Better prognosis of gastric cancer patients with high levels of tumor infiltrating lymphocytes is counteracted by PD-1 expression

Figures & data

Figure 1. Immunohistological staining for CD3⁺, CD4⁺, and CD8⁺ cells in 400-fold magnification.

Tumor samples were grouped into a low or high state of infiltration with CD3, CD4, or CD8 positive TILs by using the median.

Figure 2. Immunohistochemical staining for PD-1 and PD-L1 expression in tumor cells and TILs in 400fold (b,d,e) and 20fold (a,c,f) magnification.

Samples were grouped into negative or positive with a cutoff value of >1% expression or with CPS > 1.

Figure 3. (a) Overall survival depending on high CD3 positive lymphocyte infiltration state. (b) Overall survival depending on PD-1 expression on tumor-infiltrating lymphocytes. (c) Overall survival depending on high CD3 positive lymphocyte infiltration state and UICC stage. (d) Overall survival depending on PD-1 expression on TILs and on CD3 positive lymphocyte infiltration state. (e) Overall survival depending on PD-1 expression in tumor-infiltrating lymphocytes and tumor localization. Tumor samples were grouped for low versus high infiltration with CD3 positive TILs by using the median. For grouping in PD-1 and PD-L1 positive/negative, an expression ≥1% was used as a cutoff.

Table 1. Subgroup analyses: overall survival in months related to TIL infiltration state and UICC stage.

			TILs high	TILs low
			OS (Month)	OS (Month)	p
CD3
	UICC I		176.54	108.17	0.001
	UICC II		57.77	79.86	0.341
	UICC III		16.88	43.24	0.008
	UICC IV		8.05	11.94	0.221
	All		106.04	64.38	0.002
CD4
	UICC I		170.72	116.83	0.022
	UICC II		59.14	64.76	0.306
	UICC III		16.82	36.44	0.038
	UICC IV		8.05	11.94	0.221
	All		116.91	56.36	<0.001
CD8
	UICC I		168.74	118.41	0.042
	UICC II		60.12	78.55	0.508
	UICC III		26.0	28.15	0.644
	UICC IV		9.9	11.33	0.789
	All		101.53	67.34	0.016

Tumor samples were grouped for low or high infiltration with CD3^+, CD4⁺, or CD8⁺ TILs by using the median. Differences in overall survival were compared by Kaplan Meier using log rank.

Table 2. Univariate and multivariate survival analyses including CD3, CD4, CD8 positive infiltration state and PD-1 and PD-L1 positive infiltration state.

		Univariate						Multiple Cox Regression (after forward/[backward] selection)
Clinical Factor	Mean Survival		CI	p			HR		CI	p
Total patients	83.78		74.7–92.86
Age ≤ 62 years > 62 years	97.85 65.86		84.97–110.72 54.51–77.2	<0.001			1.027 1.025‡		1.009–1.046 1.007–1.043	0.004 0.006
Sex Female Male	96.82 72.92		81.88–111.76 62.09–83.75	<0.005			0.790		0.540–1.154	0.222
UICC stage UICC I UICC II UICC III UICC IV	154.83 70.16 26.65 10.97		140.48–169.18 56.53–83.79 16.61–36.68 8.93–12	<0.001			2.484 2.339‡		1.964–3.142 1.839–2.976	<0.001 <0.001
R R0 R1 R2 Palliative*	116.82 23.91 7.0 Censored		105.42–128.22 8.53–39.28 7.0–7.0	<0.001			2.399 2.394‡		1.318–4.369 1.329–4.313	0.004 0.004
Lauren Intestinal Diffuse Mixed	95.96 69.83 69.34		82.01–109.91 56.78–82.86 51.39–87.4	0.002			1.182		0.913–1.530	0.205
MMR dMMR pMMR	47.22 79.78		28.16–66.28 70.15–89.39	0.025			0.953		0.552–1.646	0.864
CD3 High Low	106.04 64.38		90.73–121.34 53.43–75.32	0.002			0.315 [0.326‡		0.109–0.908 0.124–0.858	0.032 0.023]
CD8 High Low	101.53 67.34		86.49–116.56 55.8–78.97	0.016
CD4 High Low	116.92 56.36		101.16–132.67 78.95–100.08	<0.001
PD-L1 Immune Cells Positive Negative Tumor Cells Positive Negative	63.03 83.25 119.56 68.23		49.19–76.86 71.29–95.21 90.57–148.56 59.05–77.53	0.428 0.003			0.848		0.474–1.519	0.580
PD-1 Immune Cells Positive Negative Tumor Cells Positive Negative	56.51 95.07 175.79 58.96		45.99–67.02 80.32–109.82 152.29–199.28 50.78–67.13	0.001 <0.001			0.925		0.552–1.552	0.768
UICC*CD3							1.683 [1.949‡		1.039–2.725 1.242–3.059	0.034 0.004]
PD-1*CD3							1.556 1.522‡		0.690–3.508 1.065–2.174	0.286 0.021

CI: confidence interval; HR: hazard ratio. Multivariate analysis: n = 210. TILs were grouped into high or low by a median cutoff. Mean survival is shown in months of survival. All factors significant in univariate analysis were analyzed in a multivariate cox-regression model. The reference categories for multivariate analysis are “low infiltration” for CD3, CD4 and CD8, “low CD3 infiltration and UICC I + II” for interaction between CD3 and UICC stage, “low CD3 infiltration and PD-1 expression negative” for interaction between PD-1 expression and CD3, “patient age < 62 y” for age, “male” for sex, “positive” for MMR status, “UICC I” for UICC stage, “R0” for R, “negative” for PD-1 and PD-L1 and “intestinal type” for Laurén classification. CD4⁺ and CD8⁺ TILs infiltration as well as PD-1 infiltration in tumor cells were excluded from multivariate analysis due to high statistical correlation.

* The resection state of the palliative cases was censored in multivariate confounder analysis, because of strong statistical interaction with UICC stage.