| Part Ⅰ Exploring the Prognostic Value of the Lung Immune Prognostic Index in Patients with LA-NSCLCObjective:Previous study demonstrated that the baseline lung immune prognostic index(LIPI)was a potential biomarker which could identify advanced non-small cell lung cancer(NSCLC)patients who would benefit from treatment with immune checkpoint inhibitor(ICI).However,a recent study found that the LIPI might be an important prognostic biomarker irrespective of treatment modality for patients with metastatic NSCLC.It remains unclear whether LIPI is associated with long-term outcomes in unresected stage Ⅲ NSCLC.Therefore,this study aimed to investigate the prognostic value of LIPI in locally advanced NSCLC(LA-NSCLC)patients.Methods and Materials:Patients with LA-NSCLC treated with definitive radiotherapy(RT)in Cancer Hospital Chinese Academy of Medical Sciences from January 2000 to December 2017 were retrospectively reviewed.According to previous study,pretreatment derived neutrophils/(leukocytes minus neutrophils)ratio(dNLR)and lactate dehydrogenase(LDH)made up the LIPI and divided it into two groups(good,0 score;intermediate-poor,1 or 2 scores).The endpoints include overall survival(OS),progression-free survival(PFS),locoregional relapse-free survival(LRRFS),and distant metastasis-free survival(DMFS).Kaplan-Meier method and Cox hazards regression analysis were used to explore associations between the LIPI and LA-NSCLC prognosis.Propensity score matching(PSM)was conducted to balance the confounding variables.Results:A total of 1079 patients were eligible for analysis.Patients with an intermediate-poor pretreatment LIPI had significantly inferior OS,PFS,LRRFS,and DMFS than those with a good LIPI(median OS,19.0 vs 25.0 months,Log-rank P<0.001;median PFS,10.0 vs 13.0 months,Log-rank P=0.001;median LRRFS,13.0 vs 18.0 months,Log-rank P<0.001;median DMFS,15.0 vs 17.0 months,Log-rank P=0.002).According to multivariate analysis,it was also found that the LIPI was an independent prognostic marker for OS(P=0.026,hazard ratio[HR]=1.19,95%CI:1.02-1.40),PFS(P=0.024,HR=1.18,95%CI:1.02-1.36),and LRRFS(P=0.009,HR=1.22,95%CI:1.05-1.41)in patients with inoperable LA-NSCLC.PSM analysis further verified that intermediate-poor LIPI was an independent prognostic factor for shorter survivals(OS,PFS and LRRFS).In subgroup analyses,the intermediate-poor LIPI was remained significantly associated with increased risks of death in male patients,who had smoking history,with KPS>80.Similar results were observed in PFS.In addition,the intermediate-poor LIPI also increased the risk of progression in early diagnosed patients with squamous carcinoma.Among patients receiving concurrent chemoradiotherapy,the intermediate-poor LIPI status was no longer associated with inferior survival(5-y OS:34.5%vs 29.5%,HR=1.12,95%CI:0.84-1.49,Log-rank P=0.422)compared with sequential chemoradiotherapy(5-y OS:16.0%vs 26.8%;HR=1.33;95%CI:1.02-1.74;Log-rank P=0.035)or radiotherapy alone(5-y OS:8.2%vs 14.5%,HR=1.44,95%CI:1.12-1.85,Log-rank P=0.004).Conclusion:LIPI is a simple and promising prognostic marker for patients with unresectable LA-NSCLC.Further prospected studies are warranted to validated these findings.Part Ⅱ Exploring the association between tumor-infiltrating lymphocytes and clinical outcomes of inoperable LA-NSCLC patients undergoing concurrent chemoradiotherapyObjective:As an important component of the tumor microenvironment,tumor-infiltrating lymphocytes(TILs)have been reported to play a prognostic and immune efficacy predictive role in many solid tumors.Currently,the effect of TILs on inoperable locally advanced non-small cell lung cancer(LA-NSCLC)patients undergoing concurrent chemoradiotherapy(cCRT)remains unclear.This study aims to analyze the relationship between TILs and CD8+TILs and clinical outcomes of inoperable stage Ⅲ NSCLC patients.Methods and Materials:The clinicopathological data of patients with inoperable LA-NSCLC undergoing cCRT in Cancer Hospital Chinese Academy of Medical Sciences from 2016 to 2017 were collected.Paraffin-embedded specimens were available and sufficient for testing.TILs in tumor stroma were evaluated by hematoxylin-eosin staining and two scoring standards were included.One is based on the recommendation for TILs evaluation in breast cancer surgical specimens,that is,the proportion of TILs in tumor stroma.The other is a modified method for TILs evaluation in biopsy specimens,that is,the proportion of TILs in tumor tissue.Using CD8 antibody detected CD8 TILs by immunohistochemistry.The optimal clinical cut-off values of TILs were determined by X-Tile software.The endpoints include overall