Genetic mutations in archival ocular biopsies from histiocytic orbital disease lesions

Abstract

Purpose : The classification of Langerhans cell histiocytosis (LCH) as a neoplastic disorder is based on lesional mutations in genes of the MAPK activation pathway, with BRAF and MAP2K1 mutations found in a majority of cases. LCH is one of several clinically variable histiocytic disorders characterized by lesional cell phenotypes, usually involving bone, skin, or lung, but may arise in any organ/tissue, including orbit. The purpose of this study was to test orbital histiocytic lesions for pathogenic mutations known in LCH.

Methods : Formalin-fixed paraffin-embedded (FFPE) biopsies were sectioned and lesion-rich areas dissected for genomic DNA extraction. Mutational analysis was performed by massively parallel sequencing utilizing multiplexed PCR combined with sequencing on an Illumina platform. A panel of target exons and flanking intronic sequences of 124 cancer-related genes, including BRAF and MAP2K1, was used.

Results : Fifteen biopsies from 13 subjects (7 female; 6 male) were included. Orbital disease diagnoses included juvenile xanthogranuloma (n=6), xanthogranuloma (n=4) and granulomatous inflammation consistent with, or suggestive of, Erdheim-Chester disease (n=3). Biopsy tissues included eyelid (n=7), orbit (n=6), lacrimal sac (n=1) and conjunctiva (n=1). FFPE specimens were stored from <1-18 years. Subject age at biopsy ranged from <1-78 years. Race/ethnicity of subjects included Caucasian/non-Hispanic (n=9), Hispanic (n=3) and Native American (n=1). Sequencing was successful on 13/15 samples. No pathogenic mutations were detected. No variants were found in 3 samples and 10 samples exhibited 1-3 variants known to be either benign or of unknown clinical consequence. One sample had no observed lesional tissue and the remaining samples exhibited 10-75% lesion. DNA Deamination levels ranged from background to 20% and was higher in the older archived samples.

Conclusions : In this series of orbital histiocytic disease no known pathogenic mutations were found. The archived samples were amenable to DNA extraction and sequencing but relatively high levels of DNA deamination in long-term stored samples could limit mutation detection. The possibility remains that neoplastic mutations contribute to histiocytic orbital lesion development, but larger studies are needed to determine the extent this may occur. It is important to identify individuals with pathogenic mutations who could benefit from targeted therapies.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.