Efficacy of epidermal growth factor receptor inhibitors in combination with chemotherapy in advanced non-small cell lung cancer: A meta-analysis of randomized controlled trials

The role of a combination of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) and chemotherapy for non-small-cell lung cancer (NSCLC) has not been well established. To clarify this problem, we performed a meta-analysis with 15 studies identified from PubMed, EMBASE and the Cochrane Library. We found that the combined regimen had a significant benefit on progression-free survival (PFS) (hazard ratio (HR) = 0.80; 95% CI = 0.71–0.90; P < 0.001) and the objective response rate (ORR) (RR = 1.35; 95% CI = 1.14–1.59; P < 0.001). However, the combined regimen had no significant impact on overall survival (OS) (HR = 0.96; 95% CI = 0.90–1.03; P = 0.25). Subgroup analysis showed significantly higher OS advantages in EGFR mutation positive patients (P = 0.01), never smokers (P = 0.01), Asian patients (P = 0.02), patients receiving second-line treatment (P < 0.001), and those receiving a sequential combination of EGFR-TKIs and chemotherapy (P = 0.005). The combination regimen showed a higher incidence of grade 3–4 toxicities (leucopenia, neutropenia, febrile neutropenia, anemia, rash, fatigue and diarrhea). In summary, the combination of EGFR-TKIs plus chemotherapy in advanced NSCLC achieved a significantly longer PFS and a higher ORR but not longer OS. Well-designed prospective studies are needed to confirm these findings.


IntroductIon
Lung cancer is the leading cause of cancer death worldwide [1].The majority of new cases are advanced non-small-cell lung cancer (NSCLC) at the time of diagnosis, and palliative therapy with platinum-based doublets are the standard therapy [2].However, no doublet regimen has proven to be superior, and survival outcomes are poor [3].Therefore, novel agents are urgently needed for this disease, and epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) are among the most widely used agents to serve this purpose.
Therefore, we performed a meta-analysis of randomized controlled trials to comprehensively examine the efficacy and safety of EGFR-TKIs in combination with chemotherapy for the treatment of advanced NSCLC and to find the most effective combinatorial strategy.

rEsuLts study selection and characteristics
In the present study, 1,235 articles were identified by the initial search strategy.Through reading the study titles and abstracts, 1,120 articles were removed.After we reviewed the full texts of the 46 potentially eligible articles in detail and identified articles through conference, 15 trials meeting the inclusion criteria were included for the final analysis.A flowchart depicting the study selection is shown in Figure 1.Among these 15 trials, 5,861 patients with advanced NSCLC were investigated.The characteristics of the 15 trials are shown in Table 1.

Progression-free survival
The PFS analysis was based on 14 trials.The meta-analysis showed that the EGFR-TKI combinations significantly reduced the risk of disease progression compared with EGFR-TKIs or chemotherapy alone (HR = 0.80; 95% CI = 0.71-0.9;P < 0.001) (Figure 2).Subgroup analysis was conducted according to the EGFR mutation status, smoking status, line of treatment, dose schedules and ethnicity (Figure 3).Subgroup analysis showed that the EGFR-TKI combination was associated with a lower risk of disease progression in never smokers (HR = 0.51; 95% CI = 0.40-0.65;P < 0.001).However, EGFR-TKIs did not show a treatment advantage in smoking patients.In addition, the combination group showed a significant improvement in PFS compared to the group receiving chemotherapy alone (HR = 0.76; 95% CI = 0.63-0.91;P < 0.002), but this difference was not statistically significant compared to EGFR-TKIs alone (HR = 0.94; 95% CI = 0.86-1.01;P = 0.10) (Supplementary Figures S1-S2).

objective response rate
Data for the objective response rate (ORR) were available from all 15 trials.The results of the collaboration analysis showed heterogeneity among the various studies (I 2 = 71%, P < 0.05); thus, randomeffects model was employed for the analysis.The metaanalysis demonstrated that the ORR of the EGFR-TKI plus chemotherapy group was significantly higher than the EGFR-TKI-or chemotherapy-alone group (RR = 1.35, 95% CI = 1.14-1.59;p < 0.001) as shown in Figure 6.

