Dynamics of Early Serum Tumour Markers and Neutrophil-to-Lymphocyte Ratio Predict Response to PD-1/PD-L1 Inhibitors in Advanced Non-Small-Cell Lung Cancer

Figures & data

Table 1 Baseline Characteristics of Patients

Median Age Years (Range)		63 (36–85)
Age (n, %)	<65 years	73 (58.87)
	≥65 years	51 (41.13)
Gender (n, %)	Female	35 (28.23)
	Male	89 (71.77)
BMI (n, %)	<18.5 kg/m^2	3 (2.46)
	18.5–23.9 kg/m^2	61 (50.00)
	>23.9 kg/m^2	58 (47.54)
Histology (n, %)	Adenocarcinoma	70 (56.45)
	Squamous carcinoma	40 (32.26)
	Adenosquamous carcinoma	3 (2.42)
	Large cell carcinoma	5 (4.03)
	Unknown	6 (4.84)
TNM stage (n, %)	III	32 (25.81)
	IV	92 (74.19)
ECOG PS (n, %)	0	25 (20.16)
	1	98 (79.03)
	2	1 (0.81)
Smoking status (n, %)	Never	46 (37.10)
	Former	59 (47.58)
	Current	19 (15.32)
Metastasis sites (n, %)	Bone	14 (14.89)
	Lymph node	30 (31.91)
	Lung	36 (38.30)
	Brain	7 (7.45)
	Liver	4 (4.26)
	Adrenal gland	3 (3.19)
Radiotherapy (n, %)	Yes	44 (35.48)
	No	80 (64.52)
PD-L1 expression (n, %)	Not available	89 (71.77)
	<50%	21 (16.94)
	1–49%	10 (8.06)
	Negative	4 (3.23)
Line of immunotherapy (n, %)	1	31 (25.00)
	2	63 (50.81)
	≥3	30 (24.19)
ICI (n, %)	PD-1	107 (86.29)
	PD-L1	17 (13.71)
Treatment type (n, %)	Monotherapy	37 (29.84)
	Combination therapy	87 (70.16)
Immune-related adverse events (n, %)	Yes	119 (95.97)
	No	5 (4.03)
Leading STM (n, %)	CEA	57 (51.35)
	CYFRA 21–1	36 (32.43)
	NSE	12 (10.81)
	CA19-9	2 (1.80)
	CA125	4 (3.61)
Baseline CEA (n, %)	Normal (<5 ng/mL)	47 (43.52)
	Increased (≥5 ng/mL)	61 (56.48)
Baseline CYFRA 21–1 (n, %)	Normal (<3.3 ng/mL)	23 (24.47)
	Increased (≥3.3 ng/mL)	71 (75.53)
Baseline NSE (n, %)	Normal (<16.3 ng/mL)	49 (56.98)
	Increased (≥16.3 ng/mL)	37 (43.02)
Baseline CA19-9 (n, %)	Normal (<37 U/mL)	13 (59.09)
	Increased (≥37 U/mL)	9 (40.91)
Baseline CA125 (n, %)	Normal (<35 U/L)	20 (54.05)
	Increased (≥35 U/L)	17 (45.95)
Baseline NLR (n, %)	Decreased (<5)	97 (78.23)
	Increased (≥5)	27 (21.77)

Note: Data are given as absolute number and percent within the respective group unless otherwise specified.

Abbreviations: BMI, body mass index; TNM, tumor node metastasis; ECOG PS, Eastern Cooperative Oncology Group performance status; ICI, immune checkpoint inhibitor; PD-1, programmed cell death-1; PD-L1, programmed cell death-ligand 1; STM, serum tumor markers; CEA, carcinoembryonic antigen; CYFRA 21–1, cytokeratin-19 fragments; NSE, neuron specific enolase; CA19-9, carbohydrate antigen 19–9; CA125, carbohydrate antigen 125.

Table 2 Associations Between Clinicopathological Features and Leading STM, NLR, and Combination Score

Variables	Category	Leading STM		P	NLR		P	Combination Score			P
		Decreased	Increased		Decreased	Increased		0	1	2
Age (years)	<65	24	33	0.069	58	15	0.687	21	28	8	0.050
	≥65	26	17		42	9		25	11	7
Gender	Female	11	16	0.260	29	6	0.696	11	12	4	0.828
	Male	39	34		71	18		35	27	11
BMI (kg/m^2)	18.5–23.9	26	21	0.414	47	14	0.247	24	15	8	0.387
	<18.5 or >23.9	24	27		52	9		22	23	6
Histology	Adenocarcinoma	31	27	0.220	62	8	0.004	30	22	6	0.088
	Non-adenocarcinoma	15	22		32	16		12	16	9
TNM stage	III	14	13	0.822	29	3	0.097	13	13	1	0.143
	IV	36	37		71	21		33	26	14
ECOG PS	0	11	4	0.050	21	4	0.781	11	2	2	0.040
	≥1	39	46		79	20		35	37	13
Smoking status	Never	17	20	0.534	37	9	0.964	16	14	7	0.698
	Former/Current	33	30		63	15		30	25	8
Metastasis	No	12	13	0.817	27	3	0.136	11	13	1	0.130
	Yes	38	37		73	21		35	26	14
Radiotherapy	Yes	17	18	0.834	31	13	0.033	16	9	10	0.011
	No	33	32		69	11		30	30	5
PD-L1 expression	<50%	7	7	0.210	18	3	0.533	7	6	1	0.388
	≥50%	8	2		10	0		8	2	0
PD-L1 status	Positive	15	9	1.000	27	5	1.000	14	8	2	1.000
	Negative	3	1		4	0		3	1	0
Line of immunotherapy	<3	37	36	0.822	75	19	0.669	35	26	12	0.588
	≥3	13	14		25	5		11	13	3
ICI	PD-1	45	44	0.749	88	19	0.320	41	36	12	0.467
	PD-L1	5	6		12	5		5	3	3
Mono-immunotherapy	Yes	13	10	0.476	28	9	0.361	13	5	5	0.124
	No	37	40		72	15		33	34	10
Immune-related adverse events	Yes	46	49	0.362	96	23	1.000	42	39	14	0.132
	No	4	1		4	1		4	0	1

