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Keywords:

  • tyrosine kinase inhibitors;
  • non–small cell lung cancer;
  • B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11;
  • Bim;
  • epidermal growth factor receptor;
  • EGFR;
  • polymorphism

Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES

BACKGROUND

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are widely used for the treatment of patients with advanced non–small cell lung cancer (NSCLC) who have EGFR mutations. Recent studies have indicated that some patients with positive mutations were refractory to EGFR TKIs if they harbored a B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (Bim) deletion polymorphism. The objective of the current work was to retrospectively study the Bim deletion polymorphism in Chinese patients with NSCLC and its correlation with the efficacy of EGFR TKIs.

METHODS

Distribution of the Bim polymorphism was detected using polymerase chain reaction analysis and direct sequencing of DNA from peripheral neutrophils in samples from 352 patients with NSCLC. Of the 352 patients, 166 who received TKI therapy and had an activating mutation identified were involved in further analysis. Progression-free survival (PFS) was the primary endpoint of the subsequent analyses, and the incidence of the Bim polymorphism and its relation to clinical benefit from EGFR TKIs also were investigated.

RESULTS

In total, 45 of 352 patient samples (12.8%) had the Bim deletion polymorphism, which was distributed randomly with regard to various clinical characteristics. In patients with EGFR mutations who received treatment with TKIs, the median PFS and the median objective response rate were 4.7 months and 25%, respectively, for those with the Bim deletion polymorphism versus 11 months (P = .003) and 66% (P = .001), respectively, for those with wild-type Bim. Cox regression analysis identified Bim status (P = .016) and sex (P = .002) as independent factors predicting clinical benefit from EGFR TKIs in patients with EGFR-mutated NSCLC.

CONCLUSIONS

The incidence of the Bim deletion polymorphism was approximately 13% in this study, and it was associated with a poor clinical response to EGFR TKIs in patients who had NSCLC with EGFR mutations. Cancer 2014;120:2299–2307. © 2014 American Cancer Society.


INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES

Lung cancer is the leading cause of major cancer incidence and mortality worldwide.[1, 2] With the introduction of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs), approaches to non–small cell lung cancer (NSCLC) treatment have changed dramatically in the last decade.[3, 4] Compared with traditional platinum-based doublet chemotherapy, EGFR TKIs like erlotinib and gefitinib have produced amazing effects in patients with EGFR-mutant NSCLC by prolonging progression-free survival (PFS) significantly while also improving quality of life.[5, 6] However, the efficacy of TKIs is not consistent for every patient. Approximately 10% of patients have primary resistance to TKIs, even those who have EGFR mutations; and others who respond at first eventually will develop acquired (secondary) resistance (in approximately 1 year).[7-9] However, the mechanism of primary resistance remains unclear, because it is a complicated, multifactor process involving a nonsensitive mutation, gene polymorphism, a cancer stem cell subpopulation, and related protein (eg, amphiregulin) expression.[10-13] Thus, investigations are needed to further understand and overcome these possible mechanisms.

Bim, also known B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (BCL2L11), is a member of the Bcl-2 family gene that encodes the protein Bim. By binding to all members of the prosurvival Bcl-2 subfamily with high affinity, Bim serves as a key element in promoting apoptosis.[14, 15] A previous study demonstrated that, in EGFR-TKI–sensitive cell lines, Bim triggered apoptosis through the intrinsic caspase pathway under EGFR-TKI treatment.[16] In addition, another study indicated that a Bim germline deletion polymorphism was correlated intensively with primary resistance to TKI therapy.[17] That deletion polymorphism led to impaired expression of BH3-containing Bim isoforms, which compromised the therapeutic efficacy of EGFR TKIs in patients with NSCLC who had EGFR mutations. It is noteworthy that this mutation was observed only in individuals of East Asian ethnicity, indicating its racially heterogeneous nature.

