Clinical impact of rebiopsy among patients with epidermal growth factor receptor-mutant lung adenocarcinoma in a real-world clinical setting

INTRODUCTION

Lung cancer is still the leading cause of cancer-related death worldwide.¹ According to the histological classification, non-small cell lung cancer (NSCLC) accounts for over 80% of lung cancer cases,² and adenocarcinoma is the most common subtype worldwide.³ Although immunotherapy, such as immune checkpoint inhibitors (ICIs), has recently been highlighted,⁴ epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are recommended as a first-line therapy for patients with advanced adenocarcinoma with EGFR mutation.^{5, 6}

Unfortunately, acquired resistance occurs in the majority of patients treated with EGFR-TKIs, with various resistance mechanisms.^7-9 A point mutation in exon 20 (T790M mutation) accounts for over half of the acquired resistance to EGFR-TKIs,^{10, 11} which has led to the development of osimertinib, a third-generation EGFR-TKI.¹² Thus, it is important to identify T790M mutations in patients who are suspected to have acquired resistance to EGFR-TKIs in a clinical setting. There is growing evidence regarding repeat biopsy.^13-15 However, in many previous studies, the number of patients was relatively small, or the main focus was on feasibility or diagnostic accuracy. In addition, there is a lack of data on the prevalence and clinical characteristics of acquired T790M mutation in a real-world setting.

Here, we investigated the risk factors of acquired T790M mutation among patients with lung adenocarcinoma with EGFR mutation who were treated with EGFR-TKIs. The intent was to identify the clinical impact of rebiopsy in a real-world clinical setting.

Population cohort

This multicenter, retrospective cohort study involved “the lung cancer patients who were EGFR mutation positive in Korea” cohort. Data were gathered from The Korean Academy of Tuberculosis and Respiratory Disease. From January 2007 to June 2017, adenocarcinoma patients with EGFR mutation who underwent rebiopsy during the follow-up period and were treated with EGFR-TKIs were included in this study. Among the eligible patients, those who did not undergo an EGFR mutation test, had an interval of more than one month between biopsy and EGFR detection, or whose information about the EGFR mutation test was unknown, were excluded. Patients who initially had T790M mutation were also excluded (Figure 1).

A standardized protocol was used to collect data regarding age, gender, smoking history, clinical stage at the time of initial diagnosis, brain metastasis, repeat biopsy performed during the follow-up period, and treatment information regarding conventional chemotherapy and EGFR-TKIs. The repeat biopsy information included the date and site of biopsy, histological diagnosis, timing correlation between biopsy and EGFR-TKI treatment, EGFR mutation status, and its detection method. The previous TKI treatment duration was set as the sum from the start date to the end date of each TKI administration. The study protocol was approved by the Institutional Review Board of Asan Medical Center (approval number: 2018-0092). Requirements for written informed consent were waived due to the retrospective nature of the present study and the use of anonymized patient data.

Evaluation of EGFR mutations

The peptide nucleic acid (PNA)-mediated PCR clamping method is the gold standard for the detection of EGFR mutations. The method uses formalin-fixed paraffin-embedded tumor tissue obtained via tissue biopsy. Previously, direct sequencing (DS) methods were commonly used to compare amplified sequences with GenBank EGFR through PCR. Recently, Mutyper methods have been used, in which circulating-free tumor DNA (ctDNA) is detected in plasma after collecting blood into an EDTA anticoagulation bottle. As this cohort was recruited from multiple institutions, various EGFR mutation detection techniques were used. They included direct sequencing and the aforementioned PNA clamping method and Mutyper as mentioned above. We used the PNAClamp EGFR, PANAqPCR, and PANAMutyper EGFR mutation detection kits (all from Panagene) to detect the various EGFR mutations.

Statistical analysis

All values are expressed as median and interquartile range (IQR) for continuous variables or percentages for categorical variables. Student's t-test or the Mann-Whitney U test was used to examine continuous data. A chi-square test or Fisher's exact test was used to examine the categorical data. Logistic regression and Cox regression analysis were performed to identify the factors affecting T790M mutation. Furthermore, variables with p < 0.2 in the univariate analysis were entered into the multivariate models. Survival analysis was performed using Kaplan-Meier analysis and log-rank test. IBM SPSS statistics v.20 (IBM, Armonk) was used for analyses. A statistically significant difference was defined as a p-value < 0.05.

