Prognostic significance of tumor budding and single cell invasion in gastric adenocarcinoma

Figures & data

Table 1 Associations between invasion types and clinicopathological characteristics of 296 patients with gastric adenocarcinoma

Variables	Tumor budding			Single cell invasion			Large cell invasion
	(−)	(+)	P-value	(−)	(+)	P-value	(−)	(+)	P-value
Age (years)			0.362			0.690			0.164
≤60	67 (22.6%)	72 (24.3%)		50 (16.9%)	89 (30.1%)		120 (40.5%)	19 (6.4%)
>60	84 (28.4%)	73 (24.7%)		60 (20.3%)	97 (32.8%)		126 (42.6%)	31 (10.5%)
Sex			0.441			0.640			0.180
Male	113 (38.2%)	114 (38.5%)		86 (29.1%)	141 (47.6%)		185 (62.5%)	42 (14.2%)
Female	38 (12.8%)	31 (10.5%)		24 (8.1%)	45 (15.2%)		61 (20.6%)	8 (2.7%)
T classification			<0.001			<0.001			0.005
T1 + T2	63 (21.3%)	21 (7.1%)		55 (18.6%)	29 (9.8%)		78 (26.4%)	6 (2.0%)
T3 + T4	88 (29.7%)	124 (41.9%)		55 (18.6%)	157 (53.0%)		168 (56.8%)	44 (14.9%)
N classification			<0.001			<0.001			0.002
N0	66 (22.3%)	31 (10.5%)		60 (20.3%)	37 (12.5%)		90 (30.4%)	7 (2.4%)
N1 + N2 + N3	85 (28.7%)	114 (38.5%)		50 (16.9%)	149 (50.3%)		156 (52.7%)	43 (14.5%)
M classification			0.005			0.001			<0.001
M0	127 (42.9%)	102 (34.5%)		97 (32.8%)	132 (44.6%)		201 (67.9%)	28 (9.5%)
M1	24 (8.1%)	43 (14.5%)		13 (4.4%)	54 (18.2%)		45 (15.2%)	22 (7.4%)
Pathological stage			<0.001			<0.001			0.003
Stage I + II	77 (26.0%)	30 (10.1%)		68 (23.0%)	39 (13.2%)		98 (33.1%)	9 (3.0%)
Stage III + IV	74 (25.0%)	115 (38.9%)		42 (14.2%)	147 (49.7%)		148 (50.0%)	41 (13.9%)
Tumor differentiation degree			<0.001			<0.001			0.075
Grade I + I–II	76 (25.7%)	123 (41.6%)		52 (17.6%)	147 (49.7%)		160 (54.1%)	39 (13.2%)
Grade II + III	75 (25.3%)	22 (7.4%)		58 (19.6%)	39 (13.2%)		86 (29.1%)	11 (3.7%)
Tumor budding						<0.001			<0.001
(−)	–	–		96 (32.4%)	55 (18.6%)		137 (46.3%)	14 (4.7%)
(+)	–	–		14 (4.7%)	131 (44.3%)		109 (36.8%)	36 (12.2%)
Large cell invasion			<0.001			0.001
(−)	137 (46.3%)	109 (36.8%)		102 (34.5%)	144 (48.6%)		–	–
(+)	14 (4.7%)	36 (12.2%)		8 (2.7%)	42 (14.2%)		–	–
Single cell invasion			<0.001						0.001
(−)	96 (32.4%)	14 (4.7%)		–	–		102 (34.5%)	8 (2.7%)
(+)	55 (18.6%)	131 (44.3%)		–	–		144 (48.6%)	42 (14.2%)
Fibrosis			0.098			0.321			0.022
(+)	63 (21.3%)	69 (23.3%)		47 (15.9%)	85 (28.7%)		103 (34.8%)	29 (9.8%)
(++)	53 (17.9%)	55 (18.6%)		38 (12.8%)	70 (23.6%)		92 (31.1%)	16 (5.4%)
(+++)	35 (11.8%)	21 (7.1%)		25 (8.4%)	31 (10.5%)		51 (17.2%)	5 (1.7%)
Mitosis			<0.001			<0.001			<0.001
(−)	100 (33.8%)	42 (14.2%)		83 (28.0%)	59 (19.9%)		133 (44.9%)	9 (3.0%)
(+)	51 (17.2%)	103 (34.8%)		27 (9.1%)	127 (42.9%)		113 (38.2%)	41 (13.9%)
Smoking or drinking history			0.993			0.300			0.688
Yes	52 (17.6%)	50 (16.9%)		42 (14.2%)	60 (20.3%)		86 (29.1%)	16 (5.4%)
No	99 (33.4%)	95 (32.1%)		68 (23.0%)	126 (42.6%)		160 (54.1%)	34 (11.5%)

Notes: Degree of tumor differentiation: grade I, poorly differentiated; grade II, moderately and poorly differentiated; grade III, moderately differentiated; IV, well differentiated. P<0.05 was considered significant.

