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Global Translational Medicine Role of HTS in cancer therapeutics

evaluated through phenotypic screening (e.g., Azacitidine, and metabolism) studies are conducted. In this review, we
Fulvestran, Nelarabine, and Vorinostat) and 17 through briefly explore screening methods, their challenges, and
target-based screening (e.g., Bortezomib, Gefitinib, benefits in HTS. In addition, various recent methodologies
Imatinib, Sorafenib, and Sunitinib). are discussed, surrounding topics on developing successful
Despite the incorporation of new methodologies, the experimental assays, implementation through primary and
rate of successful drug screening remains notably low. This secondary screens, and target identification. We also discuss
the identification of the most efficacious drugs using these
is attributed to the molecular heterogeneity underlying approaches, particularly in the context of cancer.
disease mechanisms, which varies within and between
patients across various disease types. 12-16 These challenges 2. Target-based screening
highlight the limitations of contemporary drug discovery
strategies. Therefore, “one drug, one target” paradigm is Target-based biochemical assays are based on defined/
usually followed to reduce unwanted off-target side effects. known molecular targets, which are used to find a lead
However, this approach does not take into consideration compound from the library that can efficiently induce/
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the complexities of mechanisms, which are in fact very inhibit the target’s activity. Target-based screening utilizes
complicated and controlled by various factors inside genomics information to identify the targets causing disease,
the cells. Highly potent, single-target treatments may which is already available through previous phenotypic
demonstrate weak clinical efficacy when compared to studies. Genomic studies also provide functional targets
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multi-target drugs. 17,18 Nevertheless, most effective drugs to understand mechanism of action (MOA). Therefore,
are multi-target ligands, and their efficacy is affected by the target-based approach is simpler, direct, and specific
the diverse molecular mechanisms. 19-22 Some examples because of prior knowledge of drug’s MOA availability,
of multi-target, anti-cancer drugs include lapatinib and which can be easily utilized to understand the interaction of
duvelisib. Lapatinib is a reversible, ATP‑competitive drug with the target in a relatively easy manner as compared
inhibitor of the human epidermal growth factor receptor to phenotype-based screening approaches. Various
2 (HER2) and epidermal growth factor receptor (EGFR) methods have been employed to identify MOA including
tyrosine kinases. Duvelisib, on the other hand, is a protein–protein interactions, examination of structural and
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dual inhibitor of PI3K-δ and PI3K-γ. Both drugs have receptor-mediated protein targets, and regulatory factors.
demonstrated promising clinical efficacy in advanced Target-based drug discovery relies on two of the most
hematologic malignancies. 24 popular target classes: enzymes and G‑protein‑coupled
receptors (GPCRs). GPCRs, recognized as the largest family
HTS demands the development of robust assays that of targets for approved drugs, engage in direct interactions
provide high signal-to-noise ratios, which can be applied to with numerous chemical entities, initiating molecular
small volumes as well. Therefore, cell-based in vitro assays interactions in the extracellular milieu. Another druggable
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are more relevant to biological phenotypes in predicting target is ion channels by virtue of its coupling with the
the therapeutic response in a quick and effective manner. plethora of physiological consequences. 35-42
Cell‑based assays include in vitro toxicity assay, RNA
The hallmark biochemical assays involve the assessment
interference (RNAi), second messenger, cell proliferation, of enzymatic activity for cell growth, proliferation,
and reporter assays. Cell‑based assays offer several differentiation, and metabolism. Biochemical assays include
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advantages over cell-free biochemical assays in various enzymatic kinase assays, voltage-gated ion channels, and
aspects. They are more cost-effective, closely resemble the serine/cysteine proteases FRET, fluorescence correlation
clinical physiological state, and provide real selectivity spectroscopy, fluorescence intensity distribution analysis,
for compounds that can cross the cell membrane to reach and in vitro transcription assays. The main purpose
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target sites. 26-30 Both screening methods have immensely of these assays is to select small molecules from the
contributed to drug discovery by producing high-quality compound libraries based on their affinity to bind with
data (Figure 1, Tables 1 and 2). 31
purified target protein of interest and inhibit or induce
Decision of following both phases parallelly or which enzymatic activity in vitro as per the hypothesis being
stream of assays would be performed is dependent on tested. A crucial aspect of kinase assay development is the
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the information available about the disease target and the selection of an appropriate “readout” involving inhibition
availability of specific library against that target. Eventually, of ligand-receptor complex formation, reduction of
all primary hit compounds that pass through Step I enzymatic activity, or change in cellular phenotype, which
(primary screening) proceed to Step II for cell-based assays, further depend on several factors such as the amount of
where in vitro pharmacokinetics and pharmacodynamics enzymes, the type of cell lines, the type of antibodies, and
(bioavailability, solubility, permeability, protein binding, the reference compounds. Furthermore, these assays must

Volume 3 Issue 1 (2024) 4 https://doi.org/10.36922/gtm.2448