PD-L1 Protein Expression and Gene Amplification Correlate with the Clinicopathological Characteristics and Prognosis of Lung Squamous Cell Carcinoma

Figures & data

Table 1 Classification of PD-L1 Protein Expression Level in LUSC

PD-L1 Expression Status	PD-L1 Expression Levels	Staining Type
No PD-L1 Expression	TPS<1%	Tumor cells showing partial or complete cell membrane staining (≥ 1 +) accounted for less than 1% of viable tumor cells.
PD-L1 Expression (Low to medium)	1%≤TPS≤49%	Tumor cells with partial or complete cell membrane staining (≥ 1 +) accounted for 1–49% of viable tumor cells.
High PD-L1 Expression	TPS≥50%	Tumor cells with partial or complete cell membrane staining (≥ 1 +) accounted for more than 50% of viable tumor cells.

Figure 1 PD-L1 protein expression and gene amplification in lung squamous cell carcinoma. (A and B) Representative images of PD-L1 protein expression. (A) Low to moderate expression of PD-L1 (1%≤TPS≤49%) × 400. (B) High expression of PD-L1 protein (TPS> 50%) × 400. (C and D) Representative images of PD-L1 gene amplification analysed by FISH. (C) PD-L1 gene amplified Ratio > 2, parts of PD-L1 gene (red fluorescence) were expressed in clusters ×1000. (D) PD-L1 gene without amplification×1000.

Table 2 Relationship Between PD-L1 Gene Amplification and Clinicopathological Features of Lung Squamous Cell Carcinoma

Variable	Total	PD-L1 Amplification				PD-L1 Expression
		No	Yes	χ^2	p-value	<1%	1–49%	≥50%	χ^2	p-value
Total	164	153	11			86	47	31
Gender
Female	18	15	3	3.495	0.073	6	8	4	3.283	0.194
Male	146	138	8			80	39	27
Age (years)
≤60	87	79	8	6.462	0.176	37	30	20	7.300	0.026
>60	77	74	3			49	17	11
Smoking
Yes	101	94	7	0.728	0.885	60	26	15	5.496	0.064
No	63	59	4			26	21	16
Tumor size
T1	4	4	0	9.847	0.340	1	2	1	20.450	<0.001
T2	75	72	3			28	33	14
T3	85	77	8			57	12	16
Histological grade
1	44	40	4	1.802	0.531	26	10	8	1.347	0.853
2	49	45	4			24	15	10
3	71	68	3			36	22	13
Stage
I	39	39	0	52.897	<0.001	27	9	3	15.391	0.017
II	84	81	3			45	23	16
III	37	32	5			13	12	12
IV	4	1	3			1	3	0
Lymph node metastasis
Yes	71	61	10	49.639	0.001	14	34	23	53.774	<0.001
No	93	92	1			72	13	8
Metastasis
Yes	4	1	3	30.973	<0.001	1	3	0	4.436	0.109
No	160	152	8			85	44	31
Relapse
Yes	7	7	0	1.501	0.468	3	3	1	0.725	0.696
No	157	146	11			83	44	30

Table 3 Correlation Between PD-L1 Gene Amplification and Protein Expression

PD-L1 Amplification	PD-L1 Expression
	<1%	1–49%	≥50%	r=0.786	P<0.001
Yes	0	4	7
No	86	44	23

