A diagnostic algorithm using EGFR mutation-specific antibodies for rapid response EGFR-TKI treatment in patients with non-small cell lung cancer

Kawahara, Akihiko; Taira, Tomoki; Azuma, Koichi; Tominaga, Masaki; Hattori, Satoshi; Kawahara, M; Abe, Hideyuki; Yamaguchi, Tomohiko; Akiba, Jun; Takamori, Shinzo; Hayashi, Akira; Kage, Masayoshi

doi:10.1016/j.lungcan.2012.07.002

Cited by 19 publications

(33 citation statements)

References 17 publications

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“…Therefore, a more extensive research is required to reach a consensus. The results of the present study and other reports [15]–[19], [23] suggest that scoring criteria based on the staining intensity of the membrane and/or cytoplasm of tumor cells and the percentage of the staining area (which was divided into four grades: 0, 1+, 2+ and 3+) may be the best of all available scoring systems. The agreement between IHC-based and molecular-based assays was calculated using Cohen κ in accordance with different levels of immunohistochemical grading.…”

Section: Discussionsupporting

confidence: 61%

“…Some researchers also obtained a score for expression by multiplying the staining intensity by the percentage of staining area (0–300 or 400), and respectively advocated a score of >10 or >20 to be categorized as positive, but an appreciable difference in sensitivities (42.2–100%) and specificities (77–100%) was observed [20]–[22]. Recently, Kawahara et al proposed that immunostaining should be classified as positive (score of 2+), negative (score of 0) or equivocal (score of 1+), indicating questionable, negative or weak expression, respectively, which can obtain a sensitivity of 81.4%, specificity of 97.5%, positive predictive value (PPV) of 94.6%, and negative predictive value (NPV) of 90.6% [23]. Consensus of a universally accepted criterion for “positive” is lacking, which severely hinders the clinical use of EGFR mutation-specific antibody to detect EGFR mutations.…”

Section: Introductionmentioning

confidence: 99%

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Ascertaining an Appropriate Diagnostic Algorithm Using EGFR Mutation-Specific Antibodies to Detect EGFR Status in Non-Small-Cell Lung Cancer

et al. 2013

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BackgroundEpidermal growth factor receptor (EGFR) mutation status is the most valuable indicator in the screening of non-small-cell lung cancer (NSCLC) patients for tyrosine kinase inhibitor (TKI) therapy. Accurate, rapid and economical methods of detecting EGFR mutations have become important. The use of two mutation-specific antibodies targeting the delE746-A750 mutation in exon 19 and L858R mutation in exon 21 makes this task possible, but the lack of consensually acceptable criteria for positive results limits the application of this antibody based mutation detection.MethodsWe collected 399 specimens from NSCLC patients (145 resection specimens, 220 biopsy specimens, and 34 cytology specimens) whose EGFR mutation status had been detected by TaqMan PCR assay. Immunohistochemical (IHC) analyses using EGFR mutation-specific antibodies were employed for all samples. After staining and scoring, the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were calculated in accordance with different levels of positive grades in comparison with the results of PCR-based assay.ResultsIn IHC-based analyses, 144 cases were scored 0, 104 cases were scored 1+, 103 cases were scored 2+, and 48 cases were scored 3+. With the molecular-based results were set as the “gold standard”, the prevalence of mutation was 6.94% (10/144), 23.08% (24/104), 67.96% (70/103) and 100% (48/48), respectively, for samples with scores 0, 1+, 2+ and 3+. When score 3+ was considered positive, the specificity and PPV were 100%; if only score 0 was considered negative, 93.06% NPV was obtained.ConclusionPatients with score 3+ have a perfect PPV (100%), and may accept TKI treatment directly without any molecular-based assays. Patients with score 0 had high NPV (93.06%), which could reach 97.22% when the detection of total EGFR was applied. However, samples with score 1+ or 2+ are unreliable and need further verification of EGFR mutation status by molecular-based assays.

