Search for: All records

Synthetic matrices with dynamic presentation of cell guidance cues are needed for the development of physiologically relevant in vitro tumor models. Towards the goal of mimicking prostate cancer progression and metastasis, we engineered a tunable hyaluronic acid-based hydrogel platform with protease degradable and cell adhesive properties employing bioorthogonal tetrazine ligation with strained alkenes. The synthetic matrix was first fabricated via a slow tetrazine-norbornene reaction, then temporally modified via a diffusion-controlled method using trans-cyclooctene, a fierce dienophile that reacts with tetrazine with an unusually fast rate. The encapsulated DU145 prostate cancer single cells spontaneously formed multicellular tumoroids after 7 days of culture. In situ modification of the synthetic matrix via covalent tagging of cell adhesive RGD peptide induced tumoroid decompaction and the development of cellular protrusions. RGD tagging did not compromise the overall cell viability, nor did it induce cell apoptosis. In response to increased matrix adhesiveness, DU145 cells dynamically loosen cell-cell adhesion and strengthen cell-matrix interactions to promote an invasive phenotype. Characterization of the 3D cultures by immunocytochemistry and gene expression analyses demonstrated that cells invaded into the matrix via a mesenchymal like migration, with upregulation of major mesenchymal markers, and down regulation of epithelial markers. The tumoroids formed cortactin positive invadopodia like structures, indicating active matrix remodeling. Overall, the engineered tumor model can be utilized to identify potential molecular targets and test pharmacological inhibitors, thereby accelerating the design of innovative strategies for cancer therapeutics. 

While self-assembly is relatively well-known and widely used to form hierarchical structures and thin film coatings, controlled assembly is less known and utilized. Our prior work has demonstrated the concept of controlled assembly of macromolecules such as star polymers (MW ~383 kDa, hydrodynamic radius R ~ 13.8 nm) in droplets. The present work extends this concept to smaller molecules, in this case, poly(ethylene glycol) bis-tetrazine (PEG-bisTz, Mn 8.1 kDa, R ~1.5 nm). The key to control molecular assembly is to first deliver ultrasmall volumes (sub-fL) of solution containing PEG-bisTz to a substrate. The solvent evaporates rapidly due to the minute volume, thus forcing the assembly of solute, whose overall size and dimension are dictated by the initial liquid geometry and size. Using pre-patterned surfaces, this work revealed that the initial liquid shape can be further tuned, and as such we could control the final assembly of solute such as PEG-bisTz molecules. The degree of control is demonstrated by varying the micropatterns and delivery conditions. This work demonstrates the validity of controlled assembly for PEG-bisTz, and as such enables 3D nanoprinting of functional materials. The technology has promising applications in nanophotonics, nanoelectronics, nanocomposite materials, and tissue engineering.