Targeted sequencing reveals distinct pathogenic variants in Chinese patients with lung adenocarcinoma brain metastases

Abstract

Lung cancer is the most common type of malignancy to metastasize to the brain, with the median survival time of patients being 6-11 months. In the present study, the aim was to compare the actionable gene mutation profiles of primary lung adenocarcinoma (LC) samples and LC brain metastasis (LCBM) samples through targeted sequencing. Next generation sequencing (NGS) of 13 formalin-fixed, paraffin-embedded LC samples and 15 LCBM samples was performed using a customized OncoAim™ cancer panel and OncoAim™ RNA fusion panel on the MiSeq platform. The OncoAim™ cancer panel pipeline and OncoAim™ RNA fusion panel pipeline were used for bioinformatic analysis. Together, 43 variants were observed in 7 genes from the 28 cancer samples. The mutated genes of LCBM were tumor protein (TP)53, epidermal growth factor receptor (EGFR), catenin β1, phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit α, mothers against decapentaplegic homolog 4, Kirsten rat sarcoma viral oncogene homolog (KRAS) and proto-oncogene B-Raf, which were exhibited in 10/15 (66.7%), 6/15 (40.0%), 3/15 (20.0%), 2/15 (13.3%), 2/15 (13.3%), 1/15 (6.7%) and 1/15 (6.7%) of samples, respectively. The mutated genes of LC were TP53, EGFR and KRAS, which were exhibited in 11/13 (84.6%), 5/13 (38.5%) and 2/13 (18.2%) of samples, respectively. echinoderm microtubule associated protein like 4-anaplastic lymphoma kinase rearrangements were present in 1 LCBM sample. For 2 LC samples and 1 LCBM sample, no genetic alterations were observed. The NGS data also revealed a novel 4-codon deletion of TP53 (p.V166_H169del) and a novel TP53 splice site mutation (7577157-63del TACTCAG). Further potentially actionable mutations were detected in LCBM, indicating a high degree of genetic heterogeneity between the LC and LCBM samples that were analyzed. The present study demonstrated that NGS provides an improved approach for the discovery of potentially actionable mutations and the understanding of the mechanisms underlying tumor progression and evolution.

Keywords: brain metastases; gene mutations; lung adenocarcinoma; next-generation sequencing; targeted therapy.

Figure 1.

Heat map depicting the somatic mutations identified in each sample. Red indicates variants, whereas gray indicates no variants detected. The y-axis indicates the gene name and the x-axis indicates the identification of the matched pairings, including 13 samples of LC and 15 samples of LCBM. LCBM, lung adenocarcinoma brain metastasis; TP53, tumor protein 53; EGFR, epithelial growth factor receptor; CTNNB1, catenin-β1; KRAS, Kirsten rat sarcoma viral oncogene; PIK3CA, phosphatidylinositol-4,5-bisphosphate 3-kinase; BRAF, proto-oncogene B-Raf; SMAD4, mothers against decapentaplegic homolog 4.

Figure 2.

(A) Mutations identified in 28 samples and their distribution. (B) Distribution of samples according to variants identified. TP53, tumor protein 53; EGFR, epithelial growth factor receptor; CTNNB1, catenin-β1; KRAS, Kirsten rat sarcoma viral oncogene; PIK3CA, phosphatidylinositol-4,5-bisphosphate 3-kinase gene; BRAF, proto-oncogene B-Raf; SMAD4, mothers against decapentaplegic homolog 4.

Figure 3.

RNA fusion panel tests revealed only EML4(13)-ALK(20) in one sample of lung adenocarcinoma brain metastasis. EML4, echinoderm microtubule associated protein like 4; ALK, anaplastic lymphoma kinase. The blue bar represents the Cris, and the pink bar represents the Watson strand of DNA in the reference genome (hg19).

Figure 4.

Variant frequency detected by next generation sequencing in 28 formalin-fixed, paraffin-embedded samples sub-grouped by LC and LCBM. CTNNB1, PI3KCA, SMAD4, BRAF and EML4-ALK mutations were present in the metastatic lesions in 3/15, 2/15, 2/15, 1/15 and 1/15 LCBM cases, but not in primary LC. LCBM, lung adenocarcinoma brain metastasis; LC, lung adenocarcinoma; TP53, tumor protein 53; EGFR, epithelial growth factor receptor; CTNNB1, catenin-β1; KRAS, Kirsten rat sarcoma viral oncogene; PIK3CA, phosphatidylinositol-4,5-bisphosphate 3-kinase gene; BRAF, proto-oncogene B-Raf; EML4, echinoderm microtubule associated protein Like 4; ALK, anaplastic lymphoma kinase; SMAD4, mothers against decapentaplegic homolog 4.

Figure 5.

Sanger sequencing chromatograms. (A) TP53-p.V166_H169del and (B) TP53-wild-type; (C) TP53-7577157-63del TACTCAG and (D) TP53-wild-type. TP53, tumor protein 53.