Medical Student Research Fellowship for Summer 2011

Mentor: Rolf Brekken, PhD                                  
Department:   Sugical Oncology 
Room number: NB8.22
Mail Code: 8593                  
Phone number: x85151       
E-mail: Rolf.Brekken@utsouthwestern.edu               
Project title: DDRs               

Human subjects IRB approved project number (where applicable):      

Animal subjects IRB approved project number (where applicable):APN 0974-06-01-1

Project Type (patient-based research, animal-based research, or basic research; this characterization is only to permit a general classification for grouping similar types of projects): Basic Research

Brief Description of Project:

The extracellular matrix (ECM) is the non-cellular constituent of the tumor microenvironment and as such it is a significant contributor to every facet of tumor biology. Collagen I, the major component of ECM, binds to integrins and discoidin domain receptors (DDR1, DDR2), which are expressed broadly by multiple cell types in pancreatic ductal adenocarcinoma (PDAC). Collagen-induced signaling affects the biology of multiple pathways relevant for tumor progression, including transforming growth factor beta (TGFβ). TGFβ typically suppresses the growth of early stage tumors but response to this cytokine switches during the evolution of the tumor such that invasion and metastasis are promoted by TGFβ in late stage disease. This paradigm is exemplified by PDAC a collagen-rich, desmoplastic disease where TGFβ is known to enhance tumor progression and promote chemoresistance. One mechanism by which TGFβ drives tumor progression is induction of epithelial to mesenchymal transition (EMT), a process that is dependent upon simultaneous collagen signaling. SPARC is a secreted glycoprotein that binds to collagen and is required for appropriate collagen deposition into the ECM. We propose that SPARC reduces collagen binding to the cell surface and inhibits collagen-induced activation of DDRs. We hypothesize that reduced tumor cell SPARC expression, via epigenetic regulation, results in elevated collagen signaling through DDR1 and DDR2, which facilitates TGFβ-induced EMT and chemoresistance. An attractive target for therapy is inhibition of collagen receptor signaling. This proposal will address the function of SPARC in reducing TGFβ promotion of PDAC and explore strategies to inhibit collagen receptor activity. The function of the abundant ECM and collagen in PDAC is not understood. This project will create a new paradigm focused on the collagen-rich ECM as an active participant in disease progression and will also determine whether inhibition of collagen receptor activity blunts EMT and chemoresistance in PDAC. This is consistent with the overall goal of the Brekken laboratory to demonstrate how the microenvironment of PDAC promotes tumor progression.

The project will initially consist of in vitro experiments. Gaurab will participate in the following:

• Determining if SPARC interferes with collagen-induced DDR activation and signaling
• Tissue culture
• Demonstration of collagen activation of DDR 1 or 2 via immunoprecipitation and subsequent Western blotting for phospho-tyrosine
• Identification of active Src via Western blotting post stimulation of DDR-expressing cells with collagen, also probing for other signaling molecules (e.g., Akt, Erk)
• Production and purification of wild-type and mutant SPARC (these constructs are already in house and the purification strategies are established)
• Determining if SPARC reduces EMT of pancreatic cancer cells plated on collagen +/- TGFβ
•  qPCR and immunocytochemistry for markers of EMT

Future studies (beyond the summer of 2011) will encompass animal work. When applicable Gaurab will be added to the APN listed above.

Overall this encompasses a significant effort in my laboratory and Gaurab will be working with a postdoc (Lee Rivera) as well as myself to learn the required techniques and perform the experiments. He will be given more independence as he perfects techniques. I will stress controls and repetition of experiments. 

Previous Research Activities or Publications with Medical Students:

I have participated as a collaborator with Megan Wachsmann, a medical student in the MD-Masters program. Megan is working under the direction of Dr. Ellen Vitetta where she is studying the effect of combination of anti-ICAM and anti-VEGF therapeutic antibodies in pancreatic cancer.