Naphthyridine‐Benzoazaquinolone: Evaluation of a Tricyclic System for the Binding to (CAG)_n Repeat DNA and RNA

Abstract

The expansion of CAG repeats in the human genome causes the neurological disorder Huntington's disease. The small-molecule naphthyridine-azaquinolone NA we reported earlier bound to the CAG/CAG motif in the hairpin structure of the CAG repeat DNA. In order to investigate and improve NA-binding to the CAG repeat DNA and RNA, we conducted systematic structure-binding studies of NA to CAG repeats. Among the five new NA derivatives we synthesized, surface plasmon resonance (SPR) assay showed that all of the derivatives modified from amide linkages in NA to a carbamate linkage failed to bind to CAG repeat DNA and RNA. One derivative, NBzA, modified by incorporating an additional ring to the azaquinolone was found to bind to both d(CAG)9 and r(CAG)9 . NBzA binding to d(CAG)9 was similar to NA binding in terms of large changes in the SPR assay and circular dichroism (CD) as well as pairwise binding, as assessed by electron spray ionization time-of-flight (ESI-TOF) mass spectrometry. For the binding to r(CAG)9 , both NA and NBzA showed stepwise binding in ESI-TOF MS, and NBzA-binding to r(CAG)9 induced more extensive conformational change than NA-binding. The tricyclic system in NBzA did not show significant effects on the binding, selectivity, and translation, but provides a large chemical space for further modification to gain higher affinity and selectivity. These studies revealed that the linker structure in NA and NBzA was suitable for the binding to CAG DNA and RNA, and that the tricyclic benzoazaquinolone did not interfere with the binding.

Understanding the Contributions of Conformational Changes, Thermodynamics, and Kinetics of RNA–Small Molecule Interactions

“…133 However, a single-cycle method and corresponding binding models have been described that can be used for such systems. 135,136 Additionally, standard multicycle experiments can potentially account for incomplete dissociation by adding a corresponding term in the dissociation model. 137 A second challenge facing SPR analysis of RNA-small molecule interactions is related to the requirement to immobilize on the binding partners.…”

“…133,137 Mass-transport limitation can also be mitigated by adjusting the flow rate. 136 When possible, it may be good practice to evaluate RNA-small molecule interactions in both immobilization formats to check for the consistency of data.…”

A Dimeric 2,9‐Diamino‐1,10‐phenanthroline Derivative Improves Alternative Splicing in Myotonic Dystrophy Type 1 Cell and Mouse Models

“…The bivalent effect is proven to be useful to improve the binding affinity of molecules to several multivalent targets, including r(CUG) repeats . According to the success of previous examples, in addition to our own experience in the development of DNA‐ and RNA‐binding molecules, the dimeric molecule DDAP (Figure b), in which two DAP molecules were connected at the primary amino group by an amide linkage, was developed. Herein, we report the synthesis of DDAP , its binding properties to r(CUG) repeats, and its effects on pre‐mRNA splicing in vitro and in vivo.…”

A Ligand That Targets CUG Trinucleotide Repeats

“…The desirable characteristics of small molecules targeting the trinucleotide repeat (TNR) are high affinity and selectivity for the targeted trinucleotide repeat sequence. In previous reports, we developed small molecules selectively binding to the CAG/CAG and CGG/CGG motifs, which appeared in the hairpin secondary structure of CAG and CGG repeats, respectively . Naphthyridine‐azaquinolone NA bound selectively and strongly to the CAG/CAG motif in the hairpin structure of the CAG repeats.…”