Three-Dimensional Printed Abdominal Imaging Windows for In Vivo Imaging of Deep-Lying Tissues

The ability to microscopically image diseased or damaged tissue throughout a longitudinal study in living mice would provide more insight into disease progression than having just a couple of time points to study. In vivo disease development and monitoring provides more insight than in vitro studies as well. In this study, we developed permanent 3D-printed, surgically implantable abdominal imaging windows (AIWs) to allow for longitudinal imaging of deep-lying tissues or organs in the abdominal cavity of living mice. They are designed to prevent organ movement while allowing the animal to behave normally throughout longitudinal studies. The AIW also acts as its own mounting bracket for attaching them to a custom 3D printed microscope mount that attaches to the stage of a microscope and houses the animal inside. During the imaging of the living animal, cellular and macroscopic changes over time in one location can be observed because markers can be used to find the same spot in each imaging session. We were able to deliver cancer cells to the pancreas and use the AIW to image the disease progression. The design of the AIWs can be expanded to include secondary features, such as delivery and manipulation ports and guides, and to make windows for imaging the brain, subcutaneous implants, and mammary tissue. In all, these 3D-printed AIWs and their microscope mount provide a system for enhancing the ability to image and study cellular and disease progression of deep-lying abdominal tissues of living animals during longitudinal studies

Inhibition of Geranylgeranyl Diphosphate Synthase is a Novel Therapeutic Strategy for Pancreatic Ductal Adenocarcinoma

Rab proteins play an essential role in regulating intracellular membrane trafficking processes. Rab activity is dependent upon geranylgeranylation, a post-translational modification that involves the addition of 20-carbon isoprenoid chains via the enzyme geranylgeranyl transferase (GGTase) II. We have focused on the development of inhibitors against geranylgeranyl diphosphate synthase (GGDPS), which generates the isoprenoid donor (GGPP), as anti-Rab agents. Pancreatic ductal adenocarcinoma (PDAC) is characterized by abnormal mucin production and these mucins play important roles in tumor development, metastasis and chemo-resistance. We hypothesized that GGDPS inhibitor (GGDPSi) treatment would induce PDAC cell death by disrupting mucin trafficking, thereby inducing the unfolded protein response pathway (UPR) and apoptosis. To this end, we evaluated the effects of RAM2061, a potent GGDPSi, against PDAC. Our studies revealed that GGDPSi treatment activates the UPR and triggers apoptosis in a variety of human and mouse PDAC cell lines. Furthermore, GGDPSi treatment was found to disrupt the intracellular trafficking of key mucins such as MUC1. These effects could be recapitulated by incubation with a specific GGTase II inhibitor, but not a GGTase I inhibitor, consistent with the effect being dependent on disruption of Rab-mediated activities. In addition, siRNA-mediated knockdown of GGDPS induces upregulation of UPR markers and disrupts MUC1 trafficking in PDAC cells. Experiments in two mouse models of PDAC demonstrated that GGDPSi treatment significantly slows tumor growth. Collectively, these data support further development of GGDPSi therapy as a novel strategy for the treatment of PDAC

Pancreatic Cancer: Novel Therapy, Research Tools, and Educational Outreach

Since the 1980’s, legislatures have invested immense effort into strategies to capitalize on the United States’ global lead in basic and translational research. This is largely motivated by the discrepancy the US faces in converting its world leading research discoveries and innovations into commercial products, relative to other countries. This truncated productivity leaves the US out of substantial GDP growth and reduces public access to new innovations. Proponents of government and university owned patents reason that enhanced public private partnerships narrow the gap between the technology transfer of basic research discoveries and accessible commercial products. With these concepts of technology transfer in mind, we set out to develop translationally relevant technologies to improve PDAC patient outcomes. First, we then partnered with pharmaceutical scientists and medicinal chemists to evaluate novel small molecule inhibitors and combined therapeutic approaches to treating PDAC. Second, we engineered a research tool, our Abdominal Imaging Window (AIW), which facilitates in vivo longitudinal deep tissue microscopy of pancreatic tumors and microenvironment throughout disease progression. Then, we collaborated with experts in the field of fluorescent imaging to develop tumor specific contrast agents that could be used in real time intraoperatively to improve surgical resection outcomes. Lastly, we investigated tools for strategic Science, Technology, Engineering, and Mathematics (STEM) outreach programing for not only recruiting future scientists, but for building a community of trust and understanding of scientific practices in the next generation. In our investigation of novel combination therapeutics, we found that inhibition of CDK5 with small molecule inhibitor CP had the potential to reduce orthotopically implanted murine pancreatic adenocarcinomas of clinically relevant size to radiographically undetectable when used in combination with gemcitabine. These previously tumor bearing mice remained in remission and showed durable treatment free response of up to 40 days. Given this profound response, we pursued a multifaceted investigation into the effects of CDK5 inhibition on the tumor microenvironment, including the effects on perineural invasion and tumor associated neural migration. To do so, we had to first engineer an Abdominal Imaging Window which would allow us to perform longitudinal intravital microscopy of tumor progression and neural invasion via multiphoton microscopy. These well tolerated AIWs provided macro and microscopic detail of PDAC progression longitudinally, and brought to the market a novel mechanism for performing upright microscopy in vivo