Epidermal Growth Factor Receptor Sequencing Research Articles

Discordance of Epidermal Growth Factor Receptor Mutation between Brain Metastasis and Primary Non-Small Cell Lung Cancer.

BackgroundThe aim of this study was to compare epidermal growth factor receptor (EGFR) mutations between non-small cell lung cancer (NSCLC) and corresponding brain metastases (BMs) in Korea society.MethodsFrom 2011 to 2016, a total of 74 patients underwent surgical resection of a metastatic brain tumor from NSCLC. Among them, we performed retrospective analysis for 46 patients who underwent EGFR sequencing of primary NSCLC tissues.ResultsAmong these 46 cases, 18 (39.1%) cases showed EGFR mutation in primary lung cancer. Detected mutation sites were exon 19 (8 cases), exon 21 (6 cases), exon 18 (1 cases), and multiple mutations (3 cases). In 18 cases of BM, EGFR mutation studies were done. Among them, 8 (25.6%) cases showed mutation on exon 19 (5 cases) or exon 21 (3 cases). To compare EGFR mutation status between primary lung cancer and BM, 18 paired tissues from both NSCLC and matched BM were collected. Four (22.5%) patients were discordant for the status of EGFR between primary and metastatic sites.ConclusionEGFR mutations were significantly discordant between primary tumors and corresponding metastases in a significant portion of NSCLC. In treatment of BM of EGFR mutant metastatic NSCLC, due to possibility of discordance, pathologic confirming through brain biopsy is recommended.

PUB034 Profile of Lung Cancer in Developing Country: Single Centre Experience? Changing Paradigm

Adenocarcinoma is the commonest histological subtype of non-small cell lung cancer (NSCLC) in most of the Western countries. However, in India squamous cell carcinoma has been reported as the commonest histological type in most of the series. The aim of the study is to analyze the clinico-pathological profile , treatment outcome and survival of NSCLC treated at our center. We analyzed 364 NSCLC registered at our centre over a period of three years. They were evaluated for their clinical and pathological profiles, treatment received and outcome. We also analyzed epidermal growth factor receptor (EGFR) in 250 patients for which formalin-fixed paraffin-embedded tissues was available. EGFR sequencing was performed with ABI PRISM 310 genetic analyzer. Median age was 55 years with a male:female ratio of 5:1. 70% of patients were smokers. Adenocarcinoma was the commonest histological subtype after the pathology review. Among NSCLC, 75% cases were of stage IV. 40% of patients had mutation in one of the four exons characterized. Patients whose EGFR mutational status was not available at presentation before the start of treatment were started on chemotherapy. If EGFR mutational analysis was available and mutations were present, the patients were started on either upfront tyrosine kinase inhibitor (TKI), 30% or if on chemotherapy arm were allowed to finish six cycles and then start with maintenance TKIs, 40%. The median progression free survival for patients with and without mutations was 12 months and 8 months. A median PFS of 16 months was seen in the mutation-positive group that received both chemotherapy followed by switch maintenance with TKIs versus 9 months in the group that received only TKI. This analysis suggests that adenocarcinoma , the commonest histological subtype in India, Most of the patients present at advanced stage and outcome remains poor. The prevalence of EGFR mutations in this population of NSCLC patients was 42% with exon 19 mutation being the most common.

Clinicopathologic features of non-small cell lung cancer in India and correlation with epidermal growth factor receptor mutational status

We performed retrospective analysis of 106 patients with lung cancer for which formalin-fixed paraffin-embedded tissues was available. Their epidermal growth factor receptor (EGFR) mutation status and treatment outcomes are described. All patients with confirmed non-small cell lung cancer (NSCLC) during Jan 2008 to Dec 2010 were included. EGFR sequencing was performed with ABI PRISM 310 genetic analyzer. Forty-two (39.6%) patients had mutation in one of the four exons characterized. Patients whose EGFR mutational status was not available at presentation before the start of treatment were started on chemotherapy, n = 46 (43.39%). If EGFR mutational analysis was available and mutations were present, the patients were started on either upfront tyrosine kinase inhibitor (TKI), n = 15 (14.15%) or if on chemotherapy arm were allowed to finish six cycles and then start with maintenance TKIs, n = 26 (24.52%). The median progression free survival for patients with and without mutations was 11 months (95% CI,7-14) and 9 months (95% CI,7-10) respectively. A median PFS of 14 months (95%CI, 12-16) was seen in the mutation-positive group that received both chemotherapy followed by switch maintenance with TKIs versus 8 months (95%CI, 7-8 months) in the group that received only TKI. The prevalence of EGFR mutations in this population of NSCLC patients was 39.6% with exon 19 mutation being the most common. The observed benefit of addition of chemotherapy over TKI in EGFR mutation-positive group raises the question, can we offer the therapy of chemotherapy-TKI combination to EGFR mutation-positive lung cancer patients as shown in the present study.

