0
Original Research: LUNG CANCER |

Assessment of Epidermal Growth Factor Receptor Mutation by Endobronchial Ultrasound-Guided Transbronchial Needle Aspiration* FREE TO VIEW

Takahiro Nakajima, MD; Kazuhiro Yasufuku, MD, PhD, FCCP; Makoto Suzuki, MD, PhD; Kenzo Hiroshima, MD, PhD; Rieko Kubo, MS; Sherif Mohammed, MD; Yohei Miyagi, MD, PhD; Shoichi Matsukuma, PhD; Yasuo Sekine, MD, PhD; Takehiko Fujisawa, MD, PhD
Author and Funding Information

*From the Departments of Thoracic Surgery (Drs. Nakajima, Yasufuku, Suzuki, Mohammed, Sekine, and Fujisawa, and Ms. Kubo) and Diagnostic Pathology (Dr. Hiroshima), Graduate School of Medicine, Chiba University, Chiba, Japan; and the Division of Molecular Pathology and Genetics (Drs. Miyagi and Matsukuma), Kanagawa Cancer Center Research Institute, Kanagawa, Japan.

Correspondence to: Takehiko Fujisawa, MD, PhD, Department of Thoracic Surgery, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670, Japan; e-mail: fujisawat@faculty.chiba-u.jp



Chest. 2007;132(2):597-602. doi:10.1378/chest.07-0095
Text Size: A A A
Published online

Background: The presence of somatic mutations in epidermal growth factor receptor (EGFR) predicts the effectiveness of EGFR tyrosine kinase inhibitors (TKIs). It would be ideal if an EGFR mutation could be detected in biopsy samples, since the majority of non-small cell lung cancer patients are inoperable at the time of presentation. We have reported the usefulness of endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) for the lymph node staging of lung cancer. EBUS-TBNA enables the sampling of histologic cores, which can be used for genetic analysis.

Methods: The purpose of this study was to develop and analyze the feasibility of detecting EGFR mutations in samples obtained by EBUS-TBNA. Forty-six patients with primary lung cancer in whom metastatic adenocarcinoma in the hilar and/or mediastinal lymph node was diagnosed by EBUS-TBNA were enrolled into the study. DNA was extracted from paraffin-embedded samples, and the EGFR mutation was analyzed in exons 19 and 21 using a newly developed loop-hybrid mobility shift assay. The results were confirmed by direct sequencing.

Results: Forty-three cases were eligible for analysis and in 11 cases, EGFR mutation (25.6%) was detected; one case was an in-frame deletion (E746-A750del) of exon 19, nine cases were point mutations (L858R) of exon 21, and one case was a double point mutation (L858R+L861V). All cases with EGFR mutations were confirmed by direct sequencing.

Conclusions: EGFR mutation can easily be detected in metastatic lymph nodes sampled by EBUS-TBNA. EBUS-TBNA allows genetic evaluations of tumor cells within the lymph node and may provide us with indications for EGFR-TKI therapy in the near future.

Figures in this Article

Although platinum-based combined chemotherapy for advanced lung cancer is widely accepted, the efficacy for this approach is only about 30%.1The newly developed epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) such as gefitinib and erlotinib have shown dramatic effects in some patients with primary lung cancer.2Clinical studies have revealed significant variability in the clinical and pathologic factors associated with the patient response to gefitinib. It is also known that this drug is very effective in female, nonsmoking patients of East Asian origin with adenocarcinoma histology.35 On the other hand, some of the patients experience fatal interstitial pneumonia as an adverse side effect.6 Some studies34 have shown that the effect of EGFR-TKIs is well correlated with various EGFR mutations.

Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is a new modality for the evaluation of mediastinal and hilar lymph node metastasis from lung cancer.68 Compared to other diagnostic modalities, EBUS-TBNA is a real-time procedure that enables multiple biopsies with high-quality histologic cores. It is associated with minimal complications.610 In this study, we examined the somatic genetic alterations in paraffin-embedded lymph node biopsy specimens obtained by EBUS-TBNA from patients with primary lung cancer.

