Design And Synthesis Of Novel Quinazolinones As PI3Kδ Inhibitors And Preliminary Biological Evaluations

Hematological malignancy is a malignant tumor originating from hematopoietic tissue,which features with abnormal increasing or decreasing of white blood cells,increaseing number of eosinophils,and diffusing infiltration of neutrophils.The etiology of hematological malignancies has not yet been fully elucidated,but is related to genetic,immunodeficiency,environmental and chemical drug infections.At present,the main treatment methods are radiation therapy,combined chemotherapy,bone marrow transplantation and surgical resection.Although some pathological symptoms can be improved,traditional cytotoxic drugs are more toxic and have side effects like bone marrow suppression and immune destruction.Therefore,the development of small molecule targeting drugs becomes the main concerns.PI3Kδ is mainly expressed in hematopoietic cells,and is involved in B cell receptor-mediated signaling,which plays an important role in B cell activation and proliferation.PI3Kδ phosphorylates phosphatidylinositol-4,5-bisphosphate(PIP2)to form phosphatidylinositol-3,4,5-triphosphate(PIP3).As a second messenger,PIP3 recognizes the N-terminal domain of protein kinase,phosphorylates at the AKT Ser473 site,and activates the downstream signaling pathways,thereby regulating multiple biological processes of cells.PI3Kδ inhibitors can inhibit the phosphorylation of AKT and downstream signaling proteins and affect tumor cell proliferation.PI3Kδ inhibitors can also inhibit the production of cytokines and inflammatory factors,such as IL-2,IL-4,IL-6,so as to inhibit cell migration and reduce inflammatory infiltration.Therefore,selective PI3Kδ inhibitors is of great significance for the treatment of hematological malignancies and immune cell-mediated diseases.In the previous research,based on the crystal structure of PI3Kδ and the conformational restriction drug design strategy,our group have designed and synthesized a series of novel quinazolinone selective PI3K8 inhibitors and discovered two compounds with better selectivity than the marketed drug CAL-101.In this study,we like to change the hinge region binding elements to design novel PI3Kδ inhibitors.Since 1H-pyrazolo[3,4-b]pyridin-3-amine is appeared in a number of drugs in clinic and those in clinical trials for a wide therapeutic purpose,such as anti-inflammatory,anti-cancer,and anti-depression,we introduced the 3-aminopyrazolopyridine core as a hinge binding group in the inhibitor design,and expected the pyridine nitrogen to form a key hydrogen bonding with the hinge VAL828.In addition,the substituents at the 5-position of 3-aminopyrazolopyrimidine,such as fluorine,bromo,aromatic ring and aromatic heterocyclic ring,are expected to interact with the affinity hydrophobic pocket.Moreover,the phenyl,2,6-dimethylphenyl,trifluoromethyl are introduced at the 3-position of the quinazolinone ring to induce a conformational change of the ATP-binding pocket of the PI3K protein,resulting in the formation of a specificity pocket between Trp760(PI3Kδ numbering)and Met752.The 2-hydroxynicotinonitrile was used to construct the aminopyrazolopyridine core.After the 2-hydroxynicotinonitrile was first converted to 2-chloronicotinonitrile,and then reacted with hydrazine hydrate to generate the 1H-pyrazolo[3,4-b]pyridin-3-amine.Bromination of 2-hydroxynicotinonitrile provided the 5-bromo-2-hydroxynicotinonitrile,which was transformed into 5-bromo-1H-pyrazolo[3,4-b]pyridin-3-amine in a similar way.The 1H-pyrazolo[3,4-b]pyridin-3-amine,5-bromo-1H-pyrazolo[3,4-b]pyridin-3-amine and 5-fluoro-1H-pyrazolo[3,4-b]pyridin-3-amine were acylated at the 3-position and then at the 1-position to give the diacylated 1H-pyrazolo[3,4-b]pyridin-3-amine intermediates,respectively.The 6-fluoro-2-nitrobenzoic acid was reacted with aniline,2,6-dimethylaniline and 2-trifluoromethylaniline to form the corresponding amides.The nitro groups in the amides were reduced to amines and then reacted with 2-cloroacetyl chloride to form the 2-(chloromethyl)-5-fluoro-3-phenylquinazolin-4(3H)-one,2-(chloromethyl)-3-(2,6-dimethylphenyl)-5-fluoroquinazolin-4(3H)-one,and 2-(chloromethyl)-5-fluoro-3-(2-(trifluoromethyl)phenyl)quinazolin-4(3H)-one.The nucelophilic substitution of the quinazolinone chlorides with the the diacylated 1H-pyrazolo[3,4-b]pyridin-3-amine intennediates,respectively,and removing the aceyl group at the pyrazol 1-position simulataneously,generated the corresponding 5-fluoro-4-oxo-3,4-dihydroquinazolin-2-yl)methyl)acetamides.After the removal of the acetyl group at the pyrazol 3-position,the series I compounds were prepared.The Suzuki coupling of N-(5-bromo-1H-pyrazolo[3,4-b]pyridin-3-yl)-N-((3-(2,6-dimethylphenyl)-5-fluoro-4-oxo-3,4-dihydroquinazolin-2-yl)methyl)acetamide and N-(5-bromo-1H-pyrazolo[3,4-b]pyridin-3-yl)-N-((3-(2-trifluoromethylphenyl)-5-fluoro-4-oxo-3,4-dihydroquinazolin-2-yl)methyl)acetamide with a series of arylboronic acids generated the series II compounds.After the removel of the acetyl group in series II compounds,the series III were prepared.In PI3K8 kinase inhibitory assay,most of these compounds showed little inhibitory effects,only compound 20d showed moderate 64.7%inhibitory rate at the concentration of 1 μM.In summary,we designed and synthesized a series of novel quinazolinone derivatives as PI3K8 inhibitors using the 3-aminopyrazolopyrimidine as the hinge binding element.Most of the compounds have no activity on PI3K8,indicating the compounds design should be improved.However,compound 20d has considerable effect on PI3Kδ,which provides a base for further structural modifications.