Neuroendocrine Tumors of the Endometrium: a Case Series with Clinico-Pathological Analysis and Review of Literature

Abstract

Neuroendocrine carcinomas are rare aggressive tumors that are either mixed with endometrial carcinomas or pure neuroendocrine carcinoma. They show at least one or two neuroendocrine markers by immunohistochemistry. Here, we describe a case series of six cases that arise from endometrium. The mean age of presentation was 57 years with the most common type being mixed endometrioid and neuroendocrine carcinoma. Immunohistochemistry showed all cases positive for Pan-cytokeratin and INSM-1 while the majority were positive for synaptophysin and chromogranin. At least two neuroendocrine markers were positive in all cases in > 20% of tumor cells. The majority of the cases presented at FIGO (International Federation of Gynecology and Obstetrics) Stage III & IV. Three patients had a recurrence, one patient had bone metastasis, one patient died, and one patient was free of disease. Four patients with Stage IV disease were not alive for more than 18 months. A multidisciplinary approach is required for these aggressive tumors for better management of patients.

Introduction

Endometrial cancer is one of the most prevalent cancers among women globally, accounting for approximately 7% of all new cancer diagnosis and 4% of cancer-related deaths [1, 2]. The incidence of neuroendocrine tumors is < 1% of all endometrial cancers [3]. Neuroendocrine tumors (NET) of the endometrium are rare tumors with aggressive behavior and short overall survival [4]. The most common site of NET in the gynecological tract is the cervix followed by the ovary, uterine corpus, and vulva/vagina [5]. These tumors comprise two groups; one group is mixed carcinoma with endometrioid or serous carcinoma and the other group is pure neuroendocrine carcinoma [6]. The pure neuroendocrine component is divided into NET grade 1/2, small cell carcinoma, and large cell neuroendocrine carcinoma in the latest version of WHO Blue Book of Gynecological tract [5, 7]. NET grade 1 and grade 2 tumors show a ribbon-like, trabecular, nested, or glandular pattern with monotonous small round to oval cells with stippled chromatin. Grade 1 tumors show rare mitotic activity while grade 2 tumors can show 5–10 mitosis/2 mm². These tumors in the endometrium are extremely rare and found in only a few case reports [8, 9]. Small cell neuroendocrine carcinoma shows sheets of highly atypical cells with a high N:C ratio, scant cytoplasm, brisk mitotic activity, and necrosis. Large cell neuroendocrine carcinoma shows large atypical cells with a high N:C ratio with vesicular chromatin and prominent nucleoli and shows neuroendocrine differentiation.

Materials and Methods

We analyzed six cases of neuroendocrine carcinoma in our institute in a period of 3 years (2021–2024).

The diagnosis of small cell neuroendocrine carcinoma, large cell neuroendocrine carcinoma, or mixed carcinoma comprising both neuroendocrine-non neuroendocrine was made according to latest World Health Organization Blue Book guidelines.

Immunohistochemistry was performed in all cases. The antibody used were PAX-8 (Clone EP331, Sigma Aldrich, Dilution 1:100), WT-1 (Clone SC06-41, Thermo Fisher Scientific, Dilution 1:250), Pan-CK (Clone AE1, Sigma Aldrich, Dilution 1:100), p53 (Clone DO7, Agilent DAKO, Dilution 1:100), p16 (Clone D25, Sigma Aldrich, Dilution 1:100), Estrogen Receptor (Clone SP1, Thermo Fisher Scientific, Dilution 1:200), Progesterone Receptor (Clone 1E2, Ventana), Synaptophysin (Clone RM258, RevMab Biosciences, Dilution 1:200), Chromogranin (Clone RM385, RevMab Biosciences, Dilution 1:5000), INSM1 (Clone CL11630, Thermo Fisher Scientific, Dilution 1:100), INI-1 (Clone 25/BAF-47, Vitro Master Diagnostica, Dilution 1:50), BRG1 (Clone 1N12, Sigma Aldrich, Dilution 1:100).

