Elsevier

Clinical Breast Cancer

Volume 23, Issue 2, February 2023, Pages 199-210
Clinical Breast Cancer

Full Length Article
AMACR Expression is a Potential Diagnostic Marker in Apocrine Lesions of Breast, and is Associated with High Histologic Grade and Lymph Node Metastases in Some Invasive Apocrine Breast Cancers

https://doi.org/10.1016/j.clbc.2022.11.012Get rights and content

Abstract

Background

Carcinoma with apocrine differentiation (AC) is a subtype of breast carcinoma with apocrine features in >90% of the tumor. Molecular studies demonstrate AC has high expression of androgen receptor (AR) mRNA. Pure AC lack estrogen receptor (ER), progesterone receptor (PR), and express AR, with variable human epidermal growth factor 2 (HER2) status. Currently, in triple negative AC, no targetable therapies or specific diagnostic markers exist.

Materials and Methods

α-Methylacyl CoA racemase (AMACR) expression was investigated as a marker of apocrine differentiation using a single-plex immunoperoxidase stain, and a novel AMACR/p63 dual stain in a subset of cases, across 1) benign apocrine lesions (apocrine metaplasia, adenosis) 2) apocrine DCIS (ADCIS), 3) AC/ invasive ductal carcinoma (IDC) with apocrine features, 4) non-apocrine triple negative breast cancer (TNBC) and 5) IDC, no special type. A sub-set of cases were evaluated by tissue microarray.

Results

AMACR expression was increased in both AC and ADCIS, with minimal expression in benign breast tissue, TNBC and IDC, NST cases. In invasive cases, those with positive AMACR (>5% positivity) were significantly associated with higher histologic grade (P = .006), initial N stage (chi squared 0.044), and lack of ER or PR expression (both P < .001), with no correlation with overall survival. Analysis of TCGA breast cancer datasets revealed AMACR expression was significantly higher in molecularly defined apocrine carcinomas relative to basal and luminal subtypes. Moreover, high AMACR expression predicted worse relapse-free and distant-metastasis free survival, among both ER-/PR-/Her2- and ER-/PR-/Her2+ breast cancer cohorts (log-rank P = .081 and .00011, respectively).

Conclusion

AMACR represents a promising diagnostic and prognostic marker in apocrine breast lesions. Further study is needed to determine the biologic and clinical significance of this protein in AC.

Introduction

Apocrine differentiation in breast lesions, defined by cuboidal or columnar cells with low nuclear-cytoplasmic ratio, abundant eosinophilic cytoplasm, prominent apical granules, and round nuclei with pale chromatin and often prominent nucleoli, are commonly encountered in daily breast pathology practice.1 The histologic spectrum of these lesions includes apocrine metaplasia, apocrine adenosis, ductal hyperplasia, carcinoma in situ, and invasive carcinoma. Invasive apocrine breast carcinomas (AC) are rare – by some estimates 0.3% to 4% of all invasive ductal carcinomas (IDC), and have been defined as lesions showing >90% apocrine morphology in tumor cells.2 Historically, AC has been categorized as a subset of IDC of no-special type.3 Recently, however, the field has now moved to classify these lesions via molecular rather than morphologic criteria, as a subset of the “triple negative” breast cancers lacking appreciable steroid receptor expression (estrogen receptor [ER], progesterone receptor [PR], and human epidermal growth factor 2 [HER2]), with positivity for the androgen receptor (AR).4,5 A recent study of invasive apocrine carcinomas in a large national database showed that while these cancers more likely to present with aggressive clinicopathologic features than non-apocrine carcinomas, breast cancer-specific survival was the same, and among triple-negative apocrine carcinomas, survival was better than non-apocrine triple negative carcinomas.6 Therefore, accurate diagnosis of these lesions is of paramount importance for practicing surgical pathologists.

Apocrine morphology may be encountered in tumors from organ systems outside of the breast, including primary cutaneous apocrine (sweat gland) carcinoma of the skin, apocrine salivary duct carcinoma, papillary renal cell carcinoma, urothelial cell carcinoma in situ, and rare reports of apocrine carcinoma arising within ovarian teratoma.7, 8, 9 In patients presenting with axillary lymphadenopathy or an axillary mass of unknown primary, without adequate clinical history, differentiating primary cutaneous apocrine carcinoma and metastatic apocrine carcinoma of the breast may be virtually impossible on morphologic or immunohistochemical grounds alone – representing an important challenge for practicing breast surgical pathologists.10 We recently encountered such a case at our institution, which stained strongly and diffusely for the traditional apocrine markers AR and gross cystic disease fluid protein 15 (GCDFP15), as well as α-methylacyl CoA racemase (AMACR; Figure 1). Additional immunohistochemical markers supported an invasive breast primary, including diffuse GATA3 and cytokeratin 7 positivity and negativity for cytokeratin 20, TTF-1, PSA, CA IX, mammaglobin, S100, SOX10, and PAX8, and p63, with absence of myoepithelial markers p63, CK5/6 and calponin as well (not shown). Ultimately, primary apocrine breast carcinoma was favored.