survival(OS),progression-free survival(PFS),locoregional relapse-free survival(LRRFS),and distant metastasis-free survival(DMFS).Results:In this study,40 paraffin specimens of inoperable LA-NSCLC patients who received cCRT were collected,of which 34 were available for TILs analysis.For the proportion of TILs in the tumor strorna,results showed no significant difference between high-density TILs(>20%)and low-density TILs(≤20%)regarding the distribution of age,smoking history,histological type,and TNM stage(all P>0.05).Although it was observed that high-density TILs tended to improve OS(median OS,30 vs 15 months,Log-rank P=0.061),PFS(median PFS,19 vs 9 months,Log-rank P=0.164),LRRFS(median LRRFS,21 vs 9 months,Log-rank P=0.173),and DMFS(median DMFS,30 vs 15 months,Log-rank P=0.072)of patients,multivariate analysis indicated that TILs was not an independent prognostic factor for LA-NSCLC patients(all P>0.05).When using the proportion of TILs in tumor tissue for scoring,this study found that the distribution of high-density TILs in patients with squamous carcinoma was higher than that in patients with adenocarcinoma(P=0.048).The results of survival analysis demonstrated that high-density TILs(>7%)significantly reduced the risk of death(median OS,35 vs 17 months,Log-rank P=0.020),disease progression(median PFS,19 vs 8 months,Log-rank P=0.016),local recurrence(median LRRFS,21 vs 8 months,Log-rank P=0.030),and distant metastasis(median DMFS,30 vs 12 months,Log-rank P=0.007)compared with low-density TILs(TILs≤7%).In addition,the multivariate analysis suggested that the high-density TILs was an independent marker for good survival in patients with inoperable stage Ⅲ NSCLC who received cCRT(OS:hazard ratio[HR]=0.30,95%CI:0.12-0.75,P=0.010;PFS:HR=0.27,95%CI:0.11-0.67,P=0.005;LRRFS:HR=0.26,95%CI:0.10-0.65,P=0.004;DMFS:HR=0.31,95%CI:0.12-0.78,P=0.013).For the CD8+TILs,29 patients were available for analysis.The results of multivariate analysis showed that positive expression of CD8+TILs(CD8+TILs>0)was an independent factor for good OS(HR=0.17,95%CI:0.05-0.58,P=0.005),PFS(HR=0.17,95%CI:0.05-0.57,P=0.004),LRRFS(HR=0.19,95%CI:0.06-0.67,P=0.008),and DMFS(HR=0.25,95%CI:0.08-0.77,P=0.015)in LA-NSCLC patients.Conclusion:TILs,assessed by a modified method,is a simple and effective prognostic biomarker for patients with inoperable stage Ⅲ NSCLC receiving cCRT.Besides,the positive expression of CD8+TILs was also associated with good OS,PFS,LRFFS,and DMFS in patients.A larger sample size of research is warranted to further validate these findings.Part Ⅲ Study on the PD-L1 Expression in LA-NSCLC Patients Receiving Concurrent Chemoradiotherapy and its Relationship with PrognosisObjective:The immune checkpoint inhibitors(ICIs)have made breakthroughs in many malignant tumors,especially changing the treatment models of unresectable locally advanced non-small cell lung cancer(LA-NSCLC).However,there is still a lack of effective prognostic markers for LA-NSCLC.Programmed death ligand 1(PD-L1)is considered as a predictive biomarker for the immunotherapy of NSCLC.In recent years,some studies reported the prognostic effect of PD-L1 in NSCLC patients,but the conclusions were inconsistent.This study mainly investigated the PD-L1 expression detected by the rabbit monoclonal antibody SP263 and its association with overall survival(OS)in LA-NSCLC patients with non-EGFR mutated undergoing concurrent chemoradiotherapy(cCRT).Methods and Materials:The basic information of patients with unresectable LA-NSCLC diagnosed by histopathology in Cancer Hospital Chinese Academy of Medical Sciences from January 2016 to December 2017 was collected.The rabbit monoclonal antibody SP263 was used for immunohistochemical analysis to detect the expression of PD-L1 in tumor tissues.Setting the Tumor Proportion Score(TPS)as an evaluation criterion,that is,the percentage of positive tumor cells with any intensity of membrane stainng.The TPS>1%was defined as PD-L1 positive,and the associations between PD-L1≥1%and PD-L1≥25%with the clinical characteristics and clinical outcome of LA-NSCLC patients were evaluated respectively.Fisher’s exact test was used to compare the differences between the groups,the Kaplan-Meier method was used to calculate overall survival(OS),and the Log-Rank test was used to compare the survival outcomes.Results:A total of 19 patients with NSCLC were eligible for analysis.Among them,13 patients(68.4%)had PD-L1 positive expression,and 4 patients(21.1%)had PD-L1 expression greater than or equal to 25%.No significant difference was observed between different PD-L1 expressions regarding the distribution of age,smoking history,pathological