Biomarkers to biology: dissecting the collaborative pathways that drive collective invasion and metastasis Restricted; Files & ToC

Abstract

Metastasis is the leading cause of cancer-related mortality, and cellular heterogeneity within the bulk tumor propels tumor progression and metastasis. Invasive tumor cells utilize collective invasion – whereby cohesive groups of cells invade into the adjacent stroma or circulation while maintaining cell-cell contacts – to escape the primary tumor, navigate the vasculature, and pilot to secondary sites. These packs consist of distinct cellular subpopulations – leaders found at the forefront and followers which trail behind – that exhibit heterogeneous genetic, epigenetic, and phenotypic characteristics that drive invasive behavior. The mechanisms that enable subpopulation communication to facilitate emergence and persistence as a cohesive unit remain largely unclear. Additionally, whether cooperation among these subpopulations is essential for metastatic seeding, adaptation, and colonization has yet to be elucidated. To this end, we investigate real-time intercellular communication within actively invading non-small cell lung cancer (NSCLC) packs by integrating Spatiotemporal Genomic and Cellular Analysis (SaGA) with single-cell RNA sequencing to reveal that NSCLC packs are composed of heterogeneously cycling subpopulations utilizing distinct yet cooperative pathways to enhance invasion dynamics. We demonstrate that the follower subpopulation produces and secretes TGFΒ1 to stimulate invasion and pack cohesion via both autocrine and paracrine signaling. We determine that TGFΒ1 downstream signaling varies across subpopulations and governs differential functional responses, including proliferation and JAG1-dependent invasion. To elucidate how these subpopulations influence metastatic colonization, we utilize the follower-specific surface protein Interleukin 13 receptor alpha 2 (IL13R⍺2) as a biomarker to label leader-like and follower-like subpopulations across NSCLC lines and patient-derived populations. We demonstrate a diversity across these NSCLC cell samples which includes IL13R⍺2 heterogeneous (both positive and negative subpopulations) and homogeneous (100% negative) populations and single-cell sequencing indicates IL13R⍺2 heterogeneous cells upregulate fatty acid response mechanisms. This finding complements our CRISPR IL13R⍺2 knockout experiments where the rate-limiting enzyme critical for lipid biosynthesis is depleted upon loss of IL13R⍺2. In vivo, we establish increased metastasis within IL13R⍺2 heterogeneous populations, and single-cell sequencing suggests this may be due to an enhanced resistance to death by ferroptosis. Together, we elucidate a novel intercellular communication network active during collective invasion and enhanced metastases enabled by IL13R⍺2-specific subpopulation heterogeneity.

Table of Contents

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About this Dissertation

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Biochemistry, Cell & Developmental Biology
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Biology, Cell Biology, Molecular
Palavra-chave	Cancer, Heterogeneity, Metastasis
Committee Chair / Thesis Advisor	Adam Marcus, Emory University
Committee Members	Cheng-Kui Qu, Emory University Michael Koval, Emory University Ken Moberg, Emory University Dorothy Lerit, Emory University

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