the PITX2 methylation status (PMR = 12) was confirmed in a randomly selected cohort (n = 60) and validated (n = 145) prospectively with disease-free survival (DFS) at the 10-year follow-up. DFS was significantly different between the PMR ≤ 12 versus the PMR > 12 group with a hazard ratio (HR) of 2.74 (p < 0.001) in the validation cohort and also for the patient subgroup treated additionally with endocrine therapy (HR 2.47; p = 0.001). Conclusions: Early-stage lymph node-positive breast cancer patients with low PITX2 methylation do benefit from adjuvant anthracycline-based chemotherapy. Patients with a high PITX2 DNA methylation ratio, approximately 30%, show poor outcome and should thus be considered for alternative chemotherapy regimens.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Clinical Validation of PITX2 DNA Methylation to Predict Outcome in High-Risk Breast Cancer Patients Treated with Anthracycline-Based Chemotherapy

et al. 2018

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“…The Therascreen PITX2 RGQ PCR assay , developed by Perkins et al (2018) in conjunction with the Therawis Diagnostics Company, analyzes the methylation status of the PITX2 gene obtained from the DNA of FFPE biospecimens. PITX2 methylation is assessed by methylation-specific real-time PCR and exploits the quantitative PCR (qPCR) oligonucleotide hydrolysis principle of two TaqMan probes labeled with different fluorescent dies (FAM ™ for fully methylated and HEX ™ for fully unmethylated DNA) in combination with methylation nonspecific primers to measure the methylation status of the target sequences of PITX2 gene promoter 2 in bisulfite-treated DNA.…”

Section: In Vitro Diagnostic Tests Based On Epigenetic Biomarkersmentioning

confidence: 99%

“…The Therascreeen PITX2 RGQ PCR assay can be easily adopted in clinical laboratories that already run other Therascreen assays commercialized by Qiagen. The complete workflow is streamlined throughput for a medium sample with highly reliable and robust readouts and can be performed in a time of 2 working days (Perkins et al, 2018).…”

Section: In Vitro Diagnostic Tests Based On Epigenetic Biomarkersmentioning

confidence: 99%

Epigenetic IVD Tests for Personalized Precision Medicine in Cancer

et al. 2019

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Epigenetic alterations play a key role in the initiation and progression of cancer. Therefore, it is possible to use epigenetic marks as biomarkers for predictive and precision medicine in cancer. Precision medicine is poised to impact clinical practice, patients, and healthcare systems. The objective of this review is to provide an overview of the epigenetic testing landscape in cancer by examining commercially available epigenetic-based in vitro diagnostic tests for colon, breast, cervical, glioblastoma, lung cancers, and for cancers of unknown origin. We compile current commercial epigenetic tests based on epigenetic biomarkers (i.e., DNA methylation, miRNAs, and histones) that can actually be implemented into clinical practice.

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Clinical performance of an analytically validated assay in comparison to microarray technology to assess PITX2 DNA-methylation in breast cancer

Schricker

Napieralski

Noske

et al. 2018

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Significant evidence has accumulated that DNA-methylation of the paired-like homeodomain transcription factor 2 (PITX2) gene can serve as a prognostic and predictive biomarker in breast cancer. PITX2 DNA-methylation data have been obtained so far from microarray and polymerase chain reaction (PCR)-based research tests. The availability of an analytically validated in vitro methylation-specific real-time PCR assay format (therascreen PITX2 RGQ PCR assay) intended for the determination of the percent methylation ratio (PMR) in the (PITX2) promoter 2 prompted us to investigate whether the clinical performance of these different assay systems generate comparable clinical outcome data. Mathematically converted microarray data of a previous breast cancer study (n = 204) into PMR values leads to a PITX2 cut-off value at PMR 14.73. Recalculation of the data to experimentally equivalent PMRs with the PCR PITX2 assay leads to a cut-off value at PMR 12 with the highest statistical significance. This cut-off predicts outcome of high-risk breast cancer patients to adjuvant anthracycline-based chemotherapy (n = 204; Hazard Ratio 2.48; p < 0.001) comparable to microarray generated results (n = 204; Hazard ratio 2.32; p < 0.0001). The therascreen PITX2 RGQ PCR assay is an analytically validated test with high reliability and robustness and predicts outcome of high-risk breast cancer patients to anthracycline-based chemotherapy.

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therascreenPITX2 RGQ PCR assay for the assessment of PITX2 DNA-methylation status to investigate the role of the transcription factor PITX2 and the regulation of the Wnt/ß-catenin pathway in pathophysiological processes.

Cited by 4 publications

References 0 publications

Clinical Validation of PITX2 DNA Methylation to Predict Outcome in High-Risk Breast Cancer Patients Treated with Anthracycline-Based Chemotherapy

Clinical Validation of PITX2 DNA Methylation to Predict Outcome in High-Risk Breast Cancer Patients Treated with Anthracycline-Based Chemotherapy

Epigenetic IVD Tests for Personalized Precision Medicine in Cancer

Clinical performance of an analytically validated assay in comparison to microarray technology to assess PITX2 DNA-methylation in breast cancer

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