Epidermal Growth Factor Recetor mutation study for 5 years, in a population of patients with non-small cell lung cancer

IntroductionIn 2006, the Vila Nova de Gaia/Espinho Hospital Centre Pulmonary Oncology Unit started performing EGFR (Epidermal Growth Factor Receptor) mutation sequencing in selected patients with NSCLC and systematically in all patients since 2010, regardless of histology, smoking habits, age or sex. The aim of this study was to characterize the group of patients that carried out the sequencing between 2006-2010, to determine EGFR mutation frequency, to evaluate the overall survival and the survival after the use of tyrosine kinase inhibitors (TKI), in patients who performed this therapy in second and third line, knowing the EGFR mutation status. MethodsDescriptive statistical analysis of patients who did EGFR sequencing in 2006-2010 and of overall survival in patients treated with TKI as 2nd and 3rd line therapy. Record of the material available for analysis and average delay of exam results, according to the material submitted. ResultsThe sequencing was performed in 374 patients, 71,1% males, 67,1% non/ex-smokers, 32,9% smokers, 57,8% adenocarcinoma and 23,5% squamous cell carcinoma (SCC). The mutation was detected in 49 patients (13,1%). In all studied patients, the mutation rate was 9% in males and 23% in females. Median overall survival after erlotinib use of was 14 months for patients with positive EGFR mutation versus 6 months in not mutated patients (p = 0.003). ConclusionOur group had an overall mutation rate of 13.1% with female, non-smokers, adenocarcinoma histology predominance. In selected patients (2006/2009), the mutation rate was 16%, in not selected patients (2010) the mutation rate was 10.4%. This study has permitted a better understanding of the EGFR mutation rate in the Portuguese population as welll as an evaluation of the patients survival after the use of of tyrosine kinase inhibitors, in second and third line therapy with previous knowledge of the EGFR mutational status. Statistical significant differences in survival were found in the two patient groups (EGFR mutated and non mutated).The EGFR mutation research should be performed in all patients with NSCLC, giving the possibility to a considerable number of patients to perform a first line treatment with TKI (EGFR mutated patients) and the advantage of performing other chemotherapy schemes, when progression occurs.

Heterogeneity of Epidermal Growth Factor Receptor Status and Mutations of KRAS/PIK3CA in Circulating Tumor Cells of Patients with Colorectal Cancer

Molecular characterization of circulating tumor cells (CTCs) is pivotal to increasing the diagnostic specificity of CTC assays and investigating therapeutic targets and their downstream pathways on CTCs. We focused on epidermal growth factor receptor (EGFR) and genes relevant for its inhibition in patients with colorectal cancer (CRC). We used the CellSearch® system for CTC detection in peripheral blood samples from 49 patients with metastatic CRC (mCRC) and 32 patients with nonmetastatic CRC (nmCRC). We assessed EGFR expression in 741 CTCs from 27 patients with mCRC and 6 patients with nmCRC using a fluorescein-conjugated antibody with the CellSearch Epithelial Cell Kit. DNA of a single CTC isolated by micromanipulation was propagated by whole-genome amplification and analyzed by quantitative PCR for EGFR gene amplification and sequencing for KRAS (v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog), BRAF (v-raf murine sarcoma viral oncogene homolog B1), and PIK3CA (phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit α) mutations. At least 2 CTCs were detected in 24 of 49 patients with mCRC and 7 of 32 patients with nmCRC. In 7 of 33 patients, CTCs with increased EGFR expression were identified. Heterogeneity in EGFR expression was observed between CTCs from the same patient. EGFR gene amplification was found in 7 of 26 CTCs from 3 patients. The investigated BRAF gene locus was not mutated in 44 analyzed CTCs, whereas KRAS mutations were detected in 5 of 15 CTCs from 1 patient and PIK3CA mutations in 14 of 36 CTCs from 4 patients. Molecular characterization of single CTCs demonstrated considerable intra- and interpatient heterogeneity of EGFR expression and genetic alterations in EGFR, KRAS, and PIK3CA, possibly explaining the variable response rates to EGFR inhibition in patients with CRC.