Patients and EBUS-TBNA Tissue Sampling

From December 2003 to April 2006, metastatic adenocarcinoma of hilar and/or mediastinal lymph node histologic type was diagnosed in 46 patients using EBUS-TBNA samples. A pathologist reviewed all cases and confirmed the presence of nodal tissue and cancer tissue in each sample. Recut sections of the paraffin-embedded samples yielded tumor cells in samples from 43 of the 46 patients, and these patients were enrolled in the study. In five cases, the corresponding DNA from the resected lung cancer tissue, the adjacent nonneoplastic lung tissue, the lymph node with metastasis, and the lymph node without metastasis was analyzed. To examine the genetic homology between the primary tumor and the metastatic lymph node, we evaluated eight other cases that had previously been analyzed for the mutation status of EGFR.11 These cases included three cases of resected lung cancer tissue and metastatic lymph node with a known exon 19 deletion and five cases with a known exon 21 point mutation.

We have developed a method for obtaining histologic samples by EBUS-TBNA. Briefly, a dedicated 22-gauge needle equipped with an internal sheath is used. After the initial puncture, the internal sheath is used to clean out the internal lumen that is clogged with the bronchial tissue. The internal sheath is removed, and negative pressure is applied using a syringe. The needle is moved back and forth inside the tumor. Finally, the needle is retrieved, and the internal sheath is used once again to push out the histologic core.7

DNA Extraction

A total of two sections of paraffin-embedded biopsy specimen with a thickness of 10 μm were obtained. DNA was extracted from paraffin-embedded tumor cells (Pinpoint Slide DNA Isolation System; Zymo Research Corp; Orange, CA) according to the manufacture’s instructions.

Polymerase Chain Reaction for Exon 19 and Exon 21

We evaluated the EGFR tyrosine kinase domain, exons 19 and 21, using the newly developed loop-hybrid mobility shift assay and confirmed the results by direct sequencing.12 EGFR exon 19 and 21, which encode the tyrosine kinase domain, were amplified by nested polymerase chain reaction (PCR). The primers and amplification conditions for the loop-hybrid mobility shift assay were based on a report by Matsukuma et al.12 The PCR was performed in a total volume of 20 μL (Platinum PCR SuperMix; Invitrogen; Carlsbad, CA).

Loop-Hybrid Mobility Shift Assay

The resulting PCR amplicons for the loop-hybrid mobility shift assay were mixed with specific oligo probes (loop-hybrid [LH] probe) designed for this assay and incubated in a thermal cycler in a total volume of 5 μL including a 500 nmol/L LH probe. The probe sequence and incubation conditions have been previously reported.12 One of the probes designed for the detection of exon 19 in-frame deletions is also complimentary for detecting wild-type EGFR exon 19 sequences. When there is an in-frame deletion in the exon, the probe forms a loop to the mutated sequence, which retards the electrophoretic mobility of the band compared with the nonmutant band. The probe for the detection of exon 21 point mutation is complimentary for wild-type EGFR exon 21 sequences, but in this case the probe was shortened by seven bases. The probe forms a seven-base loop to the wild-type sequence. Thus, when there is an exon 21 point mutation, the probe forms an eight-base loop, resulting in an altered mobility compared with the normal band. We used 10% of polyacrylamide gels (c10-PAGEL Precast Gels; ATTO; Tokyo, Japan) for electrophoresis.

Direct Sequence

The resulting PCR amplicons for the direct sequence were purified (Rapid PCR Purification System; Marligen; Ijamsville, MD) and sequenced (BigDye Terminator kit and ABI Prism 310 DNA Analyzer; Applied Biosystems; Foster City, CA) according to the manufacturer’s instructions. Mutations were examined in both forward and reverse directions.

Ethics Committee Approval

The study was approved by the ethics committee of Chiba University, Graduate School of Medicine (No. 119). Written consent was obtained from all patients. All samples were coded and managed independently.