Notes: Data are given as absolute number and testing for significance between the subgroups was accomplished using a chi square test, a P value of < 0.05 was regarded statistically significant.

Abbreviations: BMI, body mass index; TNM, tumor node metastasis; ECOG PS, Eastern Cooperative Oncology Group performance status; ICI, immune-checkpoint inhibitors; PD-1, programmed cell death-1; PD-L1, programmed cell death-ligand 1; STM, serum tumor markers; NLR, neutrophil-to-lymphocyte ratio.

Figure 1 ORR and DCB from the decreased leading STM versus the increased leading STM, and from the decreased NLR versus the increased NLR. ORR and DCB for patients in the overall cohort with different combination scores, and for patients in the cohort of the Third Affiliated Hospital of Shandong First Medical University with different combination scores.

Abbreviations: STM, serum tumour marker; NLR, neutrophil-to-lymphocyte ratio; ORR, objective response rate; DCB, durable clinical benefit.

Figure 2 Kaplan–Meier survival curves for progression-free and overall survival according to the leading STM and the NLR. Kaplan–Meier survival curves for progression-free and overall survival according to the combination scores in the overall cohort and in the cohort of the Third Affiliated Hospital of Shandong First Medical University.

Abbreviations: NR, not reached; STM, serum tumour markers; NLR, neutrophil-to-lymphocyte ratio; PFS, progression-free survival; OS, overall survival; HR, hazard ratio; CI, confidence interval.

Figure 3 DCB for stable or progressive patients with different combination scores (A). Kaplan–Meier survival curves for progression-free (B) and overall survival (C) according to the combination scores of stable or progressive patients.

Abbreviations: NR, not reached; DCB, durable clinical benefit; PFS, progression-free survival; OS, overall survival; CI, confidence interval.

Figure 4 Computed tomography (CT) scans at baseline and at the first and subsequent radiological assessments for 3 representative NSCLC patients treated with PD-1/PD-L1 inhibitors. The first patient (Case 1) with a combination score of 0 (low combination score) had achieved a partial response at the first radiological evaluation and subsequently achieved a complete response (at 14 weeks) with long-term survival (alive; progression-free survival: 30.9 months; overall survival: 30.9 months). The second patient (Case 2) with a combination score of 0 (low combination score) had stable disease at the first radiological evaluation and subsequently exhibited a partial response (at 13 weeks) with long-term survival (alive; progression-free survival: 19.1 months; overall survival: 21.7 months). The third patient (Case 3) with a combination score of 2 (high combination score) had progressive disease at the first radiological evaluation and subsequently exhibited continuous progression (at 12 weeks) with short-term survival (death; progression-free survival: 2.3 months; overall survival: 5.0 months). The arrows indicate lesions in the lungs.