Estudo da mutação do recetor do fator de crescimento epidérmico, durante 5 anos, numa população de doentes com cancro do pulmão de não pequenas células

IntroductionIn 2006, the Vila Nova de Gaia/Espinho Hospital Centre Pulmonary Oncology Unit started performing EGFR (Epidermal Growth Factor Receptor) mutation sequencing in selected patients with NSCLC and systematically in all patients since 2010, regardless of histology, smoking habits, age or sex. The aim of this study was to characterize the group of patients that carried out the sequencing between 2006-2010, to determine EGFR mutation frequency, to evaluate the overall survival and the survival after the use of tyrosine kinase inhibitors (TKI), in patients who performed this therapy in second and third line, knowing the EGFR mutation status. MethodsDescriptive statistical analysis of patients who did EGFR sequencing in 2006-2010 and of overall survival in patients treated with TKI as 2nd and 3rd line therapy. Record of the material available for analysis and average delay of exam results, according to the material submitted. ResultsThe sequencing was performed in 374 patients, 71,1% males, 67,1% non/ex-smokers, 32,9% smokers, 57,8% adenocarcinoma and 23,5% squamous cell carcinoma (SCC). The mutation was detected in 49 patients (13,1%). In all studied patients, the mutation rate was 9% in males and 23% in females. Median overall survival after erlotinib use of was 14 months for patients with positive EGFR mutation versus 6 months in not mutated patients (p = 0.003). ConclusionOur group had an overall mutation rate of 13.1% with female, non-smokers, adenocarcinoma histology predominance. In selected patients (2006/2009), the mutation rate was 16%, in not selected patients (2010) the mutation rate was 10.4%. This study has permitted a better understanding of the EGFR mutation rate in the Portuguese population as welll as an evaluation of the patients survival after the use of of tyrosine kinase inhibitors, in second and third line therapy with previous knowledge of the EGFR mutational status. Statistical significant differences in survival were found in the two patient groups (EGFR mutated and non mutated).The EGFR mutation research should be performed in all patients with NSCLC, giving the possibility to a considerable number of patients to perform a first line treatment with TKI (EGFR mutated patients) and the advantage of performing other chemotherapy schemes, when progression occurs.

Effusion Immunocytochemistry as an Alternative Approach for the Selection of First-Line Targeted Therapy in Advanced Lung Adenocarcinoma

Tumor tissue is often not obtainable or suitable for molecular-based epidermal growth factor receptor (EGFR) mutational analysis in advanced non-small-cell lung cancer (NSCLC). This retrospective and single-institution study was conducted to evaluate the role of effusion immunocytochemistry using two EGFR mutant-specific antibodies for the detection of relevant EGFR mutations in NSCLC, along with the selection of candidates for first-line therapy with EGFR tyrosine kinase inhibitors (TKIs). Immunocytochemistry using two antibodies binding specifically to the major forms of mutant EGFR, L858R, and E746-A750 deletion (delE746-A750), was performed on cell blocks of malignant pleural effusion (MPE) from 78 patients with lung adenocarcinoma, who received first-line EGFR TKIs. The yield of EGFR-mutation detection and prediction of response rate and progression-free survival to TKI treatment by immunocytochemistry were compared with those by clinical characteristics and EGFR sequencing using cell-derived RNA from MPEs. Of the 78 MPE samples, direct sequencing using cell-derived RNA identified L858R mutation in 42 cases, deletions in exon 19 in 12 cases (delE746-A750 in eight cases), other types of mutations in three cases, and wild-type EGFR in 21 cases. Effusion immunocytochemistry with these two mutant-specific antibodies exhibited a sensitivity of 71% and 88% and a specificity of 86% and 96% for identifying predefined L858R and delE746-A750 mutations, respectively. Effusion immunocytochemistry provided a superior prediction of tumor response and progression-free survival to first-line EGFR TKIs than did clinical characteristics like sex and smoking status. Patients whose effusion immunocytochemistry showed a reaction to either of the two antibodies had a comparable TKI response rate (67% versus 72%) to those with EGFR mutations assessed by direct sequencing from cell-derived RNA. Effusion immunocytochemistry could be introduced into clinical practice to identify more NSCLC patients likely to have benefit from first-line TKI treatment, especially for those without adequate tissue for molecular-based EGFR analysis.

