Emerging Approaches for the Identification of Protein Targets of Small Molecules - A Practitioners’ Perspective

Abstract

Small-molecule (SM) leads in the early drug discovery pipeline are progressed primarily based on potency against the intended target(s) and selectivity against a very narrow slice of the proteome. So, why is there a tendency to wait until SMs are matured before probing for a deeper mechanistic understanding? For one, there is a concern about the interpretation of complex -omic data outputs and the resources needed to test these hypotheses. However, with recent advances in broad endpoint profiling assays that have companion reference databases and refined technology integration strategies, we argue that data complexity can translate into meaningful decision-making. This same strategy can also prioritize phenotypic screening hits to increase the likelihood of accessing unprecedented target space. In this Perspective. we will highlight a cohesive process that supports SM hit prosecution, providing a data-driven rationale and a suite of methods for direct identification of SM targets driving relevant biological end points.

2‐Sulfonylpyrimidines Target the Kinesin HSET via Cysteine Alkylation

“…In BT‐549 breast carcinoma cells, treatment with 5 led to a threefold increase in the amount of mitotic cells (Figure S3C‐S3D). This phenotype is prominent for small molecules that target microtubules . However, compound 5 did not affect the in vitro tubulin polymerization (Figure E) and thus, most likely does not target tubulin/microtubules.…”

Mycobacterium tuberculosis Shikimate Pathway Enzymes as Targets for the Rational Design of Anti-Tuberculosis Drugs

“…Accordingly, there has been a revival on the interest of phenotypic drug discovery approaches based on their potential to address the incompletely understood complexity of diseases, their promise of delivering first-in-class drugs and major advances in the tools for cell-based phenotypic screening [8]. Ideally, phenotypic drug discovery campaigns utilize physiologically relevant models that are linked to patient-derived biology, which in turn may self-select small molecule phenotypic hits affecting pathways and protein targets most relevant to the disease of interest [9]. Although phenotypic drug screening interrogates a larger molecular target space, primary hits are rarely target-specific [8,10].…”

“…Moreover, phenotypic screening efforts may be followed by identification of target(s) and, if amenable, heterologous expression, purification, protein assay and structural determination to help guide medicinal chemistry efforts (closing full circle). In both cases, small molecule prioritization from target-based drug discovery and phenotypic drug discovery would benefit from a deeper appreciation of the mode of action and target engagement profile [9].…”

Target identification of natural medicine with chemical proteomics approach: probe synthesis, target fishing and protein identification

“…[16][17][18] However, developing a natural product-based new medicine from random moiety modification is a lengthy and costly process, due in part to the difficulties associated with comprehensively understanding their mechanism of action (MOA) as well as side effects. 19,20 Interactions with intracellular protein targets are the foundation through which natural products exert their pharmacological activity. Therefore, target identification is the initial key step for the discovery and development of new natural medicines, 21,22 as this allows the determination of the MOA and side effects.…”