Although the importance of the Bim deletion polymorphism in patients with NSCLC who receive EGFR-TKI treatment has garnered much attention, the distribution and clinicopathologic characteristics of this polymorphism in Chinese patients with NSCLC and its prognostic value for EGFR-TKI outcomes remain to be determined. Therefore, the objective of this study was to gain a more comprehensive understanding of the Bim polymorphism in Chinese patients with NSCLC and especially to determine the efficacy of TKI therapy in the subgroup of patients who have EGFR mutations.

MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES

Patients and Samples

This was a retrospective study inspired by the work of Ng and colleauges.[17] In total, 352 consecutive patients with NSCLC who were hospitalized at Shanghai Pulmonary Hospital from January 1, 2008 to March 31, 2013 were identified in our database. All patients met the following inclusion criteria: they signed an informed consent form; they had histologically confirmed NSCLC; they were aged >18 years; and they had adequate formalin-fixed, paraffin-embedded (FFPE) tissue available to detect EGFR mutations, including sufficient neutrophils to determine their Bim polymorphism status. Before starting any treatment, a complete medical history interview, physical examination, laboratory tests, and radiology examinations were carried out for each patient. In total, 239 patients, regardless of line of treatment, received 250 mg gefitinib or 150 mg erlotinib daily until they developed either disease progression or intolerable toxicity.

DNA Exaction and EGFR Mutation Detection

DNA from tumor tissue was extracted using the DNeasy Blood and Tissue Kit or the QIAamp DNA FFPE Tissue Kit (both from Qiagen, Hilden, Germany) according to the manufacturer's protocol. The Amplification Refractory Mutation System (ARMS) was used to detect EGFR mutations using the Human EGFR Gene Mutations Fluorescence Polymerase Chain Reaction Diagnostic Kit (AmoyDx, Xiamen, China).

Bim Genotyping and Direct Sequencing

All samples were amplified by polymerase chain reaction (PCR) to detect Bim polymorphisms using the following primer sequences: wild-type (WT) Bim forward primer, 5′-ACTGTAAAACGACGGCCAGTCCTCATGATGAAGGCTAACTCAA-3′; and reverse primer, 5′-ACCAGGAAACAGCTATGACCAACCTCTGACAAGTGACCACCA-3′. For the Bim deletion polymorphism, the forward primer sequence was the same as that used for WT Bim, and the reverse sequence was 5′-ACCAGGAAACAGCTATGACCGGCACAGCCTCTATGGAGAACA-3′ (for details, see Fig. 1) The reaction condition was 95°C for 10 minutes followed by 40 cycles at 94°C for 30 seconds, 60°C for 30 seconds, and 72°C for 30 seconds; and a final extension at 72°C for 10 minutes using the ExTaq Polymerase premix PCR Kit (TaKaRa Bio Inc., Shiga, Japan). PCR products (173 base pairs [bp] for the Bim deletion polymorphism and 216 bp for WT Bim) were then separated on a 3% agarose gel with nucleic acid dye by electrophoresis and were purified before direct sequencing.

image

Figure 1. The B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (Bim) deletion polymorphism was measured using polymerase chain reaction analysis and direct sequencing. (a) This is an illustration of the primer for the Bim deletion polymorphism located at chromosome 2. (b) The results from agarose gel electrophoresis revealed a wild-type (WT) band (216 base pairs [bp]) and a deletion (DEL) band (173 bp). (c) The results from direct sequencing confirmed the Bim deletion polymorphism in samples.

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Data Collection and the Evaluation of TKI Clinical Efficacy

Clinical data were recorded in detail systemically when patients were hospitalized for the first time. Smoking status was defined as follows: never smokers were those who had consumed <100 cigarettes in their whole life, and smokers were those who had smoked >100 cigarettes in their whole life (including former and current smokers). Pathologic diagnoses were reviewed by 3 experienced pathologists independently, and disease stages were classified according to the seventh edition the International Association for the Study of Lung Cancer TNM classification system. Tumor response was reviewed every 6 weeks based on Response Evaluation Criteria in Solid Tumors version 1.1. PFS was the primary endpoint of this study, because we mainly wanted to evaluate the role played by the Bim polymorphism in EGFR-TKI therapy. PFS was calculated from the date TKI treatment was initiated to the date of either disease progression or death from any cause. Primary resistance was defined according to a previous report by Jackman et al as disease progression in <6 months without any evidence of response when receiving TKIs.[18] Because of the possibility that the predictive power of the Bim polymorphism might be diluted by the poor response to TKIs among patients who had WT EGFR, only patients who had activating EGFR mutations were entered into the subsequent analyses of PFS and Bim polymorphisms.

