Introduction
Gastric hepatoid adenocarcinoma was reported for the first time by Ishikura et al. [1, 2, 3, 4]. Subsequently, Matsunoh et al. proposed the gastric carcinoma with enteroblastic differentiation [5]. Nowadays, AFP-producing gastric adenocarcinoma is histologically classified into three categories, namely yolk sac, hepatoid, and enteroblastic types [6]. Among them, enteroblastic type seems to be associated with hepatoid type in some tumours. Gastric adenocarcinoma with hepatoid or enteroblastic differentiation (GAHED) is a rare neoplasm, and reported cases have been limited to date. In this article, we performed a clinicopathological study of 10 cases with GAHED.
Material and methods
This study was approved by the ethical committee of Kochi Red Cross Hospital (No.238). Two hundred and nineteen cases were selected from pathology files of surgically resected gastric cancers by partial or total gastrectomy from January 2011 to December 2015 in my hospital. The patients consisted of 10 cases with GAHED and 209 cases without GAHED. Surgically resected gastric tissues for light microscopy were fixed in 10% buffered formalin, processed by conventional methods, and stained with haematoxylin and eosin stain and periodic acid-Schiff stain with diastase pretreatment (d-PAS) for the identification of hyaline globules. Immunohistochemistry was performed using an Ventana Benchmark Ultra autostainer (Roche). Antibodies against AFP (C3, 1 : 50, Novocastra Laboratories Ltd, Newcastle, UK), Hep Par1(Hepatocyte) (OCHI1E5, 1 : 200, DAKO, Glostrup, Denmark), Glypican-3 (IG12, pre-diluted, Nichirei, Tokyo, Japan) and SALL4 (6E3, prediluted, Roche, Tokyo, Japan), D2-40 (D2-40, 1 : 50, DAKO, Glostrup, Denmark), HER2 (4B5, pre-diluted, Roche, Tokyo, Japan) were employed in the present study. EBER in situ hybridisation (Roche, Tokyo, Japan) was performed in all tumours. Appropriate positive control specimens were used as positive controls for immunohistochemistry and EBER in situ hybridisation. Clinicopathological features (sex, gross findings, lymphatic invasion, and venous invasion) of the GAHED and non-GAHED groups were investigated and compared with each other, based on pathologic reports and clinical information from electronic medical reports. The gross findings were subdivided into five categories, namely Type 0 (superficial), 1 (protruding), 2 (ulcerative and localised), 3 (ulcerative and infiltrating type), 4 (diffusely infiltrating), and 5 (unclassified). Type 0 (superficial) was divided into three categories, namely 0-I (superficial and protruding), 0-II (superficial and flat), and 0-III (superficial and encavated). The frequency of metastasis to lymph node, liver, lung, peritoneum, and bone was compared between both groups within the time from initial diagnosis to end of follow-up. The disease-free survival (DFS) of the GAHED group was also compared with that of the non-GAHED group using Kaplan-Meier method and the long-rank test. All p values were two sided and p < 0.05 was considered to be significant. The statistical analysis was performed with BellCurve (Tokyo, Japan). Fluorescence in situ hybridisation (FISH) of the HER2 gene was performed according to the manufacturer’s protocol in two cases. Pretreatment was carried out using a VP-2000 Processor (Abbott Molecular, Tokyo, Japan), and hybridisation was performed using ThermoBrite (Abbott Molecular Tokyo, Japan). FISH signals were observed using an Axio Imager 2 Upright Microscope (Zeiss, Tokyo, Japan) and analysed using MetaCyte Lite (MetaSystems, Altlussheim, Germany) with Isis (Zeiss). When the ratio of HER2/CEP17 was more than 2.0, we considered it to be positive for HER2 gene amplification.
