Investigation of cancer specific DNA changes using oxford nanopore technology (ONT) and more traditional sequencing technologies in the surveillance of patients with neuroendocrine neoplasms

Masato Ahsan; Shailesh Gohil; Narendra Reddy; Rebecca Allsopp; Karen Page; Jacqui Shaw; Miles Levy

doi:10.1530/endoabs.109.P185

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Endocrine Abstracts (2025) 109 P185 | DOI: 10.1530/endoabs.109.P185

SFEBES2025 Poster Presentations Neuroendocrinology and Pituitary (48 abstracts)

Investigation of cancer specific DNA changes using oxford nanopore technology (ONT) and more traditional sequencing technologies in the surveillance of patients with neuroendocrine neoplasms

Masato Ahsan ^1,2 , Shailesh Gohil ^1,2 , Narendra Reddy ^1,2 , Rebecca Allsopp ² , Karen Page ² , Jacqui Shaw ² & Miles Levy ^1,2

Author affiliations

¹The University Hospitals of Leicester NHS trust, Leicester, United Kingdom; ²University of Leicester, Leicester, United Kingdom

Background: Circulating tumour DNA (ctDNA) found in the plasma or serum of individuals with cancer presents a non-invasive means of accessing tumour DNA. This ’liquid biopsy’ facilitates the examination of various DNA aspects including fragmentation, chromosomal alterations, sequence mutations, and epigenetic modifications. Aberrant methylation of specific promoter regions tends to be a consistent hallmark of cancer. This consistency makes ctDNA methylation amenable to design a clinical assay for neuroendocrine neoplasm.

Subject and Design of study: We plan investigate samples from patients with neuroendocrine neoplasm (NEN) with germline mutation and sporadic mutations and NENs of unknown primary origin. This will be a longitudinal, observational, cohort, pilot study.

ctDNA methylation via Oxford Nanopore sequencer: Nanopore sequencing is the only sequencing technology that offers real-time analysis for rapid insights in fully scalable formats, and can analyse native DNA or RNA, and sequence any length of fragment to achieve short to ultra-long read lengths. We aim to isolate cell-free DNA from 4 ml of blood plasma using the MagMAX Cell-free DNA Isolation Kit on the Kingfisher Flex instrument. For Library preparation we will follow an established Oxford Nanopore Technique protocol using Ligation Sequencing Kit V14 (SQK-LSK114) chemistry, using 10 ng cfDNA input to load on PrometION Flow Cell. In individual patients, change in ctDNA methylation analysis over time will be plotted and compared against change in disease burden on imaging and changes in current biomarker performed as per usual care, using RECIST criteria and, depending on data generated, sensitivity analysis will be completed.

Conclusion: There is limited information on the clinical utility of DNA methylation analysis in the detection and surveillance of NETs. We hope to use third generation nanopore sequencing to investigate circulating DNA methylation to establish if this correlates to disease activity in patients with NENs and will publish our results next year.