Publication bias
In the present meta-analysis, no publication bias for PFS and OS was found according to Begg's test (P = 0.101 and P = 0.583; Figure 7A-7B).

dIscussIon
Although platinum-based doublet therapy remains the mainstay of treatment for most patients with advanced NSCLC [27], EGFR-TKIs have assumed an increasingly important role, particularly in patients harbouring EGFRactivating mutations [28].However, the combination of chemotherapy and EGFR-TKIs has been long debated.
To derive a more precise estimate of the effectiveness of EGFR-TKIs in combination with chemotherapy, we systematically reviewed the published studies and carried out a meta-analysis.The meta-analysis demonstrated that the combination of EGFR-TKIs plus chemotherapy in advanced NSCLC achieved significantly longer PFS and higher ORR.The reason may be that the combination regimen enhances anti-proliferative and cytotoxic activities, as demonstrated in human NSCLC cell lines and tumor models [29][30].However, our results showed that there was no statistically significant difference between the two groups in OS.The reason may be that the differences in OS are potentially affected by the subsequent treatment options.Although the survival data were not statistically significant difference between the two arms, there was a trend in favour of the combination arm.
In the subgroup of patients with EGFR mutations, an improvement in OS was observed for the combination arm.Our results were consistent with the findings of a previous study, which also demonstrated that the addition of EGFR-TKIs to chemotherapy significantly prolonged OS in patients harbouring EGFR mutations [31].However, no significant difference in OS was noted in EGFR mutation negative patients.These findings demonstrated that the EGFR status may serve as a biomarker to identify patients who can benefit the most from combination therapy and further emphasized the need to test the mutation status at the time of diagnosis.Unfortunately, the EGFR-mutation status was assessed in only a few patients enrolled in eligible trials.Therefore, this result should be interpreted with caution.A head-to-head study is needed to define the value of the combination therapy in the patients with EGFR mutations.Previous studies have suggested that never smokers and Asian patients are more likely to harbour EGFR mutations and benefit more from EGFR-TKIs [32][33].Furthermore, a history of never smoking was a significant independent predictor for survival in EGFR-TKI treatment [34].Similarly, this meta-analysis showed that the addition of EGFR-TKIs to chemotherapy had an improvement in the never smoker and Asian-dominant groups.
Two methods of drug delivery were adopted in the combination group, including concurrent and intercalated administration.Previous studies have confirmed a lack of efficacy on the concurrent administration of EGFR-TKIs and chemotherapy [18][19].In our subgroup analysis, we found that concurrent administration did not confer a survival benefit to patients with advanced NSCLC.Concurrent administration may not be effective because of TKI-induced, G1-phase cell-cycle arrest [35].Our results are consistent with those in a previous systematic review [36].The second approach was to administer EGFR-TKIs as intercalated therapy.Our results showed that the intercalated regimen improved the PFS and OS.When EGFR-TKI and chemotherapy are given in a sequentially intercalated way, thus achieving pharmacodynamic separation of the two agents, the inhibitory drug interaction could be avoided [37].Moreover, EGFR-TKIs were administered not only as a sequential intercalated regimen during chemotherapy but also as maintenance therapy after the end of chemotherapy in intercalated therapy.Subsequent maintenance treatment possibly reduced the effect of the intercalated administration in our pooled analysis because TKI maintenance therapy has been shown to be beneficial in survival [38][39].Despite this OS benefit in the second-line treatment, only 3 trials were evaluated for OS, and the results should be treated with caution.
In 2013, OuYang et al. [40] also analyzed the treatment effect of the combined regimen on PFS and OS.This meta-analysis was based on only 8 studies.Furthermore, it did not include data from second-line therapy for advanced NSCLC.This study demonstrated that the combined regimen resulted in superior PFS  (HR = 0.81, 95% CI 0.69-0.95,P = 0.01).The results were consistent with those of our study.Another recent meta-analysis compared chemotherapy plus erlotinib with chemotherapy alone.This also showed an improvement in PFS from the combined regimen, but it did not improve in OS [31].However, that meta-analysis did not include another important EGFR-TKI, gefitinib, and it did not compare the ORR differences in the two arms.
This meta-analysis had several limitations.First, all of data were extracted from published studies, possibly resulting in publication bias.Second, the EGFR-mutation status was only assessed in a few patients enrolled in eligible trials.Third, all of the clinical trials included in this meta-analysis were conducted in various countries with patients of different ethnicities; these differences may have selected for biases.Fourth, the quality of the included studies may slightly affect the pooled results.
In summary, our study indicated that EGFR-TKIs combined with chemotherapy present a viable therapy for patients with advanced NSCLC.Importantly, the present study suggests that there is a larger magnitude of benefit for Asians, never smokers, and EGFR mutation patients and further suggests that intercalated therapy is the most effective combinatorial strategy.

