Fragment‐Based Drug Discovery for RNA Targets

Lundquist, Kasper P.; Panchal, Vipul; Gotfredsen, Charlotte Held; Brenk, Ruth; Clausen, Mads Hartvig

doi:10.1002/cmdc.202100324

Cited by 20 publications

(14 citation statements)

References 76 publications

(309 reference statements)

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“…While structure-based computational methods like molecular docking are regularly used for protein targets, most identified RNA-binding small molecules, however, emerged from high-throughput or fragment-screening campaigns and derivatization of a known ligand or serendipity rather than structure-based design by molecular docking. ,− One challenge in this regard is the low number of available 3D RNA structures compared to the number of proteins deposited in the protein data bank (PDB), often additionally struggling with low resolution. However, these structures already show clearly that the limited chemical diversity of just four RNA bases compared to 20 canonical amino acids in proteins can still result in many different folds, for example, hairpins, G-quadruplexes, multiway junctions, (pseudo)knots, L-shaped tRNAs, triple helices, ribozymes, and the ribosome complex .…”

Section: Introductionmentioning

confidence: 99%

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ₁-Riboswitch

Kallert

Fischer

Schneider

et al. 2022

J. Chem. Inf. Model.

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Targeting RNA with small molecules is an emerging field. While several ligands for different RNA targets are reported, structure-based virtual screenings (VSs) against RNAs are still rare. Here, we elucidated the general capabilities of protein-based docking programs to reproduce native binding modes of small-molecule RNA ligands and to discriminate known binders from decoys by the scoring function. The programs were found to perform similar compared to the RNA-based docking tool rDOCK, and the challenges faced during docking, namely, protomer and tautomer selection, target dynamics, and explicit solvent, do not largely differ from challenges in conventional protein-ligand docking. A prospective VS with the Bacillus subtilis preQ1-riboswitch aptamer domain performed with FRED, HYBRID, and FlexX followed by microscale thermophoresis assays identified six active compounds out of 23 tested VS hits with potencies between 29.5 nM and 11.0 μM. The hits were selected not solely based on their docking score but for resembling key interactions of the native ligand. Therefore, this study demonstrates the general feasibility to perform structure-based VSs against RNA targets, while at the same time it highlights pitfalls and their potential solutions when executing RNA–ligand docking.

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Section: Introductionmentioning

confidence: 99%

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ₁-Riboswitch

Kallert

Fischer

Schneider

et al. 2022

J. Chem. Inf. Model.

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“…We consistently observe that SPR can successfully measure binding affinities of low-molecular weight, weakly binding (fragment) ligands. Fragment screens have been widely adopted, and growing evidence supports that fragment-based screening is well-suited for targeting RNA. ,, The orthogonal methods outlined in the introduction yield valuable information about RNA–ligand binding, including thermodynamics (ITC); RNA structure, ligand engagement, and conformational rearrangements (chemical probing); and atomic-resolution binding modes (NMR). SPR surpasses these methods in throughput and efficient use of material.…”

Section: Discussionmentioning

confidence: 99%

RNA–Ligand Interactions Quantified by Surface Plasmon Resonance with Reference Subtraction

Arney

Weeks

2022

Biochemistry

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Structured RNAs bind ligands and are attractive targets for small-molecule drugs. A wide variety of analytical methods have been used to characterize RNA−ligand interactions, but our experience is that most have significant limitations in terms of material requirements and applicability to complex RNAs. Surface plasmon resonance (SPR) potentially overcomes these limitations, but we find that the standard experimental framework measures notable nonspecific electrostatic-mediated interactions, frustrating analysis of weak RNA binders. SPR measurements are typically quantified relative to a non-target reference channel. Here, we show that referencing to a channel containing a non-binding control RNA enables subtraction of nonspecific binding contributions, allowing measurements of accurate and specific binding affinities. We validated this approach for small-molecule binders of two riboswitch RNAs with affinities ranging from nanomolar to millimolar, including low-molecular-mass fragment ligands. SPR implemented with reference subtraction reliably discriminates specific from nonspecific binding, uses RNA and ligand material efficiently, and enables rapid exploration of the ligand-binding landscape for RNA targets.

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“…Indeed, the assumption from previous studies is that fragments bind their targets with modest affinities (μM to mM) and must be lead-optimized for bioactivity. 13 To study the limits of physicochemical properties of compounds that bind RNA and to test the assertion that fragment-like compounds must be lead optimized for bioactivity, a 2500-member RNA-focused fragment-like library was constructed (molecular weight ≤ 300 Da), selected by possessing chemical features that are found in known RNA binders. These compounds were then tested for their RNAbinding capacity in a massively parallel format to define their binding landscapes.…”

Section: ■ Introductionmentioning

confidence: 99%

Low-Molecular Weight Small Molecules Can Potently Bind RNA and Affect Oncogenic Pathways in Cells

et al. 2022

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RNA is challenging to target with bioactive small molecules, particularly those of low molecular weight that bind with sufficient affinity and specificity. In this report, we developed a platform to address this challenge, affording a novel bioactive interaction. An RNA-focused small-molecule fragment collection (n = 2500) was constructed by analyzing features in all publicly reported compounds that bind RNA, the largest collection of RNA-focused fragments to date. The RNA-binding landscape for each fragment was studied by using a library-versus-library selection with an RNA library displaying a discrete structural element, probing over 12.8 million interactions, the greatest number of interactions between fragments and biomolecules probed experimentally. Mining of this dataset across the human transcriptome defined a drug-like fragment that potently and specifically targeted the microRNA-372 hairpin precursor, inhibiting its processing into the mature, functional microRNA and alleviating invasive and proliferative oncogenic phenotypes in gastric cancer cells. Importantly, this fragment has favorable properties, including an affinity for the RNA target of 300 ± 130 nM, a molecular weight of 273 Da, and quantitative estimate of drug-likeness (QED) score of 0.8. (For comparison, the mean QED of oral medicines is 0.6 ± 0.2). Thus, these studies demonstrate that a low-molecular weight, fragment-like compound can specifically and potently modulate RNA targets.

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Fragment‐Based Drug Discovery for RNA Targets

Cited by 20 publications

References 76 publications

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ₁-Riboswitch

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ₁-Riboswitch

RNA–Ligand Interactions Quantified by Surface Plasmon Resonance with Reference Subtraction

Low-Molecular Weight Small Molecules Can Potently Bind RNA and Affect Oncogenic Pathways in Cells

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Fragment‐Based Drug Discovery for RNA Targets

Cited by 20 publications

References 76 publications

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ1-Riboswitch

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ1-Riboswitch

RNA–Ligand Interactions Quantified by Surface Plasmon Resonance with Reference Subtraction

Low-Molecular Weight Small Molecules Can Potently Bind RNA and Affect Oncogenic Pathways in Cells

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Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ₁-Riboswitch

Protein-Based Virtual Screening Tools Applied for RNA–Ligand Docking Identify New Binders of the preQ₁-Riboswitch