Label-Free Real-Time Microarray Imaging of Cancer Protein–Protein Interactions and Their Inhibition by Small Molecules

Abstract

A rapid optical microarray imaging approach for anticancer drug screening at specific cancer protein-protein interface targets with binding kinetics and validation by a mass sensor is reported for the first time. Surface plasmon resonance imager (SPRi) demonstrated a 3.5-fold greater specificity for interactions between murine double minute 2 protein (MDM2) and wild-type p53 over a nonspecific p53 mutant in a real-time microfluidic analysis. Significant percentage reflectivity changes (Δ%R) in the SPRi signals and molecular-level mass changes were detected for both the MDM2-p53 interaction and its inhibition by a small-molecule Nutlin-3 drug analogue known for its anticancer property. We additionally demonstrate that synthetic, inexpensive binding domains of interacting cancer proteins are sufficient to screen anticancer drugs by an array-based SPRi technique with excellent specificity and sensitivity. This imaging array, combined with a mass sensor, can be used to study quantitatively any protein-protein interaction and screen for small molecules with binding and potency evaluations.

One-step methodology for the direct covalent capture of GPCRs from complex matrices onto solid surfaces based on the bioorthogonal reaction between haloalkane dehalogenase and chloroalkanes

“…Anchoring protein-based techniques 1 – 5 are central to biomarker detection, proteomic screens, drug discovery and the systemic study of biological mechanisms at the molecular level. 6 These methods require the immobilization of target proteins onto solid scaffolds.…”

A Multiplexed Microfluidic Platform for Bone Marker Measurement: A Proof-of-Concept

“…Despite the recent improvements in the development of multiplexed microfluidic-based systems, especially in personalized cancer diagnostics, a clear challenge still resides in the integration and operation of such microfluidic platforms for the measurement of different biomarkers in clinical practice [ 14 , 15 , 16 ]. As such, a few attempts have been made in the past for the development of new screening assay methods for bone turnover markers (BTMs); yet none have succeeded to fully satisfy the criteria needed for their application in clinical practice [ 17 ].…”

Electrochemical and Surface-Plasmon Correlation of a Serum-Autoantibody Immunoassay with Binding Insights: Graphenyl Surface versus Mercapto-Monolayer Surface

“…The experimentally obtained SPR sensograms (Figure S6A,B) for different concentrations of GADA were fit into a 1:1 bimolecular kinetic model. , The equations presented in the Supporting Information were used to calculate the association-rate ( k a ), the dissociation-rate ( k d ), and the binding-constant ( K D ) parameters (Table ).…”