Pancreatic cancer represents the 10th most common cancer diagnosis, yet the 4th most common estimated cause of death. The only potential curative therapy for pancreatic cancer is surgical resection; however, few patients have tumors which can be respected. Pancreatic ductal adenocarcinoma, which represents 90% of pancreatic cancers, is particularly aggressive, since it rapidly metastasizes and often expresses several growth factors and signaling components that permit rapid growth. Alternative therapies are desperately needed, as there have been no recent medical advances for treatment of pancreatic adenocarcinoma. Immunotherapy by adoptive transfer of engineered T cells can mediate cancer regression and overcome evasive mechanisms by which tumors avoid immune responses. Chimeric antigen receptors (CAR) are engineered molecules that are a fusion of an antibody-derived antigen-binding motif and intracellular signaling domains, and can recognize tumor antigens independently of the major histocompatiblity complex, expression of which is often lost by tumor cells. At Discovery Genomics, Inc. (DGI), we are developing the Sleeping Beauty transposon system to generate T cells for autologous adoptive T cell therapy. Here we propose to use the SB system for engineering T cells to express CAR recognizing ?V?6, an integrin that is highly expressed on pancreatic cancer cells. In addition, we will stably co-express a chimeric receptor (4??), consisting of a fusion of the IL-4 receptor ? extracellular domain and the endodomain of the common ?- receptor, which is a component of both IL-2 and IL-15 receptors. Expression of the 4?? receptor by anti-??V?6 T cells will allow selective expansion of CAR positive cells. Binding of interleukin-4 to the 4?? receptor will activate a proliferative signal, thus stimulating the T cells to divide. The engineered T cells will be immunophenotypically characterized by flow cytometry. Antitumor cytotoxic activity will be assessed in vitro by measuring cytokine secretion and degranulation after exposure of engineered T cells to ?V?6 positive pancreatic cancer cells and the ability of CAR-expressing T cells to lyse ?V?6 positive pancreatic cancer cells. Results from these experiments will provide an assessment of the effectiveness of SB-engineered T cells in killing pancreatic cancer cells. Subsequent studies in animal models of pancreatic cancer will lead to a clinical trial for the treatment of pancreatic adenocarcinoma by administering human T-cells genetically engineered ex vivo using the Sleeping Beauty transposon system.
Public Health Relevance Statement: Public Health Relevance: In this grant application, studies are proposed to develop a new approach for treating pancreatic cancer by non-viral gene therapy using an integrating DNA element (a transposon) called "Sleeping Beauty". The experiments described in the proposal will establish conditions to genetically engineer lymphocytes as a cellular therapy to kill pancreatic cancer cells.
Project Terms: Adoptive Transfer; Adult; advanced disease; Affinity; Alternative Therapies; Animal Model; Antibodies; antigen binding; Antigen Receptors; Applications Grants; Autologous; base; Binding (Molecular Function); Biological Assay; cancer diagnosis; Cancer Etiology; cancer regression; cancer therapy; Cause of Death; CD28 gene; CD3 Antigens; CD8B1 gene; Cell Degranulation; Cell Line; Cell Therapy; Cells; cellular engineering; Cessation of life; Chronic Lymphocytic Leukemia; Clinical Treatment; Clinical Trials; Coculture Techniques; Complex; cytokine; Cytolysis; cytotoxic; cytotoxicity; Data; Detection; Development; Diagnosis; Disease; Disease remission; DNA; Drug Formulations; Early Diagnosis; Effectiveness; Elements; Engineering; Enzyme-Linked Immunosorbent Assay; Epithelial; Excision; Exposure to; Extracellular Domain; Fc domain; flexibility; Flow Cytometry; gene therapy; Genetic Engineering; Genomics; Growth Factor; Human; Human Engineering; IgG4; Immune response; Immunotherapy; improved; In Vitro; in vitro activity; Integrins; Interleukin-15; Interleukin-2; Interleukin-4; Killings; Lead; Ligands; Lymphocyte; Malignant neoplasm of pancreas; Malignant Neoplasms; Measures; Mediating; Medical; Neoplasm Metastasis; neoplastic cell; Neuroblastoma; non-viral gene therapy; novel strategies; Operative Surgical Procedures; outcome forecast; Pancreatic Adenocarcinoma; pancreatic cancer cells; Pancreatic Ductal Adenocarcinoma; pancreatic neoplasm; Patients; Peptides; peripheral blood; Phase I Clinical Trials; Plasmids; pre-clinical; Preclinical Testing; Proteins; public health relevance; rapid growth; receptor; receptor binding; research study; Resectable; Signal Transduction; Sleeping Beauty; Survival Rate; Symptoms; System; Systems Integration; T cell therapy; T-Cell Receptor; T-Lymphocyte; Testing; Time; Tissues; Transmembrane Domain; Transposase; tumor; Tumor Antigens; Viral Vector