In all cases immunohistochemistry for MLH-1 (Clone OT3C1, Thermo Fisher Scientific, Dilution 1:1000), PMS-2 (Clone OTI2G5, Thermo Fisher Scientific, Dilution 1:150), MSH-2 (Clone FE11, Sigma Aldrich, Dilution 1:500), and MSH-6 (Clone JA-8426, Thermo Fisher Scientific, Dilution 1;100) were performed.

Neuroendocrine differentiation was defined by immunohistochemical expression of at least two of the three markers used (synaptophysin, chromogranin, and INSM1).

Semi-quantitative assessment of immunohistochemistry was performed [3].

Negative: No tumor cell staining.

Focally positive: < 10% tumor cells positive.

Patchy positive: 11–50% tumor cells positive.

Diffusely positive: > 50% tumor cells positive.

Results

The summary of the 6 cases has been compiled in Table 1.

Table 1.

Clinical summary of all cases

CASES	Age	Clinical features	Tumor size	Procedure	Histology	NE %	FIGO	Treatment	Follow up
Case 1	58	Bleeding	4.5	TAH + BSO + LND	Endometrioid + SCNC	30	IIIC	Surgery + CT	NED 36 months
Case 2	75	Bleeding	6.5	TAH + BSO + LND	Serous + SCNC	50	IVB	Surgery + CT	Recurrence at 18 months, 45 months
Case 3	51	Abdominal discomfort	4.2	TAH + BSO + LND	Pure SCNC	100	IVB	Surgery + CT	DOD 12 months
Case 4	60	Bleeding	4.3	TAH + BSO + LND	Endometrioid + SCNC	40	IIIC	Surgery + CT	Recurrence at 9 months
Case 5	45	Bleeding	7.5	TAH + BSO + LND	Endometrioid + SCNC	60	IVB	Surgery + CT	Recurrence at 6 months
Case 6	53	Bleeding	3.0	TAH + BSO + LND	Serous + LCNC	45	IVB	Surgery + CT	Metastasis to bone (distal femur) after 1.5 years

N/A, not available; TAH + BSO, total abdominal hysterectomy + bilateral salpingo-oophorectomy; SCNC, small cell neuroendocrine carcinoma; LCNC, large cell neuroendocrine carcinoma; CT, chemotherapy; NED, no evidence of disease; DOD, dead of disease

The mean age at diagnosis is 57 years (45–77 years). Most patients presented with bleeding and abdominal discomfort.

Tumor size ranged from 2.6 to 7.5 cm.

Of the six patients, one was a consult case where surgery of five patients was undergone in our hospital.

Total abdominal hysterectomy with bilateral salpingectomy and lymph node dissection was performed in all six cases.

Tumor tissue was fixed in 10% neutral buffered formalin, routinely processed, and paraffin-embedded. Five microns sections were cut for H&E staining and immunohistochemistry. Immunohistochemistry was performed on the Ventana Benchmark autostainer using various antibodies.

Morphology showed three cases of mixed endometrioid and neuroendocrine carcinoma (50%), two cases of mixed serous and neuroendocrine (33.3%), and 1 case of pure neuroendocrine carcinoma (16.7%).

Mixed Endometrioid and Neuroendocrine Tumor

Endometrioid carcinoma showed grade 2 and grade 3 morphology (Fig. 1a, b) with back-to-back glandular pattern and high-grade nuclear cytology along with neuroendocrine component (Fig. 1c, d).