Statistical Analysis

PFS was plotted and calculated using the Kaplan-Meier method, and differences between groups were compared using the log-rank test. Differences in distribution of the Bim deletion polymorphism according to patients' baseline characteristics were compared using the Pearson chi-square test or the Fisher exact test. Cox regression was used to analyze independent factors. Hardy-Weinberg equilibrium was tested using the Pearson chi-square test. All analyses were carried out using PASW version 18.0 (SPSS Statistics, IBM, New York, NY). The 2-sided significance level was set at P < .05.

Ethics

This study was approved by the Ethics Committees of Shanghai Pulmonary Hospital Affiliated with Tongji University and was carried out in accordance with the World Medical Association's Declaration of Helsinki. Informed consent was obtained from each patient.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES

Clinicopathologic Characteristics

In total, 352 patients with histologically confirmed NSCLC who were hospitalized at Shanghai Pulmonary Hospital from January 1, 2008 to March 31, 2013 were selected for Bim polymorphism evaluation. All patients were of Chinese Han ethnicity. The median age at diagnosis was 59 years (age range, 32-81 years), 255 patients were never smokers, and 97 were smokers. There were 273 patients with adenocarcinoma, 39 with squamous NSCLC, 27 with adenosquamous NSCLC, and13 with NSCLC-not otherwise specified. There were 124 WT EGFR mutations, 122 EGFR exon 19 deletions, 102 EGFR exon 21 substitution of lysine with arginine at amino acid position 858 (L858R) mutations, and 4 other rare mutations (including mutations of EGFR exons 18 and 20). Only 1 patient in the group with EGFR mutations had a coexistent substitution of threonine with methionine at EGFR amino acid position 790 (T790M) mutation. Patient characteristics are detailed in Table 1. One hundred sixty-six of 352 patients who received EGFR TKIs (regardless of the line of treatment) and had an activating mutation participated in additional analyses of Bim polymorphisms and TKI efficiency.

Table 1. Distribution of the Bim Polymorphism in the Overall Study Population With Non–Small Cell Lung Cancer
 No. of Patients (%) 
  Bim Polymorphism Status 
VariableAll, n = 352Positive, n = 45Negative, n = 307P
  1. Abbreviations: Bim, B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (BCL2L11); EGFR, epidermal growth factor receptor; L858R, substitution of lysine with arginine at EGFR amino acid position 858; TKIs, tyrosine kinase inhibitors.

  2. a

    This P value is for the comparison between adenocarcinoma histology and nonadenocarcinoma histology (adenosquamous, squamous, and others).

  3. b

    Others included those who had non–small cell lung cancer, not otherwise specified and those who had large cell lung carcinoma.

  4. c

    This P value is for the comparison between mutated EGFR versus wild-type EGFR.

  5. d

    Others included those who had EGFR mutations in exons 18 and 20.

  6. e

    Others included those who received chemotherapy and radiotherapy.

Age, y    
Median [range], y59 [32-81]59 [39-75]59 [32-81].586
<65254 (72.2)34 (75.6)220 (71.7) 
≥6598 (27.8)11 (24.4)87 (28.3) 
Sex   .499
Men173 (49.1)20 (44.4)153 (49.8) 
Women179 (50.9)25 (55.6)154 (50.2) 
Smoking history   .374
Never smoked255 (72.4)30 (66.7)225 (73.3) 
Smoker97 (27.6)15 (33.3)82 (26.7) 
Histology   .211a
Adenocarcinoma273 (77.6)33 (73.3)240 (78.2) 
Squamous39 (11.1)7 (15.6)32 (10.4) 
Adenosquamous27 (7.6)3 (6.7)24 (7.8) 
Othersb13 (3.7)2 (4.4)11 (3.6) 
EGFR mutation   .608c
Wild type124 (35.2)17 (37.8)107 (34.8) 
Exon 19 deletion122 (34.7)15 (33.3)107 (34.8) 
Exon 21 L858R102 (30)12 (26.7)90 (29.3) 
Othersd4 (1.1)1 (2.2)3 (0.9) 
Disease stage    
IIIB42 (11.9)8 (17.8)34 (11.1) 
IV310 (88.1)37 (82.2)273 (88.9) 
Treatment    
TKIs239 (67.9)28 (62.2)211 (68.7) 
Otherse113 (32.1)17 (37.8)96 (31.3) 

