C 21 is a kind of selective protein arginine methyltransferase 1 (PRMT1) inhibitor (IC50 = 1.8 μM). And it exhibits five-fold selectivity for PRMT1 over protein arginine methyltransferase 6 (PRMT6) and >250-fold selectivity over PRMT3 and CARM1.
The experiment results suggest that C 21 is specific for PRMT1, as demonstrated by the fact that the IC50 values for PRMT3 and CARM1 are > 500 μM. The high selectivity is consistent with the fact that the substrate specificities of PRMT1 and -3 differ substantially, and PRMT1 preferentially methylates histone H4, whereas CARM1 preferentially methylates the Nterminus of histone H3. Although C 21 also inhibits PRMT6 with micromolar potency, the compound retains a distinct ~fivefold preference for PRMT1.
Besides, the experiment results indicate that the coactivator activity of PRMT1 is almost completely inhibited even at concentrations as low as 1 μM C 21 while no inhibition of CARM1 activity was observed. The fact that no coactivator activity was observed with the catalytically deficient CARM1 mutant indicates that CARM1 coactivator activity requires its methyltransferase activity in this assay. Therefore, the lack of an effect of C 21 on the coactivator activity of CARM1 is not due to an artifact of the assay.
In comparison to other known PRMT1 inhibitors, such as arginine methyltransferase inhibitor1 (IC50=350 ± 36 μM) and the in situ generated bisubstrate analogue (IC50 = 18.5 ± 4.2 μM), C 21 is the most potent and selective PRMT1 inhibitor described to date. Thus, the development of C 21 is significant. And the fact that C 21 can selectively inhibit PRMT1 activity in cellulo indicates that this compound will be useful in discerning the role of PRMT1 in transcriptional control, RNA splicing and nuclear-cytoplasmic transport. Besides, the covalent nature of the interaction between PRMT1 and C 21 suggests that it will be possible to develop activity-based proteomic probes that selectively modify PRMT1, similarly to those that have been developed for PAD4 and other enzymes involved in human cell signaling. These compounds will be useful for identifying the factors that regulate in vivo PRMT1 activity.
Pharmacokinetics and metabolism
The analyses with C 21 revealed that this compound inactivates PRMT1 in a concentration- and time-dependent fashion. The plots of the pseudo-first-order rate constants of inactivation versus C 21 concentration are saturable and consistent with a two-step mechanism of inactivation. The KI, kinact, and kinact/KI values are ≤ 0.8 ± 0.4 μM, 3.1 ± 0.2 min-1, and 4.6 × 106 min-1 M-1. The specific residue modified by this compound is not known, but the fact that higher substrate concentrations can protect against inactivation, indicates that the enzyme inactivation is due to the modification of an active-site residue.
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