Small Molecule Screening Strategies from Lead Identification to Validation

ARTICLES DISCUSSED:
Festa, F., Labaer, J. Kinase inhibitor screening in self-assembled human protein microarrays. Journal of Visualized Experiments. 152, ...

Dana-Farber Cancer Institute

Some of the most widely used drugs, such as aspirin and penicillin, are covalent drugs. Covalent binding can improve potency, selectivity, and duration of the effects, but the intrinsic reactivity represents a potential liability and may result in idiosyncratic toxicity. For decades, the cons were believed to outweigh the pros, and covalent targeting was deprioritized in drug discovery. Recently, several covalent inhibitors have been approved for cancer treatment, thus rebooting the field. In this review, we briefly reflect on the history of selective covalent targeting, and provide a comprehensive overview of emerging developments from a chemical biology stand-point. Our discussion will reflect on efforts to validate irreversible covalent ligands, expand the scope of targets, and discover new ligands and warheads. We conclude with a brief commentary of remaining limitations and emerging opportunities in selective covalent targeting.

Recent Advances in Selective and Irreversible Covalent Ligand Development and Validation.

KRAS is the most frequently mutated oncogene found in pancreatic, colorectal, and lung cancers. Although it has been challenging to identify targeted therapies for cancers harboring KRAS mutations, KRAS can be targeted by small-molecule inhibitors that form covalent bonds with cysteine 12 (C12). Here, we designed a library of C12-directed covalent degrader molecules (PROTACs) and subjected them to a rigorous evaluation process to rapidly identify a lead compound. Our lead degrader successfully engaged CRBN in cells, bound KRASin vitro, induced CRBN/KRAS dimerization, and degraded GFP-KRAS in reporter cells in a CRBN-dependent manner. However, it failed to degrade endogenous KRAS in pancreatic and lung cancer cells. Our data suggest that inability of the lead degrader to effectively poly-ubiquitinate endogenous KRAS underlies the lack of activity. We discuss challenges for achieving targeted KRAS degradation and proposed several possible solutions which may lead to efficient degradation of endogenous KRAS.

Exploring Targeted Degradation Strategy for Oncogenic KRAS.

Small molecules continue to dominate drug discovery because of their ease of use, lower cost of manufacturing, and access to intracellular targets. However, despite these advantages, small molecules are more likely to fail in clinical trials compared with biologicals and their development remains limited to a small subset of disease-relevant 'druggable' targets. Targeted protein degradation has recently emerged as a novel pharmacological modality that promises to overcome small molecule limitations whilst retaining their key advantages. Here, we use a Strengths-Weaknesses-Opportunities-Threats (SWOT) framework to critically assess the current status of this rapidly evolving field. We expect that degrader molecules are only the beginning of a range of novel targeting modalities that hijack existing endogenous cellular machineries to chemically redirect biological targets and pathways. Therefore, this piece may offer a roadmap for enhancing development of both degraders and related modalities.

Critical Assessment of Targeted Protein Degradation as a Research Tool and Pharmacological Modality.

Targeted covalent inhibitors have regained widespread attention in drug discovery and have emerged as powerful tools for basic biomedical research. Fueled by considerable improvements in mass spectrometry sensitivity and sample processing, chemoproteomic strategies have revealed thousands of proteins that can be covalently modified by reactive small molecules. Fragment-based drug discovery, which has traditionally been used in a target-centric fashion, is now being deployed on a proteome-wide scale thereby expanding its utility to both the discovery of novel covalent ligands and their cognate protein targets. This powerful approach is allowing 'high-throughput' serendipitous discovery of cryptic pockets leading to the identification of pharmacological modulators of proteins previously viewed as "undruggable". The reactive fragment toolkit has been enabled by recent advances in the development of new chemistries that target residues other than cysteine including lysine and tyrosine. Here, we review the emerging area of covalent fragment-based ligand discovery, which integrates the benefits of covalent targeting and fragment-based medicinal chemistry. We discuss how the two strategies synergize to facilitate the efficient discovery of new pharmacological modulators of established and new therapeutic target proteins.

Fragment-based covalent ligand discovery.

[This corrects the article DOI: 10.1039/D0CB00222D.].

Correction: Fragment-based covalent ligand discovery.

Twenty years after the publication of the first draft of the human genome, our knowledge of the human proteome is still fragmented. The challenge of translating the wealth of new knowledge from genomics into new medicines is that proteins, and not genes, are the primary executers of biological function. Therefore, much of how biology works in health and disease must be understood through the lens of protein function. Accordingly, a subset of human proteins has been at the heart of research interests of scientists over the centuries, and we have accumulated varying degrees of knowledge about approximately 65% of the human proteome. Nevertheless, a large proportion of proteins in the human proteome (∼35%) remains uncharacterized, and less than 5% of the human proteome has been successfully targeted for drug discovery. This highlights the profound disconnect between our abilities to obtain genetic information and subsequent development of effective medicines. Target 2035 is an international federation of biomedical scientists from the public and private sectors, which aims to address this gap by developing and applying new technologies to create by year 2035 chemogenomic libraries, chemical probes, and/or biological probes for the entire human proteome.

Milka Kostic

Department of Cancer Biology,
Chemical Biology Program

Dana-Farber Cancer Institute

Small Molecule Screening Strategies from Lead Identification to Validation

Milka Kostic

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Cancer Biology,Chemical Biology Program

Dana-Farber Cancer Institute

Small Molecule Screening Strategies from Lead Identification to Validation

Department of Cancer Biology,
Chemical Biology Program