Abbreviations: T, tumor; N, node; M, metastasis.

Figure 1 Microscopic observations for gastric adenocarcinoma (HE stain).

Notes: (A) Tumor budding (arrows) was defined as a cluster of tumor cells composed of fewer than five tumor cells and was evaluated at 100× magnification. (B) Tumor budding (arrows) evaluated at 400× magnification. (C) Single cell invasion indicated by arrows at 400× magnification. (D) Large cell invasion was defined as a tumor cell whose nucleus diameter was quadruple the size of a small nearby lymphocyte. Indicated by arrows at 400× magnification. (E) Mitosis indicated by arrows at 400× magnification. (F) Fibrosis was confirmed if the area of fibrosis was >60% of the microscopic field. Evaluated at 100× magnification.
Abbreviation: HE, hematoxylin and eosin.

Figure 2 Overall survival (OS) figures pertaining to tumor budding (A), single cell invasion (B), large cell invasion (C), and mitotic count (D) in gastric adenocarcinoma.

Notes: (A) Kaplan–Meier survival curve for tumor budding in patients with gastric adenocarcinoma. The OS of cases with high-grade tumor budding was shorter than for cases exhibiting low-grade budding (P<0.001). (B) Patients with single cell invasion had shorter OS compared to those without single cell invasion (P<0.001). (C) Patients with large cell invasion had shorter OS compared to those without large cell invasion (P<0.001). (D) Patients with high mitotic count had shorter OS compared to those without high mitotic count (P=0.035).

Figure 3 Survival curves according to different classification systems in gastric adenocarcinoma.

Notes: (A) Kaplan–Meier survival curve of WHO classification in patients with gastric adenocarcinoma. Papillary carcinoma showed statistically significant differences with tubular carcinoma subtype (P=0.001) but not with other subtypes (mucinous carcinoma, P=0.628; signet ring cell carcinoma, P=0.353; low-differentiated carcinoma, P=0.550; and undifferentiated carcinoma, P=0.528). P, T, M, S, L, and U represent papillary carcinoma, tubular carcinoma, mucinous carcinoma, signet ring cell carcinoma, minimally differentiated carcinoma, and undifferentiated carcinoma, respectively. (B) According to the Lauren classification, diffuse-type patients have unfavorable prognosis (P<0.001). I and D represent intestinal-type and diffuse-type gastric adenocarcinoma, respectively. (C) According to the Gosrki classification, the well-differentiated grades I and II had higher survival rates than the poorly differentiated grades III and IV. Grade I showed statistically significant differences compared with grades II, III, and IV (P=0.023, P<0.001, and P=0.002). I: well-differentiated and mucin-poor areas of tumor. II: well-differentiated and mucin-rich parts of the tumor. III: poorly differentiated and mucin-poor parts of the tumor. IV: poorly differentiated and mucin-rich parts of the tumor. (D) According to the new pathological classification, grade I showed statistically significant difference from grades II, III, and IV (P=0.001, P<0.001, and P<0.001). I: patients without tumor budding, single cell invasion, and large cell invasion. II: patients with single cell invasion. III: patients with tumor budding and single cell invasion. IV: patients with tumor budding, single cell invasion, and large cell invasion.
Abbreviation: WHO, World Health Organization.