Patient characteristics

A total of 352 patients with adenocarcinoma and EGFR mutations were included in this study. The median follow-up period was 22 months (interquartile range [IQR]: 15–36 months). The baseline characteristics of patients are summarized in Table 1. The median age was 58 years (IQR: 52–68 years); 34.9% patients were male and 29.0% were ever-smokers. Most patients (90.9%) initially had stage IV adenocarcinoma and the lungs were the most common initial biopsy site (67.5%). Regarding EGFR mutation, exon 19 deletion was the most common (60.8%), followed by L858R (34.7%), L861Q (5.4%), and G719X (2.3%). A total of 80.6% of patients were treated with an EGFR-TKI. They initially included gefitinib, erlotinib, or afatinib. Approximately 13.4% received cytotoxic chemotherapy.

Acquired T790M mutation

All eligible patients underwent repeat biopsy during the follow-up period. The median duration from initial biopsy to rebiopsy was 17 months (IQR 10–29 months). T790M mutation was identified in 156 (41.9%) patients at the time of rebiopsy. The comparison of characteristics according to the T790M mutation status at the time of rebiopsy is shown in Table 2. There were no significant differences in demographics, brain metastasis, rebiopsy site, and initial TKI treatment between the two groups. However, patients with T790M mutation had a longer duration between initial biopsy to rebiopsy (median, 19 vs. 16 months, p = 0.008) compared to the patients without T790M mutation. According to the initial EGFR profile, exon 19 deletion was more frequent in patients with T790M mutation (67.3 vs. 55.6%, p = 0.028), whereas L858R was less frequent in the T790M mutation group (28.8 vs. 39.3%, p = 0.043).

Treatment information according to T790M mutation status at the time of rebiopsy is summarized in Table S1. Although there were no significant differences in number, type, and the best response to EGFR-TKIs between the two groups, patients with T790M mutation had a longer EGFR-TKI treatment duration (median 15 vs. 11 months, p < 0.001) compared to patients without T790M mutation. Among the 156 patients with T790M mutation, 66 were treated with the third-generation TKI, osimertinib. In 90 patients who were not treated with osimertinib after repeat biopsy, 29 (32.2%) received conservative management, 24 (26.7%) participated in clinical trials, 18 (20%) received cytotoxic chemotherapy, and 19 (21.1%) received different types of EGFR-TKI.

Risk factors of acquired T790M mutation

To identify the risk factors of acquired T790M mutation, we performed logistic regression analysis (Table 3) and Cox regression analysis (Table 4). Univariate logistic analysis showed that the presence of exon 19 deletion (odds ratio [OR], 1.643; 95% CI: 1.061–2.545; p = 0.026) and the absence of L858R (OR, 0.627; 95% CI, 0.400–0.982; p = 0.042) were associated with T790M mutation. The previous EGFR-TKI treatment duration (OR, 1.039; 95% CI: 1.019–1.058; p < 0.001) was also associated with T790M mutation. Multivariate logistic analysis showed that previous EGFR-TKI treatment duration (OR, 3.580; 95% CI: 2.257–5.678; p < 0.001) was the only factor independently associated with T790M mutation. In addition, the presence of exon 19 deletion displayed a trend toward association with T790M mutation (OR, 1.509; 95% CI: 0.955–2.386; p = 0.078).

The univariate Cox proportional hazard model showed that only brain metastasis at the initial biopsy was associated with an acquired T790M mutation (hazard ratio [HR], 1.439; 95% CI: 1.035–2.001; p = 0.030). In the multivariate Cox proportional hazard model, brain metastasis tended to be associated with T790M mutation. Patients with brain metastasis had a shorter duration between initial biopsy and rebiopsy (median, 15 vs. 19 months; p = 0.003) compared to patients without brain metastasis (Table S2). Patients with brain metastasis also had a shorter EGFR-TKI treatment duration (median, 12 vs. 14 months; p = 0.009) than patients without brain metastasis.

Survival analysis

The T790M mutation positive group (n = 156) had an improved survival (median survival period: 43 months vs. not reached, p = 0.001) than the T790M mutation negative group (n = 196, Figure 2(a)). Even if the start date was the date of rebiopsy for the survival analysis, the T790M mutation positive group still had an improved survival (median survival period: 12 vs. 18 months, p = 0.004) than the T790M mutation negative group (Figure 2(b)).

Among the patients with T790M mutation at the time of rebiopsy, the osimertinib user group (n = 90) had a better one-year survival (68.7 vs. 58.3%, p = 0.048) than the osimertinib nonuser group (n = 66, Figure 3). The baseline and clinical characteristics in patients with T790M mutation treated with osimertinib are shown in Table S3. There were no significant differences in the clinical characteristics between the two groups.