EML4-ALK Translocation Predicts Better Outcome in Lung Adenocarcinoma Patients with Wild-Type EGFR

The echinoderm microtubule-associated protein like 4-anaplastic lymphoma kinase (EML4-ALK) fusion represents a novel target in a subset of non-small cell lung cancer, especially adenocarcinoma. EML4-ALK fusion is mutually exclusive with epidermal growth factor receptor (EGFR) mutations. To understand the impact of EML4-ALK on the prognosis of non-small cell lung cancer, we examined EML4-ALK fusion in lung adenocarcinoma from patients with wild-type EGFR and analyzed their clinical treatment outcomes. Lung adenocarcinoma patients with malignant pleural effusions having wild-type EGFR and measurable target lesions were enrolled for EML4-ALK analysis by reverse transcription-polymerase chain reaction and direct sequencing. Demographic data, EML4-ALK status, and survival data were analyzed. We also performed fluorescence in situ hybridization on some available tumor samples to validate the PCR result. In addition, K-ras mutation was analyzed for patients without EML4-ALK fusion genes. A total of 116 patients with wild-type EGFR sequencing results had complete clinical data for analysis. No patients received ALK inhibitor therapy. There were 39 patients (34%) with the EML4-ALK fusion gene. The concordance rate between reverse transcription-polymerase chain reaction and fluorescence in situ hybridization was 85%. The K-ras mutation rate for patients without EML4-ALK fusion gene was 6.5%. By multivariate analysis, patients who had better performance status (p < 0.001) and EML4-ALK translocation (p = 0.017) had longer overall survival. Comparing patients with tumors harboring variant 1 with those harboring nonvariant 1 EML4-ALK fusion genes, there were no significant differences in clinical factors and survival outcome. For lung adenocarcinoma patients with wild-type EGFR, EML4-ALK translocation is associated with longer overall survival.

Inherited Germline T790M Mutation and Somatic Epidermal Growth Factor Receptor Mutations in Non-small Cell Lung Cancer Patients

Five cases of non-small cell lung cancer, associated with germline transmission of epidermal growth factor receptor (EGFR)-T790M mutation, have been reported1; these patients had family histories of lung cancer. The activity of gefitinib was tested in only two patients, who were both refractory to this drug.2 Herein, we describe a family of European descent in which two family members had non-small cell lung cancer associated with germline transmission of T790M mutation and who were treated with gefitinib (Figure 1A). The proband was a 72-year-old female, never-smoker, who was diagnosed with an adenocarcinoma of the lung with bilateral pulmonary lesions. This patient was initially treated with chemotherapy for four courses, resulting in stable disease for 6 months, as determined by radiological examination. When the disease progressed, treatment with oral gefitinib at the dose of 250 mg/d was initiated, resulting in a partial response lasting for 9 months; subsequently, the patient developed pleural and cerebral progression of disease and died within 5 months. A 74-year-old never-smoker sister of the proband was diagnosed with a poor differentiated locally advanced (stage IIIB for pleural effusion) carcinoma of the lung and was treated with a first-line chemotherapy, resulting in a partial response lasting for 12 months, after which new bone metastases were observed. As second-line treatment, oral gefitinib 250 mg/d was started, and a partial response was obtained that is lasting from 45 months; the patient is still alive and on treatment with gefitinib. No other members of this family developed lung cancer. DNA was isolated from peripheral blood mononuclear cells or paraffin-embedded tumor samples, using the microDNA-kit (Qiagen, Hilden, Germany). Nested polymerase chain reaction to amplify EGFR (exons 18–21) and K-RAS (exon 1) and sequencing of polymerase chain reaction products on a ABI-3100 Genetic Analyzer was performed. The EGFR-T790M mutation was found in the germline DNA of both patients (Figure 1B), whereas no mutations were detected in the EGFR exons 18, 19, and 21. In contrast, no EGFR mutations were observed in the two daughters of proband and in the two sons of individual II. In the tumor tissue of individual I, we identified the deletion of EGFR E746-A750 in exon 19 but no EGFR mutations in the exons 18, 20, and 21 (Figure 1C). In the tissue of individual II, no EGFR mutations in the exons 18, 19, and 21 were found, whereas the exon 20 could not be studied because no sufficient tumor tissue was present. No mutations were observed in codons 12 and 13 of K-RAS exon 1 in the tissues of both patients. Our data support the hypothesis that an inherited T790M mutation confers an enhanced susceptibility to develop lung tumors3; moreover, the presence of a T790M germline mutation does not necessarily predict for resistance to EGFR-tyrosine kinase inhibitors. The responses to gefitinib could possibly be due to the absence of T790M and K-RAS mutations and the occurrence of EGFR-activating mutations in the tumor tissues. However, other mechanisms, such as increased gene copy number,4 may be responsible for response to gefitinib in the case of individual II.

