YZL-51N functions as a selective inhibitor of SIRT7 by NAD+ competition to impede DNA damage repair

Summary The NAD+-dependent deacetylase SIRT7 is a pivotal regulator of DNA damage response (DDR) and a promising drug target for developing cancer therapeutics. However, limited progress has been made in SIRT7 modulator discovery. Here, we applied peptide-based deacetylase platforms for SIRT7 enzymatic evaluation and successfully identified a potent SIRT7 inhibitor YZL-51N. We initially isolated bioactive YZL-51N from cockroach (Periplaneta americana) extracts and then developed the de novo synthesis of this compound. Further investigation revealed that YZL-51N impaired SIRT7 enzymatic activities through occupation of the NAD+ binding pocket. YZL-51N attenuated DNA damage repair induced by ionizing radiation (IR) in colorectal cancer cells and exhibited a synergistic anticancer effect when used in combination with etoposide. Overall, our study not only identified YZL-51N as a selective SIRT7 inhibitor from insect resources, but also confirmed its potential use in combined chemo-radiotherapy by interfering in the DNA damage repair process.

A mixture of purified His-SIRT7, H3K18ac peptide, NAD + and serial concentrations of (0-30 mM) NAM were incubated at 37°C for 90 min.Insert: Dose-dependent inhibitory effect of NAM on SIRT7 activity as measured by FDL assay.IC50 value is approximately 2.291 mM in this system.The biotinylated SIRT1/2/6 proteins were immobilized onto the surface of SSA biosensors.0-40 µM YZL-51N were allowed to flow through the chip at room temperature in PBST buffer (pH 7.4).Kinetic parameters and affinities were calculated using Octet Data Analysis software version 7.0 (Fortebio)., where (Dx)1 represents the dose of YZL-51N alone that inhibits the growth of cells by x% and (Dx)2 is the dose of etoposide alone that inhibits the growth of cells by x% (as shown in Table 1).

Figure S4 .
Figure S4.Chemical structure of NAM and inhibitory effects on SIRT7 deacetylase activity, related to Figure 1.

Figure S5 .
Figure S5.The inhibitory effects of compounds on SIRT7 deacetylase activity measured by FDL assays, related to Figure 1.

Figure S8 .
Figure S8.SIRT7 mutations decreased the binding affinity with YZL-51N and impaired its deacetylase activity, related to Figure 4.

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Figure.S9 Molecular docking of YZL-51N and NAD + to SIRT1 and SIRT6 by using AutoDock 4.2, related to Figure 4. (A) Three dimensional structures of ligand-protein prediction showed NAD + (Left in panel) or YZL-51N (Right in panel) bound to SIRT1 and interacted with potential amino acids.(B) NAD + (Left in panel) or YZL-51N (Right in panel) bound to SIRT6 and interacted with potential amino acids.

Figure S11 .
Figure S11.Cellular thermal shift assay used to detect the thermostability of SIRT1 and SIRT6 incubated with or without 20 µM YZL-51N, related to Figure 5.
Figure S13.YZL-51N and IR combination against colorectal cancer cells, related to Figure 7.

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Figure S27. 1 H NMR spectrum of synthetic compound 2 in CDCl3, related to Figure 3.

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Figure S29. 1 H NMR spectrum of synthetic compound 6 in CDCl3, related to Figure 3.