Pancreatic ductal adenocarcinoma is a devastating disease in dire need of improved therapies targeted at specific signaling pathways. Strategies to molecularly profile aberrant pancreatic tissue and inform targeted therapeutic decisions would be of immense value in patient treatment. However, molecular profiling is extremely challenging since biopsied patient tissue is a complex mixture of normal and malignant pancreatic cells. Furthermore, there is a growing understanding that mutations and gene expression alone do not tightly correlate with clinical response. In the current application, a multidisciplinary research team proposes to develop a state-of-the-art, single-cell, platform technology to measure the catalytic activity of sentinel kinases within the KRAS pathway and gene expression through RNA sequencing. The investigators will optimize and validate microsampling and microelectrophoresis methods to assay single cells, simultaneously overcoming the challenges of cellular heterogeneity and sample-size limits. Novel reporters of kinase activity within KRAS-outflow signaling pathways will be designed and new methods and instrumentation combining single-cell capillary electrophoresis with efficient RNA capture will be pioneered. Human tumor samples maintained in murine xenografts will be assayed to gain unique insights into tumor properties not currently addressable. The work will directly link mRNA production with the catalytic activity of kinases in individual tumor cells derived from patients. The technology will enable questions such as which kinase signaling patterns drive the classical vs the basal phenotypes of pancreatic adenocarcinoma and whether a single tumor possesses a mixture of classical and basal-type cells. The data and the insights gained from implementation of this technology will provide a new approach for clinical assays with the potential for a profound impact on therapeutic strategies in the emerging field of precision medicine.
The purpose of the proposed research is to develop and implement an assay system for the combined study of gene expression and signal transduction pathways in human pancreatic cancer cells. The goal of this work is to enhance our understanding of these important aspects of the molecular basis of this deadly cancer and to lead to improved treatments.