Comparison of Reverse Transcription-Polymerase Chain Reaction, Immunohistochemistry, and Fluorescence In Situ Hybridization Methodologies for Detection of Echinoderm Microtubule-Associated Proteinlike 4-Anaplastic Lymphoma Kinase Fusion-Positive Non-Small Cell Lung Carcinoma Implications for Optimal Clinical Testing
Comparison of Reverse Transcription-Polymerase Chain Reaction, Immunohistochemistry, and Fluorescence In Situ Hybridization Methodologies for Detection of Echinoderm Microtubule-Associated Proteinlike 4-Anaplastic Lymphoma Kinase Fusion-Positive Non-Small Cell Lung Carcinoma Implications for Optimal Clinical Testing
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DOI：
10.5858/arpa.2011-0321-oa
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发表时间：
2012-07-01
影响因子：
4.6
通讯作者：
Layfield, Lester J.
中科院分区：
文献类型：
--
作者：
Wallander, Michelle L.;Geiersbach, Katherine B.;Layfield, Lester J.
背景：棘皮微管相关蛋白样间变性淋巴瘤激酶(EML4-ALK)基因融合在3%至13%的非小细胞肺癌中被检测到。准确检测EML4-ALK融合对于适当的治疗选择是至关重要的。目的：比较逆转录聚合酶链式反应(RT-PCR)、免疫组织化学(IHC)和荧光原位杂交(FISH)检测EML4-ALK融合的方法。设计：选择46例肺腺癌，进行野生型表皮生长因子受体(EGFR)状态检测(野生型，42例；突变型，4例)。用IHC(DAKO；克隆ALK1)、FISH(雅培分子；LSI ALK裂解)和RT-PCR(变异体1和3a/b)进行检测。结果RT-PCR、FISH和IHC检测到4%(2/46)的标本中-EML4-ALK变异体3a/b。FISH和IHC对变异体3a/b的检测结果完全一致。EML4-ALK变异体1的检测方法之间没有一致性。RT-PCR方法在20%(9/46)的标本中检测到变异体1。鱼类观察者对变体1的一致性很差，因为只有11%(1/9)的标本被所有3个观察者评分为阳性。IHC对检测变异体1的敏感性也很差，因为9份样本中只有1份(11%)评分为阳性。总体而言，在富含野生型EGFR状态的腺癌中，EML4-ALK变异体1和3a/b的频率为24%(11/46)。一个EML4-ALK变异体1融合被发现与EGFR外显子21突变共存。结论-FISH解释在观察者中表现出很大的差异。RT-PCR方法是检测EML4-ALK融合的最敏感和最不主观的方法。(ARCH Pathol Lab Med.2012年；136.796803DOI：10.5858/arpa.2011-0321OA)
Context.-Echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase (EML4-ALK) gene fusions are detected in 3% to 13% of non-small cell lung carcinomas. Accurate testing for detection of EML4-ALK fusions is essential for appropriate therapy selection.Objective.-To compare reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemistry (IHC), and fluorescence in situ hybridization (FISH) methodologies for detection of EML4-ALK fusions.Design.-Forty-six pulmonary adenocarcinomas were selected with enrichment for wild-type epidermal growth factor receptor (EGFR) status (wild type, n = 42; mutant, n = 4). Specimens were tested by IHC (Dako; clone ALK1), FISH (Abbott Molecular; LSI ALK break apart), and RT-PCR (variants 1 and 3a/b).Results.-EML4-ALK variant 3a/b was detectable by RTPCR, FISH, and IHC in 4% (2 of 46) of specimens. Complete agreement among FISH and IHC reviewers was obtained for variant 3a/b. No concordance existed among methodologies for the detection of EML4-ALK variant 1. The RT-PCR method detected variant 1 in 20% (9 of 46) of specimens. Agreement among FISH viewers was poor for variant 1 because only 11% (1/9) of specimens were scored as positive by all 3 viewers. The sensitivity of IHC for detection of variant 1 was also poor because only 1 of 9 samples (11%) was scored as positive. Overall, the frequency of EML4-ALK variants 1 and 3a/b was 24% (11 of 46) in adenocarcinomas enriched for wild-type EGFR status. One EML4-ALK variant 1 fusion was found to coexist with an EGFR exon 21 mutation.Conclusions.-The FISH interpretation demonstrated great variability among observers. The RT-PCR method was the most sensitive and least-subjective methodology for detection of EML4-ALK fusions. (Arch Pathol Lab Med. 2012; 136: 796-803; doi: 10.5858/arpa.2011-0321-OA)