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Section: Discussionsupporting

confidence: 61%

Section: Introductionmentioning

confidence: 99%

Ascertaining an Appropriate Diagnostic Algorithm Using EGFR Mutation-Specific Antibodies to Detect EGFR Status in Non-Small-Cell Lung Cancer

et al. 2013

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“…These methods include Sanger sequencing, single strand conformation polymorphism (SSCP) [6], co-amplification at lower denaturation temperature-PCR (COLD PCR) [7], immunohistochemistry with EGFR -mutation specific antibodies [8], peptide nucleic acid-locked nucleic acid (PNA-LNA) PCR clamp assays, real-time PCR (RT-PCR) methods [9] and next generation sequencing [10]. However, none of the methods that have been used so far are ideal, and each of these methods has limitations that mostly relate to the following characteristics of cancer samples: (i) diversified types of tumor samples available for analysis (surgical, biopsied), (ii) contamination with normal cells, (iii) the genetic heterogeneity of tumors, (v) the often low frequency of the analyzed mutations, (iv) degradation of DNA, and (v) damage to or modification of DNA [the two latter factors also apply to formalin-fixed, paraffin-embedded (FFPE) samples, which are the most frequently available samples].…”

Section: Introductionmentioning

confidence: 99%

The Use of a Two-Tiered Testing Strategy for the Simultaneous Detection of Small EGFR Mutations and EGFR Amplification in Lung Cancer

et al. 2015

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Lung cancer is the leading cause of cancer-related death worldwide. Recent progress in lung cancer diagnosis and treatment has been achieved due to a better understanding the molecular mechanisms of the disease and the identification of biomarkers that allow more specific cancer treatments. One of the best known examples of personalized therapy is the use of tyrosine kinase inhibitors, such as gefitinib and erlotinib, for the successful treatment of non-small-cell lung cancer patients selected based on the specific EGFR mutations. Therefore, the reliable detection of mutations is critical for the application of appropriate therapy. In this study, we tested a two-tiered mutation detection strategy using real-time PCR assays as a well-validated high-sensitivity method and multiplex ligation-dependent probe amplification (MLPA)-based EGFRmut+ assay as a second-tier standard-sensitivity method. One additional advantage of the applied MLPA method is that it allows the simultaneous detection of EGFR mutations and copy-number alterations (i.e., amplifications) in EGFR, MET and ERBB2. Our analysis showed high concordance between these two methods. With the use of this two-tier strategy, we reliably determined the frequency of EGFR mutations and EGFR, MET and ERBB2 amplifications in over 200 lung cancer samples. Additionally, taking advantage of simultaneous copy number and small mutation analyses, we showed a very strong correlation between EGFR mutations and EGFR amplifications and a mutual exclusiveness of EGFR mutations/amplifications with MET and ERBB2 amplifications. Our results proved the reliability and usefulness of the two-tiered EGFR testing strategy.

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“…The applicability of IHC in predicting response to EGFR-targeted therapy remains controversial. Confirming the importance of EGFR-mutational status for treatment response, positive IHC staining has been associated with longer progression-free survival compared to IHC-negative or -equivocal cases (83, 87). In addition, high mutant EGFR expression (as defined by the sum of scores for fraction and intensity) was significantly related to elevated progression-free survival but not overall survival (88) and a fraction of positive tumor cells exceeding 50% of all cells predicted better response to EGFR inhibition treatment in univariate but not multivariate analysis (89).…”

Section: Mutation-specific Antibodiesmentioning

confidence: 90%

In situ Protein Detection for Companion Diagnostics

et al. 2013

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The emergence of targeted therapies for cancer has created a need for the development of companion diagnostic tests. Assays developed in recent years are aimed at determining both the effectiveness and safety of specific drugs for a defined group of patients, thus, enabling the more efficient design of clinical trials and also supporting physicians when making treatment-related decisions. Immunohistochemistry (IHC) is a widely accepted method for protein expression analyses in human tissues. Immunohistochemical assays, used to localize and quantitate relative protein expression levels within a morphological context, are frequently used as companion diagnostics during clinical trials and also following drug approval. Herein, we describe established immunochemistry-based methods and their application in routine diagnostics. We also explore the possibility of using IHC to detect specific protein mutations in addition to DNA-based tests. Finally, we review alternative protein binders and proximity ligation assays and discuss their potential to facilitate the development of novel, targeted therapies against cancer.

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A diagnostic algorithm using EGFR mutation-specific antibodies for rapid response EGFR-TKI treatment in patients with non-small cell lung cancer

Cited by 19 publications

References 17 publications

Ascertaining an Appropriate Diagnostic Algorithm Using EGFR Mutation-Specific Antibodies to Detect EGFR Status in Non-Small-Cell Lung Cancer

Ascertaining an Appropriate Diagnostic Algorithm Using EGFR Mutation-Specific Antibodies to Detect EGFR Status in Non-Small-Cell Lung Cancer

The Use of a Two-Tiered Testing Strategy for the Simultaneous Detection of Small EGFR Mutations and EGFR Amplification in Lung Cancer

In situ Protein Detection for Companion Diagnostics

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