反转录-聚合酶链反应、免疫组织化学和荧光原位杂交方法检测棘皮动物微管相关蛋白样 4-间变性淋巴瘤激酶融合阳性非小细胞肺癌的比较：对最佳临床检测的影响。

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.

机构信息

Institute for Clinical and Experimental Pathology, ARUP Laboratories, Salt Lake City, Utah, USA.

出版信息

Arch Pathol Lab Med. 2012 Jul;136(7):796-803. doi: 10.5858/arpa.2011-0321-OA.

DOI:10.5858/arpa.2011-0321-OA

PMID:22742552

Abstract

CONTEXT

Echinoderm microtubule-associated proteinlike 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 RT-PCR, 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.

摘要

背景

在 3%至 13%的非小细胞肺癌中检测到棘皮动物微管相关蛋白样 4-间变性淋巴瘤激酶（EML4-ALK）基因融合。准确检测 EML4-ALK 融合对于选择合适的治疗方法至关重要。

目的

比较逆转录聚合酶链反应（RT-PCR）、免疫组织化学（IHC）和荧光原位杂交（FISH）方法检测 EML4-ALK 融合。

设计

选择 46 例富含野生型表皮生长因子受体（EGFR）状态的肺腺癌（野生型，n=42；突变型，n=4）。通过 IHC（Dako；ALK1 克隆）、FISH（雅培分子；LSI ALK 分离）和 RT-PCR（变体 1 和 3a/b）对标本进行检测。

结果

EML4-ALK 变体 3a/b 可通过 RT-PCR、FISH 和 IHC 在 4%（46 例中的 2 例）标本中检测到。FISH 和 IHC 观察者之间对变体 3a/b 完全达成一致。EML4-ALK 变体 1 的检测方法之间没有一致性。RT-PCR 方法在 20%（46 例中的 9 例）标本中检测到变体 1。由于只有 11%（9 例中的 1 例）的标本被所有 3 位观察者均评为阳性，因此 FISH 观察者对变体 1 的一致性较差。IHC 检测变体 1 的敏感性也较差，因为只有 9 个样本中的 1 个（11%）被评为阳性。总的来说，在富含野生型 EGFR 状态的腺癌中，EML4-ALK 变体 1 和 3a/b 的频率为 24%（46 例中的 11 例）。发现 1 例 EML4-ALK 变体 1 融合与 EGFR 外显子 21 突变共存。