Synthesis and design of macrocyclic collagen mimetic peptides for targeting the cancer-implicated DDR2 kinase

Project Summary / Abstract Collagen lays the foundation of bodily tissues, serving to strengthen, connect, and signal from the micro to the macro scale. The importance of collagen in cancer biology has been well-established: its functions in regulation of the tumor microenvironment—from increased stiffness of the extracellular matrix to dysregulation of cancer

cell signaling—influence tumor proliferation and impenetrability. Key to these effects is the interaction between fibrillar collagens and the discoidin domain receptor type 2 (DDR2), a receptor tyrosine kinase implicated in multiple human cancers. Extracellular binding of fibrillar collagen to DDR2 transduces a cell signal that activates

epithelial to mesenchymal transition, proliferation, and metastasis. This work seeks to develop synthetic collagen mimetic peptides (CMPs) for interrogation of the collagen-DDR2 interaction. As a permutated, triple helical polymer involved in biochemical signaling, collagen has potential for manipulation as a tool for modulating

protein-protein interactions. These applications have been limited by its tripartite nature, which restricts its thermal and entropic stability. Synthetic linkage and cyclization of the three collagen strands may overcome these limitations. To this end, this work focuses on the design, synthesis, biophysical/structural characterization,

and biological application of linked and cyclic CMPs targeted against DDR2 through a chemical biology approach described within two Aims. Aim 1 will evaluate the impact of strategic design strategies on the thermal and proteolytic stability of the proposed CMPs, with a goal of maximizing stability. Aim 2 will investigate the ability of

the CMPs to interact with DDR2 in vitro and modulate DDR2 signaling in cellulo. Prior work has established a method for synthesis of macrocyclic CMPs by solid phase peptide synthesis with on-resin cyclization, and demonstrated that photoreaction of CMPs using diazirine-based photocrosslinking reagents can be used to

prepare unimolecular collagen heterotrimers. Several linked/cyclic as well as linear DDR2-targeted CMPs have already been synthesized and have demonstrated appropriate thermal stability as well as capacity to interact with DDR2. Methods include organic synthesis, photoreaction, circular dichroism, collagenase assay, isothermal

calorimetry, X-ray crystallography, and mammalian cell work coupled with Western blot. This work endeavors to develop biochemical tools for modulation of the collagen interactome—concentrating on the cancer-implicated interaction between collagen and DDR2—via an innovative chemical biology approach, laying the foundation for

new discoveries surrounding the role of collagen in cancer biology with potential for applications in drug discovery. The robust, collaborative training environment at the University of Pennsylvania’s Department of Chemistry and Perelman School of Medicine’s Medical Scientist Training Program—as well as a skilled

sponsorship team with expertise in organic synthesis, collagen structure and function, receptor tyrosine kinases, crystallography, and cell biology—will support completion of the proposed fellowship work and facilitate the rigorous training of an independent physician-scientist investigator.