Surface plasmon resonance (SPR) biosensor methods are ideally suited for fragment -based lead discovery. However, generally applicable experimental procedures and detailed protocols are lacking, especially for structurally or physico-chemically challenging targets or when tool compounds are not available. Success depends on accounting for the features of both the target and the chemical library, purposely designing screening experiments for identification and validation of hits with desired specificity and mode -of -action, and availability of orthogonal methods capable of confirming fragment hits. The range of targets and libraries amenable to an SPR biosensor -based approach for identifying hits is considerably expanded by adopting multiplexed strategies, using multiple complementary surfaces or experimental conditions. Here we illustrate principles and multiplexed approaches for using flow-based SPR biosensor systems for screening fragment libraries of different sizes (90 and 1056 compounds) against a selection of challenging targets. It shows strategies for the identification of fragments interacting with 1) large and structurally dynamic targets, represented by acetyl choline binding protein (AChBP), a Cys-loop receptor ligand gated ion channel homologue, 2) targets in multi protein complexes, represented by lysine demethylase 1 and a corepressor (LSD1/CoREST), 3) structurally variable or unstable targets, represented by farnesyl pyrophosphate synthase (FPPS), 4) targets containing intrinsically disordered regions, represented by protein tyrosine phosphatase 1B (PTP1B), and 5) aggregation-prone proteins, represented by an engineered form of human tau (tau K18M). Practical considerations and procedures accounting for the characteristics of the proteins and libraries, and that increase robustness, sensitivity, throughput and versatility are highlighted. The study shows that the challenges for addressing these types of targets is not identification of potentially useful fragments per se, but establishing methods for their validation and evolution into leads.

The number of peer-reviewed publications that feature biosensor data increases every year. A search of PubMed using common technique terminology, including bio-layer interferometry (BLI), surface plasmon resonance (SPR) and grating-coupled interferometry (GCI) generated more than 2500 scientific papers from 2022. Compared to 2009, when David Myszka and Rebecca Rich presented their most recent review of biosensor literature (Rich and Myszka, 2011), this number has nearly doubled. With this increasing number of publications comes an increasing need for standardization of the way biosensor data is reported in journals to allow for replication of the experiments that were performed. Biosensor data is often poorly described in papers which makes it difficult, if not impossible, to replicate the experiment. Critical information typically missing includes sample preparation, method settings, and data evaluation details. We have also found published work in which the authors have failed to report the type of sensor that was used, or which biosensor instrumentation was used. To come to terms with this growing problem, we propose a standardization of the way biosensor data is reported in scientific journals. We call this standard STROBE, standards for reporting optical biosensor experiments.

Antibody-drug conjugates (ADCs) constitute a rapidly expanding category of biopharmaceuticals that are reshaping the landscape of targeted chemotherapy. The meticulous process of selecting therapeutic targets, aided by specific monoclonal antibodies' high specificity for binding to designated antigenic epitopes, is pivotal in ADC research and development. Despite ADCs' intrinsic ability to differentiate between healthy and cancerous cells, developmental challenges persist. In this study, we present a rationalized pipeline encompassing the initial phases of the ADC development, including target identification and validation. Leveraging an in-house, computationally constructed ADC target database, termed ADC Target Vault, we identified a set of novel ovarian cancer targets. We effectively demonstrate the efficacy of Surface Plasmon Resonance (SPR) technology and in vitro models as predictive tools, expediting the selection and validation of targets as ADC candidates for ovarian cancer therapy. Our analysis reveals three novel robust antibody/target pairs with strong binding and favourable antibody internalization rates in both wild-type and cisplatin-resistant ovarian cancer cell lines. This approach enhances ADC development and offers a comprehensive method for assessing target/antibody combinations and pre-payload conjugation biological activity. Additionally, the strategy establishes a robust platform for high-throughput screening of potential ovarian cancer ADC targets, an approach that is equally applicable to other cancer types.

The human gamma-herpesviruses Epstein–Barr virus (EBV) (HHV-4) and Kaposi’s sarcoma-associated herpesvirus (KSHV) (HHV-8) are responsible for a number of diseases, including various types of cancer. Epstein–Barr nuclear antigen 1 (EBNA1) from EBV and latency-associated nuclear antigen (LANA) from KSHV are viral-encoded DNA-binding proteins that are essential for the replication and maintenance of their respective viral genomes during latent, oncogenic infection. As such, EBNA1 and LANA are attractive targets for the development of small-molecule inhibitors. To this end, we performed a biophysical screen of EBNA1 and LANA using a fragment library by saturation transfer difference (STD)–NMR spectroscopy and surface plasmon resonance (SPR). We identified and validated a number of unique fragment hits that bind to EBNA1 or LANA. We also determined the high-resolution crystal structure of one fragment bound to EBNA1. Results from this screening cascade provide new chemical starting points for the further development of potent inhibitors for this class of viral proteins.

The inhibitor for the homophilic dimerization of P-cadherin was discovered by SPR-based screening using fragment compounds. Our SPR assays identified a specific P-cadherin binder, which was able to inhibit the cell adhesion of living CHO cells that expressed P-cadherin.