Department of Pathology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, Jian She Dong Road 1, Zhengzhou, 450052, Henan, China.
Institute of Medical and Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, 450052, China.
J Transl Med. 2021 Jul 16;19(1):308. doi: 10.1186/s12967-021-02982-4.
Complex kinase rearrangement, a mutational process involving one or two chromosomes with clustered rearrangement breakpoints, interferes with the accurate detection of kinase fusions by DNA-based next-generation sequencing (NGS). We investigated the characteristics of complex ALK rearrangements in non-small cell lung cancers using multiple molecular tests.
Samples of non-small cell lung cancer patients were analyzed by targeted-capture DNA-based NGS with probes tilling the selected intronic regions of fusion partner genes, RNA-based NGS, RT-PCR, immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH).
In a large cohort of 6576 non-small cell lung cancer patients, 343 (5.2%) cases harboring ALK rearrangements were identified. Fourteen cases with complex ALK rearrangements were identified by DNA-based NGS and classified into three types by integrating various genomic features, including intergenic (n = 3), intragenic (n = 5) and "bridge joint" rearrangements (n = 6). All thirteen cases with sufficient samples actually expressed canonical EML4-ALK fusion transcripts confirmed by RNA-based NGS. Besides, positive ALK IHC was detected in 13 of 13 cases, and 9 of 11 cases were positive in FISH testing. Patients with complex ALK rearrangements who received ALK inhibitors treatment (n = 6), showed no difference in progression-free survival (PFS) compared with patients with canonical ALK fusions n = 36, P = 0.9291).
This study firstly reveals the molecular characteristics and clinical outcomes of complex ALK rearrangements in NSCLC, sensitive to ALK inhibitors treatment, and highlights the importance of utilizing probes tilling the selected intronic regions of fusion partner genes in DNA-based NGS for accurate fusion detection. RNA and protein level assay may be critical in validating the function of complex ALK rearrangements in clinical practice for optimal treatment decision.
涉及一个或两个染色体的复杂激酶重排，这些染色体簇集的重排断点会干扰基于 DNA 的下一代测序（NGS）对激酶融合的准确检测。我们使用多种分子检测方法研究了非小细胞肺癌中复杂 ALK 重排的特征。
通过靶向捕获基于 DNA 的 NGS 对非小细胞肺癌患者的样本进行分析，该方法使用了针对融合伙伴基因选定内含子区域的探针、RNA 基于 NGS、RT-PCR、免疫组织化学（IHC）和荧光原位杂交（FISH）。
在一个包含 6576 例非小细胞肺癌患者的大样本队列中，鉴定出 343 例（5.2%）携带 ALK 重排的病例。通过 DNA 基于 NGS 鉴定出 14 例具有复杂 ALK 重排的病例，并通过整合各种基因组特征将其分为三种类型，包括基因间（n=3）、基因内（n=5）和“桥接”重排（n=6）。所有 13 例具有足够样本的病例实际上都通过 RNA 基于 NGS 证实了经典的 EML4-ALK 融合转录本的表达。此外，13 例中有 13 例的 ALK IHC 阳性，11 例中有 9 例 FISH 检测阳性。接受 ALK 抑制剂治疗的复杂 ALK 重排患者（n=6）与具有经典 ALK 融合的患者（n=36）相比，无进展生存期（PFS）无差异，P=0.9291）。
本研究首次揭示了 NSCLC 中复杂 ALK 重排的分子特征和临床结果，对 ALK 抑制剂治疗敏感，并强调了在 DNA 基于 NGS 中使用针对融合伙伴基因选定内含子区域的探针进行准确融合检测的重要性。RNA 和蛋白质水平检测可能对验证复杂 ALK 重排在临床实践中的功能以做出最佳治疗决策至关重要。
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