Clinical and pathological characterization of Epstein-Barr virus-associated gastric carcinomas in Portugal

Table 1 Antibodies used in immunohistochemistry

Protein	Primary Antibody	Dilution	Incubation	Retrieval method	Expression in gastric cells
LMP1	NCL-EBV-CS1-4, Leica, Newcastle upon Tyne, United Kingdom	1:100	3h, RT	15 min, microwave in Vector® Antigen Unmasking Solution	Cytoplasm
LMP2A	15F9, THERMO SCIENTIFIC, Fremont, United States	1:250	On, 4°C		Cytoplasm and membrane

LMP1: Latent membrane protein 1; LMP2A: Latent membrane protein 2A; RT: Room temperature; On: Overnight.

Table 2 Patients characteristics and distribution of Epstein-Barr virus in gastric cancer patients n (%)

Characteristics	All cases (n = 179)	EBV status		P value
		Negative (n = 164)	Positive (n = 15)
Gender, n = 179
Male	108 (60.3)	96 (88.9)	12 (11.1)	0.104
Female	71 (51.5)	68 (95.8)	3 (4.2)
Age, n = 179 (64.38 ± 12 yr of age)
< 65 yr of age	90 (50.3)	84 (93.3)	6 (6,7)	0.433
≥ 65 yr of age	89 (49.7)	80 (89.9)	9 (10.1)
Surgical procedure type, n = 179
Total gastrectomy	100 (55.9)	89 (89.0)	11 (11.0)	0.351
Partial gastrectomy	77 (43.0)	73 (94.8)	4 (5.2)
Esophagogastrectomy	2 (1.1)	2 (100)	-
Tumour localization, n = 179				0.087
Upper third	26 (14.5)	22 (84.6)	4 (15.4)
Middle	46 (25.7)	40 (87.0)	6 (13.0)
Lower third	107 (59.8)	102 (95.3)	5 (4.7)
Anatomical site, n = 179
Cardia	21 (11.7)	19 (95.7)	2 (4.3)	0.041
Fundus	5 (2.8)	3 (60.0)	2 (40.0)
Body	46 (25.7)	40 (87.0)	6 (13.0)
Pylorus	13 (7.3)	12 (92.3)	1 (7.7)
Antrum	94 (52.5)	90 (95.7)	4 (4.3)
Invasion pattern, n = 167
Infiltrative	88 (52.7)	83 (94.3)	5 (5.7)	0.237
Expansive	77 (46.1)	67 (87.0)	10 (13.0)
Mixed	2 (1.1)	2 (100)	-
Differentiation, n = 167
Poor	91 (54.5)	84 (92.3)	7 (7.7)	0.724
Well+Moderate	76 (45.5)	69 (90.8)	7 (9.2)
Lymphovascular invasion, n = 179
Positive	103 (57.5)	94 (91.3)	9 (8.7)	0.841
Negative	76 (42.5)	70 (92.1)	6 (7.9)
Histology WHO (2010), n = 179
Papillary adenocarcinoma	1 (0.6)	1 (100)	-	<0.001
Tubular adenocarcinoma	92 (51.4)	84 (91,3)	8 (8.7)
Mucinous adenocarcinoma	5 (2.8)	5 (100)	-
Poorly cohesive carcinoma	51 (28.5)	50 (98.0)	1 (2.0)
Mixed carcinoma	25 (14.0)	23 (92.0)	2 (8.0)
Medullary carcinoma	3 (1.7)	-	3 (100)
Undifferentiated carcinoma	1 (0.6)	1 (100)	-
Adenosquamous carcinoma	1 (0.6)	-	1 (100)
Primary tumour (T), n = 179
T1a	22 (12.3)	22 (100)	-	0.289
T1b	18 (10.1)	15 (83.3)	3 (16.7)
T2	25(14.0)	24 (96.0)	1 (4.0)
T3	69 (38.5)	60 (87.0)	9 (13.0
T4	7 (4.00)	7 (100)	-
T4a	37 (20.7)	35 (94.6)	2 (5.4)
T4b	1 (0.56)	1 (100)	-
Regional lymph nodes (N), n = 179
N0	73 (40.8)	67 (91.8)	6 (8.2)	0.024
N1	23 (12.9)	21 (91.3)	2 (8.7)
N2	31 (17.3)	24 (77.4)	7 (22.6)
N3	8 (4.5)	8 (100)	-
N3a	23 (12.8)	23 (100)	-
N3b	21 (11.7)	21 (100)	-
Distant metastasis (M), n = 179				0.18
M0	148 (82.7)	133 (89.9)	15 (10.1)
M1	12 (6.70)	12 (100)	-
Mx	19 (10.6)	19 (100)	-

P < 0.05 were considered statistical significant. EBV: Epstein-Barr virus; WHO: World Health Organization.

Table 3 Clinicopathological characteristics of Epstein-Barr virus-associated gastric cancers

ID	Gender	Age	Anatomic Site	WHO Classification	Invasion pattern	Differentiation	n	TNM Classification	LMP-1	LMP-2A
1	M	75	Body	Tubular Adenocarcinoma	Expansive	Poorly differentiated	6	T4aN2M0	-	-
2	M	80	Antrum	Tubular Adenocarcinoma	Expansive	Moderately differentiated	0	T3N0M0	-	-
3	M	56	Fundus	Tubular Adenocarcinoma	Expansive	Moderately differentiated	2	T2N1M0	-	-
4	M	80	Antrum	Tubular Adenocarcinoma	Expansive	Moderately differentiated	2	T3N0M0	-	-
5	M	64	Body	Tubular Adenocarcinoma	Expansive	Moderately differentiated	0	T3N2M0	-	(+)
6	M	56	Fundus	Tubular Adenocarcinoma	Expansive	Moderately differentiated	0	T3N0M0	-	-
7	M	69	Body	Tubular Adenocarcinoma	Expansive	Moderately differentiated	3	T3N2M0	-	Inc.
8	M	82	Cardia	Tubular Adenocarcinoma	Infiltrative	Moderately differentiated	4	T3N2M0	-	(+)
9	M	55	Body	Mixed Carcinoma	Infiltrative	Poorly differentiated	3	T3N2M0	-	(+)
10	F	66	Antrum	Mixed Carcinoma	NA	Poorly differentiated	6	T1bN2M0	-	(+)
11	F	68	Body	Medullary Carcinoma	Expansive	Poorly differentiated	1	T1bN1M0	-	-
12	M	52	Cardia	Medullary Carcinoma	Expansive	Poorly differentiated	0	T1bN0M0	-	-
13	M	44	Antrum	Medullary Carcinoma	Infiltrative	Poorly differentiated	0	T3N0M0	-	(+)
14	F	65	Pylorus	Poorly cohesive carcinoma	Infiltrative	Poorly differentiated	0	T4aN0M0	-	-
15	M	76	Body	Adenosquamous carcinoma	Expansive	NA	6	T3N2M0	-	(+)

N: Number of regional lymph nodes with metastasis; NA: Not available; (-): Negative result; (+): Positive result; Inc.: Inconclusive result; LMP1: Latent Membrane Protein 1; Latent Membrane Protein.