Glioblastoma multiforme (GBM) is the most common and malignant of all human primary brain cancers, in which drug treatment is still one of the most effective treatments. However, existing drug discovery and development methods rely on the use of conventional two-dimensional (2D) cell cultures, which have been proven to be poor representatives of native physiology. Here, we developed a novel three-dimensional (3D) brain cancer chip composed of photo-polymerizable poly(ethylene) glycol diacrylate (PEGDA) hydrogel for drug screening. This chip can be produced after a few seconds of photolithography and requires no silicon wafer, replica molding, and plasma bonding like microfluidic devices made of poly(dimethylsiloxane) (PDMS). We then cultured glioblastoma cells (U87), which formed 3D brain cancer tissues on the chip, and used the GBM chip to perform combinatorial treatment of Pitavastatin and Irinotecan. The results indicate that this chip is capable of high-throughput GBM cancer spheroids formation, multiple-simultaneous drug administration, and a massive parallel testing of drug response. Our approach is easily reproducible, and this chip has the potential to be a powerful platform in cases such as high-throughput drug screening and prolonged drug release. The chip is also commercially promising for other clinical applications, including 3D cell culture and micro-scale tissue engineering.

Glioblastoma multiforme (GBM), an extremely invasive and high-grade (grade IV) glioma, is the most common and aggressive form of brain cancer. It has a poor prognosis, with a median overall survival of only 11 months in the general GBM population and 14.6 to 21 months in clinical trial participants with standard GBM therapies, including maximum safe craniotomy, adjuvant radiation, and chemotherapies. Therefore, new approaches for developing effective treatments, such as a tool for assessing tumor cell drug response before drug treatments are administered, are urgently needed to improve patient survival. To address this issue, we developed an improved brain cancer chip with a diffusion prevention mechanism that blocks drugs crossing from one channel to another. In the current study, we demonstrate that the chip has the ability to culture 3D spheroids from patient tumor specimen-derived GBM cells obtained from three GBM patients. Two clinical drugs used to treat GBM, temozolomide (TMZ) and bevacizumab (Avastin, BEV), were applied and a range of relative concentrations was generated by the microfluidic channels in the brain cancer chip. The results showed that TMZ works more effectively when used in combination with BEV compared to TMZ alone. We believe that this low-cost brain cancer chip could be further developed to generate optimal combination of chemotherapy drugs tailored to individual GBM patients.

Complex architectures of integrated circuits are achieved through multiple layer photolithography, which has empowered the semiconductor industry. We adapt this philosophy for tissue engineering with a versatile, scalable, and generalizable microfabrication approach to create engineered tissue architectures composed of digitally specifiable building blocks, each with tuned structural, cellular, and compositional features.

Glioblastoma multiforme (GBM) is the most common and malignant primary brain tumor in adults because of its highly invasive behavior. The existing treatment for GBM, which involves a combination of resection, chemotherapy, and radiotherapy, has a very limited success rate with a median survival rate of <1 year. This is mainly because of the failure of early detection and effective treatment. We designed a novel 3-D GBM cell culture model based on microwells that could mimic in vitro environment and help to bypass the lack of suitable animal models for preclinical toxicity tests. Microwells were fabricated from simple and inexpensive polyethylene glycol material for the control of in vitro 3-D culture. We applied the 3-D micropatterning system to GBM (U-87) cells using the photolithography technique to control the cell spheroids’ shape, size, and thickness. Our preliminary results suggested that uniform GBM spheroids can be formed in 3-D, and the size of these GBM spheroids depends on the size of microwells. The viability of the spheroids generated in this manner was quantitatively evaluated using live/dead assay and shown to improve over 21 days. We believe that in vitro 3-D cell culture model could help to reduce the time of the preclinical brain tumor growth studies. The proposed novel platform could be useful and cost-effective for high-throughput screening of cancer drugs and assessment of treatment responses.

