VISTA and PD-L1 synergistically predict poor prognosis in patients with extranodal natural killer/T-cell lymphoma

Figures & data

Figure 1. Expression of PD-L1 and VISTA in tumor tissues from ENKTCL patients. A-E. Representative double immunofluorescence images of CD56 (green) and PD-L1 (red) (a), CD68 (green) and PD-L1 (red) (b), CD56 (green) and VISTA (red) (c), CD68 (green) and VISTA (red) (d), and PD-L1 (green) and VISTA (red) (e). The nucleus is labeled with DAPI (blue). Arrows indicate the colocation of indicated proteins. F. The representative IHC images of PD-L1 and VISTA from serial sections. G. The correlation between PD-L1 expression and VISTA expression depicted with scatter plot. Scale bar is 50 μm

Table 1. The correlation between PD-L1 and VISTA expression and clinicopathological characteristics

		PD-L1 expression			VISTA expression
Variables	PatientsNO.	Low	High	p	Low	High	p
Age(y)				0.412			0.666
<60	96	55	41		53	43
≥60	13	9	4		8	5
Gender				0.307			0.158
Male	74	41	33		38	36
Female	35	23	12		23	12
Location				0.325			0.039
Nasal	85	52	33		52	33
Others	24	12	12		9	15
B symptoms				0.098			0.317
Yes	49	33	16		30	19
No	60	31	29		31	29
ECOG score				0.500			1.000
0-1	105	61	44		59	46
≥2	4	3	1		2	2
Elevated LDH				0.096			0.093
Yes	75	48	27		46	29
No	34	16	18		15	19
Distal LN involvement				0.033			0.004
Yes	90	57	33		56	34
No	19	7	12		5	14
Ann Arbor stage				0.006			0.002
I-II	74	50	24		49	25
III-IV	35	14	21		12	23
NRI score				0.009			0.001
0-1	45	33	12		34	11
≥2	64	31	33		27	37
PINK				0.004			0.003
0-1	83	55	28		53	30
≥2	26	9	17		8	18
VISTA				<0.001			-
Low	70	48	13		-	-
High	39	16	32		-	-

Abbreviation: ECOG, Eastern Cooperative Oncology Group; LDH, lactate dehydrogenase; LN, lymph node, NRI, nomogram-revised risk index; PINK, prognostic index of natural killer lymphoma. p-value <0.05 marked in bold font shows statistically significant.

Table 2. Univariate analysis of prognostic factors correlated with PFS and OS in patients with ENKTCL

	PSF		OS
Variables	HR (95% CI)	p	HR (95% CI)	p
Age (y) (≥60/<60)	1.10 (0.54–2.20)	0.800	1.57 (0.77–3.19)	0.217
Gender (male/female)	0.72 (0.43–1.19)	0.200	0.77 (0.44–1.36)	0.369
Location (nasal/others)	1.93 (1.14–3.19)	0.013	1.31 (0.72–2.40)	0.378
B symptoms (yes/no)	1.30 (0.82–2.06)	0.272	1.26 (0.75–2.12)	0.388
ECOG score (≥2/0-1)	1.23 (0.39–3.92)	0.725	1.76 (0.55–5.62)	0.343
Elevated LDH (yes/no)	1.43 (0.87–2.35)	0.158	1.74 (1.01–3.00)	0.048
Distal LN involvement (yes/no)	2.17 (1.27–3.76)	0.006	1.87 (1.01–3.48)	0.048
Ann Arbor stage III–IV/I–II	2.34 (1.46–3.77)	<0.001	2.59 (1.53–4.38)	<0.001
NRI score (≥2/0-1)	1.72 (1.06–2.78)	0.027	2.24 (1.28–3.93)	0.005
PINK (≥2/0-1)	1.82 (1.09–3.02)	0.023	1.59 (0.89–2.83)	0.117
Chemotherapy (non-ANT/ANT based)	0.38 (0.22–0.65)	<0.001	0.39 (0.22–0.69)	0.003
PD-L1 expression (high/low)	1.93 (1.22–3.07)	0.005	2.44 (1.45–4.11)	0.001
VISTA expression (high/Low)	2.05 (1.29–3.25)	0.002	2.27 (1.34–3.83)	0.002

Abbreviation: ECOG, Eastern Cooperative Oncology Group; LDH, lactate dehydrogenase; LN, lymph node; NRI, nomogram-revised risk index; PINK, prognostic index of natural killer lymphoma; ANT, anthracycline; PFS, progression-free survival; OS, overall survival; HR, hazard ratio; CI, confidence interval. p-value <0.05 marked in bold font shows statistically significant.