Good response to pemetrexed in patients of lung adenocarcinoma with epidermal growth factor receptor (EGFR) mutations

There are no reports about epidermal growth factor receptor (EGFR) mutations and the responsiveness to pemetrexed treatment. In order to understand the influence of EGFR mutations on pemetrexed response, we performed EGFR sequencing of lung adenocarcinoma from patients undergoing pemetrexed treatment and analyzed their response to pemetrexed. The pemetrexed-treated lung adenocarcinoma patients with adequate specimens for EGFR sequencing and measurable target lesions were enrolled for response analysis. Demographic data, EGFR mutation status, response to pemetrexed, and survival data were collected. From January 2004 to December 2008, 156 patients with measurable target lesions and EGFR sequencing results had complete clinical data for analysis. The patients with EGFR mutations (N=93) had a better response rate (p=0.016) and longer progression-free survival (PFS) (p=0.030) than those with wild type EGFR (N=63). By multivariate analysis, those who had better performance status (p=0.013) and EGFR mutations (p=0.021) had a longer PFS. In conclusion, lung adenocarcinoma patients receiving pemetrexed with EGFR mutations have a better response rate and longer PFS than those with wild type EGFR.

Neratinib, an Irreversible Pan-ErbB Receptor Tyrosine Kinase Inhibitor: Results of a Phase II Trial in Patients With Advanced Non–Small-Cell Lung Cancer

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have had a significant impact on non-small-cell lung cancer (NSCLC) outcomes, particularly for patients with EGFR mutations. Resistance emerges after 9 to 12 months, primarily mediated by the T790M resistance mutation. We studied neratinib, an irreversible pan-ErbB TKI that may overcome T790M. Patients with advanced NSCLC underwent EGFR sequencing of available tumor tissue at enrollment. Those with > or = 12 weeks of prior TKI therapy were placed in arm A if they were EGFR mutation positive or arm B if they were wild-type. Arm C included TKI-naïve patients with adenocarcinoma and light smoking histories (< or = 20 pack-years). All patients received daily oral neratinib, initially at 320 mg but subsequently reduced to 240 mg because of excessive diarrhea. The primary end point was objective response rate (RR). One-hundred sixty-seven patients were treated: 91 in arm A, 48 in arm B, and 28 in arm C. Diarrhea was the most common toxicity; grade 3 incidence was 50% at 320 mg but improved to 25% after dose reduction. The RR was 3% in arm A and zero in arms B and C. No patients with known T790M responded. Notably, three of four patients with an exon 18 G719X EGFR mutation had a partial response and the fourth had stable disease lasting 40 weeks. Neratinib had low activity in patients with prior benefit from TKIs and in TKI-naïve patients, potentially because of insufficient bioavailability from diarrhea-imposed dose limitation. Responses were seen in patients with the rare G719X EGFR mutation, highlighting the importance of obtaining comprehensive genetic information on trials of targeted agents.