MAtErIALs And MEtHods search strategy and study selection
Two authors (Zhang MH and Guo HS) independently carried out a comprehensive systematic search for published articles using the PubMed, EMBASE, and Cochrane databases.The deadline of the included articles was September 2015.The search keywords used were as follows: "Erlotinib OR Tarceva OR Gefitinib OR Iressa or EGFR-TKI" and "non-small cell lung OR nonsmall cell lung carcinoma" and "randomized controlled trial OR controlled clinical trials, randomized".We also manually reviewed the meeting abstracts of the annual meetings of American Society of Clinical Oncology (ASCO), European Society of Medical Oncology congresses (ESMO) and the World Conference of Lung Cancer (WCLC) from 2004 to 2015.The related references from the included studies were also manually examined.

Eligibility criteria
The inclusion criteria were as follows: (1) the patients had histopathologically confirmed advanced NSCLC; (2) the combined regimen of EGFR-TKI and chemotherapy was compared with chemotherapy or EGFR-TKI alone; (3) the studies were phase II or III prospective randomized controlled clinical trials; (4) at least one of OS, PFS and ORR was evaluated; (5) the sample size was greater than 50 cases because small samples could introduce marked bias.When duplicate publications were identified, only the newest or most informative single article was selected.

data extraction and quality assessment
This meta-analysis was performed in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) Statement [41].Data were extracted independently by two reviewers (Zhang MH and Guo HS), and any disagreements between the two reviewers were resolved by consensus involving a third reviewer (Zhao S).The primary end point of this metaanalysis was OS.The secondary end points included PFS, ORR, as well as grade 3-4 adverse events.For each study, the following information was extracted: author's name, year of publication, phase, line of treatment, drug delivery, dominant ethnicity, treatment comparison, number of patients, median age, number of females, number of smokers, activating EGFR-mutant, ORR and adverse events (grade 3-4 events), hazard ratios (HRs) and 95% confidence intervals (CIs) for PFS and OS.If the HRs were not directly reported, we contacted the authors of the primary studies for additional data.If the author did not respond, we extracted data from survival curves [42].The quality of the included study was assessed using the Jadad score [43].

statistical analysis
Survival analysis was conducted using the intent-totreat (TTP) population.The risk ratio (RR) was calculated as an effect measure for ORR using the Mantel Haenszel method, and HR was calculated for PFS and OS using the inverse variance method.Statistical heterogeneity was evaluated using chi-squared test and I 2 .Statistically significant heterogeneity was defined as a chi-squared P value < 0.1 or an I 2 statistic > 50%.If heterogeneity was observed, we used a random-effects model to reduce the impact of heterogeneity on the results.If heterogeneity was not observed, a fixed-effects model was used.The potential publication bias was assessed by Begg's test.All of the statistical analyses were performed using Review Manager Version 5.2 (Revman the Cochrane Collaboration; Oxford, England) and STATA version12.0(Stata Corporation; College Station, TX, USA).P values < 0.05 were considered to indicate statistical significance.All P values and 95% CIs were two-sided.

Figure 1 :
Figure 1: Flow chart of study selection.

Figure 3 :
Figure 3: Forest Plot of subgroup Analysis for PFs.

Figure 2 :
Figure 2: Forest Plot of Meta-analysis for PFs.

Figure 4 :
Figure 4: Forest Plot of Meta-analysis for os.

Figure 5 :
Figure 5: Forest Plot of subgroup Analysis for os.

Figure 6 :
Figure 6: Forest Plot of Meta-analysis for orr.