Autapse is an unusual type of synapse generated by a neuron on itself. The ability to monitor axonal growth of single neurons and autapse formation in three-dimensions (3D) may provide fundamental information relating to many cellular processes, such as axonal development, synaptic plasticity and neural signal transmission. However, monitoring such growth is technically challenging due to the requirement for precise capture and long-term analysis of single neurons in 3D. Herein, we present a simple two-step photolithography method to efficiently capture single cells in microscale gelatin methacrylate hydrogel rings. We applied this method to capture and culture single neurons. The results demonstrated that neural axons grew and consequently formed axonal circles, indicating that our method could be an enabling tool to analyze axonal development and autapse formation. This method holds great potential for impact in multiple areas, such as neuroscience, cancer biology, and stem cell biology.

Glioblastoma (GBM) is the most common form of primary brain tumor with a high infiltrative capacity, increased vascularity, and largely elusive tumor progression mechanism. The current GBM treatment methods do not increase the patient survival rate and studies using two-dimensional (2D) cell cultures and in vivo animal models to investigate GBM behavior and mechanism have limitations. Therefore, there is an increasing need for in vitro three-dimensional (3D) models that closely mimic in vivo microenvironment of the GBM tumors to understand the underlying mechanisms of the tumor progression. In this study we propose to use a 3D in vitro model to overcome these limitations, using poly (ethylene glycol) dimethyl acrylate (PEGDA) hydrogel-based microwells and co-culture GBM (U87) cells and endothelial cells (HUVEC) in the 3D microwells to provide a 3D in vitro simulation of the tumor microenvironment. Furthermore, we investigated the gene expression differences of co-cultures by quantitative real-time PCR. Our results suggested that the relative expression profiles of tumor angiogenesis markers, PECAM1/CD31, and VEGFR2, in co-cultures are consistent with in vivo GBM studies. Furthermore, we suggest that our microwell platform could provide robust and useful 3D co-culture models for high-throughput drug screening and treatment of the GBM.

Angiogenesis is an indispensable mechanism in physiological and pathological development of tumors that requires an adequate blood supply. Therefore, understanding the angiogenesis mechanism of tumors has become an important research area to develop reliable and effective therapies for the treatment of tumors. Although several in vivo and in vitro models were developed and used to study the underlying mechanism of angiogenesis, they showed limited success. Therefore, there is an urgent need to build a stable and cost-effective three-dimensional (3D) in vitro angiogenesis model to investigate the tumor formation. In this study, we designed a 3D in vitro angiogenesis model based on gelatin methacrylate (GelMA) hydrogel microwells to mimic an in vivo-like microenvironment for co-cultured glioblastoma and endothelial cells. Our results confirmed the in vitro formation of microtubules during the angiogenic process. We believe that our cost-effective platform can be used for the high-throughput screening of anti-angiogenesis drugs and even for the development of better treatment strategies.

Glioblastoma (GBM) is the most aggressive brain tumor, with 12-15 months median survival time despite current treatment efforts. Among the alternative treatment approaches that have gained acceptance over the last decade is the use of replication-competent oncolytic adenoviruses, which are promising due to their relatively low toxicity and tumor-specific targeting. Three-dimensional (3D) tumor models can mimic the physiological microenvironment of GBM tumors and provide valuable information about the interaction between tumor cells and adenoviruses. Therefore, robust in vitro 3D tumor models are critical to investigate the mechanisms underlying tumor progression and explore the cytotoxicity effect of the adenovirus on tumor cells. In this study, we used a hydrogel microwell platform to generate in vitro 3D GBM spheroids and studied their interactions with the Delta-24-RGD adenovirus. The results showed that the cultured 3D spheroids were successfully infected by the Delta-24-RGD. A significant cell lysis was observed. Cell viability was decreased approximately 37%, 54% and 65% with 10, 50, and 100 MOIs, respectively. The infection of the Delta-24-RGD was found more effective on 3D spheroids when compared to 2D monolayer cell culture. These results implicate that our hydrogel microwell platform could provide a promising 3D model to investigate the oncolytic potential of the viruses in vitro.