Casey J. Krusemark
Ph.D. Biochemistry, 2007, University of Wisconsin--Madison (under Peter Belshaw)
B.S. Chemistry, B.S. Crop Science, 2001, University of Illinois--Urbana-Champaign (under Jack Widholm)
Our work centers on two nascent technologies that not only borrow from the structure of nature’s bioactive molecules but also from nature's approaches to synthesize molecules and assess them for function. The application of these technologies is focused on advances in medicinal chemistry.
DNA-ecoded chemical libraries. In the first area, we build synthetic small molecule libraries that are covalently linked encoding DNA scaffolds. Within the past few years, DNA-encoded chemical libraries (DELs) have been adopted at every major pharmaceutical company for hit generation and lead discovery. Several successful cases have been reported yielding multiple clinical candidates. The DEL approach has the ability to generate and assay small molecule libraries of much greater complexity (present demonstrations up to 1010 molecules) than by any other approach. Rather than screening molecules in discrete assays, DELs can be assessed collectivly for binding to a protein target by a selection assay, all together, in a single tube. The cost, effort, and infrastructure required are reduced dramatically, particularly in comparison to traditional high-throughput screening (HTS). This approach enables medicinal chemistry to capitalize on many of the amazing capabilities of molecular biology, such has high detection sensitivity via DNA amplification (PCR) and massively parallel analysis of DNAs in next-generation DNA sequencing. Our work in this area involves development of novel libraries and selection strategies for discovery of protein-protein interaction inhibitors that are mediated by short linear peptide motifs, with a particular focus on protein kinases and chromodomains. We are using these newly developed approaches to develop therapeutic lead molecules for application in treating cancer, Alzheimer's disease, and malaria.
DNA-linked probes for protein assays conducted via DNA analysis. In the second area, we have developed an evolution-inspired assay approach to detect sample stimuli using DNA-linked molecules as activity probes. We have developed DNA-encoded probes for detection of several enzymatic activities, as well as synthetic ligand binding, which allow activity detection through DNA sequence analysis. We work to apply this approach in proteomic activity profiling and in screening of small molecules for enzyme inhibition. Developments in genetic analysis technologies, particularly DNA sequencing, have been transformative to biomedical research. In contrast to genomic information, the barrier to accessing proteomic information, particularly enzyme activity, is dramatically higher. As aberrant enzyme activities are consistently observed in disease, this information is critical for appropriate diagnosis and treatment. As with DNA-encoded libraries, these approaches allow protein assays to harness the power of DNA analysis technologies. Current efforts are focused on developing robust assays for protein targets directly in cell lysates for small molecule screening and in using libraries of protein kinase substrates to understand resistance mechanisms in drug-resistant breast cancer (in collaboration with Dr. Michael Wendt).
PHRM 824 Principles of Pathophysiology and Drug Action
MCMP 208 Biochemistry for Pharmaceutical Sciences I
NINDS R21 (1NS101535) (02/01/2017-01/31-2019)
(PIs: Hockerman, GH and Krusemark, CJ) Differential modulation of Cav1.2 and Cav1.3)
NIGMS R35 (PI: Krusemark, CJ)
Denton, K. E., Wang, S., Gignac, M. C., Milosevich, N., Hof, F., Dykhuizen, E. C., Krusemark, C. J., "Robustness of In Vitro Selection Assays of DNA-encoded Peptidomimetic Ligands to CBX7 and CBX8." SLAS Discovery, 2018, 23, 417-428.
Kim, D., Jetson, R. R., Krusemark, C. J., "A DNA-assisted Immunoassay for Enzyme Activity via a DNA-linked, Activity-based Probe." Chem. Comm. 2017, 53, 9474-9477.
Denton, K. E., Krusemark, C. J., "Crosslinking of DNA-linked ligands to target proteins for enrichment from DNA-encoded libraries." Med. Chem. Comm. 2016, 7, 2020-2027.
Jetson, R. R., Krusemark, C. J. "Sensing Enzymatic Activity by Exposure and Selection of DNA-encoded Probes." Angewandte Chemie Int. Ed. 2016, 55, 9562-9566.
Krusemark, C. J., Tilmans, N. P., Brown, P. O., Harbury, P. B., "Directed Chemical Evolution with an Outsized Genetic Code." PLoS ONE 11(8) e0154765.
Tilmans, N. P., Krusemark, C. J., Harbury, P. B., “Expedient Synthesis of a Modular Phosphate Affinity Reagent.” Bioconj. Chem. 2010, 21, 1010-1013.
Frey, B. L., Ladror, D. T., Sondalle, S. B., Krusemark, C. J., Jue, A. L., Coon, J. J., Smith, L. M., “Chemical derivatization of peptide carboxyl groups for highly efficient electron transfer dissociation." J. Am. Soc. Mass Spectrom. 2013, 24, 1710-1721.
Krusemark, C. J., Frey, B. L., Belshaw, P. B., Smith, L. M., "Modifying the Charge State Distribution of Proteins in Electrospray Ionization Mass Spectrometry by Chemical Derivatization." J. Am. Soc. Mass Spectrom. 2009, 20, 1617-1625.
Krusemark, C. J., Ferguson, J. T., Wenger, C. D., Kelleher, N. L., Belshaw, P. J., "Global Amine and Acid Functional Group Modification of Proteins." Analytical Chemistry. 2008, 80, 713-720.
Krusemark, C. J. and Belshaw, P. J., "Covalent Labelling of Fusion Proteins in Live Cells via an Engineered Receptor-Ligand Pair." Organic and Biomolecular Chemistry. 2007, 5, 2201-2204.
Krusemark, C. J.*, Lamos, S. M.*, McGee, C. J., Scalf, M., Smith, L. M., Belshaw, P. J., "Mixed Isotope Photoaffinity Reagents for Identification of Small Molecule Targets by Mass Spectrometry." Angewandte Chemie Int. Ed. 2006, 45, 4329-4333. *Authors contributed equally.