Design,synthesis And Anti-tumor Activities Of Heterocycles-fused Pyrimidine PI3K/mTOR Dual Inhibitors

The PI3K-Akt-mTOR signaling pathway plays an important role in the occurrence and development of cancer cells,including mechanisms involved in autophagy,inhibition of apoptosis,and promotion of cell metastasis.There are several inhibitors that affect this signal transduction pathway,but PI3K/mTOR dual inhibitors have the advantages of High efficacy and resistance to drug resistance.Therefore,the development of PI3K/mTOR dual inhibitors is a potential strategy for cancer treatment.According to the previous study of our research group,compound BMC-2014226748-7 has significant mTOR kinase inhibitory activity and weak PI3Kαkinase inhibitory activity.Inspired by GDC-0941,the hydrogen bond between PI3Ka and the receptor PI3Ka after the carbazole skeleton structure transition is essential for the improvement of PI3Ka kinase inhibitory activity of this compound.Therefore,in order to construct carbazole structural analogs,the NH of the thiol fragment in compound BMC-2014226748-7 and the C atom in CH=N-are linked through a 2-atom flexible chain to form a five-membered pyrazole ring,the structure of phenylpyrazole,while introducing different substituents at the 4-position of the phenyl ring,the W₁ series compounds（W-1^W-3）were designed.In order to investigate the influence of the capacity change betweenπ-conjugated systems and push-pull electrons on the stability and solubility of the compounds,the influence of the oxidation of the sulfur atom on the thiopyran ring to the antitumor activity of the compounds was further investigated in the W1 series.Based on this,a total of W₂ series compounds（W-4^W-27）were designed.According to the principle of isosteric displacement,the formed C-H hydrogen bonds can affect the conformation and the binding mode of the molecule and indirectly or directly regulate the interaction between the compound and the target.Therefore,based on the results of the activity screening,on the basis of the compound W-3,the thienopyrimidine structure was replaced with a thianthopyrimidine structure,and a total of W₃ series compounds（W-28^W-35）were designed.Based on the W₃series,in order to examine the effect of the introduction of benzene rings on the3-position of pyrazoline to the stability and solubility of compounds,and to further examine the effect of the parallel mode of thiophene rings and pyrimidine rings on the activity of compounds,a total of W₄ series compounds were designed（W-36^W-59）.Structure-based drug design has revealed that the left-expanded cyclohexane of the thienopyrimidine structure can completely occupy the ATP hydrophobic pocket region,providing the possibility to increase the strength and selectivity of the binding of the compound to the kinase protein receptor,and thus the basis of the W₄ series.We designed the W₅ series compound（W-60^W-67）.In order to observe the effect of the cyclic structure change on the antitumor activity and enzyme activity of the compound,the morpholine structure was extended to an N,N-trimethylethylenediamine structure according to the principle of electron isosteric,and a W₆ series compound was designed（W-68^W-79）.The structures of all target compounds were confirmed by ¹H-NMR and MS spectra.Using MTT colorimetric assay,A549,HepG2,and MCF-7 were used as test cell lines,GDC-0941 was a positive control drug,and 79 target compounds were tested for antitumor activity and structure-activity relationship analysis.The results showed that most of the compounds in the thiopyran-pyridines showed moderate antitumor activity.The results of enzyme activity tests showed that the 5 thianopyrimidine compounds with good antitumor activity exhibited moderate inhibitory activity against PI3Ka kinase,with inhibition rates of 48%to 78%,respectively,and the mTOR kinase inhibitory activity was weak.Among the thienopyrimidines,only the majority of the compounds in the W6 series exhibited moderate to excellent antitumor activity with IC₅₀0 values of 2.08-22.95μM,of which3 compounds had IC₅₀0 values from 7.16μM against PI3Ka kinase inhibitory activity.0.92μM,IC₅₀>10μM for mTOR kinase inhibitory activity.Overall,the inhibitory activity of the above six series of compounds on PI3Kαkinase is much higher than that of mTOR kinase.Different substituents have different inhibitory effects on tumor cells.Among them,W-73 is a potential compound.In addition,further studies on W-73 showed that the compound can inhibit the growth of A549 cells in a time-and concentration-dependent manner,and can induce apoptosis.According to the results of molecular docking model analysis,N on the pyrazoline structure forms hydrogen bonds with the amino acid residues on the PI3K alpha kinase crystal,which makes the compound and protein crystals tightly bound;N,N-trimethylethylenediamine structure and pro Water-based amino acid residues form hydrogen bonds that will moderately increase drug solubility.The 5-position phenyl ring structure is in a hydrophobic cavity that allows the compound to bind tightly to the receptor.These structures all play an important role in maintaining the activity of the compounds.The analysis of the structure-activity relationship provides optimization and transformation ideas for the study of the PI3K/mTOR dual inhibitors,pointing the way for later research.