Fig. 1.

a H&E (200 ×) shows mixed component of tumors, left upper part shows tumor cells in glandular architecture and the right lower part shows sheets of large monomorphic tumor cells. b H&E (400 ×) shows higher magnification showing endometrioid carcinoma component with closely packed glands having high-grade cytology. c H&E (200 ×) shows tumor cells in nests. d H&E (400 ×) shows a higher magnification area showing sheets of monomorphic cells with large cells and small cells intermixed and having fine chromatin

Estrogen Receptor in the endometrioid carcinoma component is diffusely strongly positive (Fig. 2a). Pan-cytokeratin shows dot-like and cytoplasmic strong positivity (Fig. 2b). Synaptophysin is diffusely positive. Many cells show dot-like positivity (Fig. 2c). Chromogranin shows dot-like and cytoplasmic positivity (Fig. 2d).

Fig. 2.

Immunohistochemistry for a Estrogen Receptor (ER) in endometrioid carcinoma component is diffusely strong positive. b Pan-cytokeratin shows dot-like and cytoplasmic strong positivity. c Synaptophysin is diffusely positive. Many cells show dot-like positivity. d Chromogranin shows dot-like and cytoplasmic positivity

Mixed Serous and Neuroendocrine Tumor

Serous carcinoma was a high-grade morphology comprising of short thick papillae with high-grade nuclear features, brisk mitosis, and necrosis (Fig. 3a).

Fig. 3.

a H&E (200 ×) shows mixed component of tumors comprising of glandular and papillary architecture and nests and sheets of small round cell tumor. b H&E (20 ×) shows large nests of tumor cells with Homer-Wright pseudorosette in between. c H&E (10 ×) shows neuroendocrine component comprising of tumor cells in ribbon-like architecture and in nests. d Higher magnification (40 ×) shows small round cell tumor with clear cytoplasm & salt and pepper chromatin

H&E shows mixed components of tumors comprising glandular and papillary architecture and nests and sheets of small round cell tumor cells with Homer-Wright pseudorosette in between (Fig. 3b–d).

Pan-cytokeratin is diffusely positive in the serous carcinoma component (Fig. 4a left upper) while the neuroendocrine component shows dot-like and cytoplasmic staining (Fig. 4a right lower). p16 is diffusely positive in both serous carcinoma and neuroendocrine carcinoma component (Fig. 4b). INSM1 is positive in neuroendocrine component (left side) while negative in serous carcinoma component (Fig. 4c). Synaptophysin is diffusely positive in neuroendocrine component (Fig. 4d).

Fig. 4.

Immunohistochemistry for a Pan-cytokeratin is diffusely positive in serous carcinoma component (left lower) while neuroendocrine component shows dot-like and cytoplasmic staining. b p16 is diffusely positive in both serous carcinoma and neuroendocrine carcinoma component. c INSM1 is positive in neuroendocrine component (left side) while negative in serous carcinoma component. d Synaptophysin is diffusely positive in neuroendocrine component

Pure Neuroendocrine Tumor

Neuroendocrine carcinoma showed two morphology-small cell and large cell types. Small cell neuroendocrine carcinoma is composed of sheets of atypical cells with a high N:C ratio, scant cytoplasm, fine-stippled chromatin, and brisk mitotic activity. Homer-wright pseudorosettes were seen. Large cell neuroendocrine morphology showed large atypical cells in nests and sheets with vesicular chromatin and prominent nucleoli and brisk mitosis. Trabecular and insular patterns were also noted (Fig. 5a–d).

Fig. 5.

Curettage from bone a H&E (200 ×) showing sheets of small round to oval blue cells, b higher magnification (400 ×) showing tumor cells have scant to moderate cytoplasm, hyperchromatic nuclei. c Review from our previous TAH + BSO Sect. (200 ×) revealed similar morphology with sheets of small round to oval blue cells. Spindling of cells is also seen. d Other areas (400 ×) show nests of tumor cells with large cells having moderate cytoplasm, round to oval nuclei

Pan-cytokeratin shows dot-like positivity (Fig. 6a). Synaptophysin shows patchy cytoplasmic positivity (Fig. 6b).p53 shows more than 90% cells strong intensely positive (Fig. 6c).SMARCA4 (BRG1) shows retained nuclear positivity in tumor cells (Fig. 6d).