Bim Deletion Polymorphism Distribution

The Bim polymorphism (2903-bp deletion) was detected by PCR and direct sequencing. Forty-five of 352 patients (12.8%) harbored this deletion polymorphism, and the incidence was consistent with a previous report.[17] Electrophoretic and sequencing results are provided in Figure 1. The median age of all Bim deletion polymorphism carriers was 59 years (range, 39-75 years). Twenty-five carriers were women, and 30 carriers were never smokers. Adenocarcinoma accounted for most of the Bim deletion polymorphism carriers. Seventeen of 45 carriers (37.8%) had WT EGFR, and the remaining carriers were positive for a mutation. Eight patients had stage IIIB disease, and 37 had stage IV disease. The clinicopathologic features of all patients are detailed in Table 1. Because it is a genetic germ-line polymorphism, its pattern of distribution did not differ significantly between the various EGFR mutation types, histologic types, sexes, smoking histories, age groups, or disease stages.

Outcomes of EGFR-TKI Treatment in Patients With Activating Mutations

Among 166 patients, 140 (84.3%) received erlotinib, and 26 (15.7%) received gefitinib. At the cutoff date of March 31, 2013, 135 patients (81.3%) had developed disease progression, and 31 were still receiving TKI therapy without progression. The overall response rate (ORR) was 62%, and the median PFS was 9.2 months (95% confidence interval [CI], 7.25-11.15 months). The clinicopathologic features and outcomes of the EGFR-mutated patients who received TKI therapy are detailed in Table 2, and data on the Bim polymorphism carriers who had mutations are provided in Table 3.

Table 2. Clinicopathologic Features and Outcomes of Enrolled Patients With Activating Mutations Who Received Treatment With Tyrosine Kinase Receptor Inhibitors
CharacteristicNo. of Patients (%)
  1. Abbreviations: Bim, B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (BCL2L11); ECOG PS, Eastern Cooperative Oncology Group performance status; EGFR, epidermal growth factor receptor; L858R, substitution of lysine with arginine at EGFR amino acid position 858.

  2. a

    Others included those who had non–small cell lung cancer, not otherwise specified and those who had large cell lung carcinoma.

  3. b

    Others included those who had EGFR mutations in exons 18 and 20.

Age: Median [range], y60 [32-81]
Sex 
Men80 (48.2)
Women86 (51.8)
Smoking history 
Never smoked125 (75.3)
Smoker41 (24.7)
ECOG PS 
03 (1.8)
1155 (93.4)
27 (4.2)
31 (0.6)
40 (0)
Histology 
Adenocarcinoma140 (84.3)
Squamous8 (4.8)
Adenosquamous9 (5.4)
Othersa9 (5.4)
EGFR mutation 
Exon 19 deletion86 (51.8)
Exon 21 L858R77 (46.4)
Othersb3 (1.8)
Bim polymorphism 
Wild type150 (90.4)
Deletion16 (9.6)
Disease stage 
IIIB24 (14.5)
IV142 (85.5)
Line of use 
First line69 (41.6)
Second line50 (30.1)
Third line or later47 (28.3)
Agent 
Erlotinib140 (84.3)
Gefitinib26 (15.7)
Response 
Complete response2 (1.2)
Partial response101 (60.8)
Stable disease29 (17.5)
Progressive disease34 (20.5)
Table 3. Clinicopathologic Features of Bim Polymorphism Carriers Who Had Activating EGFR Mutations
PatientAge, ySexSmoking HistoryHistologyStageEGFR StatusECOG PSPFS, moTherapeutic Efficiency
  1. Abbreviations: Del, deletion, ECOG PS, Eastern Cooperative Oncology Group performance status; EGFR, epidermal growth factor receptor; Ins, insertion; L858R, substitution of lysine with arginine at EGFR amino acid position 858; NSCLC-NOS, non–small cell lung cancer, not otherwise specified; PD, progressive disease; PFS, progression-free survival; PR, partial response; S768I, substitution of serine with isoleucine at EGFR amino acid position 768; SD, stable disease; T790M, substitution of threonine with methionine at EGFR amino acid position 790.

