Semi-quantitative Immunohistochemical Assay versus oncotype DX® qRT-PCR Assay for Estrogen and Progesterone Receptors

An Independent Quality Assurance Study

James A Kraus; David J Dabbs; Sushil Beriwal; Rohit Bhargava

Disclosures

Mod Pathol. 2012;25(6):869-876. 

In This Article

Materials and Methods

We identified 464 breast cancer cases at Magee-Womens Hospital (from the year 2008 and 2009), which were analyzed by Oncotype DX® by qRT-PCR with supplemental reporting of ER/PR expression units. The Oncotype DX® assay was performed on one tissue block from an appropriately fixed (8–72 h) resection specimen in each case. On all cases a tumor tissue block was sent for Oncotype DX® testing to Genomic Health on clinician's request. Subsequent analysis including qRT-PCR for hormone receptors was performed at Genomic Health. According to the company's reporting criteria, a tumor is considered ER positive with expression units of ≥6.5 and PR positive with expression units of ≥5.5.

Immunohistochemical hormone receptor analysis was performed on corresponding core-needle biopsies at the time of initial diagnosis. ER expression was assessed using the antibody clone SP1 and PR using clone 1E2. Immunohistochemical staining for both antibodies was performed using iVIEW detection on the Benchmark XT system (Ventana, Tuscon, AZ). The cases with discordant immunohistochemistry and RT-PCR results were re-examined by immunohistochemistry on the same tissue block that was sent for oncotype DX® testing to exclude any possibility of tissue heterogeneity accounting for the discordance.

Hormone receptor immunohistochemical semi-quantitation was determined using the modified H-score. The score consists of the sum of the percent of tumor cells staining multiplied by an ordinal value corresponding to the intensity level (0=none, 1=weak, 2=moderate, and 3=strong). With four intensity levels, the resulting score ranges from 0 (no staining in the tumor) to 300 (diffuse intense staining of the tumor).[4,5] Examples of ER and PR immunohistochemical stains, and corresponding H-score calculations are shown in Figures 1a–d. In accordance to ASCO/CAP guidelines, an H-score of ≥1 was considered a positive result for both ER and PR.

Figure 1.

Modified H-score examples (range: 0 to 300). Case (a) (strong ER positive; H-score 300), 0: 0%, 1+: 0%, 2+: 0%, 3+: 100%. ER H-score=(0 × 0)+(1 × 0)+(2 × 0)+(3 × 100)=300. Case (b) (heterogeneous but still strong PR positive; H-score 205), 0: 25%, 1+: 5%, 2+: 10%, 3+: 60%. PR H-score=(0 × 25)+(1 × 5)+(2 × 10)+(3 × 60)=205. Case (c) (moderate ER positive; H-score 135), 0: 10%, 1+: 50%, 2+: 35%, 3+: 5%. ER H-score=(0 × 10)+(1 × 50)+(2 × 35)+(3 × 5)=135. Case (d) (weak PR positive; H-score 30), 0: 85%, 1+: 5%, 2+: 5%, 3+: 5%. PR H-score=(0 × 85)+(1 × 5)+(2 × 5)+(3 × 5)=30.

RT-PCR and immunohistochemistry results were compared qualitatively (positive/negative), and quantitatively using Pearson correlation.

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