Patient Characteristics

The clinical characteristics of the 43 patients are listed in Table 1 . Thirty patients (69.8%) were men. All patients had lung cancer with adenocarcinoma histology. The median age was 64 years (range, 44 to 85 years). Three patients had stage IIB disease, 19 patients had stage IIIA disease, 10 patients had stage IIIB disease, 3 patients had stage IV disease, and 8 patients had postoperative recurrences. Two patients with stage IIB disease underwent thoracotomy, and the other 41 patients underwent chemotherapy and/or radiotherapy.

EGFR Mutation

Preliminary studies via the loop-hybrid mobility shift assay in five patients who had undergone surgical resection revealed three patients with an EGFR mutation at exon 21. Mutations were detected from both primary lung tumor and metastatic lymph nodes, and were not detected from nonneoplastic lung tissue and lymph nodes without metastasis. The homology of the primary lung tumor and the metastatic lymph node was confirmed by examining the EGFR gene expression by both loop-hybrid mobility shift assay and direct sequencing (data not shown).

EGRR Mutation in EBUS-TBNA Samples

One exon 19 in-frame deletion (one woman) and 10 exon 21 point mutations (five women) were detected by this newly developed method. All cases, including 1 exon 19 in-frame deletion (E746-A750del) and 10 exon 21 point mutations (L858R, 9 mutations; L858R+L861V, 1 mutation) were confirmed by direct sequencing (Figs 1, 2 , Table 2 ). The cases without mutations were also confirmed by direct sequencing.

Gefitinib Treatment Response

In this study, two patients with exon 21 point mutations were treated with gefitinib. In one case, gefitinib administration was discontinued due to the condition of the patient. The other patients was a 67-year-old woman with multiple mediastinal lymph node metastases along with a malignant pericardial effusion. Two weeks after the administration of gefitinib, all of the metastatic sites decreased in size dramatically. The patient has been in stable condition for 17 months after the administration of gefitinib (Fig 3 ).

In this study, we sought to develop and analyze the feasibility of detecting EGFR mutations in samples obtained by EBUS-TBNA. We looked at the EGFR mutation status within EBUS-TBNA samples obtained from metastatic mediastinal and/or hilar lymph nodes in patients with adenocarcinoma of the lung. In 11 of 43 cases, an EGFR mutation was detected. We also showed that histologic cores obtained by EBUS-TBNA can be used for molecular analysis.

EBUS-TBNA is a new approach for the evaluation of mediastinal and hilar lymph node metastasis in patients with lung cancer.610 A recent study8 comparing EBUS-TBNA, positron emission tomography, and CT scanning for lymph node staging of lung cancer revealed that EBUS-TBNA has a high sensitivity as well as specificity for mediastinal and hilar lymph node staging in patients with lung cancer. Importantly, there were no complications associated with EBUS-TBNA in the study.8 The great advantage of EBUS-TBNA is that it is a real-time procedure, which can be performed repeatedly under local anesthesia to obtain multiple samples. This allows for the possibility of analysis of other gene mutations or verifying the mutation in question1314 by performing a repeat EBUS-TBNA. This is particularly useful when we encounter patients who do not respond to a certain EGFR-TKI.

Another advantage of EBUS-TBNA may relate to the composition of the histologic material obtained for genetic analysis. Prior reports on EGFR mutation analysis in nonsurgical materials such as biopsy specimens, pleural effusions, and serum have noted problems of contamination by nontarget normal cells within the samples.1518 Clinical samples may contain a contaminating amount of mutated genes and a large number of wild-type genes; thus, EGFR mutation analysis sometimes requires microdissection or a highly sensitive assay.19 In our present study, histologic cores obtained by EBUS-TBNA consisted mainly of tumor cells, blood constituent, and a small amount of lymph node tissue. Compared to the number of tumor cells, there was relatively little representation of normal or reactive tissue within the samples. This minimizes the problem of contamination by excessive nontarget normal cells.

