Endosalpingiosis Is Negative for GATA3: A Useful Marker for Distinguishing Metastatic Breast Carcinoma From a Benign Mimicker

This study was approved by the institutional review board. Two cases of endosalpingiosis involving axillary lymph nodes were identified from a previous study of benign axillary lymph node inclusions.¹¹  Additional, consecutive cases of endosalpingiosis during a 1-year period were identified by searching the pathology data system for archival specimens whose diagnoses contained the term endosalpingiosis. Cases with a diagnosis of “atypical endosalpingiosis” were excluded, as were cases lacking archival hematoxylin-eosin–stained sections or available formalin-fixed, paraffin embedded (FFPE) blocks. Histologic slide re-review to confirm the diagnosis of benign endosalpingiosis within normal tissue was performed for all cases. Clinicopathologic features including patient sex, age, concurrent diagnosis of malignancy, and anatomic location were recorded.

GATA3 immunostaining was performed on unstained sections cut fresh from archival FFPE blocks, using a mouse monoclonal antibody (clone L50-823, Cat No. CM405B, BioCare Medical Systems, Pacheco, California) at a dilution of 1:100. Immunostaining was performed on an automated immunohistochemistry system (BenchMark Ultra, Ventana Medical Systems, Tucson, Arizona). Briefly, tissue sections were loaded into the autostainer where they were deparaffinized and hydrated, with heat-induced epitope retrieval using CC1 solution (EDTA pH9, Ventana) for 64 minutes. After cooling and rinsing with wash buffer, primary antibody was applied for 44 minutes. Signals were detected with horseradish peroxidase-labeled secondary antibody followed by DAB chromogen, using the manufacturer's instructions (Cat No. 760-500, UltraView DAB, Ventana). Nuclear GATA3 labeling in endosalpingiosis was recorded as percentage and intensity labeling, with any labeling greater than 1% considered positive. All sections of endosalpingiosis contained adjacent benign lymphocytes, which served as an internal control for GATA3 labeling. Any incidental GATA3 labeling in adjacent normal tissues was also recorded.

A total of 15 cases of incidental, benign endosalpingiosis were identified. All 15 patients were female, with a mean age of 51 years (range, 36–73 years). Approximately half (n = 7, 47%) of the endosalpingiosis cases involved lymph nodes (5 pelvic lymph nodes, 2 axillary lymph nodes), and approximately half (n = 8, 53%) involved pelvic soft tissues or gynecologic organs (3 uterine myometrium, 2 paratubal soft tissue, 2 periadnexal soft tissue, and 1 pelvic sidewall). Four patients had a concurrent diagnosis of malignancy, including 2 patients with breast carcinoma, 1 patient with high-grade ovarian serous carcinoma, and 1 patient with ovarian endometrioid carcinoma with squamous differentiation. One additional patient had a concurrent intermediate-grade Sertoli-Leydig cell tumor of the ovary. The 2 cases of axillary endosalpingiosis occurred within axillary sentinel lymph nodes excised during surgical management (staging and treatment) in patients with primary breast carcinoma, both of which were ER⁺ invasive mammary carcinoma of no special type (ie, invasive ductal carcinoma). The 2 cases of endosalpingiosis involving axillary sentinel lymph nodes were also positive for PAX8, ER, and pancytokeratin by immunohistochemistry. PAX8 immunohistochemistry was not performed on the 13 subdiaphragmatic cases of endosalpingiosis, as it is well established that benign Müllerian epithelia are positive for PAX8.^12–14  Likewise, WT1 immunohistochemistry was not performed as WT1 is expected to be positive in benign fallopian tube epithelium.^15,16 

In all cases involving lymph nodes, the endosalpingiosis inclusions were located either within the capsule or in the subcapsular region. Two cases were multifocal. The lymph nodes containing endosalpingiosis were uninvolved by metastatic carcinoma. In all 15 cases, the endosalpingiosis inclusion was composed of cytologically bland ciliated and nonciliated, low cuboidal to columnar cells (Figure 1, A). In all cases, the endosalpingiosis showed no desmoplastic stromal response.

GATA3 immunohistochemistry was negative in all foci of endosalpingiosis (Figure 1, B through D), with intact strong nuclear labeling seen in normal lymphocytes in all cases. In addition, strong and diffuse (100%) GATA3 labeling was noted in an incidental periadnexal Walthard cell rest (ie, benign urothelial cell metaplasia) (Figure 2, A and B). Weak and focal (<5%) GATA3 labeling was noted in the normal fallopian tube epithelium of 2 cases (Figure 2, C and D) and in the normal endometrial stroma of 2 cases. Normal ovarian stroma, endometrial glands, myometrium, and endothelium were negative for GATA3.

