Abstract P6-01-06: Feasibility of the PROSIGNA® multigene test in core biopsies and comparison to corresponding surgical breast cancer sections
Authors: Prat Aleix , Galván Patricia , Buckingham Wesley , Vidal Maria , Díaz Sherley , Nuciforo Paolo , Ferree Sean , Adamo Barbara , Ramon y Cajal Santiago , Peg Vicente 
Published: May 2015

Abstract
Background: The PROSIGNA® (PAM50) gene assay has been validated on formalin-fixed paraffin embedded (FFPE) surgical resection specimens (SRS) to identify the intrinsic subtypes of breast cancer and to estimate the 10-year risk of recurrence (ROR) in post-menopausal patients treated with adjuvant endocrine therapy. However, demonstration of the ability to perform PAM50 assay in diagnostic core biopsy specimens (CBS) before primary surgery and/or systemic therapy could be clinically useful. The objectives of this study were to 1) evaluate the feasibility of performing the PAM50 assay in CBS and 2) compare the PAM50 results from paired CBS and SRS.
Methods and materials: Baseline tissue surface area, cellularity and RNA yield (obtained after ∼10 FFPE 10μm sections) were determined in CBS from 30 newly diagnosed breast cancer patients. The tissue volume requirements determined from these samples reflected the lower 95% confidence limits of a minimum RNA concentration of &gt;20ng/µL. The RNA yield and assay pass rate of the established tissue volume requirements were then tested in 30 independent CBSs. Intrinsic subtype concordance, and ROR score variability, were determined from 1) multiple extractions of the same CBS (10 independent cases for a total of 84 extractions) and 2) multiple CBS of the same tumor (30 independent cases for a total of 79 CBS). To test the PAM50 assay concordance between paired CBS and SRS, the following PAM50 data were evaluated in an independent and retrospective set of 33 paired samples: 4-class subtype classification (Luminal A, Luminal B, HER2-enriched and Basal-like), 3-class subtype classification (Luminal A/B, HER2-enriched and Basal-like), ROR score (0-100), proliferation score and the correlation to each subtype centroid. Correlation and concordance between CBS and SRS were estimated using Pearson coefficients and multi-rater kappa values, respectively.
Results: Baseline median surface area, cellularity and RNA yield concentration were 10.2 mm2, 45% and 155.3 ng/µl, respectively. Correlation of surface area and cellularity with RNA yield concentration was low (Person coefficient &lt;0.25). Minimum tissue requirements were determined based on surface area: &gt;12 mm2 = 2 10-micron slides, 6-12 mm2 = 4 slides; &lt;6 mm2 = 8 slides. Subtype calls on multiple extractions from the same CBS were 98% concordant (82/84) and the average ROR score standard deviation (SD) was 2.2 units. Subtype calls on different CBS of the same tumor were 94% concordant (74/79) and the average ROR score SD was 6.8 units. All 7 discordant cases were between Luminal A and B calls. Comparison between paired CBS and SRS revealed correlations of ≥0.90 (range 0.90-0.98) for ROR scores, proliferation scores and subtype centroid correlations. Intrinsic subtype concordance between paired CBS and SRS was 87% (kappa=0.81) and 97% (kappa= 0.91) for the 4-subtype and 3-subtype classifications, respectively. Finally, the overall PAM50 assay pass rate in CBS was &gt;95%.
Conclusions: The PAM50 assay in CBS is feasible and measurements are comparable with surgical resections, which suggest that PAM50 can be performed on diagnostic core biopsy tissues.
Citation Format: Aleix Prat, Patricia Galván, Wesley Buckingham, Maria Vidal, Sherley Díaz, Paolo Nuciforo, Sean Ferree, Barbara Adamo, Santiago Ramon y Cajal, Vicente Peg. Feasibility of the PROSIGNA® multigene test in core biopsies and comparison to corresponding surgical breast cancer sections [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P6-01-06.