# Single-cell mitochondrial mutation lineage tracing of non-dysplastic and dysplastic Barrett's esophagus

> **NIH NIH R01** · UNIVERSITY OF PENNSYLVANIA · 2024 · $467,659

## Abstract

Project Summary
The incidence of esophageal adenocarcinoma and its premalignant metaplastic precursor Barrett's esophagus
(BE) have both displayed marked increases in incidence in the US in recent decades. The progression to
esophageal adenocarcinoma occurs through a transformation in which non-dysplastic BE becomes dysplastic
BE and acquires the earliest histological features of cancer. Currently, we do not know the molecular drivers of
this transition and how it occurs in human patients. The primary objective of this proposal is to identify the earliest
events underlying the transition to dysplasia as these will yield new therapeutic and diagnostic targets. Our
proposal leverages technological advances in single-cell biology using mitochondrial mutation lineage tracing
combined with RNA-sequencing to track clonality and RNA profiles in single cells. Our preliminary data
demonstrates the technological approach and reveals that non-dysplastic BE emerges through multiple clones
that can generate all BE cell types. However, our preliminary data from dysplastic BE revealed the opposite
result. Dysplasia emerged from non-dysplasia BE through a single clone with a distinguishing transcriptional
profile. Using our single-cell and clonal tracking data, we identified the Wnt pathway, including Wnt genes LGR5,
NOTUM, and DKK1, as differentially expressed in dysplasia. Further supporting the Wnt pathway, we also
uncovered an APC mutation occurring in the same sample. Thus, our central hypothesis is that the
transformation from non-dysplastic to dysplastic Barrett's esophagus emerges from a single clone with genetic
and transcriptional changes that can be tracked to identify new targets for dysplasia. To test this hypothesis, we
will implement a comprehensive and rigorous experimental approach that couples single-cell RNA-Seq,
mitochondrial mutation-based lineage tracing, and 3D organoids for functional investigation into the mechanisms
of dysplasia. The proposed studies will leverage these techniques to track the clonal evolution and RNA profiles
of single cells in non-dysplasia and dysplastic BE (Aim 1) and to provide mechanistic exploration into the role of
the Wnt pathway in the emergence of dysplasia (Aim 2). This study leverages the research team's diverse and
complementary expertise in cancer single cell genomics and bioinformatics, clinical and basic approaches to BE,
and in vitro epithelial model systems to develop a robust platform for tracking cell fate in human tissues that will
provide unprecedented molecular insight into the development of dysplasia in BE. Our expected outcomes are
a molecular characterization of the earliest changes underlying the transformation to dysplasia in BE. Our work
will have broad positive impact as it will reveal both therapeutic targets as well as diagnostic markers for patients
with BE who will progress to esophageal adenocarcinoma. This work ultimately aims to dramatically decrease
the number of patients who are ...

## Key facts

- **NIH application ID:** 10823733
- **Project number:** 1R01DK135729-01A1
- **Recipient organization:** UNIVERSITY OF PENNSYLVANIA
- **Principal Investigator:** Amanda Brooke Muir
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $467,659
- **Award type:** 1
- **Project period:** 2024-08-01 → 2028-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10823733

## Citation

> US National Institutes of Health, RePORTER application 10823733, Single-cell mitochondrial mutation lineage tracing of non-dysplastic and dysplastic Barrett's esophagus (1R01DK135729-01A1). Retrieved via AI Analytics 2026-06-12 from https://api.ai-analytics.org/grant/nih/10823733. Licensed CC0.

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