Table 4 Univariate Prognostic Factor Analysis by Log Rank Test

Variable	Total	Disease-Free Survival			Overall Survival
		Mean ± SE (Months)	95% CI	P	Mean ± SE (Months)	95% CI	P
Gender
Female	18	38.726±4.682	29.550–47.903	0.458	52.434±5.124	42.391–62.478	0.358
Male	146	42.153± 1.641	38.936–45.370		56.618±1.757	53.173–60.062
Age (years)
≤60	87	38.142± 2.114	33.999–42.284	0.046	51.473± 2.026	47.149–55.707	0.006
>60	77	45.851±2.168	41.601–50.101		61.394± 2.351	56.787–66.001
Smoking
Yes	101	43.377±2.018	39.422–47.333	0.221	57.998±2.181	48.347–58.373	0.153
No	63	39.372±2.406	34.656–44.089		53.360±2.558	53.723±62.273
Tumor status
T1	4	46.500±10.618	25.688–67.312	0.155	60.250±9.630	41.376–79.124	0.169
T2	75	45.797±2.127	41.627–49.967		60.478±2.302	55.967–64.989
T3	85	38.053±2.216	33.709–42.379		52.235±2.388	47.555–56.915
Histological grade
1	44	42.243±3.224	35.924–48.562	0.689	57.26±03.602	50.200–64.319	0.555
2	49	40.361±2.653	35.161–45.561		54.681±2.814	49.165–60.198
3	71	42.350±2.311	37.820–46.880		56.449±2.461	51.626–61.272
Stage
I	39	49.736±2.760	44.327–55.145	<0.001	66.524±2.816	61.004–72.043	<0.001
II	84	43.933±1.976	40.059–47.807		57.701±2.037	53.708–61.695
III	37	30.872±3.271	24.461–37.283		44.554±3.819	37.068–52.039
IV	4	14.500±3.610	7.425–21.575		26.000±6.364	13.527–38.473
Lymph node
Yes	71	36.594±2.414	31.863–41.325	0.026	49.510±2.635	44.344–54.675	0.015
No	93	45.531±1.853	41.898–49.163		60.844±1.920	57.081–64.606
Metastasis
Yes	4	14.500±3.610	7.425–21.575	<0.001	26.000±6.364	13.527–38.473	<0.001
No	160	42.309±1.540	39.291–45.327		56.753±1.646	53.527–59.979
Relapse
Yes	7	42.143±8.221	26.029–58.257	0.901	55.000±8.608	38.129–71.871	0.926
No	157	41.771±1.579	38.677–44.865		56.199±1.696	52.875–59.522
PD-L1 expression
<1%	86	49.281±1.801	45.751–52.811	<0.001	64.291±1.992	60.388–68.195	<0.001
1–49%	47	38.652±2.801	33.161–44.143		52.892±2.847	47.311–58.473
≥50%	31	23.925±2.221	19.572–44.143		36.359±2.297	31.858–40.860
PD-L1 amplification
No	153	43.934±1.507	40.980–46.889	<0.001	58.184±1.636	54.978–61.391	<0.001
Yes	11	11.909±1.522	8.925–14.893		27.909±3.942	20.184–35.634
PD-L1 >50%
And gene amplification (+)	8	11.625±1.658	8.376–14.874	<0.001	29.875±4.812	20.443–39.307	0.130
And gene amplification (-)	22	28.461±2.454	23.650–33.271		38.885±2.608	33.774–43.996
PD-L1 <50%
And gene amplification (+)	3	12.667±4.055	4.719–20.615	<0.001	22.667±7.055	57.756–64.380	<0.001
And gene amplification (-)	131	46.269–1.591	43.149–49.388		61.068±1.690	59.275–66.725

Table 5 Cox Multivariate Survival Analysis in LSUC Patents

Variable	Disease-Free Survival			Overall Survival
	HR	95% CI	P	HR	95% CI	P
Age (years)	0.823	0.576–1.175	0.283	0.823	0.576–1.175	0.179
Stage			0.001			0.184
Stage (1)	0.219	0.043–1.101	0.065	0.315	0.063–1.577	0.160
Stage (2)	0.306	0.066–1.418	0.130	0.416	0.093–1.853	0.250
Stage (3)	0.838	0.180–3.897	0.822	0.691	0.157–3.042	0.625
Lymph node metastasis	2.696	1.521–4.779	0.001	0.494	0.284–0.861	0.013
PD-L1 expression			<0.001			<0.001
PD-L1 expression (1)	0.168	0.088–0.320	<0.001	0.050	0.010–0.249	<0.001
PD-L1 expression (2)	0.415	0.231–0.746	0.003	0.228	0.048–1.077	0.062
PD-L1 amplification	0.102	0.038–0.268	<0.001	2.552	1.547–4.210	<0.001

Figure 2 Kaplan-Meier survival curves of PD-L1 amplification and expression in patients with lung squamous cell carcinoma. (A and C) DFS curve of patients with lung squamous cell carcinoma. (B and D) OS curve of patients with lung squamous cell carcinoma. (E) DFS curve of PD-L1 expression >50% and gene amplification. (F) OS curve of PD-L1 expression >50% and gene amplification. (G) DFS curve of PD-L1 expression<50% and gene amplification. (H) OS curve of PD-L1 expression <50% and gene amplification.