Table 2 Univariate Cox regression analysis of clinical and pathological characteristics

Variables	HR	95% CI	P-value
Age (years)
≤60	1.000	Reference	–
>60	1.556	1.116–2.168	0.009
Sex
Male	1.000	Reference	–
Female	0.746	0.497–1.121	0.159
T classification
T1	1.000	Reference	<0.001
T2	5.162	1.471–18.117	0.010
T3	10.607	3.320–33.886	<0.001
T4	15.324	4.823–48.683	<0.001
N classification
N0	1.000	Reference	<0.001
N1	2.700	1.417–5.147	0.003
N2	4.234	2.426–7.390	<0.001
N3	6.233	3.750–10.360	<0.001
M classification
M0	1.000	Reference	–
M1	2.844	2.027–3.989	<0.001
Pathological stage
Stage I	1.000	Reference	<0.001
Stage II	2.767	0.961–7.964	0.059
Stage III	10.972	4.432–27.163	<0.001
Stage IV	16.701	6.666–41.845	<0.001
Tumor differentiation degree
Grade I	1.000	Reference	0.110
Grades I–II	0.729	0.484–1.095	0.128
Grade II	0.690	0.470–1.014	0.059
Grade III	0.228	0.032–1.640	0.142
Tumor budding
(−)	1.000	Reference	–
(+)	2.260	1.617–3.159	<0.001
Large cell invasion
(−)	1.000	Reference	–
(+)	2.466	1.700–3.578	<0.001
Single cell invasion
(−)	1.000	Reference	–
(+)	3.553	2.349–5.374	<0.001
Fibrosis
(+)	1.000	Reference	–
(++)	0.767	0.538–1.092	0.141
(+++)	0.517	0.315–0.848	0.009
Mitosis
(−)	1.000	Reference	–
(+)	1.419	1.022–1.971	0.037
Smoking or drinking history
No	1.000	Reference	–
Yes	1.350	0.969–1.881	0.076
WHO classification
Papillary gastric carcinoma	1.000	Reference	0.013
Tubular adenocarcinoma	0.440	0.267–0.726	0.001
Mucinous gastric carcinoma	0.863	0.475–1.567	0.628
Signet ring cell carcinoma	0.748	0.404–1.382	0.353
Low-differentiated adenocarcinoma	0.844	0.485–1.470	0.550
Undifferentiated adenocarcinoma	1.158	1.158–0.739	0.523
Lauren classification
Intestinal type	1.000	Reference	–
Diffuse type	2.563	1.789–3.673	<0.001
Goseki classification
I	1.000	Reference	<0.001
II	1.890	1.093–3.267	0.023
III	2.634	1.684–4.119	<0.001
IV	2.202	1.350–3.594	0.002
Pathological classification
I	1.000	Reference	–
II	2.820	1.568–5.071	0.001
III	3.907	2.375–6.425	<0.001
IV	6.028	3.345–10.865	<0.001

Abbreviations: CI, confidence interval; HR, hazard ratio; WHO, World Health Organization; T, tumor; N, node; M, metastasis.

Table 3 Multivariate Cox regression analysis of clinical and pathological characteristics

Factors	Category	HR	95% CI	P-value
Model for tumor budding
Age (years)	≤60	1.000	Reference	–
	>60	1.578	1.121–2.221	0.009
Large cell invasion	(−)	1.000	Reference	–
	(+)	1.566	1.051–2.333	0.028
Tumor budding	(−)	1.000	Reference	–
	(+)	1.568	1.044–2.354	0.030
Mitosis	(−)	1.000	Reference	–
	(+)	0.878	0.611–1.263	0.484
Pathological stage	I	1.000	Reference	–
	II	2.280	0.786–6.616	0.129
	III	8.855	3.514–22.317	<0.001
	IV	12.699	4.959–32.520	<0.001
Tumor differentiation degree	Grade I	1.000	Reference	–
	Grades I–II	0.827	0.541–1.264	0.380
	Grade II	1.146	0.726–1.810	0.557
	Grade III	0.639	0.085–4.794	0.663
Model of single cell invasion
Age (years)	≤60	1.000	Reference	–
	>60	1.677	1.192–2.358	0.003
Large cell invasion	(−)	1.000	Reference	–
	(+)	1.699	1.145–2.522	0.008
Single cell invasion	(−)	1.000	Reference	–
	(+)	2.385	1.562–3.787	<0.001
Mitosis	(−)	1.000	Reference	–
	(+)	0.789	0.547–1.136	0.203
Pathological stage	I	1.000	Reference	–
	II	2.068	0.708–6.038	0.184
	III	7.237	2.846–18.401	<0.001
	IV	10.259	3.962–26.564	<0.001
Tumor differentiation degree	Grade I	1.000	Reference	–
	Grades I–II	0.802	0.530–1.215	0.298
	Grade II	1.144	0.752–1.740	0.529
	Grade III	0.712	0.095–5.354	0.741

Abbreviations: CI, confidence interval; HR, hazard ratio.