660ManSmokerAdenocarcinomaIV19DelL/T790M111.0PR
1669ManSmokerAdenocarcinomaIV19 Del17.3SD
2648WomanNeverLarge cellIIIB20 Ins20.9PD
3052ManSmokerAdenocarcinomaIVL858R/S768I11.3PD
8945WomanNeverAdenocarcinomaIVL858R18.0PR
11462WomanNeverAdenocarcinomaIVL858R14.7SD
11639WomanNeverAdenocarcinomaIIIBL858R14.1SD
14260WomanNeverAdenocarcinomaIV19 Del16.2PR
14351ManSmokerAdenocarcinomaIV19 Del11.2PD
14449ManSmokerSquamousIV19 Del12.4PD
16249WomanNeverNSCLC-NOSIV19 Del17.0SD
16459ManSmokerAdenocarcinomaIVL858R12.4PD
17148ManNeverAdenocarcinomaIVL858R19.2PR
18562ManSmokerAdenocarcinomaIV19 Del13.0PD
19960WomanNeverAdenocarcinomaIV19 Del15.0SD
21148WomanNeverAdenocarcinomaIIIB19 Del17.7SD

Association Between Bim Polymorphism and TKI Efficiency

Because of the low response rate and short PFS among patients with WT EGFR (data not shown), it was possible that the predictive power of the Bim polymorphism could be diluted. Therefore, additional analyses were conducted only in the subgroup with EGFR mutations. In this group, the median PFS was 11 months (95% CI, 8.48-13.52 months) for patients with WT Bim and 4.7 months (95% CI, 2.94-6.46 months) for those who had the Bim deletion polymorphism (P = .003). The ORR for patients who had the Bim deletion polymorphism versus those who had WT Bim was 25% versus 66%, respectively (P = .001), and the disease control rate was 62.5% versus 81.3%, respectively (P = .076).

An analysis of the impact of the EGFR exon 19 deletion and the EGFR L858R mutation on Bim deletion polymorphism carriers also was performed. However, the results did not indicate statistical significance. For patients who harbored an EGFR exon 19 deletion, PFS was 11.3 months (95% CI, 7.74-14.86 months); and, for those who had the L858R mutation, the PFS was 8 months (95% CI, 5.66-10.34 months; P = .372). However, in their counterparts who had the Bim deletion polymorphism, the PFS was 7.3 months (95% CI, 4.72-11.86 months) and 4.1 months (95% CI, 2.06-6.14 months; P = .377), respectively. These data are summarized in Table 4 and Figure 2.

Table 4. A Brief Summary of Responses to Epidermal Growth Factor Receptor (EGFR) Tyrosine Kinase Receptor Treatment and Bim Status in Patients With EGFR Mutations
 Bim Polymorphism Status 
VariablePositive, n = 16Negative, n = 150P
  1. Abbreviations: Bim, B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (BCL2L11); CI, confidence interval; CR, complete response; DCR, disease control rate; mPFS, median progression-free survival; ORR, overall response rate; PD, progressive disease; PR, partial response; SD, stable disease.