Noninvasive Detection of EGFR T790M in Gefitinib or Erlotinib Resistant Non–Small Cell Lung Cancer

Tumors from 50% of epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer patients that develop resistance to gefitinib or erlotinib will contain a secondary EGFR T790M mutation. As most patients do not undergo repeated tumor biopsies we evaluated whether EGFR T790M could be detected using plasma DNA. DNA from plasma of 54 patients with known clinical response to gefitinib or erlotinib was extracted and used to detect both EGFR-activating and EGFR T790M mutations. Forty-three (80%) of patients had tumor EGFR sequencing (EGFR mutant/wild type: 30/13) and seven patients also had EGFR T790M gefitinib/erlotinib-resistant tumors. EGFR mutations were detected using two methods, the Scorpion Amplification Refractory Mutation System and the WAVE/Surveyor, combined with whole genome amplification. Both EGFR-activating and EGFR T790M were identified in 70% of patients with known tumor EGFR-activating (21 of 30) or T790M (5 of 7) mutations. EGFR T790M was identified from plasma DNA in 54% (15 of 28) of patients with prior clinical response to gefitinib/erlotinib, 29% (4 of 14) with prior stable disease, and in 0% (0 of 12) that had primary progressive disease or were untreated with gefitinib/erlotinib. EGFR T790M can be detected using plasma DNA from gefitinib- or erlotinib-resistant patients. This noninvasive method may aid in monitoring drug resistance and in directing the course of subsequent therapy.

EGFR mutations are detected comparably in cytologic and surgical pathology specimens of nonsmall cell lung cancer

Somatic mutations in the epidermal growth factor receptor (EGFR) are present in approximately 10% of nonsmall cell lung cancers, and higher in never-smokers, women, and Asians. Small in-frame deletions in exon 19 ( approximately 45%) and L858R mutation in exon 21 ( approximately 40%) predict response to treatment with tyrosine kinase inhibitors, whereas some others herald resistance. Direct sequencing of tumor DNA detects all EGFR mutations, but is limited by interference from nonmalignant cells within the samples. Concern over such interference has discouraged testing cytologic samples, but the adequacy of cytologic specimens for EGFR sequencing has not been studied. EGFR sequencing of surgical and cytologic specimens at Brigham and Women's Hospital over the past 2 years was reviewed. Of 239 specimens, 227 (95%) were surgical, and 12 (5%) were cytologic (fine needle aspirations, pleural fluids, bronchial washings, and bronchoalveolar lavages). Sixty-three (28%) surgical specimens showed EGFR mutations, whereas 143 (63%) were negative, 8 (3.5%) failed, and 14 (6.2%) were inconclusive (negative result in a heterogeneous sample). Seven (58%) cytologic specimens showed EGFR mutations, whereas 4 (33%) were negative, and 1 (8.3%) was inconclusive. Cytologic specimens were more likely to have a mutation than surgical specimens (P = .02). There was no significant difference in the frequency of inconclusive results. Cytologic specimens are suitable for EGFR sequencing and show comparable sensitivity for mutation detection as compared with surgical specimens. The suitability of a sample should be determined on a case-by-case basis, and cytologic samples should not be dismissed as inadequate without a thorough review.

Functional Analysis of Epidermal Growth Factor Receptor (EGFR) Mutations and Potential Implications for EGFR Targeted Therapy

Epidermal growth factor receptor (EGFR) mutations in non-small cell lung cancer (NSCLC) might be predictive for clinical response to EGFR inhibitor treatment. However, retrospective analyses of EGFR mutations in clinical trials have shown inconclusive results and the effect of EGFR sequencing in NSCLC is still controversial. Because the vast majority of EGFR mutations described have not been functionally characterized, simple correlation of mutational status and treatment response may not provide reliable information about the predictive value of EGFR mutations. Thus, we aimed to characterize a comprehensive panel of clinically observed EGFR mutations. A panel of 30 EGFR mutations was cloned and characterized for kinase activity and the ability to confer growth factor independence. Interestingly, 4 of 30 EGFR mutations showed no kinase activity even after ligand stimulation and were not able to confer growth factor independence. Ba/F3 cells expressing activating EGFR mutants were then used to test the efficacy of EGFR inhibitors in a cell proliferation assay. IC(50) values were calculated for gefitinib, erlotinib, and AEE788. We show that the sensitivity of EGFR mutations toward different inhibitors varies significantly, thus establishing a comprehensive sensitivity profile for each inhibitor. EGFR mutations identified in NSCLC patients display distinct biological features. The variability in kinase activity, transforming potential, and sensitivity to EGFR inhibitors has to be considered in clinical studies aiming to correlate mutational status and drug response. The identification of comprehensive drug resistance profiles opens the opportunity to test alternative EGFR inhibitors in vitro.