To improve the sensitivity of detecting EGFR mutations and to reduce the cost and time of analysis, many screening methods have been developed.1921 The loop-hybrid mobility shift assay that was used in this study is a newly developed method for screening the existence of EGFR exon 19 and 21 mutations.12 This method can easily detect in-frame deletion mutations in exon 19 and the hot-spot mutation L858R in exon 21 of EGFR that is associated with the therapeutic response to EGFR-TKIs such as gefitinib.12 Compared with other screening methods, the loop-hybrid mobility shift assay does not require a second PCR. This hybridization method may have the advantage of reducing the artifact and pseudopositive results associated with excessive amplification. These advantages, taken together with the simplicity and sensitivity of the technique, make it a preferred method for biopsy and diagnosis.

Since < 30% of patients with lung cancer at the time of presentation have an indication for surgery, the ability to develop a biological analysis using nonsurgical biopsy samples will become very important for the future of lung cancer treatment. This study demonstrates that EGFR mutation status can be analyzed from samples obtained from metastatic lymph nodes by EBUS-TBNA. EBUS-TBNA samples will possibly provide other molecular biological information, which will be useful for the treatment of advanced and recurrent lung cancer.

Abbreviations: EBUS-TBNA = endobronchial ultrasound-guided transbronchial needle aspiration; EGFR = epidermal growth factor receptor; LH = loop-hybrid; PCR = polymerase chain reaction; TKI = tyrosine kinase inhibitor

This study was supported in part by Grant-in-Aid for Scientific Research No. 18790986 (to Dr. Nakajima) and No. 18591540 (to Dr. Suzuki) from the Ministry of Education, Culture, Sports, Science and Technology of Japan. Dr. Suzuki was also supported by the Emphasis Research Project by expenditure at the discretion of the president of the Chiba University in 2005.

The authors have reported to the ACCP that no significant conflicts of interest exist with any companies/organizations whose products or services may be discussed in this article.

Table Graphic Jump Location
Table 1. Patient Characteristics
Figure Jump LinkFigure 1. Detection of a deletion mutation in EGFR exon 19 with loop-hybrid mobility shift assay and direct sequencing. Loop-hybrid mobility shift assay results of PCR products from the EBUS-TBNA case of deletion mutant alleles E746-A750del (lane case 2) and the normal allele (lane x) treated with the control LH probe for exon 19 are shown. N = negative control; P = positive control; bp = base pair.Grahic Jump Location
Figure Jump LinkFigure 2. Detection of point mutations at or near the hot-spot mutation of EGFR exon 21 of L858R. The electropherogram obtained after the loop-hybrid mobility shift assay using LH probe for exon 21 in DNA from EBUS-TBNA samples is shown. The detection of heterozygous mutations in EGFR exon 21 of L858R (CTG > CGC, lane case 10) and L858R+L861V (CTG > CGG+CTG > GTG, lane case 6) are shown. Lane x is the normal allele. See the legend of Figure 1 for abbreviations not used in the text.Grahic Jump Location
Table Graphic Jump Location
Table 2. Patient Characteristics With EGFR Mutations*
* 

F = female; M = male; PR = partial response; SD = stable disease; LN = lymph node.

Figure Jump LinkFigure 3. Chest CT scan findings in a patient successfully treated with gefitinib. A 67-year-old woman with multiple mediastinal lymph node metastases along with malignant pericardial effusion was treated with gefitinib. Top, A: CT scan shows enlarged stations 2R, 6, 4L, and 5 before the administration of gefitinib. Middle, B: two weeks after the administration of gefitinib, all of the enlarged mediastinal lymph nodes decreased in size dramatically. Bottom, C: the patient has been in stable condition for 17 months after the administration of gefitinib.Grahic Jump Location
Wakelee, H, Belani, CP (2005) Optimizing first-line treatment options for patients with advanced NSCLC.Oncologist10(suppl),1-10
 
