WO2016127455A1 - Pyrimidine derivative, cytotoxic agent, pharmaceutical composition and use thereof - Google Patents

Abstract

Provided are a pyrimidine derivative as shown by formula (I) and a variety of crystal forms or a pharmaceutically acceptable salt thereof, wherein R₁, R₂ and R₀ are as stated in the description, and are used for preparing a cytotoxic agent and an anti-tumour drug.

Description

Pyrimidine derivatives, cytotoxic agents, pharmaceutical compositions and applications thereof

The PI3K/Akt/mTOR signaling pathway is a very important pathway in the cell signaling pathway, which regulates many important physiological functions of cells, such as cell growth, proliferation, differentiation, migration, survival, angiogenesis, glucose metabolism, and Fluctuations in cell membranes ^{[1, 2]} .

PI3K kinase is a complex family of lipases, including IA, IB, II, III and IV (PI3K-associated kinases). IA type PI3k is activated by receptor tyrosine kinases (RTK), including a regulatory subunit p85 and a catalytic subunit p110. According to different catalytic subunits, IA type PI3k is divided into p110α, β and δ. Subtype. Type IB PI3k has only one p110γ subtype, which includes an a p110γ catalytic subunit and a regulatory subunit of p101, which are activated by G protein-coupled receptors. Type I PI3K can catalyze the phosphorylation of the hydroxyl group at the 3 position of 4,5-phosphatidylinositol (4,5)-bisphosphate (PIP2) to form a signal molecule 3,4,5-phosphatidylinositol triphosphate Salt (phosphatidylinositol (3,4,5)-trisphosphate, PIP3), PIP3 activates downstream PDK1 and Akt kinases to signal transmission. The concentration of PIP3 in cells is also regulated by PTEN (Phosphatase and tensin homolog deleted on chromosome ten). The role of PTEN is to antagonize the activation of downstream pathway by PI3K. Its most important function is to play at the 3 position of phosphoinositide. Dephosphorylation plays a role in dephosphorylation of the signal molecule PIP3 produced by PI3K back to PIP3 to reverse-regulate the PI3K signal ^[1,3,4] . Type II PI3K includes three subtypes of PI3K-C2α, PI3K-C2β and PI3K-C2γ, which are capable of phosphorylating phosphatidylinositol and 4-phosphatidylinositol phosphate but not phosphorylating 4,5-phosphatidylinositol diphosphate Salt (PIP2) ^[5] . Type III PI3K has only one member, Vps34, which phosphorylates phosphatidylinositol to produce 3-phosphatidylinositol phosphate, a function associated with protein transport and autophagy ^[6] . Type IV PI3K contains catalytic regions similar to other PI3K kinases, members of which include mTOR, ATR, ATM, DNA-PK ^[4,7] .

The dysregulation or abnormal activity of the PI3K signaling pathway is closely related to the occurrence of many diseases. Mutations in p110α due to mutations have been found in a variety of tumors, and mutations in the PIK3CA gene encoding p110α have been found in more than 30% of solid tumors ^[7] in 40% of ovarian cancers and 50% of cervical cancers. A mutation in PIK3CA was found ^[8] . P110δ and γ are key enzymes in the leukocyte signaling system and can treat inflammation and autoimmune-related diseases by inhibiting their activity ^[9] . There is evidence that p110β plays a key role in tumorigenesis due to PTEN deletion, and down-regulation of PIK3CB gene encoding p110β in vivo and in vitro inhibits PI3K signaling and inhibits tumor growth ^[10] . In recent years, with the deepening of research on the structure and function of PI3K, the role of PI3K in many diseases has become more and more clear, and PI3K has become a very promising drug target. Therefore, the development of PI3K-targeted inhibitors has become a hot spot in the field of medicinal chemistry and a large number of PI3K inhibitors have been reported, and many compounds have been in clinical research. At present, PI3K inhibitors that have been reported are mainly classified into the following categories:

1. Non-selective type I PI3K inhibitor

LY294002 and wortmannin (structure shown in Figure 1) are the first generation of non-selective type I PI3K inhibitors, both of which inhibit the p110 catalytic subunit. LY294002 is a class of reversible ATP competitive inhibitors developed by Eli Lilly, which not only inhibits type I PI3K but also inhibits mTOR and DNA-PK. Wortmannin is a natural product isolated from fungi that acts by covalent interaction with the Lys802 residue of p110α and the Lys833 residue of p110γ. Due to their toxic side effects, poor pharmacological properties and lack of selectivity, there is no further clinical development ^[11,12] .

XL147 (structure shown in Figure 1) is a new generation of type I PI3K inhibitor with inhibitory activities against p110α, β, δ and γ of IC50=39, 383, 23 and 36 nM and no inhibition of VPS34, DNA-PK and mTOR, respectively. active. In an in vitro tumor cell inhibition assay, XL147 is able to inhibit PI3K signaling in tumor cells, which can cause tumor growth to slow down and contract in preclinical breast, lung, ovarian, and prostate cancer models. XL147 has entered the clinical trial stage, and will be evaluated in the clinical stage for the safety and efficacy of single or combined drugs for solid tumors, lymphoma and metastatic breast cancer ^[1] .

DGC-0941 (structure shown in Figure 1) is a class of orally effective type I PI3K inhibitors with inhibitory activities for p110α, β, δ, and γ of IC50 = 3, 33, 3, and 75 nM, respectively, and tested for inhibition of AKT phosphate in cells. In the test, IC50=28nm ^[11] . GDC-0941 inhibits U87MG, PC3, SKOV-3, IGROV-1, Detroit 562, HCT116, SNUC2B and LoVo cell lines with lower IC50 values. GDC-0941 also plays a very good role in the athymic mouse U87MG and IGROV-1 transplantation models. Currently, GDC-0941 is also in the clinical research stage ^[13] .

BKM120 (structure shown in Figure 1) is a potent type I PI3K inhibitor with inhibitory activities for p110α, β, δ, and γ of IC50=52, 166, 116, and 261nM, respectively, which exhibits for type III and IV PI3K. It has a lower inhibitory activity and is at least 50-fold selective for mTO R and other protein kinases such as HER1, JAK2 and PDK1. BKM120 is effective in reducing the phosphorylation of AKT in both cell and tumor transplantation models. It also inhibits BCR and cell chemotaxis and increases the sensitivity of chronic myeloid leukemia cells to bendamustine and fludarabine phosphate. And the toxicity to normal lymphocytes is small ^[14] . At present, BKM120 is used in the treatment of acute lymphoblastic leukemia, acute myeloid leukemia, combined with INC424 for the treatment of myelofibrosis, and rituximab for the treatment of B-cell lymphoma in the first phase of clinical trials (http://www.clinicaltrials .gov/).

Second, selective PI3K inhibitors

CAL-101 (Idelalisib) (structure shown in Figure 1) is a class of p110δ selective inhibitors developed by Gilead. The inhibitory activities against p110α, β, δ and γ are IC50=820, 565, 89 and 2.5 nM, respectively. The selectivity of p110α for other subtypes is between 40 and 300 times. It has a good clinical effect on many malignant blood diseases such as relapsed or refractory chronic myeloid leukemia, acute myeloid leukemia, non-Hodgkin's lymphoma and multiple myeloma ^[1] . On July 23, 2014, the US FDA approved the launch of Idelalisib (trade name: Zydelig) developed by Gilead, and Idelalisib became the first PI3K inhibitor to be marketed. The FDA approved three indications for Idelalisib: rituximab (Rituxan) in combination with relapsed chronic lymphocytic leukemia (CLL), monotherapy for recurrent follicular B cells, and non-Hodgkin's lymphoma (FL) And recurrent small lymphocytic lymphoma (SLL).

AS-252424 (structure shown in Figure 1) is a class of p110γ selective inhibitors that exhibits more than 300-fold selectivity for other type I PI3K isoforms other than p110γ, and IC50 for p110α, β, δ, and γ, respectively. =935, 20000, 2000 and 33nM. It has low inhibitory activity against serine/threonine and tyrosine kinases (IC50 > 10 μM, 78 and 79 μM, respectively). Although AS-252424 has a relatively high clearance rate (2.3 L/kg*h) and a relatively short oral half-life (t _1/2 =1h) in rats, it is orally administered in a murine model of peritonitis caused by sulphur. Administration of a dose of 10 mg/kg reduced the aggregation of neutrophils by 35% ^[3] .

PIK-75 (structure shown in Figure 1) is a p110α selective inhibitor with IC50 = 0.3 nm for p110α and 130–2800 times for other PI3K subtypes. In the A373, HeLa, A549, MCF7 and MCF7ADR-res cell lines, the anti-proliferation test showed activity below the micromolar level and was also effective in the HeLa human cervical cancer transplantation model test ^[15] .

TG100-115 (structure shown in Figure 1) is a class of selective p110δ and γ inhibitors with inhibitory activities on p110α, β, δ and γ of IC50=1300, 1200, 83 and 235nM, respectively. Most notable is its selectivity, which inhibits the other 136 protein kinases by less than 50% at 1 μM ^[3] . It has a good inhibitory effect on edema and inflammation in myocardial infarction involving vascular endothelial growth factor and platelet activating factor; it has a very important role in promoting endothelial cell mitosis after tissue injury in ischemic brain injury, TG100- 115 has no disruption to this process; it has been reported to be extremely effective in protecting the heart and reducing infarction in animal models of myocardial infarction ^[4] .

Third, PI3K / mTOR dual inhibitor

The imidazoquinazoline compound NVP-BEZ235 (structure shown in Figure 1) is a dual inhibitor of PI3K/mTOR, which has inhibitory activities on p110α, β, δ, and γ and mTOR of IC50=4, 75, 7 and 5 nM and 20.7 nm, respectively. . This compound is effective in blocking the overactive PI3K signal by arresting the cell cycle in the G1 phase, and it is well tolerated. In addition, NVP-BEZ235 also inhibits the abnormal activation of the PI3K/mTOR pathway by the oncogenic mutations of p110α (E545K and H1047R). It has shown promising results in the evaluation of its effectiveness in combination with the traditional cytotoxic agent doxorubicin and vincristine. It is already in the clinical evaluation stage for the treatment of advanced solid tumors and breast cancer ^{[1] ]} .

XL-765 (structure shown in Figure 1) is a dual inhibitor of PI3K/mTOR with activity at low nanomolar levels. Its inhibitory activities against p110α, β, δ and γ and mTOR are IC50=39, 113, 9 and 43 nM, respectively. And 157nm. In a recent study, it was found that a combination of XL-765 and an autophagy-inducing drug can increase its anti-tumor activity in pancreatic cancer. The combination of XL-765 and chemotherapeutic drugs has been in clinical trials, and the data show that patients have a terminal half-life between 2-15 hours with patient-to-patient variability ^[13] . The cessation or contraction of tumor growth was observed in various human cancer cell transplantation models such as prostate cancer, breast cancer, lung cancer, and ovarian cancer in mice. Clinical phase I trial data on solid tumors alone showed that 12 of 83 patients were stable for 16 weeks or more; 7 patients were stable for 24 weeks or more. The most frequently observed side effects were increased liver enzyme activity, rash and gastrointestinal discomfort ^[16] .

GDC-0980 (structure shown in Figure 1) is a morphine derivative which acts as a dual inhibitor of PI3K/mTOR in phase II clinical research by replacing the indazole ring on GDC-0941 with 2-aminopyrimidine. Synthetic, this substitution increases the inhibitory activity of mTOR. GDC-0980 has shown good activity in various cancer cell lines such as prostate cancer, breast cancer and lung cancer, but it has lower activity on melanoma and pancreatic cancer, which may be related to KRAS in these two tumors. Related to BRAF resistance markers. It showed potent inhibition in both PC3 and MCF-7 neo/HER2 cell transplantation models ^[13] .

Wyeth discovered a series of triazole pyrimidine compounds, of which PKI-402 (structure shown in Figure 1) is a dual inhibitor of p110α/mTOR with inhibitory activities on p110α and mTOR of IC50=1.4nM and 1.7nM, respectively. In vitro cell experiments, PKI-402 can induce apoptosis and show high anti-proliferative activity, and has anti-tumor effects in many tumor transplantation models ^[1] . The mechanism of inhibition of PKI-402 production in MDA-MB-361 cell line was found to inhibit phosphorylation of Akt Ser473 and Thr308 in vivo and in vitro, and also inhibit phosphorylation of target molecules downstream of Akt; PKI-402MDA-MB -361 cell line apoptosis is dependent on the caspase-3 channel ^[13] .

