Analytic validation of a clinical-grade PTEN immunohistochemistry assay in prostate cancer by comparison with PTEN FISH

Abstract

PTEN loss is a promising prognostic and predictive biomarker in prostate cancer. Because it occurs most commonly via PTEN gene deletion, we developed a clinical-grade, automated and inexpensive immunohistochemical assay to detect PTEN loss. We studied the sensitivity and specificity of PTEN immunohistochemistry relative to 4-color fluorescence in situ hybridization (FISH) for detection of PTEN gene deletion in a multi-institutional cohort of 731 primary prostate tumors. Intact PTEN immunostaining was 91% specific for absence of PTEN gene deletion, (549/602 tumors with 2 copies of the PTEN gene by FISH showed intact expression of PTEN by immunohistochemistry) and 97% sensitive for presence of homozygous PTEN gene deletion (absent PTEN protein expression by immunohistochemistry in 65/67 tumors with homozygous deletion). PTEN immunohistochemistry was 65% sensitive for presence of hemizygous PTEN gene deletion, with protein loss in 40/62 hemizygous tumors. We reviewed the 53 cases where immunohistochemistry showed PTEN protein loss and FISH showed 2 intact copies of the PTEN gene. On re-review, there was ambiguous immunohistochemistry loss in 6% (3/53) and failure to analyze the same tumor area by both methods in 34% (18/53). Of the remaining discordant cases, 41% (13/32) revealed hemizygous (n=8) or homozygous (n=5) PTEN gene deletion that was focal in most cases (11/13). The remaining 19 cases had 2 copies of the PTEN gene by FISH, representing truly discordant cases. Our automated PTEN immunohistochemistry assay is a sensitive method for detection of homozygous PTEN gene deletions. Immunohistochemistry screening is particularly useful to identify cases with heterogeneous PTEN gene deletion in a subset of tumor glands. Mutations, small insertions or deletions and/or epigenetic or microRNA-mediated mechanisms may lead to PTEN protein loss in tumors with normal or hemizygous PTEN gene copy number.

PTEN loss in prostatic adenocarcinoma correlates with specific adverse histologic features (intraductal carcinoma, cribriform Gleason pattern 4 and stromogenic carcinoma)

“…[8][9][10][11][12] While PTEN loss at the genomic level may be evaluated by different techniques, PTEN immunohistochemistry (IHC) is a simple, yet sensitive and specific approach to detect this underlying genetic abnormality. 7,[15][16][17] The goal of this study, therefore, was to evaluate for correlations between PTEN loss by immunohistochemistry and specific histologic features validated as predictive of an aggressive clinical course.…”

Use of multicolor fluorescence in situ hybridization to detect deletions in clinical tissue sections

“…Based on recent studies using diagnostic needle biopsies, it has been proposed that an initial analysis of PTEN expression could be carried out using immunohistochemistry. Thereafter, regions of tumor or suspicious areas in the biopsy that have reduced expression of PTEN protein, or are otherwise indeterminate by immunohistochemistry, could then be analyzed by PTEN FISH as a reflex test [18]. This general approach would be in keeping with the emerging consensus for several types of oncologic pathology tests such as those for breast [19], lung [6,20], and melanomas [21], in which the primary immunohistochemistry analysis can be followed by reflex FISH for cases that are inconclusive.…”

Biomarkers for Diagnosis and Prognosis of Prostate Cancer

“…PTEN deletion associates with poor outcome and is an established prognostic biomarker for prostate cancer. Analysis of prostatic tissue by Immunohistochemistry (IHC) or Fluorescence in situ hybridization (FISH) demonstrated that PTEN loss is associated with prostate cancer biochemical recurrence, disease progression and metastasis [58][59][60][61][62][63].…”