Synthesis of Triazole-Linked SAM-Adenosine Conjugates: Functionalization of Adenosine at N-1 or N-6 Position without Protecting Groups

Abstract

More than 150 RNA chemical modifications have been identified to date. Among them, methylation of adenosine at the N-6 position (m6A) is crucial for RNA metabolism, stability and other important biological events. In particular, this is the most abundant mark found in mRNA in mammalian cells. The presence of a methyl group at the N-1 position of adenosine (m1A) is mostly found in ncRNA and mRNA and is mainly responsible for stability and translation fidelity. These modifications are installed by m6A and m1A RNA methyltransferases (RNA MTases), respectively. In human, deregulation of m6A RNA MTases activity is associated with many diseases including cancer. To date, the molecular mechanism involved in the methyl transfer, in particular substrate recognition, remains unclear. We report the synthesis of new SAM-adenosine conjugates containing a triazole linker branched at the N-1 or N-6 position of adenosine. Our methodology does not require protecting groups for the functionalization of adenosine at these two positions. The molecules described here were designed as potential bisubstrate analogues for m6A and m1A RNA MTases that could be further employed for structural studies. This is the first report of compounds mimicking the transition state of the methylation reaction catalyzed by m1A RNA MTases.

Bisubstrate Strategies to Target Methyltransferases

“…As an extension of their previous work on bisubstrate analogues targeting N6A-RNA MTases (Figure 8), the same group described in 2020 the synthesis of novel SAM-adenosine con-jugates 16-19 mimicking the transition state of N1-methylation of adenosine residues. [25] In these bisubstrate analogues, a 1,2,3triazole ring was introduced by a copper(I)-catalyzed alkyneazide cycloaddition (CuAAC) to connect the two adenosines, the RNA substrate mimic at the N1-position (in blue) and the SAM analogue (in green) (Figure 9). These dinucleosides could be used as tools for structural studies on RNA N1-methyltransferases.…”

“…This pocket is surrounded by a few glutamic acid residues, which are highly conserved in the eleven PRMTs identified in humans and important for guanidine binding. The bisubstrate compounds were tested against several PRMTs and lysine methyltransferase G9a to assess their [25] Figure 10. Bisubstrate PRMTs inhibitors 20-21 developed by Arimondo and co-workers.…”

“…The same authors extended the original set of PRMT1 inhibitors by replacing the guanidine with a primary amine or an aryl guanidine or they changed the SAM amino acid chain to [25] Figure 10. Bisubstrate PRMTs inhibitors 20-21 developed by Arimondo and co-workers.…”

“…Figure 9. Transition state of adenosine N1-methylation and bisubstrate inhibitors 16-19 developed by Etheve-Quelquejeu and co-workers [25]. …”

Synthesis of Bisubstrate Analogues for RNA Methylation Studies using two Transition‐Metal‐Catalyzed Reactions

“…Moving forward to explore the impact of the linker of the bisubstrate, we next examined the copper(I) azide–alkyne cycloaddition reaction (CuAAC) to introduce 1,4 substituted‐triazole ring as a linker (Figure 1D). [14] However, compared to the former analogues, these conjugates lack the α‐amino acid motif of SAM.…”