Design,Synthesis And Anticancer Activity Studies Of Novel Thiopyrano[4,3-d] Pyrimidine Derivatives As PI3K/mTOR Inhibitors

Recent studies have found that the PI3K-Akt-mTOR signaling pathway which contains phosphatidylinositol 3-kinase?PI3K?and its downstream protein kinase B?Akt/PKB?and mammalian target of rapamycin?mTOR?controls many important biological processes in the development process of tumorigenesis.The signaling molecules in PI3K-Akt-mTOR pathway were present higher frequency of genetic variation,which caused the overexpression of PI3K-dependent signaling pathway,leading to tumor cell production and growth.The PI3K-Akt-mTOR signal pathway plays a key role in cell proliferation,migration,survival,and angiogenesis.Therefore,development of the inhibitors of PI3K-Akt-mTOR signal pathway has become one of the research hotspots.Compared with single-target inhibitors,the dual inhibitors of PI3K and mTOR have the advantages such as small dosage,higher efficacy,not easy to produce drug resistance in the anti-tumor test.In this paper,taking BMCL-200908069-1 as a precursor compound,and keeping one of the morpholine ring,according to the skeleton transition and bioisosteric principle,we designed and synthesized 17 new thiopyrano pyrimidine compounds?S-1^S-17?.Further investigations were carried out in details to study the effect of oxidation of sulfur atom on the antitumor activity.Therefore,sulfur was oxidized to sulfone to afford compounds S-18^S-34.On the basis of structure-activity relationship of these compounds,we replaced the aryl structure with active pharmacophore chromone moiety to obtain compounds S-35^S-54.Inspired by the PKI-587,the urea structure was introduced to give compounds S-55^S-68.Based on this,the aryl urea structure was replaced with heterocyclic amide structure to give the novel compounds S-69^S-102.The structures of target compounds were confirmed by 1H-NMR and MS spectra and some compounds also have ¹³C-NMR and HRMS spectra data.All thiopyrano pyrimidine derivatives?S-1^S-102?were tested for their cytotoxic activities on A549,PC-3 and MCF-7 cell lines by the MTT method,using GDC-0941 as positive controls.In order to determine their target,10 preferred compounds were evaluated for their activities against mTOR kinase and PI3K?kinase by the LANCE?Ultra and Kinase-Glo?Luminescent kinase assay,using BMCL-200908069-1 and PI103 as positive controls.The cytotoxicity results show that the majority of target compounds exhibit moderate to excellent cytotoxic activities with IC₅₀ value of 0.52⁸9.5?M.The results of the activity against mTOR kinase and PI3K?kinase indicated that aryl hydrazones thiopyran-pyrimidine compounds?S-2,S-5,S-8,S-19,S-22?were selective inhibitor of mTOR with IC₅₀ value of 0.8-6.93?M.Among them,the most promising compound S-5 showed strong antitumor activities against mTOR kinase with IC₅₀ values of 0.80?M which was 1.71-fold more active than BMCL-200908069-1?1.37?M?.Most chromone hydrazones compounds have good activities against tumor cells?A549,PC-3,MCF-7?,but exhibting moderate activities against mTOR and PI3K?kinase.Only compound S-44 showed good activity against mTOR and PI3K?kinase with IC₅₀ values of 1.1?M and 0.92?M.The thiopyran-pyrimidines compounds bearing the urea unit showed good inhibitory activities against different tumor cells and mTOR and PI3K?kinases.The IC₅₀ values of representative compound S-64 were 0.86?M and 0.69?M against mTOR and PI3K?kinase,which were better than the lead compound BMCL-200908069-1.The anti-tumor activities of these compounds were significantly decreased when the urea structure was replaced by amide structure.According to the results,we preliminary analyzed and discussed the structure-activity relationship of target compounds.We found that the different substituents of aryl hydrazones had a significant impact on the activities and 4-OH substitution produced the best potency.After replacing the benzene with the chromone structure,the activities of target compounds had little changed,but after the introduction of a carboxyl group on chromone structure,the PI3K?kinase activity were enhanced.According the anti-tumor activities and molecular docking results of urea unit-containing compounds,we learn that introduction of urea structure significantly increased the anti-tumor activities of the target compounds.And when the urea structure was replaced by an amide structure,the anti-tumor activities were significantly decreased,which further indicates that the urea structure is essential for anti-tumor activities.Moreover,the results show that morpholino structure play an important role in maintain the activities of these compounds.They can form hydrogen bonds with the amino acid residues of mTOR and PI3K?kinase that make them closely integrated with mTOR and PI3K?kinase.These results of structure-activity relationship could provide ideas and give us new research directions for the further study of PI3K/mTOR inhibitors.