A Novel NGS Assay to Detect Any KMT2A Fusion Transcript at Low Levels

<p>Measurable residual disease (MRD) detection is associated with cancer relapse. Leukemias with rearrangement of the lysine methyltransferase 2 A gene (<em>KMT2Ar</em>) have an adverse prognosis with more than 80 possible fusion partner genes (FPG) identified. With the advent of effective targeted therapies for <em>KMT2Ar</em> leukemia such as menin inhibitors, there is an unmet need for sensitive assays to detect these various fusions. Standard next-generation sequencing (NGS) methods cannot accurately detect genetic alterations at low frequencies because of inherent technical errors and the need for high sequencing depth. Blocker Displacement Amplification (BDA) technology enables selective detection of such rare alterations with NGS. Here, we demonstrate the effective detection of <em>KMT2Ar </em>and its various FPG with a novel NGS assay leveraging BDA. Primers were designed to flank any known exon junction of cDNA corresponding to transcripts of the gene of interest, while non-extensible oligonucleotides (blockers) span the wildtype (WT) exon junction, therefore suppressing amplification of WT while differentially amplifying fusion junctions. We tested this assay in cell line, establishing limits of detection and input and subsequently validated our findings in clinical samples from patients with <em>KMT2Ar </em>leukemia. In conclusion, FusionBDA is a novel, sensitive, and specific assay for <em>KMT2Ar</em> acute leukemia. This assay can agnostically detect <em>KMT2Ar </em>with various fusions at a predicted sensitivity of at least 0.005%, therefore allowing both MRD detection and target identification.</p>

Name thesis:degree_name

Masters of Science (MS)

Level thesis:degree_level

Thesis (MS)

Year dc:date.available

2023

Authors dc:creator

• Issa, Ghayas
• <p>0000-0002-4339-8683</p>

Contributors dc:contributor

• Andy Futreal, PhD
• Courtney DiNardo, MD
• Marina Konopleva, MD, PhD

• Menin
• KMT2A
• MRD
• Leukemia
• Medical Molecular Biology
• Translational Medical Research

Repository record dc:identifier

https://digitalcommons.library.tmc.edu/utgsbs_dissertations/1320

OAI identifier oai:identifier

oai:digitalcommons.library.tmc.edu:utgsbs_dissertations-2377