Chapter 19
CAFs and Cancer Cells Co-Migration in 3D Spheroid Invasion
Assay
Sefora Conti, Takuya Kato, Danielle Park, Erik Sahai, Xavier Trepat, and
Anna Labernadie
Abstract
In many solid tumors, collective cell invasion prevails over single-cell dissemination strategies. Collective
modes of invasion often display specific front/rear cellular organization, where invasive leader cells arise
from cancer cell populations or the tumor stroma. Collective invasion involves coordinated cellular movements which require tight mechanical crosstalk through specific combinations of cell–cell interactions and
cell–matrix adhesions. Cancer Associated Fibroblasts (CAFs) have been recently reported to drive the
dissemination of epithelial cancer cells through ECM remodeling and direct intercellular contact. However,
the cooperation between tumor and stromal cells remains poorly understood. Here we present a simple
spheroid invasion assay to assess the role of CAFs in the collective migration of epithelial tumor cells. This
method enables the characterization of 3D spheroid invasion patterns through live cell fluorescent labeling
combined with spinning disc microscopy. When embedded in extracellular matrix, the invasive strands of
spheroids can be tracked and leader/follower organization of CAFs and cancer cells can be quantified.
Key words 3D spheroid invasion, Cancer Associated Fibroblasts, Epithelial cancer cells, Collective
migration, Leader/follower cells
1 Introduction
Metastatic dissemination is responsible for most cancer-related
deaths. In the early stages of cancer spreading, tumor cells escape
from the primary site, breach through the basement membrane
(BM), infiltrate into the stroma and eventually enter the blood
circulation [1]. Increasing evidence suggests that a large number
of solid tumors invade as multicellular units within the peritumoral
stroma and in the bloodstream [2–4]. These modes of collective
migration generally involve an invasive front presenting specific
combinations of cell–cell interactions and cell–matrix adhesion
Kyra Campbell and Eric Theveneau (eds.), The Epithelial-to Mesenchymal Transition: Methods and Protocols,
Methods in Molecular Biology, vol. 2179, https://doi.org/10.1007/978-1-0716-0779-4_19,
© Springer Science+Business Media, LLC, part of Springer Nature 2021
Sefora Conti and Takuya Kato are co-authors.
243
CAFs and Cancer Cells Co-Migration in 3D Spheroid Invasion
Assay
Sefora Conti, Takuya Kato, Danielle Park, Erik Sahai, Xavier Trepat, and
Anna Labernadie
Abstract
In many solid tumors, collective cell invasion prevails over single-cell dissemination strategies. Collective
modes of invasion often display specific front/rear cellular organization, where invasive leader cells arise
from cancer cell populations or the tumor stroma. Collective invasion involves coordinated cellular movements which require tight mechanical crosstalk through specific combinations of cell–cell interactions and
cell–matrix adhesions. Cancer Associated Fibroblasts (CAFs) have been recently reported to drive the
dissemination of epithelial cancer cells through ECM remodeling and direct intercellular contact. However,
the cooperation between tumor and stromal cells remains poorly understood. Here we present a simple
spheroid invasion assay to assess the role of CAFs in the collective migration of epithelial tumor cells. This
method enables the characterization of 3D spheroid invasion patterns through live cell fluorescent labeling
combined with spinning disc microscopy. When embedded in extracellular matrix, the invasive strands of
spheroids can be tracked and leader/follower organization of CAFs and cancer cells can be quantified.
Key words 3D spheroid invasion, Cancer Associated Fibroblasts, Epithelial cancer cells, Collective
migration, Leader/follower cells
1 Introduction
Metastatic dissemination is responsible for most cancer-related
deaths. In the early stages of cancer spreading, tumor cells escape
from the primary site, breach through the basement membrane
(BM), infiltrate into the stroma and eventually enter the blood
circulation [1]. Increasing evidence suggests that a large number
of solid tumors invade as multicellular units within the peritumoral
stroma and in the bloodstream [2–4]. These modes of collective
migration generally involve an invasive front presenting specific
combinations of cell–cell interactions and cell–matrix adhesion
Kyra Campbell and Eric Theveneau (eds.), The Epithelial-to Mesenchymal Transition: Methods and Protocols,
Methods in Molecular Biology, vol. 2179, https://doi.org/10.1007/978-1-0716-0779-4_19,
© Springer Science+Business Media, LLC, part of Springer Nature 2021
Sefora Conti and Takuya Kato are co-authors.
243