The effect of cold ischemic time on the immunohistochemical evaluation of estrogen receptor, progesterone receptor, and HER2 expression in invasive breast carcinoma

Abstract

The American Society of Clinical Oncology/College of American Pathologist (ASCO/CAP) guidelines on breast cancer currently recommend a cold ischemic time of o1 h, although data are limited. Breast resection specimens were subjected to variable cold ischemic time periods (0.5, 1, 2, 3, 4, 24, and 48 h) within the refrigerator and at room temperature. The study included 25 tumors, all of which had refrigerated samples. Nonrefrigerated samples (samples at room temperature) were present on 23 cases. Hormone receptors were semiquantitatively scored using the H-score method. Human epidermal growth factor receptor 2 gene ERBB2 (HER2) was scored using the ASCO/CAP guidelines. The results were compared with the core biopsy scores, which have negligible cold ischemic time period before fixation. Mild reduction in staining for hormone receptors was judged present if the H-score on the resection specimen was between one-half and three-fourth of the H-score at core biopsy. Significant reduction was judged present when the H-score on resection was less than one-half of the core biopsy H-score. Mild reduction in HER2 staining was judged present if there was one-step reduction; and significant reduction was judged present if there was two-step reduction in staining. A true reduction was judged present only when the reduction was consistently present for the increasing time interval. A focal reduction for a particular time sample was attributed to the heterogeneity of the tumor sample. Non-refrigerated samples are affected more by prolonged cold ischemic time than refrigerated samples. Cold ischemic time period of as short as one-half hour may occasionally impact the immunohistochemical (IHC) staining for progesterone receptor. Significant reduction in IHC staining for hormone receptors, and HER2, however, generally does not result until 4 h for refrigerated samples and 2 h for non-refrigerated samples. The ASCO/CAP guideline of cold ischemic time period of o1 h is a prudent guideline to follow. Modern Pathology (2012Pathology ( ) 25, 1098Pathology ( -1105 doi:10.1038/modpathol.2012 published online 30 March 2012 Keywords: breast carcinoma; cold ischemic time; ER/PR/HER2 All invasive primary breast cancers are analyzed for at least estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 gene ERBB2 (HER2), as these markers have been shown to be the most important prognostic and predictive factors in breast cancer. The patients with ER þ primary invasive breast cancer derive considerable benefit from endocrine therapy, while ER-negative tumors do not. Similarly to ER expression, PR expression by immunohistochemistry also provides prognostic and predictive information. 1,2 Accurate determination of biomarkers is essential for the proper management of breast cancer patients. [3][4][5][6][7][8] The HER2 is overexpressed in B15-20% of breast carcinomas. 9-11 HER2 positivity is associated with higher rate of recurrence and mortality in newly diagnosed breast cancer patients, and agents that t...

Ischemia in Tumors Induces Early and Sustained Phosphorylation Changes in Stress Kinase Pathways but Does Not Affect Global Protein Levels

“…Our comprehensive proteome analysis, which greatly expands on prior studies of the consequences of cold ischemia on protein expression (15,16), revealed that none of the Ͼ10,000 proteins detected and quantified changed in abundance as a result of cold ischemia in a time span of up to one hour post-excision. However, because of the inability of conventional "bottom-up" MSbased proteomics technology to analyze the integrity of intact proteins (as samples are digested to peptides prior to analysis), we cannot exclude the possibility that limited proteolysis such as apoptotic regulation by caspases occurs at early time points of cold ischemia.…”

“…A study employing AQUA, a quantitative immunofluorescence technique, found slightly increased expression of hypoxia inducible factor but no changes in protein abundance in the four breast cancer biomarker proteins ER, PR, HER2, and Ki67 over a time span of up to 7 h, though signal reduction occurred in a subset of samples at longer intervals of up to 48 h of cold ischemia (15). In another study employing immunohistochemistry, the breast cancer biomarker proteins ER, PR, and Her2 were found to be stable for up to 2 h at room temperature (16). Evidence from RPPA studies evaluating the time course of changes induced by cold ischemia has shown that even when the total protein levels and levels of many phosphoproteins are constant, the phosphorylation stoichiometry of specific proteins can change, in some cases significantly (9,17).…”

Use of modified Magee equations and histologic criteria to predict the Oncotype DX recurrence score

“…32 In light of these concerns, several quality assurance programs have been created including ad hoc consensus conference recommendations, 33,34 and the guideline recommendations from the American Society of Clinical Oncology/College of American Pathologist for HER-2, ER, and PR testing, 35,36 which includes mandatory proficiency testing. These national efforts have led to marked improvements in the quality, reliability, and inter-laboratory agreement for these breast cancer assays, [37][38][39][40][41] and have made the use of semiquantitative immunohistochemistry results more feasible for estimating risk. These efforts in improving quality may be most helpful in reconsidering the role of Ki-67 as part of the routine panel for testing of breast cancers.…”

A systematic comparison of three commercial estrogen receptor assays in a single clinical outcome breast cancer cohort

“…Ready-to-use assays offer improved reproducibility and consistency, minimizing effects arising from reagent diversity, as well as variability that can arise from antigen retrieval. [8][9][10][11][12][13][14][15][16] Surprisingly, these platform-specific ready-to-use assays have never been directly compared using the same clinical outcome series.…”