Whole Exome Sequencing of Lacrimal Gland Adenoid Cystic Carcinoma

Abstract

Purpose: To identify genomic mutations in lacrimal gland adenoid cystic carcinoma (LGACC) samples from patients.

Methods: Genomic DNA was extracted from LGACC specimens. Whole exome sequencing (exome-seq) was conducted to screen for mutations. Capillary sequencing was performed to verify mutations in genes shared by multiple samples. Luciferase assays were used to evaluate functional consequences of NOTCH1 mutations.

Results: The mutation profile of LGACC was complicated. The most frequently mutated gene observed (28.6%) was bromodomain PHD finger transcription factor (BPTF). No mutation was identified in common cancer genes such as TP53, KRAS, and BRAF. However, mutations predicted to be functionally severe were accumulated in the Notch signaling pathway including NOTCH1 and NOTCH2, of which mutations have been reported in head/neck adenoid cystic carcinoma (ACC). Of 14 LGACC samples, five samples carry mutations in Notch pathway genes. Capillary sequencing verified all the mutations in the two NOTCH genes identified by exome-seq. Compared to the wild-type NOTCH1, three frame shifting mutations and two missense mutations (C387W and L1600Q) increased luciferase activity approximately 10- to 25-fold.

Conclusions: Major genomic mutation profiles in LGACC were uncovered by exome-seq. Although preliminary in nature, the Notch pathway could be a potential therapeutic target for LGACC.

Figure 1

A comutation plot presents the top 6 most frequently mutated genes in LGACC samples (n = 14).

Figure 2

A comutation plot shows the mutation profile of the top 25 most mutated cancer genes.

Figure 3

A side-by-side comparison of comutation plots of (A) LGACC and (B) head/neck ACC shows the mutations in LGACC, which are corresponding to the top mutated genes identified in head/neck ACC.

Figure 4

A comutation plot presents the mutations of genes involved in the Notch signaling pathway in LGACC.

Figure 5

Electropherogram traces of capillary sequencing verify various mutations identified in the genes NOTCH1 and NOTCH2 by whole exome sequencing.

Figure 6

Western blot showing the truncated Notch1 protein from frameshift mutations in (A) Notch1. Notch signaling, measured by relative luciferase activities (RLU), is activated by mutations (missense or frame shifting) in the NOTCH1 gene compared to the wild type (WT) (B).