*Please note the room change below.* BME PhD Preliminary Exam Announcement for Chun-Yi Chang (Chien-Chi Lin and Luis Solorio, Co-Chairs) Everyone is invited to attend the public presentation beginning at 10:00AM. Title: In situ generation of physiological gradients in cell-laden hydrogels Date: December 1, 2022 Time: 10:00AM Location: SL306C at IUPUI and via Zoom https://iu.zoom.us/j/87266126712 Abstract: Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease with 5-year survival rate of ~10%. A major hurdle to treating PDAC is the desmoplastic reaction in the tumor microenvironment (TME), which is a result of excess deposition of extracellular matrices (e.g., hyaluronic acid or HA) secreted by stromal cells. The desmoplastic reaction is a barrier blocking the therapeutics from entering the tumor, while allowing cancer cells to metastasize. To this end, we have developed a gelatin-based hydrogel system with biorthogonal click chemistries to mimic TME in PDAC. Gelatin was dually modified with norbornene and carbohydrazide (i.e., GelNB-CH) to form a primary gel network through thiol-norbornene chemistry and to allow dynamic stiffening via hydrazone click chemistry. The dynamic stiffening changed the cytokine and protein secretion of the encapsulated PDAC cells. Compared to gel stiffening with the bioinert control, matrix stiffening by oxidized HA (oHA) restored the spreading cell morphology and enhanced CD44 expression. To capture the heterogeneous ECM distribution in TME, we sought to create HA gradient in the hydrogel based on microporous annealed GelNB-CH microgels and the diffusion of oHA. The porous structure makes the annealed microgels more permeable for the diffusion of oHA than a nano-porous bulk gel. We expect the system can also be more permissive for cell protrusion or spheroid outgrowth. Future work will be focused on the EMT-related gene/protein expression and cell migration in response to the chemical and mechanical cues from the dynamic matrix.