Gefitinib for Epidermal Growth Factor Receptor Mutant Lung Cancers: Searching for a Weapon of Mass Destruction

In the spring of 2004, the association between somatic mutations in the epidermal growth factor receptor (EGFR) in non– small-cell lung cancer (NSCLC) and sensitivity to the EGFR tyrosine kinase inhibitor gefitinib was first described. Since that time, numerous retrospective studies have confirmed these original observations. Patients with EGFR mutations treated with gefitinib have a 60% to 90% chance of achieving a radiographic partial response and a median time to progression of approximately 12 months (Table 1). However, these were all retrospective observations that were limited by multiple potential biases including patient selection based on the availability of tumor material, differences in prior treatments, and the methods of EGFR mutation detection. In addition, other biomarkers associated with gefitinib’s clinical benefit, including increased EGFR copy number and EGFR expression as detected by immunohistochemistry, have also emerged over the last 2 years. The contribution and role of EGFR mutations remain to be definitively determined from ongoing prospective studies. In this issue, Inoue et al report the first prospective clinical trial in which chemotherapy-naive advanced NSCLC patients with somatic EGFR mutations were treated with gefitinib as their first systemic therapy. They started the study in June 2004, performed mutational analyses in 99 patients (75 patients were assessable), and found EGFR mutations in 25 of the patients (33%). The mutation frequency in this study is consistent with the previously published somatic EGFR mutation frequency in Japanese lung cancer patients. The 16 patients with EGFR mutations had a response rate of 75% and a median progression-free survival time of 9.7 months, which is consistent with the retrospective observations (Table 1). Inoue et al also found the same types of EGFR mutations as previously identified, with in-frame deletion of exon 19 being the most common type of mutation (Table 1). Despite the small sample size, the study met its primary protocol-defined end point of a 70% response rate, and this observation in a prospective trial confirms the previous retrospective observations associating EGFR mutations and the efficacy of gefitinib. What are the implications of this study? The findings of Inoue et al leave little doubt about the predictive value of EGFR somatic mutations (exon 19 deletions and L858R) for radiographic response after treatment with gefitinib. However, their study does not address whether these patients will survive longer when treated with gefitinib compared with chemotherapy. This will require a phase III trial comparing the outcome of patients with EGFR somatic mutations treated with chemotherapy with the outcome of patients treated with gefitinib. In addition, the role of adding systemic chemotherapy to EGFR tyrosine kinase inhibitors for this patient population remains to be determined and is being evaluated in ongoing studies. Inoue et al also demonstrate the feasibility of EGFR mutation detection in the routine clinical care of chemotherapy-naive patients with advanced NSCLC. The Japanese population is more likely to have a somatic EGFR mutation than the white population (30% in Japanese patients compared with 10% to 15% in white patients). This increased mutation frequency, combined with the rapid turn around time (median, 7 days) for EGFR sequencing, clearly helped this study. In the United States or Western Europe, approximately 300 patients would need to be screened (rather than the 99 patients screened in the study by Inoue et al) to find patients with sufficient tumor material for EGFR sequencing and, ultimately, patients with EGFR mutations to obtain a similar number of assessable patients. Nevertheless, these findings suggest that this may be a worthwhile effort in Western populations as well. Inoue et al found the most common types of EGFR mutations (exon 19 deletions and L858R) in their patient population. To date, more than 20 different EGFR mutations have been described. The predictive value for radiographic response to gefitinib treatment of EGFR mutations, other than exon 19 deletions or L858R, still needs to be determined from ongoing clinical studies. In the study by Inoue et al, there were four patients with NSCLC and EGFR mutations who either had stable (n 2) or progressive disease (n 2) after treatment with gefitinib. What is different about these patients compared with the patients with EGFR mutations who respond to treatment with gefitinib? There are no obvious differences in the types of EGFR mutations among these patients compared with the other 12 patients who achieved a partial response. In addition, the DNA from the tumors of the patients with stable or progressive disease did not contain the previously described EGFR T790M mutation associated with gefitinib resistance. Several recent studies have demonstrated that, both in EGFR mutant NSCLC cell lines and in patients with JOURNAL OF CLINICAL ONCOLOGY E D I T O R I A L VOLUME 24 NUMBER 21 JULY 2

Estimation of the likelihood of epidermal growth factor receptor (EGFR) mutations based on cigarette smoking history in patients with adenocarcinoma of the lung