Fig. 6.

a Pan-cytokeratin shows dot-like positivity. b Synaptophysin shows patchy cytoplasmic positivity. c p53 shows more than 90% tumor cells are intensely positive. d SMARCA4 (BRG1) shows retained nuclear positivity in tumor cells

The immunohistochemistry of all the cases has been summarized in Table 2.

Table 2.

Immunohistochemistry findings of the cases

CASES	PAX8	CK	WT1	p53	p16	ER	PR	Synapto	Chromo	INSM1	TTF-1	INI-1	BRG-1
Case 1	Diffuse positive in endometrioid component, Negative in neuroendocrine component	Pos	Neg	Wild type (endometrioid), Mutant (SCNC)	Pos	Neg	Neg	Pos (D)	Pos (P)	Pos	Neg	Ret	Ret
Case 2	Diffuse positive in serous component, Patchy positive in SCNC component	Pos	Pos	Mutant (Both component)	Pos (F)	Neg	Neg	Pos (D)	Pos (P)	Pos	Neg	Ret	Ret
Case 3	Patchy positive	Pos	Neg	Mutant	Pos	Pos (F)	Neg	Pos (D)	Pos (D)	Pos	Neg	Ret	Ret
Case 4	Diffuse positive in endometrioid component, Negative in SCNC component	Pos	Pos	Wild type (Both component)	Pos	Neg	Neg	Pos (D)	Pos (D)	Pos	Pos (F)	Ret	Ret
Case 5	Diffuse positive in endometrioid component, Negative in SCNC component	Pos	Pos	Wild type (endometrioid), Mutant (SCNC)	Pos	Neg	Neg	Pos (F)	Neg	Pos	Neg	Ret	Ret
Case 6	Diffuse positive in serous component, Negative in LCNC component	Pos	Pos	Mutant (Both component)	Pos	Neg	Pos (F)	Pos (D)	Pos (P)	Pos	Neg	Ret	Ret

Synapto, synaptophysin; Chromo, chromogranin; Neg, negative; Pos, positive; F, focal; P, patchy; D, diffuse; Ret, retained; S, scattered cells; ER, Estrogen Receptor; PR, Progesterone Receptor, SCNC- Small cell neuroendocrine carcinoma, LCNC- Large cell neuroendocrine carcinoma

The neuroendocrine component was immunopositive for at least two of the three neuroendocrine markers (Synaptophysin, Chromogranin, and INSM-1). Synaptophysin expression was seen in all cases (100%), Chromogranin in 5 cases (83.3%), and INSM-1 in all cases (100%).

Pan-cytokeratin showed dot-like and cytoplasmic positivity in all cases.

PAX-8 was diffusely positive in endometrioid and serous components while it was patchy positive in neuroendocrine components in only 2 cases. p16 was block-like positive in all cases. p53 showed diffuse strong immunoreactivity in neuroendocrine components in five cases which were called mutant (83.3% cases). INI-1 and BRG-1 were performed in all cases and all cases showed retained expression.

Two cases have shown loss of MLH-1 and PMS-2 expression. MMR protein immunohistochemistry (MLH-1, PMS-2, MSH-2, and MSH-6) was performed in the neuroendocrine component. Two cases have shown loss of MLH-1 and PMS-2 in tumor cells. One case had the morphology of small-cell neuroendocrine carcinoma while the other had the morphology of large-cell neuroendocrine carcinoma. Both patients were evaluated for Lynch syndrome. One patient had her father’s history of colon carcinoma and her sister had breast carcinoma. The other patient did not meet the criteria for Lynch syndrome.

POLE gene mutation testing was performed in all cases and was negative in all the cases.

FIGO Staging was as follows:

Stage IIIC–2 cases, Stage IVB–4 cases.

Treatment

All the patients were treated with surgery and post-operative chemotherapy.

Total abdominal hysterectomy with bilateral salpingo-oophorectomy and lymph node dissection was performed in all six cases.