Pfizer's published PF-04691502 (structure shown in Figure 1) is a dual inhibitor of p110α/mTOR whose inhibitory activities against p110α and mTOR are IC50=0.57nM and 16nM, respectively, in the in vivo transplantation model of SK103 ovarian cancer with p110α mutation. Highly effective, PF-04691502 has been used in the treatment of solid tumors in clinical trials ^[1] . .

Figure 1 PI3K inhibitor structure

references:

[1] E. Ciraolo, F. Morello and E. Hirsch. Present and Future of PI3K Pathway Inhibition in Cancer: Perspectives and Limitations [J]. Current Medicinal Chemistry, 2011, 18, 2674-2685.

[2] Matthew T. Burger, Mark Knapp, Allan Wagman, et al. Synthesis and in Vitro and in Vivo Evaluation of Phosphoinositide-3-kinase Inhibitors [J]. ACS Med. Chem. Lett. 2011, 2, 34–38 .

[3] Michael K. Ameriks, Jennifer D. Venable. Small Molecule Inhibitors of Phosphoinositide 3-Kinase (PI3K) δ and γ [J]. Current Topics in Medicinal Chemistry, 2009, 9, 738-753.

[4] Amancio Carnero. Novel inhibitors of the PI3K family [J]. Expert Opin. Investig. Drugs (2009) 18 (9): 1265-1277.

[5] Fiona M. Foster, Colin J. Traer, Siemon M. Abraham et al. The phosphoinositide (PI) 3-kinase family [J]. Journal of Cell Science, 2003, 116, 3037-3040.

[6] Williams R, Berndt A, Miller S, et al. Form and flexibility in phosphoinositide 3-kinases [J]. Biochem Soc Trans.2009;37:615–626.

[7] Peng Wu, Yongzhou Hu. Small molecules targeting phosphoinositide 3-kinases [J]. Med. Chem. Commun. 2012, 3, 1337–1355.

[8] Jaclyn LoPiccolo, Gideon M. Blumenthal, Wendy B. Bernstein, et al. Targeting the PI3K/Akt/mTOR pathway: Effective combinations and clinical considerations [J]. Drug Resistance Updates, 2008, 11: 32–50.

[9] Tom Crabbe1, Melanie J Welham, Stephen G Ward. The PI3K inhibitor arsenal: choose your weapon! [J].RENDS in Biochemical Sciences, 2007, 32: 450–456.

[10] Jo-Anne Pinson, Zhaohua Zheng, Michelle S. Miller, et al. L-Aminoacyl-triazine Derivatives Are Isoform-Selective PI3Kβinhibitors That Target Nonconserved Asp862of PI3Kβ [J].ACS Med.Chem.Lett.2013, 4: 206–210.

[11] Timothy A Yap, Michelle D Garrett, Mike I Walton, et al. Targeting the PI3K–AKT–mTOR pathway: progress, pitfalls, and promises [J]. Current Opinion in Pharmacology 2008, 8: 393–412.

[12] Bing-Hua Jiang, Ling-Zhi Liu. PI3K/PTEN Signaling in Angiogenesis and Tumorigenesis [J]. Adv Cancer Res. 2009, 102: 19–65.

[13] Ipsita PAL, Mahitosh MANDAL.PI3K and Akt as molecular targets for cancer therapy: current clinical outcomes [J]. Acta Pharmacologica Sinica, 2012 (9): 1–18.

[14] Jorge J Castillo, Meera iyengar, Benjamin Kuritzky, et al. Isotype-specifc inhibition of the

Phosphatidylinositol-3-kinase pathway in hematologic malignancies[J].OncoTargets and Therapy 2014(7):333–342.

[15] Teather J. Sundstrom, Amy C. Anderson and Dennis L. Wright. Inhibitors of phosphoinositide-3-kinase: a structure-based approach to understanding potency and selectivity [J]. Org. Biomol. Chem., 2009, 7, 840 –850.

[16] Ben Markman1, Rodrigo Dienstmann, Josep Tabernero. Targeting the PI3K/Akt/mTOR Pathway–Beyond Rapalogs [J]. Oncotarget, 2010; 1:530–543.

Summary of the invention

It is an object of the present invention to provide pyrimidine derivatives, PI3K inhibitors, pharmaceutical compositions and their use in antitumor drugs.

The present invention is implemented as follows:

a pyrimidine derivative represented by the formula (I) and a plurality of crystal forms thereof, or a pharmaceutically acceptable salt thereof,

among them,

R ₁ is hydrogen, optionally substituted aryl having C ₁ -C ₆ as a substituent, optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl, acylamino, halogen, in optionally substituted C ₁ -C ₆ linear, branched, cyclic alkyl group, a substituted aryl group, optionally substituted at C ₁ -C ₆ linear, branched, or cyclic alkyl group substituted a pyridyl group of the group, an optionally substituted C ₁ -C ₆ linear, branched, or cyclic alkyl group as a substituent of a sulfone group, optionally substituted C ₁ -C ₆ straight chain, branched or cyclic a sulfonyl group in which an alkyl group is a substituent, and an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group, pyridyl group, halogenated phenyl group, amine group, nitrile group or halogenated alkyl group as a substituent Carbonyl group, optionally substituted C ₁ -C ₆ linear, branched or cyclic alcohol group, ester group, ether group or carboxyl group;

R ₂ is hydrogen,

NHR ₂ ' or NHCONHR ₃ ;

Wherein R ₁₀ is H or an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group; said R ₂ 'is H or a C ₁ -C ₆ linear, branched alkyl group An optionally substituted amide group having a C ₃ -C ₆ cycloalkyl group as a substituent, a trisubstituted amino group having an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group as a substituent An optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group having one, two or three halogen atoms as a substituent, a C ₁ -C ₆ straight or branched alkoxy group, optionally substituted a C ₁ -C ₆ linear, branched or cyclic alcohol group, piperazinyl pyridyl, aminopyridyl, piperazinylaryl or halogen aryl; wherein piperazinyl and amine are a substituted piperazinyl and a substituted amine group optionally substituted with a C ₁ -C ₆ linear, branched or cyclic alkyl group; said R ₃ = optionally substituted C ₁ -C ₆ straight chain, a branched or cyclic alkyl group and an optionally substituted aryl group;

R ₀ is hydrogen, hydroxy or an optionally substituted C1-C6 linear, branched or cyclic alcohol group.

In the present invention, the pyrimidine derivative represented by the formula (I), and various crystalline forms thereof, or a pharmaceutically acceptable salt thereof, wherein

(1) When R ₁ = an optionally substituted aryl group, R ₂ = H, R ₀ = -OH or R ₀ 'OH, wherein R ₀ ' is an optionally substituted C ₁ - C ₆ straight chain, Chain or cyclic alkyl group;

When R ₁ = an optionally substituted aryl group, R ₀ = H,

Or NHR ₂ ', wherein R ₁₀ is H or an optionally substituted C ₁ -C ₆ straight chain, branched chain, or cyclic alkyl group; R ₂ '=H or -CONHR ₃ ; wherein R ₃ =H, a C ₁ -C ₆ straight-chain, branched alkyl group, a C ₃ -C ₆ cycloalkyl group, a trisubstituted amino group having a C ₁ -C ₆ linear or branched alkyl group as a substituent, a halogenated C ₁ - C ₆ straight, branched or cyclic alkyl, dihalo C ₁ -C ₆ straight, branched or cyclic alkyl, trihalo C ₁ -C ₆ straight, branched or cyclic alkyl ,

-OR ₈ , R ₉ OH or

among them,

When X=N, the R ₄ =H,

Or NHR ₆ , wherein R ₅ and R _{6 are} independently a C ₁ -C ₆ linear or branched alkyl group;

When X=C, the R ₄ =H,

Halogen, wherein R7=H, C ₁ -C ₆ alkyl, BOC or

The R ₈ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

The R ₉ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

(2) When R ₀ =H, R ₁ =NHCONHR ₃ ', R ₂ =CONHR ₁₁ , the R ₃ ', R ₁₁ are each independently H, an optionally substituted C ₁ -C ₆ straight chain, Chain or cyclic alkyl group;

(3) When R ₀ = H, R ₁ = H or C ₁ - C ₆ linear, branched alkyl; R ₂ = NHCONHR ₃ , R ₃ is an optionally substituted C ₁ - C ₆ straight chain, branch Chain or cyclic alkyl group;

(4) When R ₀ =H, R ₂ =NHCONHR ₃ , and R ₃ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group,

B _OC ,

-R ₂₄ -CY ₃ ,

-R ₂₅ -CY ₃ , -R ₂₇ -OH,

-R ₃₀ -OR ₃₁ , -R ₃₂ COOH,

among them,

The R ₁₁ , R ₂₀ , R ₂₁ are each independently H or an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

R ₁₂ , R ₁₃ , R _14a , R ₁₆ , R _18a , R ₂₃ , R ₂₄ , R ₂₆ , R ₂₇ , R ₂₈ , R ₂₉ , R ₃₀ , R ₃₁ , R ₃₂ , R ₃₃ , R ₃₄ , Individually independently an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

The R ₁₄ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl or trihalo C ₁ -C ₆ linear, branched, or cyclic alkyl group;

The R ₁₅ is an optionally substituted C ₁ -C ₁₂ straight chain, branched chain, cyclic alkyl group or an optional substitution with a halogen or a C ₁ -C ₆ linear, branched or cyclic alkyl group as a substituent Aromatic hydrocarbon

The R ₁₈ and R _{19 are} independently H, Boc or an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

The Y = F, Cl, Br or I;

The Z is F, Cl, Br, I or hydrogen;

In the present invention, a pyrimidine derivative and a plurality of crystal forms thereof, or a pharmaceutically acceptable salt thereof, wherein

(1) When R ₁ = benzyl, R ₂ = H, R ₀ is a hydroxyl group, methanol, ethanol, propanol, isopropanol or butanol;

When R ₁ = benzyl, R ₀ = H,

NHR ₂ ' or NHCONHR ₃ ;

among them,

The R ₁₀ is H, methyl, ethyl, propyl or butyl;

The R ₂ '=H or CONHR ₃ ;

The R ₃ =H, methyl, ethyl, propyl, isopropyl, butyl, cyclopropyl, dimethylethylamine, fluoroethyl, difluoroethyl, trifluoroethyl ,

-OR ₈ or R ₉ OH;

Wherein, when X=N, the R ₄ =H,

Or NHR ₆ , wherein R ₅ and R _{6 are} independently methyl, ethyl, propyl or butyl;

When X=C, the R ₄ =H,

Or F, where R7=H, BOC or

The R8 is a methyl group, an ethyl group, a propyl group or a butyl group;

The R9 is a methyl group, an ethyl group, a propyl group or a butyl group;

(2) When R ₀ = H, R ₁ =NHCONHR ₃ '; R ₂ =CONHR ₁₁ , each of R ₃ ', R _{11 is} independently methyl, ethyl, propyl, isopropyl or butyl ;

(3) When R ₀ = H, when R ₁ = H, methyl, ethyl, propyl, isopropyl or butyl; R ₂ = NHCONHR ₃ , R ₃ = methyl, ethyl, propyl, Isopropyl or butyl;

(4) When R ₀ = H, R ₂ = NHCONHR ₃ , R ₃ = methyl, ethyl, propyl, isopropyl or butyl,

B _OC ,

-R ₂₄ -CY ₃ ,

-R ₂₅ -CY ₃ , -R ₂₇ -OH,

-R ₃₀ -OR ₃₁ , -R ₃₂ COOH,

The R ₁₀ , R ₁₁ , R ₂₀ , R ₂₁ are each independently H or methyl, ethyl, propyl, isopropyl or butyl;

The R ₁₂ , R ₁₃ , R ₁₆ , R _18a , R ₂₃ , R ₂₄ , R ₂₆ , R ₂₇ , R ₂₈ , R ₂₉ , R ₃₀ , R ₃₁ , R ₃₂ , R ₃₃ , R ₃₄ are each independently Methyl, ethyl, propyl, isopropyl, butyl, isobutyl, cyclopentyl or cyclohexyl;

The R ₁₄ is methyl, ethyl, propyl, isopropyl, butyl, monofluoro C ₁ -C ₄ straight or branched alkyl, difluoro C ₁ -C ₄ straight or branched Alkyl, trifluoromethyl, trifluoroethyl, trifluoropropyl, trifluoroisopropyl, trifluorobutyl or trifluoroisobutyl;

The R ₁₅ is methyl, ethyl, propyl, isopropyl, butyl, isobutyl, cyclopentyl, cyclohexyl, monofluoro C ₁ -C ₄ linear, branched, cyclic alkyl , difluoro C ₁ -C ₄ straight chain, branched chain, cyclic alkyl group, trifluoro C ₁ -C ₄ straight chain, branched chain, cyclic alkyl group, with fluorine or C ₁ -C ₄ straight chain, a branched or cyclic alkyl group is a phenyl group in which a substituent is substituted, ortho, meta or unsubstituted;

The R ₁₈ , R _{19 are} independently H, methyl, ethyl, propyl, isopropyl, butyl, isobutyl or Boc;

The Y=F;

The Z = F.