Results
The clinical features of 10 cases with GAHED are summarised in Table I. The patients consisted of nine males and one female. The age of patients ranged from 52 to 82 years with a mean age of 69.5 years. The macroscopic characteristics of the 10 cases with GAHED are shown in Table II. Grossly, the tumours consisted of two early types and eight advanced types. The tumour size varied from 28 mm to 106 mm with a mean size of 52.9 mm. The microscopic features of 10 cases with GAHED are demonstrated in Table III. Histologically, all tumours were composed of various proportions of tubular (Fig. 1A), cribriform, papillary (Fig. 1B), solid, and/or trabecular (Fig. 1C) growth patterns of clear to slightly eosinophilic tumour cells. Evident hepatoid differentiation was characterised by trabecular architecture in one case (No. 10). In this tumour, giant cells were also observed. The rate of hepatoid or enteroblastic differentiation in the entire neoplastic volume ranged from 10 to 100% with a mean rate of 45%. Hyaline globules were identified in all tumours and highlighted by d-PAS stain (Fig. 1D). A summary of the immunohistochemical results is shown in Table IV. Immunohistochemically, AFP and Hep-Par1 were positive in all tumours. Glypican-3 and SALL4 were positive in 2 and 8 tumours, respectively. Interestingly, evident hepatoid carcinoma (No. 10) showed diffuse positivity for AFP (Fig. 2A), Hep Par1, Glypican-3, and SALL4 (Fig. 2B). By contrast, enteroblastic carcinoma usually showed focal positivity for AFP (Fig. 2C), Hep Par1, and SALL4 (Fig. 2D). No tumours expressed EBER. The comparison of lymphovascular invasion between both groups in the histological evaluation is shown in Table V. Lymphatic invasion more frequently occurred in the GAHED group than in the non-GAHED group (Fig. 3A), but there was statistically no significant difference of sex ratio, gross findings, and vascular invasion between both groups. Lymphatic invasion of carcinoma cells was highlighted by D2-40 immunostaining (Fig. 3B). Comparison of the metastasis to various anatomic sites is also summarised in Table V. The metastasis to lymph node, liver, or lung was more frequent in the GAHED group than in the non-GAHED group, but the metastasis to peritoneum or bone was not significantly different between both groups. Metastasis to lymph node exceeding 3 cm in maximum diameter was identified in three cases. The clinical follow-up was available in nine cases with GAHED and 189 cases with non-GAHED. The follow-up duration of the GAHED group ranged from 1 to 60 months with a mean duration of 32.4 months. Regarding the clinical outcome, 3 and 6 patients were alive without disease and alive with disease, respectively (Table I). The comparison of DFS in both groups is shown in Fig. 4. In a statistical analysis, the DFS of the GAHED group was significantly worse than that of the non-GAHED group. HER2 overexpression (3+) was identified in three tumours (Fig. 5A). A FSIH study of HER2 gene /chromosome 17 was performed in two cases (No. 6 and 7). The ratio for each case was 2.9 and 9.5, respectively, in 370 and 414 counting cells. Both cases were considered to be positive for HER2 gene amplification (Fig. 5B).
Discussion
Grossly, GAHED seems to be distinguished from usual gastric adenocarcinoma, based on the results of the present study. GAHED is morphologically characterised by various proportions of tubular, cribriform, papillary, solid, and/or trabecular growth patterns of clear to slightly eosinophilic tumour cells and intracytoplasmic possession of hyaline globules. Motoyama et al. divided AFP-producing gastric cancer into three categories, namely hepatoid, yolk sac tumour-like, and foetal gastrointestinal types [6]. However, it seems to be very difficult to clearly distinguish hepatoid differentiation from enteroblastic differentiation because both tumours have a capacity to produce AFP and some tumours admixed with each other have been describe previously. We confirmed no tumours with yolk sac tumour-like differentiation in the present tumour. Therefore, we unified these differentiations into GAHED. The elevation of the serum AFP may become a vital clue in pathological diagnosis [1, 2]. However, we found in the present study that clinicians did not measure serum AFP value in most cases. Hence, pathologists should bear in mind that the abovementioned histological features can lead to the pathological diagnosis of GAHED. It is likely that hyaline globules may be an important diagnostic clue. Whenever pathologists encounter cases suspicious for GAHED, they should try the immunostaining of AFP, Hep Par1, Glypican-3, and/or SALL4 to confirm a definite diagnosis [7, 8, 9]. This tumour seems to be associated with intestinal phenotype [1, 4, 10]. Based on the present study results, we particularly recommend the combined immunohistochemical panel of SALL4 as well as AFP and Hep Par1 for the confirmation of GAHED diagnosis because Hep Par1 usually expresses in the intestinal phenotype irrespective of hepatoid or enteroblastic differentiation, and Glypican-3 is low sensitivity (20%) of GAHED diagnosis.
In the present study, the GAHED group had a greater propensity for lymphatic invasion than the non-GAHED group, but there was no significant difference in vascular invasion between both groups. The result of lymphatic invasion is consistent with the previous report [11]. By contrast, the result of vascular invasion is not compatible with the previous result [3, 11].
Previously, the propensity to metastasise to lymph nodes and liver has been frequently described [2, 7, 11, 12]. The frequency of lung metastasis in GAHED accounts for 3.4% and is relatively low [7]. However, this is the first report showing the propensity of metastasis to lung in the GAHED group, compared with the non-GAHED group.
This tumour may have the potential to behave in an aggressive clinical fashion [2, 3, 4, 7, 8, 13, 14, 15]. The results of disease-free survival of GAHED in the present study support the previous descriptions. Motoyama et al. and Hwang et al. reported a case of gastric adenocarcinoma with choriocarcinomatous and hepatoid differentiation [6, 16]. They suggest that choriocarcinoma or hepatoid carcinoma may show the retrodifferentiation related to the unfavourable pathological differentiation [16]. Gastric neuroendocrine carcinoma also has a propensity to behave in an aggressive clinical fashion [17]. Therefore, neuroendocrine differentiation may be one of retrodifferentiation. However, there was no case with neuroendocrine differentiation in the present study.