Post-operative chemotherapy with cisplatin/carboplatin and etoposide was given for 4–6 cycles. Patients with recurrence or poor response to this first-line therapy were treated with 2nd line therapy comprising of irinotecan. Radiotherapy was not given to any of the patients.

Despite systemic chemotherapy, one patient died of the disease.

Discussion

As we go through the literature research on neuroendocrine tumors, they are rare and obviously show aggressive behavior with high recurrence rate and metastasis.

Pure neuroendocrine carcinomas are very aggressive with extremely poor prognosis and the median survival rate is 17 months which is worse compared to mixed endometrioid/serous-neuroendocrine tumors [10, 11].

We have seen in our six cases, the mean age was 57 years and the most common age group was in the 6th decade. Huntsman DG et al. have shown in their study that the most common age group was the 6th decade with the mean age group in their study being 57.4 years [11].

Vaginal bleeding was the most common clinical finding. The mean size of the tumor is 5 cm which is slightly less than described in one study which was 6 cm.

The most common histopathology described was small cell carcinoma type mixed with endometrioid type. Many previous studies have also shown that small-cell neuroendocrine carcinoma is much more prevalent than the other variants [12, 13].

The most common differential diagnosis while working up a high-grade small round cell tumor is undifferentiated/dedifferentiated carcinoma, carcinosarcoma, or Ewing sarcoma [14, 15]. The most useful immunohistochemistry are Pan-cytokeratin, synaptophysin, chromogranin, and INSM-1 to diagnose neuroendocrine carcinoma.

Three neuroendocrine markers we have used are synaptophysin, chromogranin, and INSM-1. We have not included CD56 in our panel as it is a very non-specific marker as it is expressed in many other sarcomas [16–18]. Instead, we used INSM-1 which has superior specificity [19–21]. In our case series, we have seen that INSM-1 was positive in 100% of cases. Our study highlights the utility of INSM-1 as a neuroendocrine marker in neuroendocrine carcinomas of endometrium. Synaptophysin and chromogranin were the other markers which has good sensitivity and specificity as described in literature studies [3, 4, 7].

None of the cases showed spindle cell sarcomatous areas thus the possibility of carcinosarcoma was excluded.

SMARCA-4 (BRG-1) and SMARCB-1 (INI-1) immunohistochemistry were performed in all the cases however all the tumors showed retained nuclear expression. However many undifferentiated/dedifferentiated endometrial carcinomas can show INI-1/BRG-1 retained expression and focal neuroendocrine immunopositivity [22, 23], it is sometimes difficult to differentiate these entities completely especially large cell neuroendocrine carcinoma and undifferentiated carcinoma where it is arbitrary sometimes. Some authors have proposed that at least two neuroendocrine markers should be positive in > 20% of tumor cells [24, 25]. However, diffuse immunopositivity for neuroendocrine markers with stippled chromatin and dot-like CK positivity favors neuroendocrine carcinoma. As in our case series, most of the neuroendocrine markers were diffusely immunopositive with retained INI-1/BRG-1 expression. Thus confirming the diagnosis of neuroendocrine carcinoma.

p16 showed diffuse positivity in all cases. Three cases were sent for HPV DNA testing however only one case had shown HPV DNA positive and the rest all were negative. This is one of the comprehensive studies of p16 expression in neuroendocrine carcinoma. p16 expression in neuroendocrine carcinomas has been described [3, 26]. In one of the largest series by Pocrnich CE et al. [6] of 25 cases of neuroendocrine carcinomas of the endometrium, more than half of the cases were p16 focally positive, and the rest diffusely positive. So the authors have concluded that focal p16 may indicate endometrial origin more. However, our study has shown slightly different results with 100% of cases showing diffuse strong p16 positivity. This discordant result may be due to the limited sample size of our study.

Estrogen Receptor and Progesterone Receptor immunohistochemistry were negative in neuroendocrine components in most of the cases. However endometrioid and high-grade serous components showed diffuse positivity.