Table 3 Univariate Cox Analyses of PFS and OS

Variable	Category	PFS		OS
		HR (95% CI)	P	HR (95% CI)	P
Age	≥65 vs <65	0.702 (0.361–1.364)	0.296	0.796 (0.412–1.541)	0.499
Gender	Female vs Male	0.837 (0.396–1.771)	0.642	0.663 (0.312–1.406)	0.284
BMI(kg/m^2)	<18.5 or >23.9 vs.18.5–23.9	1.032 (0.530–2.011)	0.926	1.076 (0.554–2.091)	0.829
Histology	Non-adenocarcinoma vs Adenocarcinoma	1.096 (0.578–2.081)	0.778	1.239 (0.655–2.344)	0.509
TNM stage	IV vs III	1.333 (0.608–2.925)	0.473	1.198 (0.549–2.617)	0.650
ECOG PS	≥1 vs 0	1.049 (0.510–2.158)	0.896	1.194 (0.575–2.478)	0.634
Smoking status	Former/Current vs Never	1.315 (0.650–2.657)	0.446	1.526 (0.757–3.078)	0.238
Actionable mutation	(-)/undetected vs (+)	22.777 (0.041–12,505.518)	0.331	22.628 (0.067–7697.921)	0.294
Metastasis	Yes vs No	1.222 (0.557–2.681)	0.617	1.036 (0.474–2.265)	0.929
Radiotherapy	Yes vs No	2.087 (1.098–3.965)	0.025	2.294 (1.204–4.368)	0.012
PD-L1 expression	≥50% vs 0–50%	0.226 (0.028–1.849)	0.166	0.227 (0.028–1.867)	0.168
PD-L1 status	Positive vs Negative	1.008 (0.126–8.082)	0.994	0.933 (0.114–7.609)	0.949
Line of immunotherapy	≥3 vs <3	1.299 (0.611–2.760)	0.497	1.169 (0.552–2.475)	0.683
ICI substance	PD-1 vs PD-L1	0.663 (0.303–1.451)	0.303	0.624 (0.286–1.365)	0.238
Mono-immunotherapy	Yes vs No	1.933 (1.012–3.690)	0.046	1.612 (0.841–3.087)	0.150
Immune adverse events	Yes vs No	21.511 (0.012–37,969.388)	0.421	21.115(0.003–133,441.458)	0.495
Leading STM at baseline	> the upper limit of normal vs < the upper limit of normal	0.911 (0.341–2.430)	0.852	0.981 (0.372–2.588)	0.969
Leading STM at 6 weeks	> the upper limit of normal vs < the upper limit of normal	1.548 (0.611–3.923)	0.357	1.523 (0.604 to 3.839)	0.373
Leading STM change	Decrease <20% vs Decrease ≥20%	0.201 (0.081–0.502)	0.001	0.275 (0.117–0.643)	0.003
Pretreatment NLR	≥5 vs <5	0.524 (0.264–1.040)	0.065	0.425 (0.213–0.848)	0.015
Posttreatment NLR	≥5 vs <5	0.179 (0.091–0.35)	<0.0001	0.158 (0.080–0.314)	<0.0001
NLR change	Decrease ≥20% vs Decrease <20%	0.850 (0.448–1.614)	0.620	0.892 (0.469–1.695)	0.726
	Increase ≥20% vs Increase <20%	1.720 (0.827–3.575)	0.147	1.713 (0.827–3.551)	0.148
Combination Score	0
	1	3.150 (1.021–9.716)	0.046	2.099 (0.731–6.028)	0.168
	2	21.418 (6.975–65.763)	<0.0001	17.110 (5.943–49.260)	<0.0001

Notes: Cox regression models were applied to find independent indicators associated with PFS and OS and estimate the hazard ratio (HR). Factors which were statistically significant in the univariate analysis were incorporated into the multivariate analysis. Figures are given as HR (95% CI). A P value of < 0.05 was regarded statistically significant.

Abbreviations: BMI, body mass index; TNM, tumor node metastasis; ECOG PS, Eastern Cooperative Oncology Group performance status; ICI, immune-checkpoint inhibitors; PD-1, programmed cell death-1; PD-L1, programmed cell death-ligand 1; STM, serum tumor markers; NLR, neutrophil-to-lymphocyte ratio; PFS, progression-free survival; OS, overall survival; HR, hazard ratio; CI, confidence interval.

Table 4 Multivariate Cox Analyses of PFS and OS

A. NLR and STM Multivariate Cox Analysis		PFS		OS
		HR (95% CI)	P	HR (95% CI)	P
Radiotherapy	Yes vs No	1.079 (0.432–2.691)	0.871	0.843 (0.342–2.078)	0.710
Mono-immunotherapy	Yes vs No	1.786 (0.758–4.212)	0.185	/	/
Leading STM change	Decrease <20% vs Decrease ≥20%	2.892 (0.976–8.568)	0.055	0.510 (0.193–1.350)	0.175
Pretreatment NLR	≥5 vs <5	/	/	0.805 (0.308–2.099)	0.657
Posttreatment NLR	≥5 vs <5	0.144 (0.048–0.427)	0.001	0.122 (0.042–0.352)	0.001
B. The Combination Score Multivariate Cox Analysis		PFS		OS
		HR (95% CI)	P	HR (95% CI)	P
Radiotherapy	Yes vs No	1.049 (0.417–2.643)	0.919	0.807 (0.329–1.976)	0.630
Mono-immunotherapy	Yes vs No	1.860 (0.772–4.480)	0.167	/	/
Combination Score	0
	1	3.661 (1.136–11.792)	0.030	3.183 (1.02–9.931)	0.046
	2	20.066 (5.646–71.311)	<0.0001	22.299 (6.381–77.924)	<0.0001

Notes: Cox regression models were applied to find independent indicators associated with PFS and OS and estimate the hazard ratio (HR). Factors which were statistically significant in the univariate analysis were incorporated into the multivariate analysis. Figures are given as HR (95% CI). A P value of < 0.05 was regarded statistically significant.

Abbreviations: PFS, progression-free survival; OS, overall survival; HR: hazard ratio; CI, confidence interval.

Figure 5 ROC curves for sensitivity and specificity according to leading STM change, posttreatment NLR and the combination score.

Abbreviations: STM, serum tumour marker; NLR, neutrophil-to-lymphocyte ratio; ROC, receiver operating curve; AUC, area under the curve.