A cell–ECM screening method to predict breast cancer metastasis

Abstract

Breast cancer preferentially spreads to the bone, brain, liver, and lung. The clinical patterns of this tissue-specific spread (tropism) cannot be explained by blood flow alone, yet our understanding of what mediates tropism to these physically and chemically diverse tissues is limited. While the microenvironment has been recognized as a critical factor in governing metastatic colonization, the role of the extracellular matrix (ECM) in mediating tropism has not been thoroughly explored. We created a simple biomaterial platform with systematic control over the ECM protein density and composition to determine if integrin binding governs how metastatic cells differentiate between secondary tissue sites. Instead of examining individual behaviors, we compiled large patterns of phenotypes associated with adhesion to and migration on these controlled ECMs. In combining this novel analysis with a simple biomaterial platform, we created an in vitro fingerprint that is predictive of in vivo metastasis. This rapid biomaterial screen also provided information on how β1, α2, and α6 integrins might mediate metastasis in patients, providing insights beyond a purely genetic analysis. We propose that this approach of screening many cell–ECM interactions, across many different heterogeneous cell lines, is predictive of in vivo behavior, and is much simpler, faster, and more economical than complex 3D environments or mouse models. We also propose that when specifically applied toward the question of tissue tropism in breast cancer, it can be used to provide insight into certain integrin subunits as therapeutic targets. Insight, innovation, integration We developed a high-throughput method to rapidly screen cell adhesion, motility, and growth factor responses on biomaterial surfaces. This approach is analogous to systems biology, relying on cell phenotypes in lieu of genetics. We used this technique to reveal patterns of phenotypes associated with breast cancer metastasis to possible tissue sites (bone, brain, lung). By comparing the phenotypic patterns between cell lines that metastasize to only one tissue site with heterogeneous cell lines, we provide the first method to connect in vitro phenotype to in vivo fate. This method is successful without genetic analysis, yet it also predicts outcomes related to integrin gene expression, potentially identifying new targets for tissue-specific metastasis.

Complementary, Semiautomated Methods for Creating Multidimensional PEG-Based Biomaterials

“…19 This chemistry was initially developed for single coverslips, and this method was incompatible with semiautomated applications. To optimize this chemistry for multi-well plates, we applied this same silane chemistry on a well plate-sized piece of glass (Figure 1a).…”

“…To create a 96-well plate system with precisely tailored integrin-binding ECM proteins, we modified commercially available glass with aminated silane (Figure 1a). 19 We performed XPS to ensure that the silane chemistry resulted in the appropriate functional groups after each reaction step (Figure 2a). …”

“…ECM proteins were attached to the glass surface in a 96-well plate format by modifying a procedure described previously. 19 For each plate, one 110 mm x 75 mm piece of glass was UV/ozone treated at 1 atmosphere (UV/Ozone ProCleaner, 180 micrograms/m 3 ozone level in chamber, 4.6 mW/cm 2 peak UV intensity, 10.15 W UV lamp power requirement, BioForce Nanosciences, Salt Lake City, UT) for 10 minutes to hydroxylate the surface. The glass was placed in an inverted well Figure 1.…”

“…19 We and others have demonstrated drug that response is affected by both stiffness and ECM protein composition on 2D hydrogels. [6][7]20 We have also shown that tumor spheroids encapsulated into 3D hydrogels are drug resistant compared to TCPS.…”

Polymeric hydrogels as artificial extracellular microenvironments for cancer research

“…However, it is still hard to demonstrate what mediates this tropism to physically and chemically diverse tissues. To address this issue, Peyton's group developed a simple biomaterial platform to predict breast cancer metastasis through a cell-ECM screening technique [35]. They create a simple biomaterial platform with diverse ECM components that can regulate the density of integrin binding site and ECM compositions to rapidly screen cellular activities, including cell adhesion, motility, and growth factor responses on biomaterial surfaces.…”