Table 3. Multivariate analysis of prognostic factors correlated with PFS and OS in patients with ENKTCL

	PSF		OS
Variables	HR (95% CI)	p	HR (95% CI)	p
Location (nasal/others)	1.65 (0.92–2.98)	0.094	-	-
PINK (2–3/0-1)	0.19 (0.05–0.73)	0.016	-	-
Elevated LDH (yes/no)	-	-	1.14 (0.62–2.13)	0.671
Distal LN involvement (yes/no)	2.34 (0.77–7.08)	0.134	0.71 (0.32–1.57)	0.392
Ann Arbor stage III–IV/I–II	3.70 (1.57–8.73)	0.003	2.01 (0.91–4.44)	0.083
NRI score (≥2/0-1)	1.28 (0.68–2.42)	0.443	1.78 (0.85–3.74)	0.127
Chemotherapy (non-ANT/ANT based)	0.33 (0.18–0.59)	0.001	0.27 (0.14–0.52)	0.001
PD-L1 expression (high/low)	1.46 (0.82–2.62)	0.200	1.75 (0.93–3.28)	0.081
VISTA expression (high/Low)	1.48 (0.82–2.68)	0.198	1.32 (0.68–2.55)	0.412

Abbreviation: LDH, lactate dehydrogenase; LN, lymph node; NRI, nomogram-revised risk index; PINK, prognostic index of natural killer lymphoma; ANT, anthracycline; PFS, progression-free survival; OS, overall survival; HR, hazard ratio; CI, confidence interval. p-value <0.05 marked in bold font shows statistically significant.

Figure 2. Kaplan-Meier survival curves for progression-free survival (PFS) and overall survival (OS) of patients with ENKTCL according to PD-L1 and VISTA expression. A-B. PFS for ENKTCL patients based on PD-L1 (a) and VISTA (b) expression. C-D. OS for ENKTCL patients based on PD-L1 (c) and VISTA (d) expression. E-F. PFS (e) and OS (f) for ENKTCL patients based on combination of PD-L1 and VISTA expression. Type I, PD-L1^low/VISTA^low; Type II, PD-L1^low/VISTA^high or PD-L1^high/VISTA^low; Type III, PD-L1^high/VISTA^high

Table 4. Multivariate analysis of prognostic factors correlated with PFS and OS in patients with ENKTCL

	PFS		OS
Variables	HR (95% CI)	p	HR (95% CI)	p
Location (nasal/others)	1.65 (0.92–2.97)	0.094	-	-
PINK (2–3/0-1)	0.19 (0.05–0.73)	0.016	-	-
Elevated LDH (yes/no)	-	-	1.14 (0.62–2.13)	0.670
Distal LN involvement (yes/no)	2.34 (0.77–7.08)	0.134	0.69 (0.31–1.53)	0.361
Ann Arbor stage III–IV/I–II	3.70 (1.57–8.73)	0.003	2.02 (0.91–4.47)	0.082
NRI score (≥2/0-1)	1.28 (0.68–2.42)	0.443	1.77 (0.84–3.70)	0.131
Chemotherapy Non-ANT/ANT based	0.33 (0.18–0.59)	<0.001	0.28 (0.15–0.53)	<0.001
PD-L1/VISTA expression Type III/II/I	1.47 (1.10–1.98)	0.010	1.53 (1.10–2.12)	0.012

Abbreviation: LDH, lactate dehydrogenase; LN, lymph node; NRI, nomogram-revised risk index; PINK, prognostic index of natural killer lymphoma, ANT, anthracycline, PFS, progression-free survival, OS, overall survival; HR, hazard ratio; CI, confidence interval; Type I, PD-L1^low/VISTA^low; Type II, PD-L1^low/VISTA,^high or PD-L1^high/VISTA ^low; Type III, PD-L1^high/VISTA^high. p-value <0.05 marked in bold font shows statistically significant.

Figure 3. The correlation between the expression of VISTA and PD-L1, and the density of tumor-infiltrating lymphocytes (TILs). A. The representative images of PD-L1, VISTA, CD8 and Foxp3 from serial sections from two samples, sample 1(up) and sample 2 (down). B-D. The correlations between CD8+ TILs and PD-L1 expression (b), VISTA expression (c) or novel immune classification (d) depicted with scatter plot. E-G. The correlations between Foxp3+ TILs and PD-L1 expression (e), VISTA expression (f) or novel immune classification (g) depicted with scatter plot. All data display mean ± SEM. Scale bar is 50 μm. *p < .05, ***p < .001. ns: no significance. Type I, PD-L1^low/VISTA^low; Type II, PD-L1^low/VISTA^high or PD-L1^high/VISTA^low; Type III, PD-L1^high/VISTA^high

Figure 4. The PD-L1 and VISTA expression were associated with the response to blockade of PD-1. (a). The representative images of the PD-L1 and VISTA expression in indicated groups. (b). Swimmer’s plot showing the response to PD-1 blockade in extranodal NK/T-cell lymphoma patients, who were classified into four groups as indicated. CR, complete response; PR, partial response; PD, progressive disease

Availability of data and materials

All datasets used and/or analyzed during the current study are available in the Research Data Deposit public platform (www.researchdata.org.cn, number RDDB2021001093) from the corresponding author on request.