Many prior researchers have sought to identify markers that might aid in routine identification of breast carcinoma (p63, CK5/6, GATA3, mammaglobin), as well as specific identification of primary apocrine breast carcinoma (GCDFP, AR, 15-prostaglandin dehydrogenase [15-PGDH], TRPS1, acyl-CoA synthetase medium-chain family member 1 [ACSFM1]), forkhead box protein A1 (FOXA1), and the steroid hormone receptors ER, PR, and HER2 – however, no single reliable marker with adequate sensitivity and specificity has emerged.3,5,10,11 In our study, we found strong, granular cytoplasmic expression of AMACR (P504S) in the majority (58%) of apocrine breast carcinoma cases, a tumor marker previously known for overexpression in prostatic adenocarcinoma.

AMACR is a 382-amino acid protein involved in β-oxidation of branched-chain fatty acids, which was first identified by Xu et al in 2000 using RNA subtraction and DNA microarray techniques.12 Jiang and colleagues proposed its use as a diagnostic marker in prostatic adenocarcinoma in 2004, after finding AMACR to display both high specificity and sensitivity for prostatic carcinoma, in both resection and core-needle biopsy specimens, regardless of Gleason grade.13,14 AMACR has now been in clinical use for decades as part of the so-called “PIN4” antibody cocktail (along with the basal cell markers p63 and high-molecular weight keratin).15

More recently, Nakamura and colleagues have studied AMACR in AC, finding significant expression in both invasive and in situ apocrine proliferations, and lower expression non-apocrine breast carcinomas.16 Our study expands upon these data in a large single-institution retrospective cohort study, using immunohistochemistry for AMACR as well as relevant molecular data available in the Cancer Genome Atlas (https://portal.gdc.cancer.gov). We also developed a novel dual AMACR-p63 immunohistochemical stain, which was well-suited for evaluation of AMACR (pink) and p63 (brown, a myoepithelial marker) protein expression for differentiation of invasive or in situ lesions. Our aim was to assess the potential utility of AMACR as a diagnostic tumor marker in apocrine breast lesions, and secondly to assess whether AMACR expression was associated with clinical outcome.

Section snippets

Study Cohort

Institutional Review Board approval for this study was obtained from Yale New Haven Hospital. A review of the pathology archives at our hospital was performed to identify patients with any diagnosis of apocrine lesions of the breast. All samples were obtained from breast core biopsy, lumpectomy or mastectomy specimens from the case files of the Yale Department of Surgical Pathology, with cases signed out between 2001 and 2022.

In this study, AMACR expression was evaluated and compared to that of

Results

Clinical pathologic features and diagnostic history of cases of ADCIS, AC, and TNBC, and treatment information for AC, are summarized in Table 1. Immunohistochemical staining for AMACR, AR, and GCDFP15 expression was evaluated on all cases; 20 cases of ADCIS and AC cases were evaluated using the AMACR-p63 dual stain, to assist in differentiation of in situ and invasive apocrine lesions in cases with nested and solid growth (Figure 2). Assessment of AMACR expression and clinicopathologic

Discussion

The current study identified positive AMACR protein expression, defined as >5% of cell staining, in ∼58% of AC cases, 48% of ADCIS cases, ∼4% of non-apocrine TNBC cases, 5% of IDC, NST, and no expression in normal breast tissue. In our study, although AMACR expression did not increase in a statistically significant manner across ADCIS and AC cases, it was differentially expressed in malignant apocrine lesions when compared to normal breast tissue, and a large cohort of non-apocrine invasive

Suggested reviewers

Anupma Nayak (Penn Medicine, [email protected]); Rohit Bhargava (Magee Women's Hospital of Pittsburgh, [email protected]); Paula Ginter (New York University, [email protected])

Disclosure

The authors declare that they have no relevant financial interests to disclose.

Acknowledgments

The authors would like to gratefully acknowledge Lori Charette in Yale Pathology Tissue Services for her immense technical assistance in TMA construction and apocrine case collection, Drs Yuanxin Liang and Uma Krishnamurthy for apocrine case contributions, and Dr Deepika Kumar for review of manuscript figures. MH: Yale Department of Pathology research funds; RJ: National Institute for Cancer Research (reg. No. LX22NPO5102); European Union - Next Generation EU, Programme EXCELES;

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