The pyrimidine derivative of the present invention and various crystal forms thereof or a pharmaceutically acceptable salt thereof are one of the following compounds:

The preparation method of the pyrimidine derivative of the present invention and various crystal forms thereof or a pharmaceutically acceptable salt thereof is prepared from the compound 1, and the preparation method of the compound 1 is as follows:

The cytotoxic agent of the present invention comprises the pyrimidine derivative and various crystal forms thereof or a pharmaceutically acceptable salt thereof.

The pharmaceutical composition of the present invention comprises a therapeutically effective amount of said pyrimidine derivative, and various crystal forms thereof, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

The use of the pyrimidine derivative of the present invention and various crystal forms thereof or a pharmaceutically acceptable salt thereof for the preparation of a cytotoxic agent and an antitumor drug.

The use of the cytotoxic agent of the present invention for the preparation of a medicament for abnormal alteration of PI3K kinase and an antitumor drug.

The use of the pharmaceutical composition of the present invention in the preparation of a medicament for the abnormal alteration of PI3K kinase and an antitumor drug.

In the present invention, the synthetic route of Compound 1 is as follows:

The synthesis of a urea derivative using Compound 1 involves the following two steps:

The general method for the synthesis of urea is as follows:

GL-1 (94 mg, 0.2 mmol) was dissolved in 2 mL anhydrous DCM and triethylamine &lt The mixture was cooled in an ice-salt bath, and added to phosgene (30 mg, 0.1 mmol). After stirring for 15 min in an ice salt bath, 5 eq of amine in anhydrous DCM or amine hydrochloride and 6 eq of triethylamine were added to the above. The reaction was continued overnight in the reaction mixture. Water is added to the reaction and extracted with DCM. The organic phase is washed with water and saturated sodium chloride, and the solvent is evaporated to dryness under reduced pressure. Things.

The synthetic route of piperidine derivatives is as follows:

The methods for preparing piperidine derivatives using GL-26 include the following two methods:

Process A

The GL-26 (100 mg, 0.15 mmol) was suspended in dichloromethane. EtOAc (EtOAc)EtOAc. The organic phase was washed with saturated aqueous sodium chloride and dried over anhydrous sodium sulfate.

Process B

GL-26 (100 mg, 0.15 mmol) was suspended in anhydrous dichloromethane. 3.6 eq of triethylamine was added and the mixture was cooled to 0 ° C. Then, a solution of 1.1 eq of an acid anhydride or an acid chloride in anhydrous dichloromethane was added dropwise to the reaction mixture, and the reaction was further stirred overnight or after the TLC monitoring reaction was completed, saturated sodium hydrogencarbonate and dichloromethane were added, and the organic phase was washed with water and saturated chlorine. The mixture was washed with sodium chloride, dried over anhydrous sodium sulfate and evaporated.

detailed description

Example 1

Synthesis of the compound 3-(1-benzylpiperidin-4-yl)-5-chloro-7-morphocolinylisoxazole [4,5-d]pyrimidine (Compound 1):

first step:

N-Boc-4-piperidinecarboxylic acid (230 mg, 1 mmol) and CDI (292 mg, 1.8 mmol) were mixed in a three-necked flask, air was purged with N ₂ and then 2 mL of anhydrous THF was added, and the reaction mixture was stirred at ambient temperature 1 After 2 h, nitromethane (183 mg, 3 mmol) was added to the reaction mixture and DBU (685 mg, 4.5 mmol) was added. The reaction mixture was stirred for 36 h, then diluted with ethyl acetate, and then added with 6 mL of 2N HCl solution and extracted with ethyl acetate. The organic phase was washed with water until neutral, then washed three times with saturated sodium chloride solution and dried over anhydrous sodium sulfate The ethyl acetate was removed to give a yellow-yellow product (130 mg, yield: 96%). Mp 95-97 ° C. ¹ H NMR (300 MHz, CDCl ₃ ) δ 5.36 (s, 2H), 4.14 (d, J = 13.8 Hz, 2H), 2.86-2.76 (m, 2H), 2.64 (tt, J =11.4, 3.9 Hz, 1H), 1.90–1.85 (m, 2H), 1.68–1.55 (m, 2H), 1.47 (s, 9H). ESI-MS: m/z = 172.94 [M-Boc+H] ⁺ .

The second step:

Compound 1-1 (452 mg, 1.66 mmol) and hydroxylamine hydrochloride (116 mg, 1.67 mmol) and NaHCO ₃ (140 mg, 1.67 mmol) were combined and added to 8 mL of ethanol, and the resulting suspension was reacted at 50 ° C overnight. The reaction was completed by TLC. The reaction mixture was concentrated under reduced pressure and then water and ethyl acetate. The organic phase was washed three times with water and saturated sodium chloride solution and then dried over anhydrous sodium sulfate. 2 (454 mg, cis/trans = 6/1, yield 95%), the product was directly taken to the next step without purification. Mp 147-149 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 11.62 (s, 0.11H), 11.48 (s, 0.71H), 5.35 (s, 0.3H), 5.33 (s, 1.7H) , 3.97 (d, J = 12.9 Hz, 2H), 2.93 - 2.59 (m, 2H), 2.46 (tt, J = 11.7, 3.6 Hz, 1H), 1.80-1.760 (m, 1.75H), 1.63-1.60 ( m, 0.34H), 1.40 (s, 9H), 1.40-1.29 (m, 2H). ESI-MS: m/z = 187.96 [M-Boc+H] ⁺ .

third step:

The compound obtained in the above step 1-2 (8.912 g, 31 mmol) was dissolved in 80 mL of anhydrous diethyl ether. A solution of oxalyl chloride monoethyl ester (3.966 mL, 36 mmol) in anhydrous diethyl ether was added dropwise, and then the mixture was stirred at room temperature for 24 h. The reaction solution was cooled, and triethylamine (3.923 g, 38.8 mmol) was added dropwise at 0 °C. After the dropwise addition, the mixture was stirred at room temperature for 60 hr. Column chromatography under reduced pressure gave white product 1-3 (6.614 g, yield: 58%). Mp 59-61 ° C. ¹ H NMR (300 MHz, CDCl ₃ ) δ 4.52 (q, J = 7.2 Hz, 2H), 4.21. (d, J = 13.2 Hz, 2H), 3.27 (tt, J=11.7, 3.6 Hz, 1H), 2.93 - 2.85 (m, 2H), 2.03 - 1.98 (m, 2H), 1.86 - 1.72 (m, 2H), 1.47 (s, 9H), 1.43 (t, J = 7.2 Hz, 3H) .ESI-MS: m/z = 270.05 [M-Boc+H] ⁺ .

the fourth step:

The compound 1-3 (151 mg, 0.41 mmol) obtained in the previous step was dissolved in 1 mL of methanol in methanol and stirred at room temperature for 3 h. The solvent was evaporated to dryness. Mp 184-186 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.70 (s, 1H), 8.57 (s, 1H), 4.02 (d, J = 12.9 Hz, 2H), 3.38 (tt, J) =11.7, 3.3 Hz, 1H), 2.95-2.86 (m, 2H), 2.01-1.96 (m, 2H), 1.62 - 1.49 (m, 2H), 1.41 (s, 9H). ESI-MS: m/z =241.04[M-Boc+H] ⁺ .

the fifth step:

Compound 1-4 (85 mg, 0.25 mmol) prepared in the previous step was dissolved in 0.5 mL DCM, and 1.5 mL of TFA was added and stirred at room temperature for 2 h. The solvent was evaporated to dryness <RTI ID=0.0></RTI> and the residue was taken twice with ethanol to remove residual TFA to afford pale yellow needle crystals 1-5 (89 mg, yield 100%). Mp 213-215°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.77 (brs, 2H), 8.74 (s, 1H), 8.61 (s, 1H), 3.55 (tt, J = 11.4, 3.3 Hz) , 1H), 3.41-3.37 (m, 2H), 3.14-3.07 (m, 2H), 2.19-2.14 (m, 2H), 1.97-1.83 (m, 2H). ESI-MS: m/z =241.04 [ M+H] ⁺ .

The sixth step:

Compound 1-5 (604 mg, 1.71 mmol) obtained in the previous step was suspended in acetonitrile, and 2 eq of triethylamine was added and stirred at room temperature for 2 hr. The residue was evaporated to dryness <RTI ID=0.0> The residue was taken up in 5 mL of dry EtOAc EtOAc EtOAc (EtOAc) After the completion of the dropwise addition, the reaction was stirred at room temperature, and the mixture was concentrated under reduced pressure by TLC and LC-MS, and then concentrated under reduced pressure, and then chloroform and saturated sodium carbonate solution and chloroform. The organic phase was purified by column chromatography to yield EtOAc (yield: Mp 134-136 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.70 (s, 1H), 8.57 (s, 1H), 7.36 - 7.22 (m, 5H), 3.52 (s, 2H), 3.16 (tt, J = 11.4, 3.3 Hz, 1H), 2.91 (d, J = 11.7 Hz, 2H), 2.09 (t, J = 11.7 Hz, 2H), 1.99-1.95 (m, 2H), 1.79-1.65 ( m, 2H). ESI-MS: m/z = 331.08 [M+H] ⁺ .

Step 7:

Compound 1-6 (73 mg, 0.22 mmol) prepared in the previous step was suspended in 2 mL of a mixed solvent (ethanol: water = 2:1) and cooled to 0 °C. Ammonium chloride (296 mg, 5.53 mmol) was then added, and zinc powder (145 mg, 2.22 mmol) was slowly added portionwise and stirred at room temperature for 4 h. The insoluble material was removed by suction filtration, and the filtrate was evaporated to dryness tolulululu Mp 189-191°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 7.69 (s, 2H), 7.36 (s, 1H), 7.36 - 7.21 (m, 5H), 5.16 (s, 2H), 3.50 (s, 2H), 2.89-2.65 (m, 2H), 2.75 (tt, J = 11.7, 3.6 Hz, 1H), 2.10-2.01 (m, 2H), 1.92-1.87 (m, 2H), 1.72-1.58 (m, 2H). ESI-MS: m/z = 301.08 [M+H] ⁺ .

The eighth step:

The compound 1-7 (60 mg, 0.2 mmol) obtained in the above step was dissolved in 2 mL of anhydrous THF, and then toluene (30 mg, 0.1 mmol). The reaction mixture was cooled to rt. Mp 142-144 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 11.67 (s, 2H), 11.61 (s, 1H), 10.99 (s, 1H), 7.67-7.64 (m, 2H), 7.49 -7.47(m,3H), 4.35(d,J=3.9Hz,2H), 3.46(d,J=11.7Hz,2H),3.23-3.12(m,1H),3.03-2.92(m,2H), 2.30-2.04 (m, 4H). ESI-MS: m/z = 327.11 [M+H] ⁺ .