Mitsudomi, T, Kosaka, T, Endoh, H, et al Mutations of the epidermal growth factor receptor gene predict prolonged survival after gefitinib treatment in patients with non-small-cell lung cancer with postoperative recurrence.J Clin Oncol2006;23,2513-2520
 
Lynch, TJ, Bell, DW, Sordella, R, et al Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib.N Engl J Med2004;350,2129-2139. [PubMed] [CrossRef]
 
Paez, JG, Janne, PA, Lee, JC, et al EGFR mutations in lung cancer: correlation with clinical response to gefitinib therapy.Science2004;304,1497-1500. [PubMed]
 
Pao, W, Miller, V, Zakowski, M, et al EGF receptor gene mutations are common in lung cancers from “never smokers” and are associated with sensitivity of tumors to gefitinib and erlotinib.Proc Natl Acad Sci U S A2004;101,13306-13311. [PubMed]
 
Yasufuku, K, Chiyo, M, Sekine, Y, et al Real-time endobronchial ultrasound-guided transbronchial needle aspiration of mediastinal and hilar lymph nodes.Chest2004;126,122-128. [PubMed]
 
Yasufuku, K, Chiyo, M, Koh, E, et al Endobronchial ultrasound guided transbronchial needle aspiration for staging of lung cancer.Lung Cancer2005;50,347-354. [PubMed]
 
Yasufuku, K, Nakajima, T, Motoori, K, et al Comparison of endobronchial ultrasound, positron emission tomography, and CT for lymph node staging of lung cancer.Chest2006;130,710-718. [PubMed]
 
Herth, FJ, Ernst, A, Eberhardt, R, et al Endobronchial ultrasound-guided transbronchial needle aspiration of lymph nodes in the radiologically normal mediastinum.Eur Respir J2006;28,910-914. [PubMed]
 
Herth, FJ, Eberhardt, R, Vilmann, P, et al Real-time endobronchial ultrasound guided transbronchial needle aspiration for sampling mediastinal lymph nodes.Thorax2006;61,795-798. [PubMed]
 
Suzuki, M, Shigematsu, H, Hiroshima, K, et al Epidermal growth factor receptor expression status in lung cancer correlates with its mutation.Hum Pathol2005;36,1127-1134. [PubMed]
 
Matsukuma, S, Yoshihara, M, Kasai, F, et al Rapid and simple detection of hot spot point mutations of epidermal growth factor receptor, BRAF, and NRAS in cancers using the loop-hybrid mobility shift assay.J Mol Diagn2006;8,504-512. [PubMed]
 
Kobayashi, S, Boggon, TJ, Dayaram, T, et al EGFR mutations and resistance of non-small-cell lung cancer to gefitinib.N Engl J Med2005;352,786-792. [PubMed]
 
Pao, W, Miller, VA, Politi, KA, et al Acquired resistance of lung adenocarcinomas to gefitinib or erlotinib is associated with a second mutation in the EGFR kinase domain.PLoS Med2007;2,e73
 
Shih, JY, Gow, CH, Yu, CJ, et al Epidermal growth factor receptor mutations in needle biopsy/aspiration samples predict response to gefitinib therapy and survival of patients with advanced nonsmall cell lung cancer.Int J Cancer2006;118,963-969. [PubMed]
 
Fujita, S, Mio, T, Sonobe, M, et al Accuracy of epidermal growth factor receptor mutation analysis on the basis of small biopsy specimens in patients with nonsmall cell lung cancer.Int J Cancer2006;119,1751-1752. [PubMed]
 
Kimura, H, Kasahara, K, Kawaishi, M, et al Detection of epidermal growth factor receptor mutations in serum as a predictor of the response to gefitinib in patients with non-small-cell lung cancer.Clin Cancer Res2006;12,3915-3921. [PubMed]
 
Kimura, H, Fujiwara, Y, Sone, T, et al High sensitivity detection of epidermal growth factor receptor mutations in the pleural effusion of non-small cell lung cancer patients.Cancer Sci2006;97,642-648. [PubMed]
 