MMR abnormality has been documented in previous studies of undifferentiated carcinomas [24, 27]. MMR abnormality in neuroendocrine carcinomas has also been comprehensively described in the literature [6]. In our study, we have witnessed two cases where MMR deficiency was identified by immunohistochemistry.

Ewing sarcoma /PNET (Primitive neuroectodermal tumor) is another differential diagnosis when encountering a pure neuroendocrine-like component with Homer Wright pseudorosette with small round blue cells. However, Ewing sarcoma shows a fibrillary background with usually monomorphic round to oval cells having scant cytoplasm. Ewing sarcomas show diffuse membranous immunoreactivity for CD99 and NKX2.2. To add to the confusion, Ewing sarcomas can also be positive for synaptophysin, INSM1, and chromogranin however chromogranin expression is very limited in Ewing sarcoma [28]. CD99 and NKX2.2 both can be positive in neuroendocrine tumors [29, 30]. Pan-cytokeratin dot-like positivity and Chromogranin positivity will favor neuroendocrine carcinoma because in some instances CK shows diffuse positivity in Ewing/Ewing-like sarcomas [31].

From the published case reports and series described in the literature, the ratio of advanced to early-stage disease is 1.8:1 [3, 32–36]. Another study by Atienza-Amores M et al. has shown that the ratio of advanced to early-stage disease is 1.3:1 [37]. In our limited number of six cases, all the patients were in an advanced stage.

One of the largest multi-institutional studies in Japan has shown that Stage III-IV disease and pure small-cell neuroendocrine carcinoma are associated with extremely poor prognosis [34]. Few case reports of neuroendocrine carcinoma with longer survival have also been described with surgery and additional chemotherapy and radiotherapy [38, 39].

The largest cohort study of 170 cases by Zhang J et al. has shown that surgery is the mainstay of treatment of neuroendocrine carcinoma [40]. Early-stage disease with individual post-operative therapy with chemotherapy or radiotherapy may improve overall survival. In advanced-stage disease, post-operative adjuvant therapy may improve overall survival [41].

To conclude, neuroendocrine carcinoma, either mixed with endometrial carcinoma or pure, mostly presents with advanced stage and bears poor prognosis. A multidisciplinary approach with gynecological surgery, pathology, and medical oncology may aid in the diagnosis and management of these aggressive tumors.

Abbreviations

FIGO

International Federation of Gynecology and Obstetrics

INSM-1

Insulinoma-associated protein 1

NET

Neuroendocrine tumor

N:C

Nucleus cytoplasmic ratio

PAX-8

Paired box gene-8

WT-1

Wilms’ tumor-1

Pan-CK

Pan-cytokeratin

INI-1

Integrase interactor-1

SMARCB-1

SWI/SNF related, matrix associated, actin dependent regulator of chromatin subfamily B, member 1

BRG-1

Brahma-related gene-1

SMARCA-4

SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 4

MLH-1

MutL homolog-1

PMS-2

Postmeiotic segregation increased-2

MSH-2

MutS homolog-2

MSH-6

MutS homolog-6

H&E

Hematoxylin and Eosin

CD56

Cluster differentiation 56/Neural cell adhesion molecule (NCAM)

HPV DNA

Human papilloma virus Deoxyribonucleic acid

CD99

Cluster differentiation 99

NKX2.2

NK2 Homeobox-2

Author Contribution

Conceptualization: SD, SA. Methodology: SD, AKP, JS. Formal analysis and investigation: SD, PJ, RF, AJ. Writing—original draft preparation: SD, AKP. Writing—review and editing: SD, JS. Resources: SA. Supervision: SA, RJ.

Declarations

Informed Consent

Appropriate informed consent taken.

Conflict of Interest

The authors declare no competing interests.

Study Specific Approval by the Ethics Committee

Not applicable.

Welfare of Animals if the Research Involved Animals

Not applicable.