The ninth step:

A mixture of the compound 1-8 (236 mg, 0.62 mmol) and 6 mL of phosphorus oxychloride obtained in the previous step was refluxed for 24h, then concentrated to dryness under reduced pressure. The aqueous phase was adjusted to pH = 8-9 with aqueous 6N NaOH and extracted with ethyl acetate. The organic phase was washed with water until neutral, washed with EtOAc EtOAc EtOAc EtOAc (EtOAc) A solution of morphin (55 mg, 0.63 mmol) in DCM was added dropwise to the above solution and stirring was continued for 10 min. After the TLC reaction was completed, water was added, and then the organic layer was washed with EtOAc. EtOAc (EtOAc) Mp 168-170 ° C. ¹ H NMR (300 MHz, CDCl ₃ ) δ 7.37 - 7.23 (m, 5H), 4.21 - 3.95 (m, 4H), 3.84 (t, J = 5.1 Hz, 4H), 3.57 (s) , 2H), 3.17-3.07 (m, 1H), 3.02-2.98 (m, 2H), 2.23 - 1.98 (m, 6H). ESI-MS: m/z = 414.18 [M+H] ⁺ .

Example 2

The pyrimidine derivative 4-(3-(1-benzylpiperidin-4-yl)-7-morphinolinylisoxazole [4,5-d] shown below was prepared by Suzuki coupling reaction using Compound 1. Pyrimidin-5-yl)aniline (GL-1):

Compound 1 (414 mg, 1 mmol), p-aminobenzoic acid pinacol ester (329 mg, 1.5 mmol), Pd(Pcy ₃ ) ₂ Cl ₂ (37 mg, 0.05 mmol) and CsF (456 mg, 3 mmol) were placed in a three-neck bottle. The three-necked flask was evacuated and refilled with Ar gas, and repeated three times, and then 12 mL of solvent (NMP: water = 9:1) was injected into the three-necked flask. The reaction mixture was reacted for 48 h at 100 ° C under argon gas, diluted with water and extracted with ethyl acetate. The organic phase was washed with water and saturated sodium chloride and concentrated under reduced pressure. (160 mg, yield 34%). Mp 118-119 ° C. ¹ H NMR (300 MHz, CDCl ₃ ) δ 8.26 - 8.21 (m, 2H), 7.41 - 7.26 (m, 5H), 6.75 - 6.70 (m, 2H), 4.14 - 4.11 (m, 4H), 3.88-3.85 (m, 6H), 3.63 (s, 2H), 3.30-3.13 (m, 1H), 3.07-3.03 (m, 2H), 2.42-2.09 (m, 6H). ESI-MS: m/z = 471.28 [M+H] ⁺ .

Example 3

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-methylurea (GL-2)

According to the above reaction formula, GL-2 is a yellow solid (52 mg, yield 49%) which was synthesized by the synthesis of urea with the corresponding amine hydrochloride. Mp 147-149 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.79 (s, 1H), 8.23 (d, J = 8.7 Hz, 2H), 7.52 (d, J = 8.7 Hz, 2H), 7.35–7.25 (m, 5H), 6.08 (q, J=4.8 Hz, 1H), 4.05-4.03 (m, 4H), 3.81-3.78 (m, 4H), 3.54 (s, 2H), 3.17-3.09 ( m,1H), 2.93 (d, J = 11.4 Hz, 2H), 2.66 (d, J = 4.5 Hz, 3H), 2.22-2.10 (m, 4H), 2.04 - 1.92 (m, 2H). ESI-MS :m/z=528.38[M+H] ⁺ .

Example 4

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-ethylurea (GL-3)

According to the above reaction formula, GL-3 is a yellow solid (53 mg, yield 49%) which was synthesized by the synthesis of urea with the corresponding amine hydrochloride. Mp140-141°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.70 (s, 1H), 8.24-8.20 (m, 2H), 7.52-7.48 (m, 2H), 7.35–7.22 (m, 5H) ), 6.17 (t, J = 5.5 Hz, 1H), 4.05 - 4.02 (m, 4H), 3.80 - 3.77 (m, 4H), 3.53 (s, 2H), 3.17 - 3.08 (m, 3H), 2.94 2.90 (m, 2H), 2.22 - 2.09 (m, 4H), 2.04-1.90 (m, 2H), 1.06 (t, J = 7.2 Hz, 3H). ESI-MS: m/z = 542.39 [M+H ] ⁺ .

Example 5

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)urea (GL-4)

According to the above reaction formula, GL-4 is a yellow solid (46 mg, yield 50%) which was synthesized by the synthesis of urea from a methanol solution of ammonia. Mp204-206°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.80 (s, 1H), 8.25 - 8.22 (m, 2H), 7.54 - 7.50 (m, 2H), 7.35 - 7.23 (m, 5H) ), 5.94 (s, 2H), 4.06-4.03 (m, 4H), 3.81 - 3.78 (m, 4H), 3.54 (s, 2H), 3.18-3.10 (m, 1H), 2.93 (d, J = 11.4) Hz, 2H), 2.23-2.10 (m, 4H), 2.03 - 1.91 (m, 2H). ESI-MS: m/z = 514.39 [M+H] ⁺ .

Example 6

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(2-(dimethylamino)ethyl)urea (GL-5)

According to the above reaction formula, GL-5 is a yellow solid (70 mg, yield 67%) synthesized by the synthesis of the corresponding amine. Mp 96-98 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.94 (s, 1H), 8.23 (d, J = 8.7 Hz, 2H), 7.50 (d, J = 8.7 Hz, 2H), 7.35–7.23 (m, 5H), 6.19 (t, J = 5.4 Hz, 1H), 4.06-4.03 (m, 4H), 3.81-3.78 (m, 4H), 3.54 (s, 2H), 3.20 (q, J=6.0 Hz, 2H), 3.15-3.08 (m, 1H), 2.93 (d, J = 11.1 Hz, 2H), 2.36 (t, J = 6.0 Hz, 2H), 2.20 (s, 6H), 2.20- 2.10 (m, 4H), 2.04 - 1.91 (m, 2H). ESI-MS: m/z = 585.62 [M+H] ⁺ .

Example 7

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-isopropylurea (GL-6)

According to the above reaction formula, GL-6 is a yellow solid (41 mg, yield 37%) which was synthesized by the synthesis of the corresponding amine. Mp 205-206 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.56 (s, 1H), 8.23 (d, J = 8.7 Hz, 2H), 7.49 (d, J = 8.7 Hz, 2H), 7.35–7.23 (m, 5H), 6.07 (d, J=7.5 Hz, 1H), 4.06-4.03 (m, 4H), 3.81-3.74 (m, 5H), 3.54 (s, 2H), 3.14 (tt, J=11.1, 4.2 Hz, 1H), 2.93 (d, J=11.4 Hz, 2H), 2.22–2.10 (m, 4H), 2.04-1.91 (m, 2H), 1.11 (d, J=6.6 Hz, 6H) ). ESI-MS: m/z = 556.38 [M+H] ⁺ .

Example 8

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-cyclopropylurea (GL-7)

According to the above reaction formula, GL-7 is a yellow solid (46 mg, yield 42%) which was synthesized by the synthesis of the corresponding amine. Mp 220-221 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.58 (s, 1H), 8.23 (d, J = 8.7 Hz, 2H), 7.52 (d, J = 8.7 Hz, 2H), 7.35–7.24 (m, 5H), 6.45 (d, J = 2.7 Hz, 1H), 4.06-4.03 (m, 4H), 3.81-3.78 (m, 4H), 3.54 (s, 2H), 3.18-3.10 ( m,1H), 2.93 (d, J = 10.5 Hz, 2H), 2.60-2.53 (m, 1H), 2.22-2.10 (m, 4H), 2.04-1.91 (m, 2H), 0.68 - 0.62 (m, 2H), 0.44 - 0.39 (m, 2H). ESI-MS: m/z = 554.43 [M+H] ⁺ .

Example 9

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(2-fluoroethyl)urea (GL-8)

According to the above reaction formula, GL-8 is a yellow solid (47 mg, yield 42%) which was synthesized by the synthesis of urea with the corresponding amine hydrochloride. Mp 115-117 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.84 (s, 1H), 8.24 (d, J = 9.0 Hz, 2H), 7.52 (d, J = 8.7 Hz, 2H), 7.35 –7.23 (m, 5H), 6.45 (t, J = 5.4 Hz, 1H), 4.56 (t, J = 4.8 Hz, 1H), 4.40 (t, J = 5.1 Hz, 1H), 4.06-4.03 (m, 4H), 3.81-3.78 (m, 4H), 3.54 (s, 2H), 3.47 (q, J = 5.4 Hz, 1H), 3.38 (q, J = 5.4 Hz, 1H), 3.19-3.09 (m, 1H) ), 2.93 (d, J = 11.1 Hz, 2H), 2.22-2.10 (m, 4H), 2.04 - 1.92 (m, 2H). ESI-MS: m/z = 560.38 [M+H] ⁺ .

Example 10

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(6-(4-methylpiperazin-1-yl)pyridin-3-yl)urea (GL-9)

GL-9 is a yellow solid (43 mg, yield 30%) synthesized by the synthesis of urea with the corresponding amine. Mp 144-145 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 9.00 (s, 1H), 8.54 (s, 1H), 8.30-8.26 (m, 2H), 8.16 (d, J = 3.0 Hz) , 1H), 7.71 (dd, J = 9.0, 2.7 Hz, 1H), 7.58-7.53 (m, 2H), 7.36 - 7.25 (m, 5H), 6.83 (d, J = 9.0 Hz, 1H), 4.06- 4.04 (m, 4H), 3.81-3.79 (m, 4H), 3.56 (s, 2H), 3.43 - 3.40 (m, 4H), 3.20-3.12 (m, 1H), 2.97-2.93 (m, 2H), 2.48-2.45 (m, 4H), 2.26 (s, 3H), 2.26-2.12 (m, 4H), 2.05-1.91 (m, 2H). ESI-MS: m/z = 689.78 [M+H] ⁺ .

Example 11

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-5-yl)phenyl by GL-1 )-3-(4-((2-(Dimethylamino)ethyl)(methyl)amino)phenyl)urea (GL-10)

Dissolve N ¹ -(2-(dimethylamino)ethyl)-N ¹ -methyl-1,4-phenylenediamine (116 mg, 0.6 mmol) in 4 mL anhydrous DCM and add triethylamine (182 mg) , 1.8 mmol), the resulting mixture was cooled under ice salt bath. A solution of triphosgene (89 mg, 0.3 mmol) in dry DCM was added dropwise to the reaction mixture and stirring was continued for 15 min. Then, GL-1 (94 mg, 0.2 mmol) of anhydrous DCM was added dropwise to the reaction mixture and the mixture was stirred overnight. The yellow solid GL-10 (50 mg, yield 36%) was purified by column chromatography and preparative chromatography. Mp 134-135 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.81 (s, 1H), 8.33 (s, 1H), 8.27 (d, J = 8.7 Hz, 2H), 7.56 (d, J) = 8.7 Hz, 2H), 7.35 - 7.24 (m, 7H), 6.66 (d, J = 9.3 Hz, 2H), 4.06 - 4.04 (m, 4H), 3.81-3.79 (m, 4H), 3.54 (s, 2H), 3.37 (t, J = 7.2 Hz, 2H), 3.18-3.11 (m, 1H), 2.95-2.91 (m, 2H), 2.86 (s, 3H), 2.38 (t, J = 7.2 Hz, 2H) ), 2, 23-2.11 (m, 4H), 2.20 (s, 6H), 2.05 - 1.93 (m, 2H). ESI-MS: m/z = 690.54 [M+H] ⁺ .

Example 12

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(6-(methylamino)pyridin-3-yl)urea (GL-11)

According to the above reaction formula, GL-11 is a red solid (69 mg, yield 56%) synthesized by the synthesis of the corresponding amine. Mp 149-150 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.91 (s, 1H), 8.29-8.25 (m, 3H), 8.00 (d, J = 3.0 Hz, 1H), 7.58 - 7.50 ( m, 3H), 7.36 - 7.25 (m, 5H), 6.43 (d, J = 8.4 Hz, 1H), 6.24 (q, J = 4.8 Hz, 1H), 4.07 - 4.04 (m, 4H), 3.81-3.78 (m, 4H), 3.55 (s, 2H), 3.19-3.10 (m, 1H), 2.96-2.92 (m, 2H), 2.74 (d, J = 5.1 Hz, 3H), 2.23 - 2.11 (m, 4H) ), 2.04-1.91 (m, 2H). ESI-MS: m/z = 620.55 [M+H] ⁺ .

