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TRACK 3 TRACK 3 Technical Program

chanical cues from surrounding cell types as well as physiologically relevant erogeneous cellular organization. Moreover, the tumor microenvironment
flow and extracellular matrix stiffness. In vitro models aid in the early devel- is composed of confounding factors (e.g. spatial organization of cells, 3D
opment and characterization of CNS drug candidates and allow for highly biomolecular gradients, etc.) that hinder the understanding of how and why
controlled examination of BBB function and dysfunction at the cellular level cancer cells invade throughout the surrounding tissues. These complex
as it pertains to the pathogenesis of neurobiological diseases. However, cur- characteristics must be recapitulated within physiologically relevant in vitro
rent in vitro BBB models available for preliminary drug studies have neglect- models to fully appreciate the dynamics of cancer invasion. Microengineer-
ed the inclusion of several critical parameters including (i) controlled shear ing technology provides a powerful approach to develop flexible platforms
flow that directs endothelial function in the lumen and (ii) appropriate culture with the ability to precisely control biochemical and biophysical cues with
of multiple cellular constituents of the BBB. Several microfluidic models of well-defined 3D cellular organization to mimic the native tumor microenvi-
the BBB exist in the literature, and some incorporate multiple relevant cell ronment. Current models are able to replicate the tumor architecture, how-
types. However, all existing co-culture models necessitate the incorporation ever, they were either limited by creating a 3D stroma but with a two-dimen-
of a semipermeable membrane to achieve relevant cellular geometries. sional (2D) tumor region or by incorporating a 3D stroma and tumor but with-
Furthermore, existing co-culture models pay little attention to effect of ex- out any capacity for vasculature, cell invasion, and modulation of microen-
tracellular matrix composition and stiffness on the supplementary cell types. vironmental cues (e.g. biophysical and biochemical signaling). Furthermore,
Additionally, traditional characterization of the BBB structure and function in studies of 3D cellular chemoinvasion (i.e. invasion due to a chemoattractant)
the existing in vitro models has been largely superficial. Currently, most in have been limited and most analyses have been performed in 2D.
vitro models are evaluated via a combination of transendothelial electrical Our research focuses on the development of a physiologically relevant 3D
resistance (TEER) measurements, histology for relevant proteins (ZO-1, PgP), breast tumor-stroma model on a chip using microfluidics technology and ad-
and conventional permeability assays. While these benchmarks provide vanced biomaterials (hydrogels). This platform enables a spatially organized
useful information, they are far from comprehensive enough to be used to tumor-stroma interaction model for characterization of 3D breast cancer
develop truly predictive models. We have addressed these two challenges chemoinvasion within a native like microenvironment. The device, fabricated
(the simultaneous incorporation of shear flow and direct cell contact, and from polydimethylsiloxane (PDMS), contains specific geometric regions that
the comprehensive characterization of genetic and protein specializations enable spatial compartmentalization of the tumor and stroma. Highly inva-
characteristic of brain microvasculature) with our study. Our novel in vitro sive breast cancer cells (SUM-159) were encapsulated within an ECM gel
model of the BBB is composed of microfluidic channels crafted inside a cell (Matrigel®:collagen I) and injected into a one region of the device to produce
laden hydrogel system through the use of sacrificial gelatin elements. The the “primary tumor”. This region was then surrounded by an outer collagen
hydrogel system is a tailored blend of extracellular matrix proteins previ- ECM gel to create the stromal component. This combination produced a 3D
ously demonstrated to facilitate the healthy culture of astrocytes. In addition tumor-stroma-on-a-chip model. Our studies revealed that the device was
to the development of our novel model, we have adapted a multilayered capable of high-resolution time-lapse and immunofluorescent imaging of
microfluidic endothelial culture system previously used in our lab to serve as cancer cell invasion and morphology within a 3D matrix. We modulated bio-
a dynamic transwell culture of the BBB with shear flow controlled and three chemical cues to stimulate cancer chemoinvasion throughout the tumor-stro-
cell types cultured. Using this system, we are able to measure TEER estab- ma regions. Cancer cells experienced 3D epidermal growth factor (EGF)
lishment and disruption in the presence of relevant cues. This system also gradients emanating from the stromal region, which strongly influenced their
allows us a means to compare our novel lumen device to dynamic transwell phenotype and invasive profile. Particularly, the EGF-stimulated cells demon-
microfluidic models of the BBB already present in the literature. To evaluate strated enhanced proliferation, invasive protrusions, and migratory speed.
the degree to which our novel model recapitulates relevant in vivo BBB fea- Using this platform, additional studies are planned to elucidate biomolecular
tures, we have developed primers pairs for a list of 96 PCR targets (93 BBB interactions (e.g growth factors or anti-cancer drugs) in the context of the
specific genes and 3 housekeeping genes) for use in high throughput qPCR tumor-stroma complex. Moreover, this platform will be used to investigate
analysis (Biomark Fluidigm System) that will be used in addition to traditional intracellular pathways that are known to be involved in cells migration
characterization techniques, such as TEER and histology. Thus far, we have and chemoinvasion, such as the PI3K pathway. By spatially organizing the
compared the expression levels of these targets in our model relative to tumor-stroma to better mimic the human breast cancer architecture, we
several commonly used platforms for in vitro BBB studies: static transwell envision that this work will build a foundation for a breast tumor-on-a-chip.
cultures, spheroids, and our multilayer microfluidic dynamic transwell sys- We plan to later include further complexities such as heterotypic interactions
tem. Preliminary results indicate that the type of culture system drastically and a vasculature network to work our way toward a platform that would be
influences the degree to which the endothelial cells are specialized. Current suitable for effective drug screening and personalized medicine.
work is aimed at further elucidating how individual culture conditions (pres-
ence/lack of flow, alterations in ECM stiffness, presence of cellular constitu-
ents) contribute to the expression of relevant specializations.
WEDNESDAY, FEBRUARY, 24
5:20pm Three-dimensional (3D) Invasion of Breast Cancer Cells
in a Well-Defined Tumor-Stroma Platform
3-9
Technical Presentation. NEMB2016-5967 TOWARD POINT-OF-CARE DIAGNOSTICS

Danh Truong, Alison Llave, Arizona State University, Tempe, AZ, Navarro 9:30am - 11:00am
United States, Julieann Puleo, Ghassan Mouneimne, The Univer-
sity of Arizona Cancer Center, Tucson, AZ, United States, Roger Session Organizer: Savas Tasoglu, UConn, Storrs, CT, United States
Kamm, Massachusetts Institute of Technology, Cambridge, MA,
United States, Mehdi Nikkhah, Arizona State University, Tempe, AZ, 9:30am Challenges and Opportunities for Ultrasensitive Im-
United States munoassays: Large Volume Capillaric Microfluidics and Digital
Nanodot Arrays
Breast cancer is one of the leading causes of mortalities in the United States
and will ultimately result in approximately 40,000 deaths by the end of 2015.
The majority of those deaths are due to invasion and dissemination of can- Keynote. NEMB2016-6160
cer from the primary tumor. While current regimens are able to treat local-
ized diseases, there are still few effective treatments for metastatic disease. David Juncker, McGill University, Montreal, QC, Canada
Cancer invasion is a fundamental aspect of metastasis that initiates from
the complex heterogeneous tumor microenvironment. The tumor stroma is The difficulty of making reliable ultrasensitive assays and the limitations of 45
a three-dimensional (3D) dense tissue with a distinct architecture and het- microfluidics affinity binder and complications of multiplexing will be intro-