classification,PET-CT examination,and TNM staging in LA-NSCLC patients with non-EGFR mutation receiving cCRT(Fisher’s exact test P>0.05).15 patients could be evaluated for therapeutic effect,the objective response rate(ORR)was 46.7%(7/15),and the disease control rate(DCR)was 93.3%(14/15).The ORR and DCR of patients with PD-L1 expression<1%were 75.0%(3/4)and 100.0%(4/4),respectively.The ORR and DCR of patients with PD-L1 expression>1%were 36.4%(4/11)and 90.9%(10/11),respectively.There were no statistical differences in ORR and DCR between the above two groups(P>0.05).When the PD-L1 expression was less than 25%,the ORR and DCR of patients were 50.0%(6/12)and 91.7%(11/12),respectively.When the PD-L1 expression was greater than or equal to 25%,the ORR and DCR of patients were 33.3%(1/3)and 100%(3/3),respectively.And,the ORR and DCR were not significantly different between the above two groups(P>0.05).With a median follow-up of 58 months,the median OS was 35 months for patients with PD-L1≥1%and 40 months for patients with PD-L1<1%(log-rank P=0.284).Patients with PD-L1≥25%had a median survival time of 12 months,and patients with PD-L1<25%had a median survival time of 40 months(log-rank P=0.241).Conclusion:Among inoperable LA-NSCLC patients with non-EGFR mutation receiving cCRT,there was a decreasing trend for OS in those with PD-L1 positive.A larger sample size and appropriate design are warranted to explore the prognostic value of PD-L1 expression in LA-NSCLC patients and to further explore the effect of immunotherapy on patients with different PD-L1 expression levels.Part Ⅳ Evaluating the Consistency of PD-L1 Expression Detecting by Different Antibodies in Inoperable LA-NSCLC Patients Receiving Concurrent ChemoradiationObjective:Immunotherapy has become part of the treatment in patients with non-small cell lung cancer(NSCLC),whereas different immune checkpoint inhibitors are accompanied by different programmed cell death ligand 1(PD-L1)detection reagents.Although some studies have compared the consistency of different PD-L1 detection reagents,the conclusions are still contradictory.Here,this study preliminarily explored the concordance of PD-L1 expression testing by 22C3 and SP263 in biopsy tissue of locally advanced NSCLC(LA-NSCLC)patients receiving concurrent chemoradiotherapy.Methods and Materials:The clinicopathological data and paraffin-embedded blocks of NSCLC patients undergoing cCRT in Cancer Hospital Chinese Academy of Medical Sciences from January 2016 to December 2017 were collected.PD-L1 immunohistochemical staining for 22C3 and SP263 was performed on Autostainer Link 48 platform and BenchMark ULTRA platform,respectively.Tumor Proportion Score(TPS)was used for scoring.For the staining results of the two reagents,intraclass correlation coefficient(ICC)and signed-rank test were used to analyze differences between the continuous variables.For 1%,25%,and 50%clinical cut-off values,Cohen’s Kappa analysis was conducted and overall percent agreement(OPA),positive percent agreement(PPA),and negative percent agreement(NPA)were calculated.Results:Of the 21 patients included,18 samples were available for analysis.The ICC between SP263 and 22C3 staining results was 0.759(95%CI:0.285-0.915),whereas the Wilcoxon signed-rank test showed that the TPS detected by SP263 was higher than that of 22C3(P=0.005).For 1%,25%,and 50%clinical cut-off values,TPS detected by SP263 and 22C3 showed higher concordance(κ=0.609-0.778),with OPA resulting as 88.9%,88.9%,and 94.4%.While PPA showed as 81.8%,50%and 50%,respectively,and NPA showed as 77.8%,87.5%and 94.1%,respectively.McNemar’s exact test found no differences between SP263 and 22C3 at different clinical cut-off values(all P>0.05).However,this study also found that when the TPS cut-off value was 1%,patients with positive PD-L1 expression detected by SP263 showed a trend of poor survival compared with PD-L1 negative patients(median OS:30 vs not reached,Log-Rank P=0.119).No similar trend was found in patients with positive PD-L1 expression detected by 22C3(median OS:35 vs 40 months,Log-Rank P=0.478).Conclusion:This exploratory study indicated that the PD-L1 expression score detected by SP263 was slightly higher than that of 22C3 in the biopsy tissue of LA-NSCLC patients receiving cCRT.According to the current evidence,supporting PD-L1 expression detected by 22C3 interchange with SP263 at 1%cut-off value is insufficient.Nevertheless,in terms of the 50%cut-off value,the two antibodies showed a good negative percent agreement.The above results need to be further verified by larger-sample studies. |
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