Example 13

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(pyridin-3-yl)urea (GL-12)

According to the above reaction formula, GL-12 is a yellow solid (43 mg, yield 36%) synthesized by the synthesis of the corresponding amine. Mp 212-213 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 9.14 (s, 1H), 8.97 (s, 1H), 8.62 (d, J = 2.4 Hz, 1H), 8.32-8.28 (m, 2H), 8.21 (dd, J = 4.8, 1.5 Hz, 1H), 7.99-7.95 (m, 1H), 7.61-7.57 (m, 2H), 7.36 - 7.27 (m, 6H), 4.07-4.04 (m, 4H), 3.82-3.79 (m, 4H), 3.56 (s, 2H), 3.25-3.08 (m, 1H), 3.03-2.82 (m, 2H), 2.16-1.98 (m, 6H). ESI-MS: m/z = 591.58 [M+H] ⁺ .

Example 14

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-methoxyurea (GL-13)

According to the above reaction formula, GL-13 is a yellow solid (52 mg, yield 48%) which was synthesized by the synthesis of urea with the corresponding amine hydrochloride. Mp 154-155 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 9.61 (s, 1H), 9.09 (s, 1H), 8.27 (d, J = 9.0 Hz, 2H), 7.73 (d, J = 9.0 Hz, 2H), 7.35 - 7.23 (m, 5H), 4.06-4.04 (m, 4H), 3.81-3.78 (m, 4H), 3.65 (s, 3H), 3.54 (s, 2H), 3.20-3.11 (m, 1H), 2.93 (d, J = 10.8 Hz, 2H), 2.23 - 2.11 (m, 4H), 2.04 - 1.93 (m, 2H). ESI-MS: m/z = 544.33 [M+H] ⁺ .

Example 15

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(2-hydroxyethyl)urea (GL-14)

According to the above reaction formula, GL-14 is a yellow solid (76 mg, yield 68%) which was synthesized by the synthesis of the corresponding amine. Mp 127-129°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.83 (s, 1H), 8.26-8.21 (m, 2H), 7.52-7.48 (m, 2H), 7.35 - 7.23 (m, 5H) ), 6.27 (t, J = 5.4 Hz, 1H), 4.76 (t, J = 5.4 Hz, 1H), 4.06 - 4.03 (m, 4H), 3.81 - 3.78 (m, 4H), 3.54 (s, 2H) , 3.46 (q, J = 5.7 Hz, 2H), 3.18 (q, J = 5.4 Hz, 2H), 3.15 - 3.10 (m, 1H), 2.93 (d, J = 11.4 Hz, 2H), 2.22 - 2.10 ( m, 4H), 2.04-1.93 (m, 2H). ESI-MS: m/z = 558.40 [M+H] ⁺ .

Example 16

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-phenylurea (GL-15)

According to the above reaction formula, GL-15 is a white solid (61 mg, yield 52%) which was synthesized by the synthesis of the corresponding amine. ^{mp201-202 ℃. 1 H NMR (300MHz} , DMSO-d 6) δ8.94 (s, 1H), 8.72 (s, 1H), 8.31-8.28 (m, 2H), 7.60-7.57 (m, 2H), 7.49-7.46 (m, 2H), 7.36 - 7.23 (m, 7H), 6.99 (t, J = 7.5 Hz, 1H), 4.07 - 4.04 (m, 4H), 3.82-3.79 (m, 4H), 3.54 ( s, 2H), 3.18-3.11 (m, 1H), 2.95-2.91 (m, 2H), 2.22-2.11 (m, 4H), 2.05 - 1.92 (m, 2H). ESI-MS: m/z = 590.48 [M+H] ⁺ .

Example 17

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(4-(4-methylpiperazin-1-yl)phenyl)urea (GL-16)

According to the above reaction formula, GL-16 is a yellow solid (95 mg, yield 69%) synthesized by the synthesis of the corresponding amine. Mp 216-218 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.84 (s, 1H), 8.45 (s, 1H), 8.28 (d, J = 8.7 Hz, 2H), 7.56 (d, J = 8.7 Hz, 2H), 7.35 - 7.24 (m, 7H), 6.89 (d, J = 9.0 Hz, 2H), 4.07 - 4.04 (m, 4H), 3.81-3.79 (m, 4H), 3.54 (s, 2H) ), 3.19-3.11 (m, 1H), 3.07–3.04 (m, 4H), 2.93 (d, J = 11.1 Hz, 2H), 2.47–2.44 (m, 4H), 2.22 (s, 3H), 2.22– 2.11 (m, 4H), 2.04-1.93 (m, 2H). ESI-MS: m/z = 688.44 [M+H] ⁺ .

Example 18

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(4-(pyrrolidin-1-yl)phenyl)urea (GL-17)

According to the above reaction formula, GL-17 is a yellow solid (70 mg, yield 53%) synthesized by the synthesis of the corresponding amine. Mp 221-223 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.79 (s, 1H), 8.30 (s, 1H), 8.29-8.25 (m, 2H), 7.57-7.54 (m, 2H), 7.36-7.23 (m, 7H), 6.50 (d, J = 9.3 Hz, 2H), 4.06-4.04 (m, 4H), 3.81-3.78 (m, 4H), 3.54 (s, 2H), 3.21-3.11 ( m,5H), 2.93 (d, J = 11.4 Hz, 2H), 2.23 - 2.11 (m, 4H), 2.04 - 1.92 (m, 6H). ESI-MS: m/z = 659.44 [M+H] ⁺ .

Example 19

Preparation of 4-(4-(3-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole[4,5-d]-pyrimidine--- 5-yl)phenyl)ureido)phenyl)per Pyridin-1-carboxylic acid tert-butyl ester (GL-18)

GL-18 is a yellow solid (78 mg, yield 50%) which was synthesized by the same procedure as GL-10 with the corresponding amine. Mp 156-158 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.86 (s, 1H), 8.49 (s, 1H), 8.28 (d, J = 9.0 Hz, 2H), 7.58 (d, J) = 9.0 Hz, 2H), 7.36 - 7.25 (m, 7H), 6.92 (d, J = 9.3 Hz, 2H), 4.07 - 4.04 (m, 4H), 3.82-3.79 (m, 4H), 3.55 (s, 2H), 3.47-3.44 (m, 4H), 3.19-3.10 (m, 1H), 3.03-2.99 (m, 4H), 2.96-2.92 (m, 2H), 2.23 - 2.12 (m, 4H), 2.05- 1.94 (m, 2H), 1.42 (s, 9H). ESI-MS: m/z = 774.58 [M+H] ⁺ .

Example 20

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-5-yl)phenyl by GL-18 )-3-(4-(piperazin-1-yl)phenyl)urea (GL-19)

GL-18 (45 mg, 0.058 mmol) was dissolved in 0.5 mL of DCM, then 2 mL of TFA was added, and the reaction was stirred at room temperature for 2 h. The solvent was evaporated to dryness. EtOAc was evaporated. The organic phase was concentrated under reduced pressure. Mp 156-158 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 9.04 (s, 1H), 8.67 (s, 1H), 8.27 (d, J = 8.7 Hz, 2H), 7.57 (d, J) =8.7 Hz, 2H), 7.35 - 7.23 (m, 7H), 6.88 (d, J = 9.3 Hz, 2H), 5.52 - 4.31 (brs, 1H), 4.06 - 4.03 (m, 4H), 3.81-3.78 ( m, 4H), 3.54 (s, 2H), 3.19-3.09 (m, 1H), 3.05-3.02 (m, 4H), 2.94 - 2.91 (m, 6H), 2.22 - 2.10 (m, 4H), 2.03 - 1.93 (m, 2H). ESI-MS: m/z = 674.51 [M+H] ⁺ .

Example 21

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(4-fluorophenyl)urea (GL-20)

GL-20 is a yellow solid synthesized by the synthesis of urea with the corresponding amine (68 mg, yield 45%). Mp 179-181 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.95 (s, 2H), 8.77 (s, 2H), 8.29 (d, J = 8.7 Hz, 2H), 7.58 (d, J) =8.7 Hz, 2H), 7.52–7.45 (m, 2H), 7.36–7.23 (m, 5H), 7.18–7.10 (m, 2H), 4.07-4.04 (m, 4H), 3.82-3.79 (m, 4H) ), 3.54 (s, 2H), 3.18-3.11 (m, 1H), 2.95-2.92 (m, 2H), 2.23 - 2.11 (m, 4H), 2.04 - 1.93 (m, 2H). ESI-MS: m /z=608.38[M+H] ⁺ .

Example 22

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(2,2,2-trifluoroethyl)urea (GL-21)

GL-21 is a white solid (81 mg, yield 54%) synthesized by the synthesis of urea with the corresponding amine hydrochloride. Mp 197-198 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 9.02 (s, 1H), 8.28-8.24 (m, 2H), 7.56-7.51 (m, 2H), 7.35 - 7.23 (m, 5H), 6.80 (t, J = 6.6 Hz, 1H), 4.06 - 4.03 (m, 4H), 3.99 - 3.90 (m, 2H), 3.81 - 3.78 (m, 4H), 3.54 (s, 2H), 3.19 -3.09 (m, 1H), 2.93 (d, J = 11.1 Hz, 2H), 2.10 - 2.22 (m, 4H), 2.05 - 1.93 (m, 2H). ESI-MS: m/z = 596.35 [M+ H] ⁺ .

Example 23

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]-pyrimidine by the synthesis of urea by GL-1 5-yl)phenyl)-3-(2,2-difluoroethyl)urea (GL-22)

GL-22 is a white solid (68 mg, yield 49%) which was synthesized by the synthesis of urea with the corresponding amine. Mp 185-186 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.97 (s, 1H), 8.25 (d, J = 9 Hz, 2H), 7.52 (d, J = 8.7 Hz, 2H), 7.42 –7.16(m,5H), 6.55 (t, J=6.0Hz, 1H), 6.27-5.88(m,1H), 4.05–4.03(m,1H),3.81–3.78(m,1H),3.62-3.48 (m, 4H), 3.18-3.10 (m, 1H), 2.95 - 2.91 (m, 2H), 2.10 - 2.22 (m, 4H), 2.04 - 1.19 (m, 2H). ESI-MS: m/z = 578.39[M+H] ⁺ .

Example 24

Preparation of N-isopropyl-4-(5-(4-(3-isopropylureido)phenyl)-7-morphocolinylisoxazole [4,5-d]pyrimidine by GL-1 3-yl) piperidine-1-carboxamide (GL-23)

GL-1 (80 mg, 0.17 mmol) was dissolved in 2 mL of dry DCM and triethylamine (52 mg, 0.51 mmol) Then triphosgene (31 mg, 0.1 mmol) was added and the reaction was stirred under ice bath for 15 min. A solution of 1-methylethylamine (50 mg, 0.85 mmol) in anhydrous DCM was added to the mixture and stirred overnight. Water was added, the mixture was extracted with EtOAc EtOAc m. Mp 223-225 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.59 (s, 1H), 8.23 (d, J = 8.7 Hz, 2H), 7.48 (d, J = 8.7 Hz, 2H), 6.25 (d, J = 7.8 Hz, 1H), 6.08 (d, J = 7.8 Hz, 1H), 4.09-4.05 (m, 6H), 3.81-3.74 (m, 6H), 3.37-3.31 (m, 1H) , 2.91 (t, J = 10.8 Hz, 2H), 2.04-1.83 (m, 4H), 1.11 (d, J = 3.6 Hz, 6H), 1.09 (d, J = 3.6 Hz, 6H). ESI-MS: m/z = 551.46 [M+H] ⁺ .