7069 Background: Lung adenocarcinomas with EGFR mutations demonstrate sensitivity to gefitinib or erlotinib. Unfortunately, the majority of patients with lung adenocarcinoma have inadequate tumor specimens to permit EGFR sequencing prior to treatment. Pao (Pao W. PNAS 2004) and others reported a higher incidence of EGFR mutations in never smokers. In this study we characterized features of a smoking history that predict for the presence of EGFR mutations. Methods: EGFR exons 18–24 were sequenced in 127 tumor samples using standard polymerase chain reaction techniques. Smoking histories with age at first cigarette, packs per day, number of smoking and smoke-free years were obtained. Patients were categorized as never smokers (<100 lifetime cigarettes), former smokers (quit ≥1 year ago), or current smokers (quit <1 year ago). The Fisher’s exact and Wilcoxon’s rank tests were used. The receiver operating characteristic (ROC) curve methodology assessed the ability of smoking history parameters to predict the likelihood of a mutation in EGFR. Results: We detected EGFR mutations in 17/32 never smokers (53%, 95% CI 35–71%), 8/62 former smokers (13%, 95% CI 6–24%) and 2/33 current smokers (6%, 95% CI 1–20%). The incidence of EGFR mutations and number of pack years smoked is in the table below. The areas under the ROC curves for pack years of smoking and years smoke-free are 0.84 and 0.77, respectively. Conclusions: EGFR mutations correlate with smoking history. The likelihood of EGFR mutations decreases as the number of pack years smoked increases. The optimal smoking history parameters identified are <10 pack years and >25 smoke-free years. These data can assist clinicians in assessing the probability of mutations in EGFR, and sensitivity to gefitinib or erlotinib, in patients with lung adenocarcinoma. A smoking history may provide useful information when direct sequencing is not feasible. Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration AstraZeneca, Genentech AstraZeneca AstraZeneca

A phase I trial of erlotinib and bexarotene as a targeted combination therapy for aerodigestive tract cancers

7093 Background: We previously reported the overexpression of the epidermal growth factor receptor (EGFR) and cyclin D1 as early events in lung carcinogenesis. All-trans-retinoic acid (RA) suppressed EGFR expression through a transcriptional mechanism and cyclin D1 through proteasome-dependent proteolysis. Non-classical retinoids such as the rexinoid, bexarotene (B), repress both cyclin D1 and EGFR expression but signal independent of RAR-beta, which is often deregulated in lung carcinogenesis. Combining an EGFR tyrosine kinase inhibitor, erlotinib (E), with B induced at least additive suppression of growth and cyclin D1 expression in RA-resistant human bronchial epithelial cells which had silenced RAR beta and in some lung cancer cell lines. Methods: A phase I trial of E and B was conducted in patients (pts) with advanced aerodigestive tract cancers. Primary objective - maximum tolerated dose. Secondary objectives - toxicities, efficacy, and surrogate markers of response (cyclin D1) in buccal swabs from pts at the highest dose. 3 dose levels - 1: B 300 mg/m2, E 100 mg; 2: B 300 mg/m2, E 150 mg; 3: B 400 mg/m2, E 150 mg. At least 3 pts were enrolled at each level. All pts received atorvastatin for elevated triglycerides. Results: Twenty-four pts were enrolled including 46% women and 79% with NSCLC. The median age was 61 years, 62% had first-line and 21% second line chemotherapy, one third had KPS of 70, and 67% were former smokers. Toxicities were mild with frequent hypertriglyceridemia and skin rash. No cases of pancreatitis were observed. Two DLTs were observed: CPK elevation and systemic pain. Four objective partial responses were seen in pts with NSCLC, eight pts (5 NSCLC, 3 head/neck) had stable disease. Median time to progression was 2.23 months, median overall survival was 14.1 months, and one-year survival was 59%. EGFR sequencing revealed no activating mutations in exons 18, 19, and 21 for two responding cases. Cyclin D1 protein expression was repressed in buccal swabs from 5/6 pts. Conclusions: The recommended phase II doses are erlotinib 150 mg and bexarotene 400 mg/m2 daily. This regimen is well tolerated and shows preliminary evidence of activity for the treatment of advanced aerodigestive tract cancer. Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Ligand Genentech, Ligand Genentech, Ligand