Keck Biophysics Facility

• Characterization of biomolecular interactions.

  We offer several orthogonal technologies for the characterization of biomolecular interactions in terms of affinity, specificity, stoichiometry. These tools include micro-calorimeters as well as surface plasmon resonance and bio-layer interferometry instruments, which are used for validation and optimization of drug candidates from high-throughput screens, for rational drug design and structure-activity studies.

• Characterization of exosomes, viruses, and synthetic nanoparticles.

  We offer state-of-the-art technologies, including Asymmetric Flow Field Flow Fractionation (AF4) coupled with static and dynamic light scattering, for multi-attribute characterization of biological exosomes, virus particles and liposomes as well as synthetic nano devices that are engineered for drug/gene delivery, bio-sensing and bio-imaging purposes.

• Characterization of biological and synthetic macromolecules.

  We have a sophisticated set of spectrometers and fluorimeters that can be used for examining the purity, stability, solubility and structural and conformational integrity of proteins, nucleic acids and synthetic polymers. These assays are essential to structure-activity studie and are also commonly employed for characterization of target antigens, crystallization efforts and as quality control in preclinical studies involving engineered proteins or therapeutic antibodies.

• Characterization of suspensions of small drugs, peptides and micro RNAs.

  We offer stability testing, measurements of solubility, purity, heterogeneity and ratiometric content of solutions and suspensions. These analyses are important for enhancing the quality, safety and efficiency of therapeutic reagents. Small-scale purification by HPLC or size exclusion chromatography and lyophilization services are also available.

• Imaging and quantitation of fluorescent, absorbance, phosphorescent and chemiluminescent signals from gels, blots or multiple well plates.

  These capabilities are used in a broad range of projects including quantitative protein expression and protein interaction assays in normal and malignant cells, validation of genetic profiling studies and antibody-based detection tests.

• Chen, S., Lee, L., Naila, T. et al. Structural basis of long-range to short-range synaptic transition in NHEJ. Nature 593, 294–298 (2021).

  The non-homologous end joining (NHEJ) pathway plays a key role in determining the cellular response to radiation and chemotherapy. The article from Dr. Yuan He's laboratory reported the high-resolution cryo-EM structure of two key DNA-protein complexes that are formed by human NHEJ factors. The Facility’s Laser Imager was used to support the functional relevance of these complexes

• Tyson J. MoyerFeng ChenDaniel J. ToftYves RuffVincent L. Cryns, Samuel I. Stupp Self-Assembled Peptide Nanostructures Targeting Death Receptor 5 and Encapsulating Paclitaxel As a Multifunctional Cancer Therapy, ACS Biomaterials Science & Engineering Vol 5/Issue 11 ( 2019)

  In this study by Dr.  Sammuel Stupp’s team, the Circular Dichroism Spectrophotometer in the Keck Facility was used to monitor the assembly of a nanoscale construct engineered to target death receptor 5 (DR5), which is highly expressed in many types of cancer cells. When combined with the chemotherapy drug paclitaxel, the targeting carrier showed potent antitumor activity in breast ductal carcinoma cells

• Xu Han,Yang Mei,Rama K. Mishra, Honghao Bi, Atul D. Jain,Gary E. Schiltz, Baobing Zhao,Madina Sukhanova, Pan Wang, Arabela A. Grigorescu, Patricia C. Weber, John J. Piwinski, Miguel A. Prado, Joao A. Paulo, Len Stephens,Karen E. Anderson, Charles S. Abrams, Jing Yang, and Peng Ji Targeting pleckstrin-2/Akt signaling reduces proliferation in myeloproliferative neoplasm models J Clin Invest. 133(2023)

  The Isothermal Titration Calorimeter in the Keck Facility was used in a study by Dr. Peng to determine that a putative inhibitor of pleckstrin-2Akt signaling, identified in an in silico screen, binds specifically to pleckstrin and not to 2Akt. This is important therapeutically because while pleckstrin-2 is a promising target for MPN therapeutics, small-molecule compounds that directly target Akt often result in severe side effects.

• Livia Elena Sima, Bakhtiyor Yakubov, Sheng Zhang, Salvatore Condello, Arabela A. Grigorescu, Nkechiyere G. Nwani, Lan Chen, Gary E. Schiltz, Constandina Arvanitis, Zhong-Yin Zhang, Daniela Matei Small Molecules Target the Interaction between Tissue Transglutaminase and Fibronectin. Mol Cancer Ther 18 (6): 1057–1068 (2019)

  Tissue transglutaminase (TG2) is a multifunctional protein with enzymatic, GTPase, and scaffold properties. TG2 interacts with fibronectin (FN) through its N-terminal domain, stabilizing integrin complexes, which regulate cell adhesion to the matrix. A study headed by Dr. Daniela Matei used high-throughput screening to identify several small molecule inhibitors (SMI) for the TG2/FN complex. The Facility’s service for Biophysical characterization of drug-target interactions   was employed for the characterization of SMIs binding to TG2. The data supported continued optimization of the new class of SMIs that block the TG2/FN complex at the interface between cancer cells and the tumor niche.

• Lakshmi R. Bollu, Prashant V. Bommi, Paige J. Monsen, Lijie Zhai, Kristen L. Lauing, April Bell, Miri Kim, Erik Ladomersky, Xinyu Yang, Leonidas C. Platanias, Daniela E. Matei, Marcelo G. Bonini, Hidayatullah G. Munshi, Rintaro Hashizume, Jennifer D. Wu, Bin Zhang, Charles David James, Peiwen Chen, Masha Kocherginsky, Craig Horbinski, Michael D. Cameron, Arabela A. Grigorescu, Bakhtiar Yamini, Rimas V. Lukas, Gary E. Schiltz, and Derek A. Wainwright Identification and Characterization of a Novel Indoleamine 2,3-Dioxygenase 1 Protein Degrader for Glioblastoma Journal of Medicinal Chemistry 65 (23), 15642-15662 (2022)

  A synthetic library of proteolysis targeting chimeras (PROTACs) targeting the immunosuppressive enzyme IDO1 in glioblastoma was screened. For the characterization of the lead hits, the Keck Facility; 's service for Biophysical characterization of drug-target interactions was used to assess the affinity and stability of the binary and ternary complexes formed by these PROTACs with the IDO1 target.

All manuscripts and grants presenting work supported by this core should include the following acknowledgement:

We thank the Robert H. Lurie Comprehensive Cancer Center of Northwestern University in Chicago, IL for the use of the Keck Biophysics resources. The Lurie Cancer Center is supported in part by a NCI Cancer Center Support Grant #P30 CA060553.