Asano, H, Toyooka, S, Tokumo, M, et al Detection of EGFR gene mutation in lung cancer by mutant-enriched polymerase chain reaction assay.Clin Cancer Res2006;12,43-48. [PubMed]
 
Janne, PA, Borras, AM, Kuang, Y, et al A rapid and sensitive enzymatic method for epidermal growth factor receptor mutation screening.Clin Cancer Res2006;12,751-758. [PubMed]
 
Yatabe, Y, Hida, T, Horio, Y, et al A rapid, sensitive assay to detect EGFR mutation in small biopsy specimens from lung cancer.J Mol Diagn2006;8,335-341. [PubMed]
 

Figures

Figure Jump LinkFigure 1. Detection of a deletion mutation in EGFR exon 19 with loop-hybrid mobility shift assay and direct sequencing. Loop-hybrid mobility shift assay results of PCR products from the EBUS-TBNA case of deletion mutant alleles E746-A750del (lane case 2) and the normal allele (lane x) treated with the control LH probe for exon 19 are shown. N = negative control; P = positive control; bp = base pair.Grahic Jump Location
Figure Jump LinkFigure 2. Detection of point mutations at or near the hot-spot mutation of EGFR exon 21 of L858R. The electropherogram obtained after the loop-hybrid mobility shift assay using LH probe for exon 21 in DNA from EBUS-TBNA samples is shown. The detection of heterozygous mutations in EGFR exon 21 of L858R (CTG > CGC, lane case 10) and L858R+L861V (CTG > CGG+CTG > GTG, lane case 6) are shown. Lane x is the normal allele. See the legend of Figure 1 for abbreviations not used in the text.Grahic Jump Location
Figure Jump LinkFigure 3. Chest CT scan findings in a patient successfully treated with gefitinib. A 67-year-old woman with multiple mediastinal lymph node metastases along with malignant pericardial effusion was treated with gefitinib. Top, A: CT scan shows enlarged stations 2R, 6, 4L, and 5 before the administration of gefitinib. Middle, B: two weeks after the administration of gefitinib, all of the enlarged mediastinal lymph nodes decreased in size dramatically. Bottom, C: the patient has been in stable condition for 17 months after the administration of gefitinib.Grahic Jump Location

Tables

Table Graphic Jump Location
Table 1. Patient Characteristics
Table Graphic Jump Location
Table 2. Patient Characteristics With EGFR Mutations*
* 

F = female; M = male; PR = partial response; SD = stable disease; LN = lymph node.

References

Wakelee, H, Belani, CP (2005) Optimizing first-line treatment options for patients with advanced NSCLC.Oncologist10(suppl),1-10
 
Mitsudomi, T, Kosaka, T, Endoh, H, et al Mutations of the epidermal growth factor receptor gene predict prolonged survival after gefitinib treatment in patients with non-small-cell lung cancer with postoperative recurrence.J Clin Oncol2006;23,2513-2520
 
Lynch, TJ, Bell, DW, Sordella, R, et al Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib.N Engl J Med2004;350,2129-2139. [PubMed] [CrossRef]
 
Paez, JG, Janne, PA, Lee, JC, et al EGFR mutations in lung cancer: correlation with clinical response to gefitinib therapy.Science2004;304,1497-1500. [PubMed]
 
Pao, W, Miller, V, Zakowski, M, et al EGF receptor gene mutations are common in lung cancers from “never smokers” and are associated with sensitivity of tumors to gefitinib and erlotinib.Proc Natl Acad Sci U S A2004;101,13306-13311. [PubMed]
 
Yasufuku, K, Chiyo, M, Sekine, Y, et al Real-time endobronchial ultrasound-guided transbronchial needle aspiration of mediastinal and hilar lymph nodes.Chest2004;126,122-128. [PubMed]
 
Yasufuku, K, Chiyo, M, Koh, E, et al Endobronchial ultrasound guided transbronchial needle aspiration for staging of lung cancer.Lung Cancer2005;50,347-354. [PubMed]
 
