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Metagenomic deep sequencing of aqueous fluid detects intraocular lymphomas.

ABSTRACT: INTRODUCTIONCurrently, the detection of pathogens or mutations associated with intraocular lymphomas heavily relies on prespecified, directed PCRs. With metagenomic deep sequencing (MDS), an unbiased high-throughput sequencing approach, all pathogens as well as all mutations present in the host's genome can be detected in the same small amount of ocular fluid.METHODSIn this cross-sectional case series, aqueous fluid samples from two patients were submitted to MDS to identify pathogens as well as common and rare cancer mutations.RESULTSMDS of aqueous fluid from the first patient with vitreal lymphoma revealed the presence of both Epstein-Barr virus (HHV-4/EBV) and human herpes virus 8 (HHV-8) RNA. Aqueous fluid from the second patient with intraocular B-cell lymphoma demonstrated a less common mutation in the MYD88 gene associated with B-cell lymphoma.CONCLUSIONMDS detects pathogens that, in some instances, may drive the development of intraocular lymphomas. Moreover, MDS is able to identify both common and rare mutations associated with lymphomas.

SUBMITTER: Gonzales J

PROVIDER: S-EPMC5754869 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Metagenomic deep sequencing of aqueous fluid detects intraocular lymphomas.

Gonzales John J Doan Thuy T Shantha Jessica G JG Bloomer Michele M Wilson Michael R MR DeRisi Joseph L JL Acharya Nisha N

The British journal of ophthalmology 20171109 1

<h4>Introduction</h4>Currently, the detection of pathogens or mutations associated with intraocular lymphomas heavily relies on prespecified, directed PCRs. With metagenomic deep sequencing (MDS), an unbiased high-throughput sequencing approach, all pathogens as well as all mutations present in the host's genome can be detected in the same small amount of ocular fluid.<h4>Methods</h4>In this cross-sectional case series, aqueous fluid samples from two patients were submitted to MDS to identify pa ...[more]

PMID: 29122821

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Project description:BackgroundVitreoretinal lymphoma (VRL), the most common lymphoma of the eye, is a rare form of primary CNS lymphoma (PCNSL). Most frequently a high-grade diffuse large B cell lymphoma, VRL can cause vision loss and its prognosis remains dismal: the overall survival time is 3 years after diagnosis. Radiotherapy and chemotherapy are used but remain frequently ineffective, and no standardized treatment regimen exists. Furthermore, no biologically targeted treatments, based on the genetic profile of the tumor, are available, as VRL has hitherto not comprehensively been profiled. To address these unmet needs, we hypothesized that a next generation sequencing (NGS)-based, National Cancer Institute (NCI) MATCH Trial-modified panel would be able to identify actionable genomic alterations from small-volume, intraocular liquid biopsies.Methods and findingsIn this retrospective study, we collected diluted vitreous biopsies from 4 patients with a high suspicion for VRL. Following cytological confirmation of lymphoma (all were diffuse large B cell lymphomas), we subjected genomic DNA from the biopsies to NGS, using a panel containing 126 genes (3,435 amplicons across several hotspots per gene), which was modified from that of the NCI MATCH Trial, a new trial that has matched patients with cancers that have not responded (or never responded), to investigational therapeutics based on their prioritized mutation profile rather than site of tumor origin. Using a validated bioinformatics pipeline, we assessed for the presence of actionable mutations and copy number alterations. In all four small-volume, intraocular liquid biopsies, we obtained sufficient genomic DNA for analysis, even in diluted samples in which the undiluted vitreous was used for cytology and flow cytometry. Using NGS, we found targetable heterozygous gain-of-function mutations in the MYD88 oncogene, and confirmed in our cohort the presence the L265 mutations, previously described using PCR-based assays. For the first time in VRL, we also identified the MYD88 S243N mutation. We also identified two-copy copy number losses in the tumor suppressor CDKN2A in all four cases, and one copy loss of the tumor suppressor PTEN in one sample. In one case, in which vitreous biopsies were originally read as cytologically negative, but which was confirmed as lymphoma when a lesion appeared in the brain two years later, our NGS-based approach detected tumoral DNA in the banked, original liquid biopsy.ConclusionsWe performed the first systematic exploration of the actionable cancer genome in VRL. Our NGS-based approach identified exploitable genomic alterations such as gain-of-function MYD88 oncogene mutations and loss of the tumor suppressor CDKN2A, and thus illuminates new routes to biologically targeted therapies for VRL, a cancer with a dismal prognosis. This precision medicine strategy could be used to nominate novel, targeted therapies in lymphomas and other blinding and deadly ocular, orbital, and ocular adnexal diseases for which few treatments exist.

Dataset Information

Metagenomic deep sequencing of aqueous fluid detects intraocular lymphomas.

Publications

Metagenomic deep sequencing of aqueous fluid detects intraocular lymphomas.

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