Example 25

N-methyl-4-(5-(4-(3-methylureido)phenyl)-7-morphocolinylisoxazole [4,5-d]pyrimidin-3-yl)piperidine-1 - Preparation of Formamide (GL-24)

GL-24 is a by-product isolated from the synthesis of GL-2. Mp 174-176 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.80 (s, 1H), 8.23 (d, J = 8.7 Hz, 2H), 7.51 (d, J = 8.7 Hz, 2H), 6.49 (q, J=4.5 Hz, 1H), 6.09 (q, J = 4.8 Hz, 1H), 4.06-4.0 (m, 6H), 3.81-3.78 (m, 4H), 3.40-3.32 (m, 1H) , 2.94 (t, J = 11.7 Hz, 2H), 2.64 (d, J = 4.5 Hz, 3H), 2.62 (d, J = 4.2 Hz, 3H), 2.07-2.02 (m, 2H), 1.94-1.80 ( m, 2H). ESI-MS: m/z = 495.4 [M+H] ⁺ .

Example 26

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-5-yl)phenyl by GL-1 )-3-methylthiourea (GL-25)

GL-1 (141 mg, 0.3 mmol) was dissolved in 3 mL of anhydrous dioxane, and methyl isothiocyanate (24 mg, 0.33 mmol) was added and refluxed for 18 h. Additional methyl isothiocyanate (11 mg, 0.15 mmol) was added and the reaction was refluxed for 24 h. The reaction was concentrated to dryness and water and DCM. The organic phase was washed with EtOAc (EtOAc m.) Mp 175-177 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 9.78 (s, 1H), 8.30 (d, J = 8.7 Hz, 2H), 7.84 (s, 1H), 7.55 (d, J) = 8.4 Hz, 2H), 7.35 - 7.23 (m, 5H), 4.06 - 4.02 (m, 4H), 3.81-3.80 (m, 4H), 3.54 (s, 2H), 3.20 - 3.10 (m, 1H), 2.95 - 2.91 (m, 5H), 2.22 - 1.94 (m, 6H). ESI-MS: m/z = 544.34 [M+H] ⁺ .

Example 27

4-(5-(4-(3-Methylureido)phenyl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-3-yl)piperidine-1-carboxylic acid tert-butyl ester (GL-37) and 1-methyl-3-(4-(7-morpholine-3-(piperidin-4-yl)isoxazole [4,5-d]pyrimidin-5-yl)benzene Preparation of urea) bis-trifluoroacetate (GL-26)

GL-1 (471 mg, 1 mmol) was dissolved in 4 mL DCM and triethylamine (126 mg, 1.25 mmol). A solution of methylcarbamoyl chloride (103 mg, 1.1 mmol) in DCM was slowly added to the mixture and refluxed for 3 d. The organic phase was washed with water and aq.

GL-2 (1.342 g, 2.54 mmol) was dissolved in 40 mL of DCE and then EtOAc-EtOAc (EtOAc. The reaction solution was concentrated to dryness. The methanol was evaporated to dryness under reduced pressure to give crude crystals (1. 50 mL of DCM and triethylamine (936 mg, 9.25 mmol) were added to the obtained crude product and cooled to 0 ° C under ice bath. A solution of (Boc) ₂ O (618 mg, 2.83 mmol) in DCM was added dropwise to the mixture and the mixture was stirred for 3h. After adding water, the mixture was extracted with EtOAc EtOAc EtOAc. Mp 152-154 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.22 (d, J = 9.0 Hz, 2H), 7.50 (d, J = 8.7 Hz, 2H), 6.07 (q, J = 4.5 Hz, 1H), 4.06 - 4.03 (m, 6H), 3.81 - 3.78 (m, 4H), 3.39 (tt, J = 10.8, 3.9 Hz, 1H), 3.14 - 2.86 (m, 2H), 2.66 (d, J = 4.8 Hz, 3H), 2.09-1.84 (m, 4H), 1.45 (s, 9H). ESI-MS: m/z = 538.10 [M+H] ⁺ .

GL-37 (330 mg, 0.61 mmol) was dissolved in 1 mL of DCM, 3 mL of TFA was added, and the reaction was stirred at room temperature for 2 h, evaporated to dryness under reduced pressure, and evaporated to dryness. 405 mg, yield 99%). Mp 211-213 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.89 (s, 1H), 8.80-8.76 (m, 1H), 8.61 - 8.50 (m, 1H), 8.28 (d, J = 8.7 Hz, 2H), 7.53 (d, J = 8.7 Hz, 2H), 6.21 (s, 1H), 4.07-4.04 (m, 4H), 3.82-3.79 (m, 4H), 3.55 (tt, J = 10.8) , 3.9 Hz, 1H), 3.46-3.41 (m, 2H), 3.22-3.12 (m, 2H), 2.66 (s, 3H), 2.34-2.12 (m, 4H). ESI-MS: m/z = 438.62 [M+H] ⁺ .

Example 28

Preparation of 1-(4-(3-(1-ethylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-5-yl)phenyl by GL-26 )-3-methylurea (GL-27)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile, DIPEA (71 mg, 0.55 mmol) was added, and the mixture was cooled in an ice salt bath. Ethyl iodide (29 mg, 0.18 mmol) in acetonitrile was slowly added dropwise to the reaction mixture. The reaction was stirred for 1 h, and the reaction was stirred at room temperature for 5 h. The TLC monitoring was not completed, and the reaction was carried out at 65 ° C for 11 h, and then refluxed for 8 h. Water was added The solvent was evaporated, DCM was extracted, and then extracted with _{DCM / H 2 O = 20/} 1 14 under the conditions of pH =, the organic phase was purified by column chromatography to give a yellow solid GL-27 (48mg, 69% yield). Mp 209-210 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.76 (s, 1H), 8.26 - 8.20 (m, 2H), 7.54 - 7.49 (m, 2H), 6.07 (q, J = 4.8 Hz, 1H), 4.06–4.03 (m, 4H), 3.81–3.78 (m, 4H), 3.15-3.06 (m, 1H), 2.99-2.95 (m, 2H), 2.66 (d, J = 4.8 Hz) , 3H), 2.38 (q, J = 7.2 Hz, 2H), 2.13-1.90 (m, 6H), 1.04 (t, J = 7.2 Hz, 3H). ESI-MS: m/z = 466.74 [M+H ] ⁺ .

Example 29

Preparation of 1-(4-(3-(1-(4-fluorobenzyl)piperidin-4-yl)-7-morphocolinylisoxazole [4,5-d]pyrimidine-5 using GL-26 -yl)phenyl)-3-methylurea (GL-28)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile (EtOAc, EtOAc (EtOAc) The ice salt bath was cooled, and a solution of fluorobromobenzyl bromide (35 mg, 0.18 mmol) in acetonitrile was slowly added dropwise to the reaction mixture. The mixture was stirred for 1 hour, then stirred at room temperature overnight, and the reaction was completed by TLC. After concentrating, water was added, and EtOAc was evaporated. Mp 164-165 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.25 - 8.21 (m, 2H), 7.54 - 7.49 (m, 2H), 7.41 - 7.34 (m, 2H), 7.20–7.12 (m, 2H), 6.08 (q, J=4.2 Hz, 1H), 4.06–4.03 (m, 4H), 3.81–3.78 (m, 4H), 3.52 (s, 2H), 3.13 -3.08 (m, 1H), 2.93-2,89 (m, 2H), 2.67 (d, J = 4.8 Hz, 3H), 2.14-2.10 (m, 4H), 2.04-1.91 (m, 2H). -MS: m/z = 546.44 [M+H] ⁺ .

Example 30

Preparation of 1-methyl-3-(4-(7-morpholine-3-(1-(pyridin-3-methyl)piperidin-4-yl)isoxazole [4,5-d] using GL-26 ]pyrimidin-5-yl)phenyl)urea (GL-29)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile, DIPEA (117 mg, 0.91 mmol) was added, and the mixture was cooled in ice salt, and 3-bromomethylpyridine hydrochloride (46 mg, 0.18 mmol) was added to the reaction mixture. The reaction was stirred for 1 h and stirred at room temperature overnight. The reaction mixture was concentrated under reduced pressure. EtOAc m. Mp 146-148°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.79 (s, 1H), 8.58–8.53 (m, 1H), 8.52–8.46 (m, 1H), 8.28-8.18 (m, 2H), 7.84-7.7 (m, 1H), 7.57-7.46 (m, 2H), 7.43 - 7.33 (m, 1H), 6.09 (q, J = 4.8 Hz, 1H), 4.19 - 3.91 (m, 4H) , 3.90-3.70 (m, 4H), 3.59 (s, 2H), 3.23 - 3.05 (m, 1H), 3.04 - 2.80 (m, 2H), 2.67 (d, J = 4.2 Hz, 3H), 2.22 - 1.92 (m, 6H). ESI-MS: m/z = 529.65 [M+H] ⁺ .

Example 31

Preparation of 1-methyl-3-(4-(3-(1-(methylsulfonyl)piperidin-4-yl)-7-morphinolinylisoxazole [4,5-d] by GL-26 Pyrimidin-5-yl)phenyl)urea (GL-30)

The GL-26 (100 mg, 0.15 mmol) was suspended in anhydrous THF, and triethylamine (76 mg, 0.75 mmol) was added, and the mixture was cooled in an ice bath, and a solution of methanesulfonyl chloride (29 mg, 0.18 mmol) in THF was slowly added dropwise to the reaction mixture. The reaction was further stirred for 1 h and allowed to react at room temperature for 15 h. TLC was not completely reacted and refluxed for 6 h. The solid which precipitated was filtered, and the filtrate was concentrated under reduced pressure, then water and DCM. Mp 261-263 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.25-8.22 (m, 2H), 7.53-7.5 (m, 2H), 6.08 (q, J = 4.5 Hz, 1H), 4.05-4.04 (m, 1H), 3.82-3.80 (m, 2H), 3.69-3.65 (m, 2H), 3.37-3.30 (m, 1H), 3.05-2.98 (m, 2H) , 2.93 (s, 3H), 2.66 (d, J = 4.5 Hz, 3H), 2.29-2.25 (m, 2H), 2.09-1.96 (m, 2H). ESI-MS: m/z = 516.52 [M+ H] ⁺ .

Example 32

Preparation of 1-(4-(3-(1-acetyl-4-yl)-7-morphocolinylisoxazole [4,5-d]pyrimidin-5-yl)phenyl)- using GL-26 3-methylurea (GL-31)

GL-31 is a khaki solid (70 mg, yield 97%) obtained by the procedure B from acetic acid. Mp 240-242 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.25 - 8.21 (m, 2H), 7.54 - 7.49 (m, 2H), 6.08 (q, J = 4.8 Hz, 1H), 4.44 - 4.39 (m, 1H), 4.06 - 4.03 (m, 4H), 3.96 - 3.91 (m, 1H), 3.81 - 3.78 (m, 4H), 3.50 - 3.39 (m, 1H) , 3.35–3.26(m,1H),2.91-2.82(m,1H), 2.66(d,J=4.8Hz,3H),2.22-2.12(m,2H),2.06(s,3H),1.94–1.70 (m, 2H). ESI-MS: m/z = 480.57 [M+H] ⁺ .

Example 33

Preparation of 1-methyl-3-(4-(7-morpholine-3-(1-nicotinopiperidin-4-yl)isoxazole [4,5-d]pyrimidine-5- using GL-26 Phenyl)urea (GL-32)

GL-32 is a yellow solid (58 mg, yield 72%) which was obtained from the procedure Mp 158-160 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.78 (s, 1H), 8.67 (dd, J = 4.5, 1.5 Hz, 2H), 8.26-8.23 (m, 2H), 7.89 (dt, J = 7.8, 1.8 Hz, 1H), 7.53 - 7.48 (m, 3H), 6.08 (q, J = 4.8 Hz, 1H), 4.66 - 4.44 (m, 1H), 4.06 - 4.03 (m, 4H) ), 3.82-3.79 (m, 4H), 3.75-3.61 (m, 1H), 3.57-3.48 (m, 1H), 3.46-3.27 (m, 1H), 3.27-3.06 (m, 1H), 2.66 (d) , J = 4.8 Hz, 3H), 2.39-2.09 (m, 2H), 2.04 - 1.91 (m, 2H). ESI-MS: m/z = 543.46 [M+H] ⁺ .

