Design,synthesis And Biological Activity Research Of Several Epigenetic Drugs

Epigenetics is the transmission of non-DNA sequence genetic information and does not involve changes in gene sequences.The main research on epigenetics includes DNA methylation,histone modification and histone variation.In recent years,with the deepening of people’s understanding of epigenetics,especially the research on epigenetic-related protein inhibitors,epigenetics has gradually become a hot spot in disease,especially cancer research.Although clinical epigenetics is still in the nascent stage,some epigenetic drugs have been approved for disease diagnosis and treatment.A series of epigenetic markers for prognosis have been gradually discovered.In this paper,histone methyltransferase（DOT1L）,ENL Yeats domain,and ubiquitin-specific protein 7（USP7）were selected as targets.Based on the early-stage virtual screening of the hit compounds,we designed and structurally modified the hit compounds and evaluated the relevant biological activities for the synthesized compounds.The first part of the paper mainly studies histone methyltransferase（DOT1L）inhibitors.We designed and synthesized 33 triazolothiadiazole DOT1L inhibitors,some of which showed significant DOT1L inhibitory activity,such as Model B series compounds 14b and 16a,with IC₅₀ values of 7.37μM and 7.84μM,respectively,which are comparable to the enzyme inhibitory activity of the positive control compound 8.Compound 12j from Model A series has the highest activity with IC₅₀ value of 12.88μM.When the phenyltriazolothiadiazole moiety occupies the SAM binding site,R¹ is 4-N,N-dimethyl,and the molecular tail R² is a hydrophobic group such as an alkyl chain,it may be more suitable for the molecule to be combined with Enzyme binding,and the effect of volume is small.The second part of the paper focuses on novel ubiquitin-specific protein 7（USP7）inhibitors.We designed two classes of USP7 inhibitors:one is pyrimidinones,and the other is2-aminopyridine derivatives.A total of 49 compounds were synthesized in the two series.The activity test results showed that the pyrimidinone compound CH-LJ-16 exhibited moderate USP7 inhibitory activity(IC₅₀=31.89μM),which was slightly higher than that of the selected positive control compound.While all 2-aminopyridine derivatives have 6-90%USP7 inhibitory activity in the range of 50μM,among which compounds CH-LJ-43 and CH-LJ-44 with fluorosulfonyl group have significant USP7 inhibitory activity(IC₅₀ were 17.0 and 31.3μM,respectively).Structure-activity analysis indicated that the potential interaction（such as hydrogen bond）between fluorosulfonyl and USP7 protein may be the main reason for its higher activity.In addition,we also evaluated the binding dissociation constant of compound CH-LJ-49 to USP7,which was calculated as K_D=2.4μM.Molecular simulation docking experiments showed that the binding of CH-LJ-49 to USP7 was exactly the opposite of that of the positive control GNE6640 to USP7,and this flipped binding mode suggested that the binding to Q405in the pocket was the key to maintaining the inhibitory activity of the compound,while the orientation of the amino group of 2-aminopyridine has little effect on the activity,and the existence of the 5-position bicyclic conjugated system is more conducive to the improvement of the inhibitory activity of the compound.The third part of the paper mainly studies ENL Yeats domain inhibitors.We designed and synthesized 39 ENL Yeats benzimidazole inhibitors and tested their biological activities.The results showed that compounds 3,5,12,13,14,23 and 28 had very significant inhibitory activities against ENL with IC₅₀ values below 20 n M when compared to the control compound SGC-i MLLT（1）.Structure-activity relationship（SARs）analysis showed that structural modification of SGC-i MLLT,whether introducing a new group on the indazole（13）or substitution by other large aromatic systems（28）,did not affect the ENL Inhibitory activity.In addition,compounds 13,23 and 28 have obvious inhibitory activities on MOLM-13 cells,with IC₅₀ values of 39.6μM,37.9μM and 8.3μM,respectively,and their inhibitory effect on cell growth is about 9 times stronger than that of SGC-i MLLT.Improving the lipid solubility of benzimidazole inhibitors will be beneficial to the improvement of the anti-leukemia cell activity of these compounds.The above research will contribute to the development of novel high-efficiency and low-toxicity epigenetic drugs.