Response: No. of patients   
CR02.642
PR497.002
SD623.026
PD628.076
mPFS (95% CI), mo   
Mutant EGFR4.7 (2.94-6.46)11 (8.48-13.52).003
Exon 197.3 (4.72-11.86)11.5 (9.72-13.27).053
Exon 214.1 (2.06-6.14)8.5 (4.22-12.79.033
ORR, %   
Mutant EGFR2566.001
Exon 1922.273.1.002
Exon 2133.353.2.22
DCR, %   
Mutant EGFR62.581.3.076
Exon 1966.788.5.073
Exon 2166.775.7.669
image

Figure 2. Progression-free survival (PFS) is illustrated according to tyrosine kinase inhibitor treatment status in the current study. mPFS indicates median PFS. (a) PFS is illustrated for the entire epidermal growth factor receptor (EGFR)-mutated group. (b) PFS is illustrated for patients who carried the B-cell chronic lymphocytic leukemia/lymphoma (Bcl2)-like 11 (Bim) deletion polymorphism compared with wild-type carriers among all those who had EGFR mutations. (c) PFS is illustrated for patients who had samples with the EGFR exon 19 deletion and the L858R (substitution of lysine with arginine at EGFR amino acid position 858) mutation. NSCLC indicates non–small cell lung cancer. (d) PFS is illustrated for patients who had the EGFR exon 19 deletion or the L858R mutation among those who had the B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (Bim) deletion polymorphism.

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A Cox regression analysis was also conducted including only patients in the EGFR-mutant subgroup, because the Bim polymorphism did not affect PFS significantly in patients who had WT EGFR. Univariate analysis indicated that sex (women; P = .002) and Bim status (WT; P = .004) were associated with longer PFS. Multivariate analysis demonstrated that Bim status (P = .016), together with sex (P = .001), was an independent factor in EGFR-mutated patients who received TKI treatment. All of the regression data are detailed in Table 5.

Table 5. Univariate and Multivariate Analyses of Prognostic Factors in Epidermal Growth Factor Receptor-Mutant Patients With Non–Small Cell Lung Cancer
 Univariate AnalysisMultivariate Analysis
VariableHR (95% CI)PHR (95% CI)P
  1. Abbreviations: Bim, B-cell chronic lymphocytic leukemia/lymphoma (Bcl-2)-like 11 (BCL2L11); CI, confidence interval; HR, hazard ratio; L858R, substitution of lysine with arginine at EGFR amino acid position 858; NA, not applicable; WT, wild type.

  2. a

    Smoking status was excluded in the multivariate analysis because of its high correlation with sex.

Age ≤65 y vs >65 y0.707 (0.49-1.021).0640.685 (0.467-1.003).052
Women vs men1.904 (1.212-2.392).0021.718 (1.23-2.421).002
Nonsmokers vs smokersa1.419 (0.955-2.110).083NA-
WT Bim vs polymorphism2.385 (1.313-4.330).0042.094 (1.148-3.82).016
Exon 19 deletion vs L858R1.076 (0.908-1.215).3971.127 (0.947-1.341).178

DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES

To our knowledge, the current study is the first to explore the clinical characteristics and prognostic value of EGFR-TKI outcomes among Chinese patients with NSCLC who had the Bim deletion polymorphism. The results indicated that no specific clinicopathologic feature was associated significantly with distribution of the Bim deletion polymorphism. WT Bim may be an independent factor predicting clinical benefit from EGFR TKIs in patients with NSCLC who have an activating EGFR mutation.

In the current study, we observed a Bim polymorphism rate and a distribution pattern similar to that reported previously.[17] Forty-five of 352 patients and 28 of 228 EGFR-mutated patients were identified who harbored this deletion polymorphism. Also, our data demonstrated that the Bim polymorphism was distributed randomly regardless of disease stage, histologic type, sex, or EGFR mutation status.

