The flow chart of the study selection process is shown in Figure 1. A total of 629 potentially relevant articles were identified from the PubMed database. After removing duplicate records, 602 records remained. After screening the titles and abstracts, 275 articles were excluded, including 162 nonclinical studies and 113 articles not related to afatinib. The remaining 327 articles were further reviewed by reading the full text. Among them, 292 articles were excluded, including 190 articles that focused on common EGFR mutations and 106 articles that lacked data pertaining to efficacy or PFS. Finally, 31 articles met the inclusion criteria, and another 6 articles were identified by searching the reference lists of the articles whose full texts were evaluated. Overall, 37 articles were included in this pooled analysis. They were published between 2015 and 2019 and include 31 case reports and 6 retrospective clinical studies, most of which were conducted in Asia (supplement table 1).
A total of 57 patients were included in the pooled analysis, with a median age of 60 years old, from 34 to 84 years old. Their gender distribution was basically balanced (28 males, 49.1%; 29 females, 50.1%), and most of them were Asian patients (34 Asians, 59.6%; 23 non-Asians, 40.4%). More than one-third of patients had a history of smoking (23, 40.4%). Most patients had tumor stage IV (53, 93.0%). In terms of mutation type, two-thirds of patients had a single uncommon EGFR mutation. The baseline characteristics of the patients are detailed in Table 1.
Table 1
Characteristics
|
No. of patients(n=57)
|
percentage
|
Age
|
|
|
median (range)
|
60
|
34-84
|
Gender
|
|
|
Male
|
28
|
49.1
|
Female
|
29
|
50.9
|
Ethnicity
|
|
|
Asian
|
34
|
59.6
|
Non-Asian
|
23
|
40.4
|
Smoking
|
|
|
Yes
|
23
|
40.4
|
No
|
34
|
59.6
|
Stage
|
|
|
I-III
|
4
|
7.0
|
Ⅳ
|
53
|
93.0
|
Mutation type
|
|
|
Single
|
38
|
66.7
|
Multiple
|
19
|
33.3
|
Response to TKI
|
|
|
CR
|
1
|
1.8
|
PR
|
26
|
45.6
|
SD
|
16
|
28.1
|
PD
|
14
|
24.6
|
EGFR mutation analysis
In these 57 patients, there were 31 types of uncommon EGFR mutations. As shown in Figure 2a, the top six EGFR mutation types were 20 ins, S768I, 19 del + T790M, 18 del, 19 ins and L747P, with 7 cases (12.3%), 6 cases (10.5%), 5 cases (8.8%), 3 cases (5.3%), 3 cases (5.3%), and 3 cases 5.3%, respectively. For every single mutation site, these 57 patients harbored a total of 80 EGFR mutation sites, including eight 21 exon L858R mutations, five 19 exon deletions, and 67 uncommon EGFR mutation sites. As shown in Figure 2b, in addition to L858R, 19del and T790M, the uncommon mutations with higher mutation frequencies are S768I, 20 ins, 18 del, 19 ins, C797S, G719S, G719X, L747P, L833V, and L861Q, accounting for 11.3%, 8.8%, 3.8%, 3.8%, 3.8%, 3.8%, 3.8%, 3.8%, 3.8%, 3.8%, 3.8%, and 3.8% of mutations, respectively.
Clinical outcomes
Regarding the efficacy of afatinib treatment, of the 57 patients, 1 patient had a complete response (1.8%), 26 patients had a partial response (45.6%), 16 patients had stable disease (28.1%), and 14 patients had disease progression (24.6%) (Table 1). Overall, the objective response rate to treatment with afatinib was 47.4% (Table 2). Fisher’s exact and chi-squared tests show that the number of EGFR mutations is related to the treatment response to afatinib. Patients with a single uncommon EGFR mutation are more likely to show a good tumor response after receiving afatinib than patients with multiple mutations (ORR:57.9% vs 26.3%,HR:0.260,95%CI:0.078-0.869, p=0.029). However, age (HR:2.512,95%CI:0.863-7.310, p=0.091), gender (HR:0.612,95%CI:0.2115-1.744, p=0.358), gender (HR:0.612,95%CI:0.2115-1.744, p=0.358), ethnicity (HR:0.571,95%CI:0.195-1.675, p=0.307), smoking history (HR:0.970,95%CI:0.336-2.798, p=0.955), and tumor stage (HR:1.120,95%CI:0.078-8.552, p=0.913) were not correlated with efficacy (Figure 3).
The median PFS was 7.0 months (Table 2). As shown in Figure 4a, the Kaplan–Meier curves for PFS revealed that patients with a single uncommon EGFR mutation had a longer PFS than patients with multiple mutations (HR:2.906,95%CI:1.496-5.646, p=0.002). In addition, for patients with a good treatment response, the PFS was also longer (HR: 2.902, 95% CI: 1.522-5.533, p=0.001) (Figure 4b). However, age (HR: 0.872, 95% CI: 0.471-1.617, p=0.664), gender (HR: 1.072, 95% CI: 0.580-1.984, p=0.824), ethnicity (HR: 1.375, 95% CI: 0.733-1.440, p=0.321), smoking history (HR: 0.757, 95% CI: 0.398-1.440, p=0.397), and tumor stage (HR: 0.296, 95% CI: 0.041-2.160, p=0.230) were not correlated with efficacy. (Figure 5).
Subgroup analysis
Based on the number of EGFR mutation sites, we conducted further analysis of treatment response and survival. A total of 38 patients harbored a single mutation. After treatment with afatinib, a total of 22 patients showed treatment response, with an objective response rate of 57.9% and a median PFS of 10.0 months. Further analysis revealed that the number of patients with mutations in exon 18, exon 19, exon 20, and exon 21 was 10, 10, 13, and 5, respectively; these patients had objective response rates of 70.0%, 60.0%, 46.2%, and 60.0%, respectively; and they had a median PFS of 11.0, 12.0, 7.4 and 10.9 months, respectively. For a single uncommon EGFR mutation, patients with mutations in exon 20 and exon 21 had poorer tumor responses and prognoses. In contrast, patients whose mutation sites were located on exon 18 and exon 19 had a better ORR and mPFS than the average value. For patients with multiple mutations, the ORR and mPFS were worse than among patients with a single mutation (ORR: 26.3% vs 57.9%, mPFS: 3.6 months vs 10.0 months, respectively). The ORR and PFS of patients with multiple mutations containing common mutations were the worst, at only 23.1% and 2.2 months, respectively (Table 2).
Table 2
Outcomes of different singel mutation location and different double mutation pattrens
Categories
|
No. of patients
|
OR
|
ORR
|
mPFS
|
Singel mutation
|
38
|
22
|
57.9%
|
10.0
|
Exon 18
|
10
|
7
|
70.0%
|
11.0
|
Exon 19
|
10
|
6
|
60.0%
|
12.0
|
Exon 20
|
13
|
6
|
46.2%
|
7.4
|
Exon 21
|
5
|
3
|
60.0%
|
10.9
|
Double mutation
|
19
|
5
|
26.3%
|
3.6
|
Common +
|
13
|
3
|
23.1%
|
2.2
|
Common -
|
6
|
2
|
33.3%
|
3.6
|
All
|
57
|
27
|
47.4%
|
7.0
|