Chemical Epigenetic Regulation Secondary Metabolites Derived from Aspergillus sydowii DL1045 with Inhibitory Activities for Protein Tyrosine Phosphatases

Abstract

Protein tyrosine phosphatases (PTPs) are ubiquitous in living organisms and are promising drug targets for cancer, diabetes/obesity, and autoimmune disorders. In this study, a histone deacetylase inhibitor called suberoylanilide hydroxamic acid (SAHA) was added to a culture of marine fungi (Aspergillus sydowii DL1045) to identify potential drug candidates related to PTP inhibition. Then, the profile of the induced metabolites was characterized using an integrated metabolomics strategy. In total, 46% of the total SMs were regulated secondary metabolites (SMs), among which 20 newly biosynthesized metabolites (10% of the total SMs) were identified only in chemical epigenetic regulation (CER) broth. One was identified as a novel compound, and fourteen compounds were identified from Aspergillus sydowii first. SAHA derivatives were also biotransformed by A. sydowii DL1045, and five of these derivatives were identified. Based on the bioassay, some of the newly synthesized metabolites exhibited inhibitory effects on PTPs. The novel compound sydowimide A (A11) inhibited Src homology region 2 domain-containing phosphatase-1 (SHP1), T-cell protein tyrosine phosphatase (TCPTP) and leukocyte common antigen (CD45), with IC₅₀ values of 1.5, 2.4 and 18.83 μM, respectively. Diorcinol (A3) displayed the strongest inhibitory effect on SHP1, with an IC₅₀ value of 0.96 μM. The structure-activity relationship analysis and docking studies of A3 analogs indicated that the substitution of the carboxyl group reduced the activity of A3. Research has demonstrated that CER positively impacts changes in the secondary metabolic patterns of A. sydowii DL1045. The compounds produced through this approach will provide valuable insights for the creation and advancement of novel drug candidates related to PTP inhibition.

Keywords: Aspergillus sydowii; chemical epigenetic regulation (CER); protein tyrosine phosphatases (PTPs); secondary metabolites (SMs).

Figure 1

Fermentation broth and mycelial morphology of A. sydowii DL1045 cultivated with 250 μM SAHA (b,d) and in control culture (a,c) on 10 days.

Figure 2

HPLC profile for the extracts from A. sydowii DL1045 grown in the presence of (b) 250 μM SAHA and (a) in control culture was analyzed based on UV absorption at 254 nm.

Figure 3

The heatmap and PLS-DA of metabolomics of control and 250 μM suberoylanilide hydroxamic acid (SAHA)-regulated A. sydowii DL1045. (a) Hierarchical clustering analysis (HCA) was conducted on the 200 most variably significant features depicted on a heatmap with a gradient from red (high abundance) to blue (low abundance). (b) PLS-DA scores plot results based on the LC-MS data obtained for groups of SAHA and control. (c) PLS-DA loading plot of all metabolite features. (d) PLS-DA VIP score of the top features. The compound numbers in the brackets are newly bio-synthetized metabolites identified in this study.

Figure 4

MS/MS spectrum of A4 (m/z 235.1692) in positive mode.

Figure 5

The key COSY, HMBC of A11a.

Figure 6

The interconverting geometric isomers of sydowimide A (A11).

Figure 7

Experimental and calculated electronic circular dichroism (ECD) spectra of sydowimide A (A11) (black, experimental in MeOH; red, calculated in MeOH).

Figure 8

Structures of the isolated metabolites for structural identification.

Figure 9

Global natural product social molecular network (GNPS) enabled the discovery of suberoylanilide hydroxamic acid (SAHA) derivatives. (a). Molecular network of compounds in different cultures (red in SAHA culture, purple in control culture and yellow in both cultures); (b). a cluster including 5 SAHA derivatives (m/z 249.160, 250.144, 264.159, 356.180, 496.282).

Figure 10

Representation of the binding mode of compounds A3 (a,c) and A13 (b,d) in the active site and the inactive site of Src homology region 2 domain-containing phosphatase-1 (SHP1).