Bim acts as an apoptosis facilitator in response to stress signals like DNA damage, and it functions irreplaceably in the apoptosis pathway. It is responsible for inducing apoptosis in almost all targeted therapies for various kinds of tumors (eg, lapatinib in breast cancer, imatinib in chronic myeloid leukemia).[14, 19-21] In patients who have NSCLC with a known driver mutation, the inhibition of Bim is associated with failure of targeted therapy, and Bim up-regulation is necessary in successful targeted therapy to induce apoptosis.[22-24]

By affecting alternative splicing, this Bim deletion polymorphism caused impaired expression in its proapoptosis BH3 domain, thus leading to failure of apoptosis induction. Two previous studies verified the correlation between the Bim deletion polymorphism and the insufficient efficacy of targeted therapy in the treatment of patients with chronic myelogenous leukemia using imatinib[25] and in the treatment of patients with NSCLC using EGFR TKIs.[17]

Our study confirmed the correlation between the Bim deletion polymorphism and a poor response to EGFR TKIs in patients with activating EGFR mutations. In our statistical analysis, we observed that the deletion polymorphism was related to shorter PFS for the entire study group compared with WT Bim carriers, mainly because of poor TKI efficiency in patients with EGFR mutations who had the Bim deletion polymorphism. Similarly, the ORR was lower in the deletion polymorphism carriers among EGFR-mutant patients, but not among patients with WT EGFR (data not shown). This may be attributed to the inherent poor response to TKIs and short PFS of patients with NSCLC who harbor WT EGFR. It also would dilute the predictive power of the Bim deletion polymorphism. Thus, we concluded that Bim polymorphism status may have influenced the efficacy of TKIs mainly in our EGFR-mutated patients. This conclusion was consistent with the finding from a previous study that Bim is essential for apoptosis triggered by EGFR TKIs in sensitive populations.[16]

The outcome of EGFR-mutant patients who had the exon 19 deletion appeared to be better than the outcome of those who had the L858R mutation, but the difference was not significant in our study. Although several previous studies have demonstrated that patients with the EGFR exon 19 deletion enjoyed better outcomes than patients harboring the L858R mutation,[26-28] the results from our study were consistent with several other large clinical trials[3, 29] indicating that the exon 19 deletion and the L858R mutation had an equal impact on the efficacy of EGFR TKIs. Moreover, the PFS of Bim polymorphism carriers did not differ significantly between the 2 EGFR mutation types, which indicates that Bim deletion polymorphism-induced TKI resistance may not be influenced by the 2 most common EGFR mutation types.

Recently, a study conducted in Korean patients with NSCLC did not produce significant results when comparing the efficacy of EGFR TKIs between patients with WT Bim and patients with the Bim deletion polymorphism.[30] The investigators attributed their results to population limitations and to the influence of other Bcl-2 family factors. Another report[31] demonstrated that the resistance can be circumvented by combining targeted therapy with histone deacetylase inhibition, which offers a possible explanation for the Korean results, because epigenetic factors also may influence the transcription and expression of Bim. Our study also has several limitations. First, the numbers of patients who had the deletion polymorphism, especially those who also harbored EGFR mutations, were limited. Moreover, all patients who harbored the polymorphism were heterozygous, and no homozygous carriers have been reported to date. Future research will continue to increase the population in the hope of reducing the bias to some extent and of identifying homozygous Bim polymorphism carriers. Because Bim plays a key role in TKI-induced apoptosis, we believe that not only Bim polymorphism but also other markers of Bim status, such as copy numbers or protein levels, should be taken into consideration to make a full evaluation. Further in vitro and in vivo research will be conducted to explore epigenetic factors and signal crosstalk that may have an impact on the expression or function of Bim.

In conclusion, we explored the Bim deletion polymorphism and observed that, in patients who had NSCLC with EGFR mutations, the Bim deletion polymorphism led to worse PFS, ORR, and disease control rate. Our work also indicated that this polymorphism can serve as a predictive factor in TKI therapy for NSCLC. Although the significance of Bim in targeted therapy has been noticed for some time, more work is needed to understand it better.

FUNDING SUPPORT

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES

This study was supported by grants from the National Natural Science Foundation of China (no. 811-72101) and the Key Project of the Science and Technology Commission of Shanghai Municipality (no. 11JC1 411301).

CONFLICT OF INTEREST DISCLOSURES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES

Dr. Hirsch has served on the advisory boards of Boehringer Ingelheim and Genentech/Roche.

REFERENCES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. FUNDING SUPPORT
  8. CONFLICT OF INTEREST DISCLOSURES
  9. REFERENCES