Yasufuku, K, Nakajima, T, Motoori, K, et al Comparison of endobronchial ultrasound, positron emission tomography, and CT for lymph node staging of lung cancer.Chest2006;130,710-718. [PubMed]
 
Herth, FJ, Ernst, A, Eberhardt, R, et al Endobronchial ultrasound-guided transbronchial needle aspiration of lymph nodes in the radiologically normal mediastinum.Eur Respir J2006;28,910-914. [PubMed]
 
Herth, FJ, Eberhardt, R, Vilmann, P, et al Real-time endobronchial ultrasound guided transbronchial needle aspiration for sampling mediastinal lymph nodes.Thorax2006;61,795-798. [PubMed]
 
Suzuki, M, Shigematsu, H, Hiroshima, K, et al Epidermal growth factor receptor expression status in lung cancer correlates with its mutation.Hum Pathol2005;36,1127-1134. [PubMed]
 
Matsukuma, S, Yoshihara, M, Kasai, F, et al Rapid and simple detection of hot spot point mutations of epidermal growth factor receptor, BRAF, and NRAS in cancers using the loop-hybrid mobility shift assay.J Mol Diagn2006;8,504-512. [PubMed]
 
Kobayashi, S, Boggon, TJ, Dayaram, T, et al EGFR mutations and resistance of non-small-cell lung cancer to gefitinib.N Engl J Med2005;352,786-792. [PubMed]
 
Pao, W, Miller, VA, Politi, KA, et al Acquired resistance of lung adenocarcinomas to gefitinib or erlotinib is associated with a second mutation in the EGFR kinase domain.PLoS Med2007;2,e73
 
Shih, JY, Gow, CH, Yu, CJ, et al Epidermal growth factor receptor mutations in needle biopsy/aspiration samples predict response to gefitinib therapy and survival of patients with advanced nonsmall cell lung cancer.Int J Cancer2006;118,963-969. [PubMed]
 
Fujita, S, Mio, T, Sonobe, M, et al Accuracy of epidermal growth factor receptor mutation analysis on the basis of small biopsy specimens in patients with nonsmall cell lung cancer.Int J Cancer2006;119,1751-1752. [PubMed]
 
Kimura, H, Kasahara, K, Kawaishi, M, et al Detection of epidermal growth factor receptor mutations in serum as a predictor of the response to gefitinib in patients with non-small-cell lung cancer.Clin Cancer Res2006;12,3915-3921. [PubMed]
 
Kimura, H, Fujiwara, Y, Sone, T, et al High sensitivity detection of epidermal growth factor receptor mutations in the pleural effusion of non-small cell lung cancer patients.Cancer Sci2006;97,642-648. [PubMed]
 
Asano, H, Toyooka, S, Tokumo, M, et al Detection of EGFR gene mutation in lung cancer by mutant-enriched polymerase chain reaction assay.Clin Cancer Res2006;12,43-48. [PubMed]
 
Janne, PA, Borras, AM, Kuang, Y, et al A rapid and sensitive enzymatic method for epidermal growth factor receptor mutation screening.Clin Cancer Res2006;12,751-758. [PubMed]
 
Yatabe, Y, Hida, T, Horio, Y, et al A rapid, sensitive assay to detect EGFR mutation in small biopsy specimens from lung cancer.J Mol Diagn2006;8,335-341. [PubMed]
 
NOTE:
Citing articles are presented as examples only. In non-demo SCM6 implementation, integration with CrossRef’s "Cited By" API will populate this tab (http://www.crossref.org/citedby.html).

Some tools below are only available to our subscribers or users with an online account.

Related Content

Customize your page view by dragging & repositioning the boxes below.

CHEST Journal Articles
PubMed Articles
Guidelines
Advanced breast cancer. Diagnosis and treatment.
National Collaborating Centre for Cancer | 9/18/2009
  • CHEST Journal
    Print ISSN: 0012-3692
    Online ISSN: 1931-3543