Example 34

Preparation of 1-(4-(3-(1-(4-fluorobenzoyl)piperidin-4-yl)-7-morphocolinylisoxazole [4,5-d]-pyrimidine- using GL-26 5-yl)phenyl)-3-methylurea (GL-33)

GL-33 is a white solid (57 mg, yield 68%) obtained by the synthesis of p-fluorobenzoic acid chloride. Mp 174-176°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.26-8.23 (m, 2H), 7.55 - 7.50 (m, 4H), 7.33 - 7.27 (m, 2H), 6.08 (q, J = 4.8 Hz, 1H), 4.67-4.30 (m, 1H), 4.07 - 4.04 (m, 4H), 3.91 - 3.61 (m, 5H), 3.60 - 3.44 (m, 1H) , 3.42-3.0 (m, 2H), 2.66 (d, J = 4.5 Hz, 3H), 2.36-2.08 (m, 2H), 2.02 - 1.89 (m, 2H). ESI-MS: m/z = 560.59 [ M+H] ⁺ .

Example 35

Preparation of 1-(4-(3-(1-benzoylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-5-yl)benzene using GL-26 Base)-3-methylurea (GL-34)

GL-34 is a yellow solid (77 mg, yield 95%) obtained from benzoyl chloride by the procedure B. Mp 175-177 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.26-8.22 (m, 2H), 7.54 - 7.50 (m, 2H), 7.49 - 7.42 (m, 5H), 6.08 (q, J=4.5 Hz, 1H), 4.75-4.35 (m, 1H), 4.06–4.03 (m, 4H), 3.91–3.61 (m, 5H), 3.57-3.46 (m, 1H) , 3.42-3.0 (m, 2H), 2.66 (d, J = 4.5 Hz, 3H), 2.36-2.08 (m, 2H), 2.03 - 1.91 (m, 2H). ESI-MS: m/z = 542.70 [ M+H] ⁺ .

Example 36

Preparation of 4-(5-(4-(3-methylureido)phenyl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-3-yl)piperidine-1 using GL-26 -methyl formate (GL-35)

GL-35 was obtained as a white solid (68 mg, yield: 92%). Mp 228-230 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.26-8.20 (m, 2H), 7.54 - 7.49 (m, 2H), 6.08 (q, J = 4.5 Hz, 1H), 4.06–4.03 (m, 6H), 3.81–3.78 (m, 4H), 3.64 (s, 3H), 3.42-3.37 (m, 1H), 3.14-3.05 (m, 2H), 2.66 (d, J = 4.8 Hz, 3H), 2.17-2.11 (m, 2H), 1.93 - 1.80 (m, 2H). ESI-MS: m/z = 496.45 [M+H] ⁺ .

Example 37

Preparation of 4-(5-(4-(3-methylureido)phenyl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-3-yl)piperidine-1 using GL-26 -isopropyl formate (GL-36)

GL-36 is a white solid obtained by the procedure B from isopropyl chloroformate (74 mg, yield 94%). Mp 228-229°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.78 (s, 1H), 8.24-8.20 (m, 2H), 7.53–7.48 (m, 2H), 6.08 (q, J= 4.8 Hz, 1H), 4.83 (sept, J = 6.3 Hz, 1H), 4.09 - 4.03 (m, 6H), 3.81 - 3.78 (m, 4H), 3.44 - 3.35 (m, 1H), 3.11-3.01 (m , 2H), 2.66 (d, J = 4.8 Hz, 3H), 2.13 - 2.07 (m, 2H), 1.95 - 1.82 (m, 2H), 1.23 (d, J = 6.3 Hz, 6H). ESI-MS: m/z = 524.37 [M+H] ⁺ .

Example 38

Preparation of N,N-dimethyl-4-(5-(4-(3-methylureido)phenyl)-7-morphocolinylisoxazole [4,5-d]pyrimidine- using GL-26 3-yl) piperidine-1-carboxamide (GL-38)

GL-38 was obtained as a white solid (75 mg, yield 98%) from dimethylaminocarbonyl chloride. Mp 152-154 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.26-8.21 (m, 2H), 7.53 - 7.49 (m, 2H), 6.08 (q, J = 4.8 Hz, 1H), 4.06–4.03 (m, 4H), 3.81–3.78 (m, 4H), 3.68-3.64 (m, 2H), 3.41-3.30 (m, 1H), 3.89-3.0 (m, 2H) , 2.78 (s, 6H), 2.66 (d, J = 4.5 Hz, 3H), 2.14 - 2.09 (m, 2H), 2.02 - 1.89 (m, 2H). ESI-MS: m/z = 509.34 [M+ H] ⁺ .

Example 39

Preparation of tert-butyl (2-methyl-1-(4-(5-(4-(3-methylureido)phenyl)-7-morpholinylisoxazole [4,5-d] using GL-26 And pyrimidin-3-yl)piperidin-1-yl)-1-oxopropyl-2-yl)carbamate (GL-39)

GL-39 is a yellow-white solid (117 mg, yield 84%) that was synthesized from GL-26 (150 mg, 0.23 mmol) and 2-(tert-butoxycarbonylamino)isobutyric acid. Mp 180-182°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.26-8.21 (m, 2H), 7.53 - 7.49 (m, 2H), 7.35 (s, 1H) , 6.07 (q, J = 4.8 Hz, 1H), 4.50 - 4.54 (m, 1H), 4.06 - 4.03 (m, 4H), 3.81 - 3.78 (m, 4H), 3.40 - 3.37 (m, 1H), 3.24 -2.79 (m, 1H), 2.66 (d, J = 4.5 Hz, 3H), 2.29 - 2.03 (m, 2H), 1.97 - 1.72 (m, 2H), 1.35 (s, 1H). ESI-MS: m /z=623.42[M+H] ⁺ .

Example 40

Preparation of 1-(4-(3-(1-(2-cyanoethyl)piperidin-4-yl)-7-morpholinylisoxazole [4,5-d]pyrimidine-5 using GL-26 -yl)phenyl)-3-methylurea (GL-40)

GL-40 is a yellow solid obtained by the procedure A from cyanoacetic acid (44 mg, yield: 58%). Mp 258-260 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.24 (d, J = 8.7 Hz, 2H), 7.51 (d, J = 8.7 Hz, 2H), 6.07 (q, J=4.8 Hz, 1H), 4.46-4.3 (m, 1H), 4.20-4.04 (m, 6H), 3.90-3.68 (m, 5H), 3.51–3.44 (m, 1H), 3.35– 3.27 (m, 1H), 3.01-2.94 (m, 1H), 2.66 (d, J = 4.5 Hz, 3H), 2.21-2.17 (m, 2H), 2.08-1.94 (m, 1H), 1.86-1.73 ( m, 1H). ESI-MS: m/z = 505.72 [M+H] ⁺ .

Example 41

Preparation of 1-methyl-3-(4-(7-morpholine-3-(1-(2,2,2-trifluoroethyl)piperidin-4-yl)isoxazole from GL-26 [4 ,5-d]pyrimidin-5-yl)phenyl)urea (GL-41)

GL-26 (100 mg, 0.15 mmol) was suspended in ethanol, and DIPEA (71 mg, 0.55 mmol) and trifluoroethyl trifluoromethanesulfonate (51 mg, 0.22 mmol) were added, and the reaction was refluxed for 12 h. Further, DIPEA (23 mg, 0.18 mmol) and trifluoroethyl trifluoromethanesulfonate (21 mg, 0.09 mmol) were added and the mixture was refluxed for 5 h. The reaction mixture was concentrated under reduced pressure. EtOAc m. Mp 229-230 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.25-8.21 (m, 2H), 7.54 - 7.50 (m, 2H), 6.07 (q, J = 4.8 Hz, 1H), 4.06–4.03 (m, 4H), 3.81–3.78 (m, 4H), 3.30-3.20 (q, J = 10.2 Hz, 2H), 3.18-3.11 (m, 1H), 3.08-3.02 (m, 2H), 2.66 (d, J = 4.8 Hz, 3H), 2.63 - 2.54 (m, 2H), 2.15 - 2.09 (m, 2H), 2.05-1.92 (m, 2H). ESI-MS: m /z=520.41[M+H] ⁺ .

Example 42

Preparation of 1-methyl-3-(4-(3-(1-(2-(methylsulfonyl)ethyl)piperidin-4-yl)-7-morpholinolinylisoxazole using GL-26 [4] ,5-d]pyrimidin-5-yl)phenyl)urea (GL-42)

GL-26 (100 mg, 0.15 mmol) was suspended in n-BuOH, DIPEA (71 mg, 0.55 mmol), 2-methylsulfonyl bromide (41 mg, 0.22 mmol) and NaI (41 mg, 0.27 mmol) After stirring, the reaction was completed by TLC, and filtered to give a pale yellow solid (70 mg, yield: 85%). Mp 227-228°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 9.23 (s, 1H), 8.22-8.19 (m, 2H), 7.54-7.51 (m, 2H), 6.44 (q, J = 4.5 Hz, 1H), 4.06–4.03 (m, 4H), 3.81–3.78 (m, 4H), 3.36–3.32 (t, J=6.3 Hz, 2H), 3.19–3.07 (m, 1H), 3.09 (s) , 3H), 3.04-3.0 (m, 2H), 2.78 (t, J = 6.6 Hz, 2H), 2.66 (d, J = 4.5 Hz, 3H), 2.26-2.19 (m, 2H), 2.12-1.95 ( m, 4H). ESI-MS: m/z = 544.57 [M+H] ⁺ .

Example 43

Preparation of 1-methyl-3-(4-(7-morpholine-3-(1-trifluoroacetylpiperidin-4-yl)isoxazole [4,5-d]pyrimidine- using GL-26 5-yl)phenyl)urea (GL-43)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile, and triethylamine was dissolved. The mixture was concentrated to dryness under reduced pressure. After stirring at room temperature overnight, the reaction was not completed by TLC, and the additional 1.2 eq of TFAA was immediately clarified. The reaction mixture was poured with water, EtOAc EtOAc m. Mp 203-205 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.24 - 8.21 (m, 2H), 7.53-7.50 (m, 2H), 6.07 (q, J = 4.8 Hz, 1H), 4.39-4.32 (m, 1H), 4.06–3.98 (m, 5H), 3.81–3.78 (m, 4H), 3.64–3.52 (m, 2H), 3.30–3.22 (m, 1H) , 2.66 (d, J = 4.5 Hz, 3H), 2.36-2.27 (m, 2H), 2.08-1.86 (m, 2H). ESI-MS: m/z = 534.59 [M+H] ⁺ .

Example 44

Preparation of 1-(4-(3-(1-(2-amino-2-methylpropanoyl)piperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d] using GL-39 ]pyrimidin-5-yl)phenyl)-3-methylurea trifluoroacetate (GL-44)

GL-39 (100 mg, 0.16 mmol) was dissolved in DCM, 2 mL of TFA was added, and the reaction was stirred at room temperature for 2 h, concentrated to dryness, and then concentrated to ethyl ether. %). Mp 178-180 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.88 (s, 1H), 8.24 - 8.21 (m, 2H), 8.18 (s, 3H), 7.54 - 7.51 (m, 2H) , 6.20 (q, J = 4.8 Hz, 1H), 4.49 - 4.20 (m, 2H), 4.07 - 4.04 (m, 4H), 3.81 - 3.79 (m, 4H), 3.61 - 3.52 (m, 1H), 3.40 -3.03 (m, 2H), 2.66 (d, J = 4.5 Hz, 3H), 2.24-2.20 (m, 2H), 2.00-1.98 (m, 2H), 1.62 (s, 6H). ESI-MS: m /z=523.74[M+H] ⁺ .

