# Defining leader cell biology in collective invasion

> **NIH NIH F31** · EMORY UNIVERSITY · 2020 · $8,430

## Abstract

Project Summary
Metastasis is responsible for 90% of cancer deaths, yet there is a severe lack of effective anti-metastatic targeted
therapeutics. The major mode of metastasis for many cancers is collective invasion, where heterogeneous packs
of cells travel together. Our lab has previously shown that these collective packs include specialized “leader” and
“follower” cells, and we created the first purified leader and follower cell cultures utilizing the SaGA technique
(Spatiotemporal Genomic and Cellular Analysis), which leverages a combination of live-cell confocal microscopy
and fluorescence-activated cell sorting. Using these novel cell lines, we showed that cooperativity between
leaders and followers is crucial for successful collective invasion, with leaders promoting the invasion of
followers, and followers promoting the survival and proliferation of leaders. Importantly, we now report the first
known panel of leader- and follower-specific gene mutations, identified via RNA-seq analysis. Notably, leader
cells exclusively harbor a specific missense mutation in the ARP3 protein, a crucial component of the Arp2/3
complex that drives cell motility by promoting actin polymerization. Furthermore, leader cells display increased
lamellipodia and more highly organized actin filaments compared to follower cells, suggesting increased Arp2/3
activity in leaders. Understanding how this ARP3 mutation drives the emergence and invasive biology of leader
cells could be a crucial step toward identifying therapeutic targets to inhibit metastasis in cancer patients. We
will therefore test the overall hypothesis that ARP3 mutations lead to increased Arp2/3 activity in rare cancer
cells, thereby driving the unique leader cell phenotype and promoting collective invasion. To probe this
hypothesis, we will determine 1) the molecular mechanism by which this ARP3 mutation drives the emergence
and invasive biology of leader cells in vitro, and 2) if this ARP3 mutation drives lung cancer collective invasion
and metastasis in vivo. In Aim 1 we will create stable cell lines expressing mutant ARP3, and subsequently
perform 3-D spheroid invasion assays and co-immunoprecipitation experiments to determine how this mutation
affects protein ubiquitination, Arp2/3 activation, and overall emergence of the leader cell phenotype. In Aim 2,
we will use a mouse orthotopic lung cancer xenograft model and subsequent pathological analysis to determine
the effects of this ARP3 point mutation on collective invasion and metastasis in vivo. These studies will provide
critical insight into the biological events that drive collective invasion, which could ultimately set a new paradigm
for the treatment and prevention of metastasis in patients.

## Key facts

- **NIH application ID:** 9857006
- **Project number:** 5F31CA225049-03
- **Recipient organization:** EMORY UNIVERSITY
- **Principal Investigator:** Brian Aram Pedro
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $8,430
- **Award type:** 5
- **Project period:** 2018-03-01 → 2020-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9857006

## Citation

> US National Institutes of Health, RePORTER application 9857006, Defining leader cell biology in collective invasion (5F31CA225049-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9857006. Licensed CC0.

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