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Appendix 3: Molecular Pathways, Cellular Targets and Therapies Being Studied in the MBC Grants
Dataset

A grant with a focus on invasion

Title:
The ezrin signaling network as a potential novel marker for breast cancer metastasis

Ezrin, a plasma membrane cytoskeleton linker, is required for cell survival and morphogenesis. It has been found that over-expression of
ezrin frequently occurs in invasive human breast cancer and is required for cell motility and invasion of carcinoma cells. Studies indicate
that ezrin acts co-operatively with Src in the disruption of cell-cell contacts and increased cell scattering and motility – characteristic of a
transformed phenotype. Over-activation of Src and ezrin also causes increased activation of the receptor tyrosine kinase Met, a proto-
oncogene that is frequently overexpressed in patients with high risk of metastatic disease. This transdisciplinary project will focus on
determining the role of Src/ezrin/Met activation (referred to as the Src/ezrin signalling network) at specific stages of human breast cancer
metastasis, and correlating the Src ezrin signalling network with tumour stage and grade as a possible predictor and/or treatment target
for human breast cancer metastasis.

Metastasis stage (Steeg) Invasion and motility
Hanahan/Weinberg Activating invasion & metastasis
Understanding (basic)
Research stage
Translational
Pathway Ezrin, Src, Met
Therapy/intervention none

A grant with a focus on intravasation

Title:
Microfluidic 3D Scaffold Assay for Cancer Cell Migration and Intravasation

DESCRIPTION (provided by applicant): Migration through extra-cellular matrix (ECM) and intravasation across a cellular barrier comprise
the initial, rate-limiting steps of cancer metastasis. Physiologically relevant and well-controlled models that mimic the in vivo tumor
microenvironment will enable better understanding of the initial steps of metastasis and evaluation of potential therapy efficacy. In
vivo models have physiological relevancy, yet inherently lack a high level of control. In vitro cancer migration models have high levels
of control, yet lack critical components of the tumor microenvironment. We propose a new technology, a microfluidic migration and
intravasation assay (MIA). The MIA replicates essential components of the in vivo tumor microenvironment, including a 3D ECM and a
vasculature, while providing tight control of biochemical and biophysical parameters. To further establish the MIA, we propose to use it
to investigate a specific biophysical factor - interstitial flow - which has not previously been studied in the context of metastatic disease.
The objective of the proposed work is to evaluate the metastatic potential of cancerous cells by developing the MIA and identifying
novel extent of invasion metrics (Specific Aim 1), and applying them to study the influence of interstitial flow on cancer cell metastasis
(Specific Aim 2). The MIA will have an input channel for the cancer cells, a 3D collagen gel to simulate native ECM, and an endothelial cell
(EC) layer adherent to the gel in a second channel. The configuration will permit migration of cancer cells either from the input channel
or within the gel towards the second channel. Optimized gel parameters will present appropriate chemotactic gradients and physical
parameters simulating a tumor microenvironment and inducing cancer cell migration. The EC layer will mimic the in vivo vascular barrier
allowing observation of cancer cell intravasation. Optical access from two vantage points will permit real time observation of cancer cell
migration and intravasation. The optical access combined with image processing techniques will quantify cancer cell morphological and
migratory parameters, leading to identification of novel extent of invasion metrics that will quantify the metastatic potential of cancer
cells. Finally, we will leverage the microfluidic capability of the MIA to induce interstitial flow across the gel, and quantify the effects of this
biophysical parameter on cancer cell invasion. Taken together, the two aims establish the MIA as an excellent platform for quantitative
research of molecular mechanisms governing cancer cell invasion. For example, therapies capitalizing on altered vascular morphology
near tumors would clearly benefit from using the MIA as a development platform, as the system provides a characterized EC layer in
conjunction with a well-controlled system. Future development will enable the MIA to serve as a cancer cell diagnostic device and a high
throughput drug development tool. Cancer spreads and invades through a process called metastasis, often resulting in patient death.
The metastasis process is not well understood, since there is a shortage of well-controlled models that realistically represent the tumor
microenvironment and its blood supply. This application seeks to develop a well-controlled and realistic tumor environment model to aid
cancer metastasis research and eventually provide a platform to more efficiently develop and evaluate cancer therapies.