title: Identification of BRCA1-like triple-negative breast cancers by quantitative multiplex-ligation-dependent probe amplification (MLPA) analysis of BRCA1-associated chromosomal regions: a validation study.
abstract: Triple-negative breast cancer (TNBC) with a BRCA1-like molecular signature has been demonstrated to remarkably respond to platinum-based chemotherapy and might be suited for a future treatment with poly(ADP-ribose)polymerase (PARP) inhibitors. In order to rapidly assess this signature we have previously developed a multiplex-ligation-dependent probe amplification (MLPA)-based assay. Here we present an independent validation of this assay to confirm its important clinical impact.
  
  
  
  One-hundred-forty-four TNBC tumor specimens were analysed by the MLPA-based "BRCA1-like" test. Classification into BRCA1-like vs. non-BRCA1-like samples was performed by our formerly established nearest shrunken centroids classifier. Data were subsequently compared with the BRCA1-mutation/methylation status of the samples. T-lymphocyte infiltration and expression of the main target of PARP inhibitors, PARP1, were assessed on a subset of samples by immunohistochemistry. Data acquisition and interpretation was performed in a blinded manner.
  
  
  
  In the studied TNBC cohort, 63 out of 144 (44 %) tumors were classified into the BRCA1-like category. Among these, the MLPA test correctly predicted 15 out of 18 (83 %) samples with a pathogenic BRCA1-mutation and 20 of 22 (91 %) samples exhibiting BRCA1-promoter methylation. Five false-negative samples were observed. We identified high lymphocyte infiltration as one possible basis for misclassification. However, two falsely classified BRCA1-mutated tumors were also characterized by rather non-BRCA1-associated histopathological features such as borderline ER expression. The BRCA1-like vs. non-BRCA1-like signature was specifically enriched in high-grade (G3) cancers (90 % vs. 58 %, p = 0.0004) and was also frequent in tumors with strong (3+) nuclear PARP1 expression (37 % vs. 16 %; p = 0.087).
  
  
  
  This validation study confirmed the good performance of the initial MLPA assay which might thus serve as a valuable tool to select patients for platinum-based chemotherapy regimens. Moreover, frequent PARP1 upregulation in BRCA1-like tumors may also point to susceptibility to treatment with PARP inhibitors. Limitations are the requirement of high tumor content and high-quality DNA.
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany. eva.gross@lrz.tum.de.
authors.forename: Eva
authors.initials: E
authors.lastname: Gross
authors.affiliation: Biometrics Department, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX, Amsterdam, The Netherlands.
authors.forename: Harm
authors.initials: H
authors.lastname: van Tinteren
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Li
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Raab
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Meul
authors.affiliation: Institute of Pathology, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.|Present address: Department of Pathology, Case Western Reserve University School of Medicine, University Hospitals Case Medical Center, Cleveland, OH, USA.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Avril
authors.affiliation: MRC-Holland, Willem Schoutenstraat 6, 1057 DN, Amsterdam, The Netherlands.
authors.forename: Nadja
authors.initials: N
authors.lastname: Laddach
authors.affiliation: Helmholtz Zentrum München, Institute of Pathology, Ingolstädter Landstrasse 1, D-85764, Neuherberg, Germany.
authors.forename: Michaela
authors.initials: M
authors.lastname: Aubele
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Corinna
authors.initials: C
authors.lastname: Propping
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Apostolos
authors.initials: A
authors.lastname: Gkazepis
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Manfred
authors.initials: M
authors.lastname: Schmitt
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Alfons
authors.initials: A
authors.lastname: Meindl
authors.affiliation: Department of Pathology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX, Amsterdam, The Netherlands.
authors.forename: Petra M
authors.initials: PM
authors.lastname: Nederlof
authors.affiliation: Department of Gynecology and Obstetrics, Technische Universität München, Ismaninger Strasse 22, D-81675, Munich, Germany.
authors.forename: Marion
authors.initials: M
authors.lastname: Kiechle
authors.affiliation: Department of Pathology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX, Amsterdam, The Netherlands.|Department of Molecular Pathology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX, Amsterdam, The Netherlands.
authors.forename: Esther H
authors.initials: EH
authors.lastname: Lips
mesh_terms: D000328:Adult; D000368:Aged; D019313:BRCA1 Protein / Q000235:genetics* / Q000378:metabolism; D014408:Biomarkers, Tumor; D002874:Chromosome Mapping; D003131:Combined Modality Therapy; D019175:DNA Methylation; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D008875:Middle Aged; D060885:Multiplex Polymerase Chain Reaction; D009154:Mutation; D060787:Neoplasm Grading; D000071137:Poly (ADP-Ribose) Polymerase-1 / Q000378:metabolism; D011401:Promoter Regions, Genetic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D064726:Triple Negative Breast Neoplasms / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism / Q000628:therapy; D047368:Tumor Burden
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D014408:Biomarkers, Tumor; D000071137:Poly (ADP-Ribose) Polymerase-1
keywords: BRCA1; BRCAness; DNA repair; MLPA assay; PARP1; Triple-negative breast cancer