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From dynamic combinatorial 'hit' to lead: in vitro and in vivo activity of compounds targeting the pathogenic RNAs that cause myotonic dystrophy.

Ofori LO, Hoskins J, Nakamori M, Thornton CA, Miller BL - Nucleic Acids Res. (2012)

Bottom Line: We previously used Dynamic Combinatorial Chemistry to identify the first compounds known to inhibit (CUG)-MBNL1 binding in vitro.Introduction of a benzo[g]quinoline substructure previously unknown in the context of RNA recognition, as well as other modifications, provided several molecules with enhanced binding properties, including compounds with strong selectivity for CUG repeats over CAG repeats or CAG-CUG duplex RNA.Compounds readily penetrate cells, and improve luciferase activity in a mouse myoblast assay in which enzyme function is coupled to a release of nuclear CUG-RNA retention.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Rochester, Rochester, NY 14642, USA.

ABSTRACT
The myotonic dystrophies (DM) are human diseases in which the accumulation of toxic RNA (CUG or CCUG) repeats in the cell causes sequestration of splicing factors, including MBNL1, leading to clinical symptoms such as muscle wasting and myotonia. We previously used Dynamic Combinatorial Chemistry to identify the first compounds known to inhibit (CUG)-MBNL1 binding in vitro. We now report transformation of those compounds into structures with activity in vivo. Introduction of a benzo[g]quinoline substructure previously unknown in the context of RNA recognition, as well as other modifications, provided several molecules with enhanced binding properties, including compounds with strong selectivity for CUG repeats over CAG repeats or CAG-CUG duplex RNA. Compounds readily penetrate cells, and improve luciferase activity in a mouse myoblast assay in which enzyme function is coupled to a release of nuclear CUG-RNA retention. Most importantly, two compounds are able to partially restore splicing in a mouse model of DM1.

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Hit compound 1 identified via RBDCC and molecules (2–11) synthesized in this work.
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gks298-F1: Hit compound 1 identified via RBDCC and molecules (2–11) synthesized in this work.

Mentions: RBDCC hit compound 1 (Figure 1) and related molecules identified in our initial work provided a useful demonstration of feasibility, and set the stage for building toward a compound that would be suitable for further evaluation in the biological context. To accomplish that goal, we anticipated that replacing the disulfide bridge with an olefin bioisostere would not have a dramatic impact on affinity, based on results from parallel efforts in our lab targeting an RNA sequence involved in regulating −1 ribosomal frameshifting in HIV (20). Since disulfides are easily reduced in the cytoplasm, replacing the disulfide with an olefin or alkane would facilitate cellular studies. Second, molecules containing hydrocarbon bridges of varied length would allow us to examine the effect of linker length and configuration on binding ability and selectivity. Third, we wished to explicitly examine the importance of the amino acid sequence order. Finally, as quinolines are known intercalators, at least in the DNA-binding context (21), we hypothesized that increasing the pi surface area of this group would enhance affinity. In this regard, we were surprised to discover that despite the vast amount of research conducted into the nucleic acid recognition properties and biological activity of acridine derivatives, including the use of several acridines in humans as antimicrobials (22) and chemotherapeutic agents (23), we are only aware of one mention of the closely related benzo[g]quinoline heterocycle (i.e. 2, Figure 1) in the nucleic acid recognition literature (24). Thus, synthesizing and testing derivatives incorporating this moiety would constitute the first examination of this heterocycle in the RNA binding context.Figure 1.


From dynamic combinatorial 'hit' to lead: in vitro and in vivo activity of compounds targeting the pathogenic RNAs that cause myotonic dystrophy.

Ofori LO, Hoskins J, Nakamori M, Thornton CA, Miller BL - Nucleic Acids Res. (2012)

Hit compound 1 identified via RBDCC and molecules (2–11) synthesized in this work.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3401475&req=5

gks298-F1: Hit compound 1 identified via RBDCC and molecules (2–11) synthesized in this work.
Mentions: RBDCC hit compound 1 (Figure 1) and related molecules identified in our initial work provided a useful demonstration of feasibility, and set the stage for building toward a compound that would be suitable for further evaluation in the biological context. To accomplish that goal, we anticipated that replacing the disulfide bridge with an olefin bioisostere would not have a dramatic impact on affinity, based on results from parallel efforts in our lab targeting an RNA sequence involved in regulating −1 ribosomal frameshifting in HIV (20). Since disulfides are easily reduced in the cytoplasm, replacing the disulfide with an olefin or alkane would facilitate cellular studies. Second, molecules containing hydrocarbon bridges of varied length would allow us to examine the effect of linker length and configuration on binding ability and selectivity. Third, we wished to explicitly examine the importance of the amino acid sequence order. Finally, as quinolines are known intercalators, at least in the DNA-binding context (21), we hypothesized that increasing the pi surface area of this group would enhance affinity. In this regard, we were surprised to discover that despite the vast amount of research conducted into the nucleic acid recognition properties and biological activity of acridine derivatives, including the use of several acridines in humans as antimicrobials (22) and chemotherapeutic agents (23), we are only aware of one mention of the closely related benzo[g]quinoline heterocycle (i.e. 2, Figure 1) in the nucleic acid recognition literature (24). Thus, synthesizing and testing derivatives incorporating this moiety would constitute the first examination of this heterocycle in the RNA binding context.Figure 1.

Bottom Line: We previously used Dynamic Combinatorial Chemistry to identify the first compounds known to inhibit (CUG)-MBNL1 binding in vitro.Introduction of a benzo[g]quinoline substructure previously unknown in the context of RNA recognition, as well as other modifications, provided several molecules with enhanced binding properties, including compounds with strong selectivity for CUG repeats over CAG repeats or CAG-CUG duplex RNA.Compounds readily penetrate cells, and improve luciferase activity in a mouse myoblast assay in which enzyme function is coupled to a release of nuclear CUG-RNA retention.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Rochester, Rochester, NY 14642, USA.

ABSTRACT
The myotonic dystrophies (DM) are human diseases in which the accumulation of toxic RNA (CUG or CCUG) repeats in the cell causes sequestration of splicing factors, including MBNL1, leading to clinical symptoms such as muscle wasting and myotonia. We previously used Dynamic Combinatorial Chemistry to identify the first compounds known to inhibit (CUG)-MBNL1 binding in vitro. We now report transformation of those compounds into structures with activity in vivo. Introduction of a benzo[g]quinoline substructure previously unknown in the context of RNA recognition, as well as other modifications, provided several molecules with enhanced binding properties, including compounds with strong selectivity for CUG repeats over CAG repeats or CAG-CUG duplex RNA. Compounds readily penetrate cells, and improve luciferase activity in a mouse myoblast assay in which enzyme function is coupled to a release of nuclear CUG-RNA retention. Most importantly, two compounds are able to partially restore splicing in a mouse model of DM1.

Show MeSH
Related in: MedlinePlus