To the best of our knowledge, there were no descriptions of amplification of the HER2 gene in GAHED. Therefore, this is the first report on this topic. This means that GAHED with advanced stage may be effective for molecular targeted therapy for HER2 gene. From a practical point of view, this result could supply the available information to digestive tract clinicians and patients suffering from GAHED. However, the number of patients in the present study is too small to come to a definite conclusion. Further examination in a large-scale study is required in order to elucidate the utility of molecular targeted therapy for GAHED.
In conclusion, GAHED is histologically characterised by various proportions of tubular, cribriform, papillary, solid, and/or trabecular growth patterns and intracytoplasmic possession of hyaline globules. Immunohistochemical identification of SALL4 as well as AFP and Hep Par1 is useful for the definite diagnosis of GAHED. The tumour of GAHED has a propensity of lymphatic invasion and metastasis to lymph nodes, liver, and lung, compared with the non-GAHED group. Therefore, this tumour usually behaves in an aggressive clinical fashion.
The authors declare no conflicts of interests.
References
1. Ishikura H, Fukasawa Y, Ogasawara K, et al. An AFP-producing gastric carcinoma with features of hepatic differentiation. Cancer 1985; 56: 840-848.
2. Ishikura H. Kirimoto K, Shamoto M, et al. Hepatoid adenocarcinoma of stomach. Cancer 1986; 58: 119-126.
3. Ishikura H, Kishimoto T, Andachi H, et al. Gastrointestinal hepatoid adenocarcinoma: venous permeation and mimicry of hepatocellular carcinoma, a report of four cases. Histopathology 1997; 31: 47-54.
4. Kishimoto T, Nagai Y, Kato K, et al. Hepatoid adenocarcinoma: a new clinicopathological entity and the hypothesis on carcinogenesis. Med Electron Microsc 2000; 33: 57-63.
5. Matsunou H, Konishi F, Jalal RE, et al. Alpha-fetoprotein-producing gastric carcinoma with enteroblastic differentiation. Cancer 1994; 73: 534-540.
6. Motoyama T, Aizawa K, Watanabe H, et al. Alpha-fetoprotein-producing gastric carcinomas: a comparative study of three different subtypes. Acta Pathol Jpn 1993; 43: 654-661.
7. Su JS, Chen YT, Wang RC, et al. Clinicopathological characteristics in the differential diagnosis of hepatoid adenocarcinoma: A literature review. World J Gastroenterol 2013; 19: 321-327.
8. Roberts CC, Colby TV, Batts KP. Carcinoma of the stomach with hepatocyte differentiation (hepatoid adenocarcinoma). Mayo Clin Proc 1997; 72: 1154-1160.
9. Ushiku T, Shiozaki A, Shibahara J, et al. SALL4 represents fetal gut differentiation of gastric cancer, and is diagnostically useful in distinguishing hepatoid gastric hepatoid gastric carcinoma from hepatocellular carcinoma. Am J Surg Pathol 2010; 34: 533-540.
10. Gálvez-Muñoz E, Gallego-Plazas J, Gonzalez-Orozco V, et al. Hepatoid adenocarcinoma of the stomach- a different histology for not so different gastric adenocarcinoma: a case report. Int Sem Surg Oncol 2009l; 6: 13.
11. Murakami T, Yao T, Mitomi H, et al. Clinicopathological and immunohistochemical characteristics of gastric adenocarcinoma with enteroblastic differentiation: a study of 29 cases. Gastric Cancer 2016; 19: 498-507.
12. deLorimier A, Park F, Aranha GV, et al. Hepatoid carcinoma of stomach. Cancer 1993; 71: 293-296.
13. Matsumoto K, Ueyama H, Matsumoto K, et al. Clinicopathological features of alpha-fetoprotein producing early gastric cancer with enteroblastic differentiation. World J Gastroenterol 2016; 22: 8203-8210.
14. Yang J, Wang R, Zhang W, et al. Clinicopathological and prognostic characteristics of hepatoid adenocarcinoma of the stomach. Gastroenterol Res Pract 2014; 2014: 140587.
15. Kinjo T, Taniguchi H, Kushima R, et al. Histologic and immunohistochemical analyses of alpha-fetoprotein-producing cancer of the stomach. Am J Surg Pathol 2012; 36: 56-65.
16. Hwang SW, Lee SJ, Park PE, et al. Gastric adenocarcinoma with choriocarcinomatous and hepatoid differentiation. Basic Appl Pathol 2012; 5: 22-25.
17. Deng Y, Chen X, Ye Y, et al. Histological characterization and prognostic evaluation of 62 gastric neuroendocrine carcinomas. Contemp Oncol (Pozn) 2016; 20: 311-319.
Address for correspondence
Naoto Kuroda
Department of Pathology
Kochi Red Cross Hospital
Shin-honmachi 2-13-51,
Kochi City, Kochi 780-8562, Japan
e-mail: kurochankochi@yahoo.co.jp
