The Design,Synthesis And Antitumor Activity Studies Of Novel PI3K Inhibitors

The PI3K/AKT signaling pathway,mediated by PI3K,is one of the most important signaling cascades,and play pivotal roles in cellular biological processes,including survival,growth,proliferation,differentiation and apoptosis.Disregulation of this signalling cascade has commonly been observed in a broad range of human cancer.Therefore PI3K is a well established target for anticancer therapy.Three classes of PI3Ks have been identified with the class ? PI3K being of crucial importance in tumorgenesis.The class ? PI3Ks is further divided into four isoforms including PI3K?,? ? and ? based on the differences of the catalytic and regulatory subunits.PI3K? and ? are expressed ubiquitously.PI3K? has been well established as a promising target for cancer therapy due to the prevalent mutation and amplification of PIK3CA in a diverse set of human cancers.Besides the role of activating platelets in thrombotic diseases,PI3KP was also demonstrated to play a key role in the tumorigenesis related to PTEN loss or inactivation.PI3K? is highly expressed in cells of hematopoietic origin.Aberrant activation of PI3K? could promote malignant B-cell proliferation and survival in bone marrow and lymph node microenvironments,leading to lymphoma and multiple myeloma.PI3Ky also distribute mainly in myeloid cells and is involved in inflammation and the immune disease.With the exploration for the structure and biological functions of PI3Ks.a lot of PI3K inhibitors with different scaffolds and isoform selectivity had been discovered,which are helpful for the treatment of cancer,inflammation and immune-related diseases.This study is dedicated to discover new PI3K inhibitors,and the research work could be divided into two sections which are summarized as follows:Part 1.BENC-511,a PI3K/AKT signaling pathway inhibitor,exhibited potent anti-proliferation activities against different tumor cell lines.It could block the AKT phosphorylation and induce cancer cell apoptosis.Moreover,BENC-511 showed potent oral activity against myeloma in vivo and displayed great therapeutic efficacy in a PC3-derived prostate cancer model in nude mice.Further studies demonstrated that BENC-511 showed little inhibitory effects on the PI3Ks,the target of it is still undefined.In addition,BENC-511 is not stable and easily decomposed in vivo.In order to find inhibitors with defined target,a class of coumarin derivatives had been designed.The pyridinylurea substituent were attached to the coumarin core,which were expected to act as hydrogen bond donor and/or acceptor to interact with the backbone valine residue in the hinge region of PI3Ks to improve both the potency and isoform selectivity.In the process for the construction of coumarin core,we discovered that the N-Boc group can generate a tert-butyl carbocation under the concentrated H2SO4 conditions,which can act as a rert-butylating agent and underwent the Friedel-Crafts alkylation reaction to generated the rort-butylated coumarins.This is a rather rare reaction.Different alkyl groups had also been attached to the 7-OH of the coumarins in order to investigate their effects on the activities.A total of 36 compounds had been prepared,including the 6-tert-butyl-coumarins(A series),the 7-hydroxy-coumarins with no substitution at the 6-position(B series),and the 7-alkoxyl coumarins(C series).All the compounds were subjected to in vitro antiproliferation evaluation by MTT method.In general,most of the compounds displayed good antiproliferative activities against A549,MCF-7,K562 and Hela cells.Compounds A2,A8,All,A13,B2,B7,B9,B10,B14.C4 and C7 showed better or comparable potency to BENC-511.In series A,the substituents at the urea N atom displayed diverse effects on the inhibition tumor cell growth,the halogenated phenyl,especially the 3,4-dihalogenate phenyl group,was favored for the antiproliferative activities.Compared with A series,the removal of tert-butyl group at the courmarin rings led to an improvement for the B series of compounds.In serie B,the 2,5-dimethoxyphenyl,halogenated phenyl,3,4-dihalogenated phenyl or cyclopropyl substituted urea exhibited potent activities.In series C,the introduction of different alkyl groups at the 7-OH of the coumarin generally led to decreased potency,only compound C8 exhibited better antiproliferative activites than the corresponding B8.Compounds A2,A5,A8,A9 and All were evaluated for the activities against the PI3K?,and the inhibitory rate ranged from 68.9%to 85%at the concentration of 30 ?M.Among them,A2 showed PI3K?/? selectivity and A8 showed PI3K?/?/? selectivity.A8 could block the phosphorylation of AKT,induce the activation of caspase 3 and cleavage of PARP and tumor cell apoptosis.Part 2.Aberrant activation of PI3K? leads to the B-cell malignance such as leukemia and lymphoma.CAL-101 is a potent PI3K8 selective inhibitor that had been approved for the treatment of CLL,FL and SLL.Based on the structure of CAL-101 and the "Scaffold hopping" strategy,the purine portion in CAL-101 was replced by pyrazolo[3,4-d]pyrimidin-4-amine to design novel PI3K8 inhibitors.Aromatic or heteroaromatic groups with hydrogen bond donor/acceptors were attached at C-3 position of pyrazolo[3,4-d]pyrimidine.Meanwhile,o-tolyl(D series),2,6-dimethylphenyl(E series)or cyclohexyl group(F series)was introduced at the 3-position of the quinazolinone to explore their interaction with the hydrophobic pocket.A methylene group was used as a linker between quinazolinone and pyrazolo[3,4-d]pyrimidin-4-amine to make a structural simplification through the removal of the chiral carbon in CAL-101.A total of 19 compounds were prepared.Most of the compounds exhibited excellent activities against PI3K?,and the inhibitory rate reached to 90%at the concentration of 1?M.The 2,6-dimethylphenyl derivatives E4 and E5 showed best activities,with the IC50 value of 8.6 and 8.4 nM,respectively.In general,the substituents at the C-3 position of the pyrazolo[3,4-d]pyrimidine core had much influences on the PI3K? inhibitory activities,the 5-indole or 3,4-dimethoxyphenyl sunstituents was the most favored,the 6-methoxypyridyl,3-quinolyl or 5-benzo[d][1,3]dioxolane substitutent was well tolerated,while the 6-hydroxynaphthyl or m-(trifluoromethoxy)phenyl group led to decreased potency.The phenyl group at the C-3 position of quinazolinone was important for PI3K? inhibition,the 2,6-dimethylphenyl derivatives(E series)showed the most potent activities against PI3K?,and was better than the o-tolyl derivatives(D series),while the cyclohexyl derivatives(F series)exhibited much reduced potency.Morerover,the indole derivative E4 was selective PI3K? inhibitor,the 3,4-dimethoxyphenyl derivative E5 was selective PI3K?/? inhibitor.The isoform selectivities of them were much more significant than CAL-101.It is of note that compound E4 was more than 3630-fold,390-fold and 40-fold selective for PI3K? over PI3K?,? and ?,which is,as far as we know,the most selective PI3K?inhibitor.E4 and E5 also displayed good anti-proliferation activities against SU-DHL-6 cells,with the IC50 of 0.149 and 0.222 ?M,respectively.They might be good lead compounds or drug candidates for further developments.