Breast cancer remains the most common cancer in women and the second leading cause of cancer-related deaths in women in the United States.¹⁷  Pathologic stage remains an important prognostic indicator in breast cancer. Despite evolving local treatment guidelines for axillary nodal metastases, the sentinel lymph node biopsy remains central to stage determination and subsequent patient management decisions.^18,19  Thus, accurate classification of nodal glands is critical. Misclassification of benign glandular inclusions, resulting in a false-positive diagnosis of metastatic carcinoma, could result in axillary nodal radiation, additional nodal surgery, or the addition of systemic therapy.²⁰  Although false-positive interpretations of breast sentinel lymph nodes are rare, false-positive diagnoses can occur at the time of frozen section, touch prep analysis, cytologic smear, or permanent section.^20–26 

Müllerian inclusions such as endosalpingiosis account for 1 potential source of false-positive diagnoses in lymph nodes. Müllerian inclusions most commonly occur in subdiaphragmatic locations, where their presence is not unexpected. The potential for misclassification of Müllerian inclusions is higher when they are encountered in supradiaphragmatic locations, including axillary lymph nodes. Most benign epithelial inclusions involving axillary lymph nodes take the form of mammary-type epithelium, squamous-type inclusions, and mixed glandular and squamous inclusions.²⁷  Failure to recognize endosalpingiosis as a benign Müllerian inclusion can lead to misdiagnosis as a metastatic low-grade mammary carcinoma.²⁷ 

Immunohistochemistry can be useful in separating endosalpingiosis from metastatic low-grade adenocarcinoma in histologically ambiguous cases. It is prudent to use a targeted immunopanel that includes markers of gynecologic tract origin as well as markers of mammary origin. Müllerian inclusions are immunoreactive for PAX8 and negative for GCDFP and mammaglobin,^12–14  whereas mammary carcinomas are negative for PAX8 and often immunoreactive for GCDFP and mammaglobin.^{10,11,28–30}  However, there are several potential diagnostic pitfalls. Müllerian-derived epithelium and most low-grade mammary carcinomas are strongly immunoreactive for ER.³¹  In addition, mammary carcinoma can occasionally label with WT1, a marker traditionally used to support gynecologic tract origin among adenocarcinomas.^32,33  Furthermore, mammary carcinomas may display only focal GCDFP or mammaglobin labeling, both of which are cytoplasmic and may be associated with nonspecific background labeling.¹⁰  GATA3 is a zinc-binding transcription factor, and GATA3 displays nuclear labeling by immunohistochemistry. GATA3 immunohistochemistry is sensitive although not entirely specific for mammary carcinoma, particularly those with a luminal (ie, ER⁺) phenotype.^1,2,4  However, GATA3 immunohistochemistry has not been systematically evaluated in endosalpingiosis.

Here, we evaluated GATA3 labeling in a series of 15 consecutive cases of endosalpingiosis and show that all are negative for GATA3 immunohistochemistry. Our findings support those of a recent case series by Shiino et al³⁴  that reported 2 cases of axillary nodal endosalpingiosis to be negative for GATA3. No GATA3 labeling has also been reported in a small number of endometriosis cases.^35,36  In addition, Ronaghy et al³⁶  reported GATA3 was negative in 18 samples of normal fallopian tube epithelium, from which endosalpingiosis is thought to arise. Banet et al³⁷  similarly reported that fallopian tube epithelium was typically negative in a study of GATA3 expression in 445 nonneoplastic and lesional trophoblastic tissues and tumors. We observed weak and focal (<5%) labeling for GATA3 in 2 cases of normal fallopian tube epithelium in our series, raising the possibility that some endosalpingiosis foci could potentially display weak and focal GATA3 labeling. However, this pattern of labeling would contrast with the strong and diffuse GATA3 labeling seen in ER⁺ mammary carcinomas.²  A potential challenge to the use of GATA3 immunohistochemistry in evaluating epithelial inclusions in lymph nodes is the GATA3 labeling of normal resident lymphocytes. However, this is mitigated by the histologic difference between gland-forming inclusions and singly dispersed lymphocytes; in addition, the GATA3 labeling by lymphocytes is beneficial as it serves as an internal quality control.

In summary, GATA3 immunohistochemistry is negative in endosalpingiosis, including both nodal and soft tissue inclusions, making it a useful marker to distinguish endosalpingiosis and low-grade ER⁺ mammary carcinoma. Use of a targeted immunopanel containing GATA3 and PAX8 would yield negative and positive immunohistochemical stain results in this differential diagnosis and resolve the diagnosis for histologically ambiguous intranodal glands, preventing the false-positive diagnosis of metastatic mammary carcinoma.