Example 45

Preparation of N,N-dimethyl-2-(4-(5-(4-(3-methylureido)phenyl)-7-morphocolinylisoxazole [4,5-d] using GL-26 And pyrimidin-3-yl)piperidin-1-yl)acetamide (GL-45)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile, DIPEA (71 mg, 0.55 mmol) and 2-chloro-N,N-dimethylacetamide (25 mg, 0.21 mmol). (27 mg, 0.18 mmol) was stirred for 10 h. Concentration under reduced pressure, water and EtOAc (EtOAc)EtOAc. Mp 159-160 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.24 - 8.21 (m, 2H), 7.53-7.50 (m, 2H), 6.07 (q, J = 4.5 Hz, 1H), 4.15-3.94 (m, 4H), 3.90–3.66 (m, 4H), 3.20 (s, 1H), 3.16–3.09 (m, 1H), 3.07 (s, 3H), 2.97-2.93 (m, 2H), 2.83 (s, 3H), 2.66 (d, J = 4.2 Hz, 3H), 2.33 - 2.26 (m, 2H), 2.11-1.94 (m, 4H). ESI-MS: m/z =523.80[M+H] ⁺ .

Example 46

Preparation of 2-(4-(5-(4-(3-methylureido)phenyl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-3-yl)piperidin by GL-26 Pyridin-1-yl)acetamide (GL-46)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile, and DIPEA (71 mg, 0.55 mmol) and 2-iodoacetamide (37 mg, 0.20 mmol) were added and the mixture was stirred at room temperature for 8 h. The reaction mixture was concentrated under reduced pressure. EtOAc EtOAc m. Mp 186-188°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.99 (s, 1H), 8.26-8.20 (m, 2H), 7.53-7.50 (m, 2H), 7.24 (s, 1H) , 7.13 (s, 1H), 6.25 (q, J = 4.8 Hz, 1H), 4.06 - 4.04 (m, 4H), 3.81 - 3.78 (m, 4H), 3.15 - 3.09 (m, 1H), 2.96 - 2.92 (m, 4H), 2.66 (d, J = 4.8 Hz, 3H), 2.35-2.26 (m, 2H), 2.14 - 1.99 (m, 4H). ESI-MS: m/z = 495.70 [M+H] ⁺ .

Example 47

Preparation of 1-(4-(3-(1-(2-hydroxyethyl)piperidin-4-yl)-7-morpholinylisoxazole [4,5-d]pyrimidine-5 using GL-26 -yl)phenyl)-3-methylurea (GL-47)

GL-26 (100 mg, 0.15 mmol) was suspended in ethanol, and DIPEA (71 mg, 0.55 mmol), NaI (27 mg, 0.18 mmol) and 2-bromoethanol (25 mg, 0.20 mmol) were added and refluxed for 60 h. The organic layer was concentrated under reduced pressure. EtOAc (EtOAc). Mp 198-199 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.24-8.21 (m, 2H), 7.53-7.50 (m, 2H), 6.07 (q, J = 4.5 Hz, 1H), 4.41 (t, J = 5.1 Hz, 1H), 4.06-4.03 (m, 4H), 3.81 - 3.78 (m, 4H), 3.54 (q, J = 6.0 Hz, 2H), 3.11 - 3.06 (m, 1H), 3.01-2.97 (m, 2H), 2.66 (d, J = 4.8 Hz, 3H), 2.45 (t, J = 6.3 Hz, 2H), 2.23-2.16 (m, 2H), 2.11 -1.92 (m, 4H). ESI-MS: m/z = 482.70 [M+H] ⁺ .

Example 48

Preparation of 2-(4-(5-(4-(3-methylureido)phenyl)-7-morpholinylisoxazole [4,5-d]pyrimidin-3-yl)piperidin by GL-26 Acetyl-1-yl)acetic acid ethyl ester (GL-48)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile, and DIPEA (71 mg, 0.55 mmol) and ethyl bromoacetate (36 mg, 0.22 mmol). The reaction mixture was concentrated under reduced pressure. EtOAc m. Mp 221-223 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.25-8.21 (m, 2H), 7.54-7.49 (m, 2H), 6.07 (q, J = 4.8 Hz, 1H), 4.11 (q, J = 7.2 Hz, 2H), 4.06-4.03 (m, 4H), 3.81 - 3.78 (m, 4H), 3.29 (s, 2H), 3.17 - 3.07 (m, 1H) ), 2.98-2.94 (m, 2H), 2.66 (d, J = 4.8 Hz, 3H), 2.48-2.39 (m, 2H), 2.13-1.93 (m, 4H), 1.22 (t, J = 7.2 Hz, 3H). ESI-MS: m/z = 524.45 [M+H] ⁺ .

Example 49

Preparation of 1-(4-(3-(1-(2-methoxyethyl)piperidin-4-yl)-7-morpholinylisoxazole [4,5-d]-pyrimidine by GL-26 -5-yl)phenyl)-3-methylurea (GL-49)

GL-26 (100 mg, 0.15 mmol) was suspended in ethanol, DIPEA (71 mg, 0.55 mmol), NaI (27 mg, 0.18 mmol) and 2-methoxybromoethane (27 mg, 0.20 mmol). Monitoring raw materials is no longer reduced. The reaction mixture was concentrated under reduced pressure. EtOAc (EtOAc m. Mp 208-210 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.24 - 8.21 (m, 2H), 7.53-7.50 (m, 2H), 6.07 (q, J = 4.5 Hz, 1H), 4.06-4.03 (m, 4H), 3.81 - 3.78 (m, 4H), 3.48 (t, J = 5.7 Hz, 2H), 3.26 (s, 3H), 3.16 - 3.09 (m, 1H) ), 3.03-2.98 (m, 2H), 2.66 (d, J = 4.5 Hz, 3H), 2.60-2.50 (m, 2H), 2.27-2.16 (m, 2H), 2.12-1.92 (m, 4H). ESI-MS: m/z = 496.57 [M+H] ⁺ .

Example 50

Preparation of 2-(4-(5-(4-(3-methylureido)phenyl)-7-morpholinylisoxazole [4,5-d]pyrimidin-3-yl)piperidin by GL-48 Pyridin-1-yl)acetic acid ditrifluoroacetate (GL-50)

GL-48 (110 mg, 0.21 mmol) was dissolved in MeOH, and a solution of 0.21 mL of 2N NaOH was added and refluxed for 6 h. The reaction solution was cooled, and the mixture was diluted with EtOAc EtOAc. EtOAc. The TFA was evaporated to dryness <RTI ID=0.0> Mp 240-242°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ10.05 (s, 1H), 8.87 (s, 1H), 8.28-8.25 (m, 2H), 7.54-7.51 (m, 2H) , 6.26-6.11(m,1H), 5.42(s,2H), 4.19(s,2H),4.09-4.02(m,4H),3.82–3.79(m,4H),3.75-3.43(m,3H) , 3.43-3.19 (m, 2H), 2.66 (s, 3H), 2.47-2.16 (m, 4H) .ESI-MS: m / z = 496.68 [m + H] +.

Example 51

Preparation of 1-(4-(3-(1-(2-hydroxy-2-methylpropanoyl)piperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d] using GL-26 ]pyrimidin-5-yl)phenyl)-3-methylurea (GL-51)

GL-51 is a yellow solid (75 mg, yield 95%) obtained from 2-hydroxyisobutyric acid by the procedure A. Mp 165-167°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.78 (s, 1H), 8.25-8.21 (m, 2H), 7.53-7.49 (m, 2H), 6.07 (q, J = 4.8 Hz, 1H), 5.44 (s, 1H), 5.12-4.69 (m, 1H), 4.69-4.32 (m, 1H), 4.06–4.03 (m, 1H), 3.81–3.78 (m, 1H), 3.52 -3.42 (m, 1H), 3.28-3.05 (m, 1H), 3.05-2.77 (m, 1H), 2.66 (d, J = 4.5 Hz, 3H), 2.18-2.12 (m, 2H), 2.06-1.70 (m, 2H), 1.36 (s, 6H). ESI-MS: m/z = 524.48 [M+H] ⁺ .

Example 52

Preparation of 4-(5-(4-(3-methylureido)phenyl)-7-morpholinylisoxazole [4,5-d]pyrimidin-3-yl)-N-pyridine using GL-26 -3-yl) piperidine-1-thiocarboxamide (GL-52)

GL-26 (100 mg, 0.15 mmol) was suspended in acetonitrile, 3-isothiocyanyl pyridine (50 mg, 0.37 mmol) and DIPEA (28 mg, 0.22 mmol). The organic layer was washed with EtOAc (EtOAc)EtOAc. Mp 183-185°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 9.51 (s, 1H), 8.79 (s, 1H), 8.51 (d, J = 2.4 Hz, 1H), 8.32-8.30 (m) , 1H), 8.28-8.25 (m, 2H), 7.80 - 7.76 (m, 1H), 7.54 - 7.51 (m, 2H), 7.37 (dd, J = 8.1, 4.5 Hz, 1H), 6.09 (q, J =4.5 Hz, 1H), 4.85-4.80 (m, 2H), 4.07–4.04 (m, 4H), 3.82–3.80 (m, 4H), 3.65-3.55 (m, 1H), 3.52-3.44 (m, 2H) ), 2.67 (d, J = 4.8 Hz, 3H), 2.25-1.98 (m, 4H). ESI-MS: m/z = 574.26 [M+H] ⁺ .

Example 53

Preparation of 1-methyl-3-(4-(7-morpholin-3-(1-trifluoromethanesulfonylpiperidin-4-yl)isoxazole [4,5-d]-pyrimidine by GL-26 -5-yl)phenyl)urea (GL-53)

GL-26 (137 mg, 0.21 mmol) was suspended in anhydrous THF. DIPEA (96 mg, 0.74 mmol) was added, and the mixture was cooled with ice-cooling, and a solution of trifluoromethanesulfonyl chloride (51 mg, 0.30 mmol) in THF was added dropwise to the reaction mixture. The reaction was further stirred for 1 h, stirred at room temperature overnight, and the reaction was not completely observed by TLC. The mixture was warmed to 40 ° C for 5 h and then added trifluoromethanesulfonyl chloride (40 mg, 0.25 mmol). The reaction mixture was concentrated, EtOAc (EtOAc)EtOAc. Mp 238-240°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.78 (s, 1H), 8.26-8.21 (m, 2H), 7.53-7.50 (m, 2H), 6.08 (q, J = 4.8 Hz, 1H), 4.07–4.04 (m, 4H), 3.97-3.93 (m, 2H), 3.81–3.79 (m, 4H), 3.58-3.47 (m, 3H), 2.66 (d, J=4.5 Hz) , 3H), 2.35-2.21 (m, 2H), 2.12 - 1.99 (m, 2H). ESI-MS: m/z = 570.70 [M+H] ⁺ .

Example 54

Preparation of 1-(4-(3-(1-(2-chloroacetyl))piperidin-4-yl)-7-morpholinylisoxazole [4,5-d]pyrimidine-5 using GL-26 -yl)phenyl)-3-methylurea (GL-54)

GL-54 was obtained as a yellow solid (38 mg, yield 49%) from chloroacetyl chloride. Mp 157-158°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.26-8.21 (m, 2H), 7.54-7.49 (m, 2H), 6.07 (d, J = 4.8 Hz, 1H), 4.50-4.36 (m, 3H), 4.06-3.89 (m, 5H), 3.81-3.78 (m, 1H), 3.54-3.44 (m, 1H), 3.39–3.33 (m, 1H) , 3.02 - 2.94 (m, 1H), 2.66 (d, J = 4.5 Hz, 3H), 2.16. 16.76 (m, 4H). ESI-MS: m/z = 514.71 [M+H] ⁺ .

Example 55

Preparation of 1-(4-(3-(1-(3,3-dimethylbutanoyl)piperidin-4-yl)-7-morpholinylisoxazole [4,5-d] using GL-26 And pyrimidin-5-yl)phenyl)-3-methylurea (GL-56)

GL-56 was obtained as a white solid (80 mg, yield 99%) from 3,3-dimethylbutyric acid. Mp 153-155 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.76 (s, 1H), 8.2 - 8.20 (m, 2H), 7.52 - 7.48 (m, 2H), 6.06 (q, J = 4.5 Hz, 1H), 4.60-4.47 (m, 1H), 4.13 - 4.02 (m, 5H), 3.81 - 3.78 (m, 4H), 3.52-3.40 (m, 1H), 3.31 - 3.25 (m, 1H) , 2.87-2.78 (m, 1H), 2.66 (d, J = 4.8 Hz, 3H), 2.30 (q, J = 14.1 Hz, 2H), 2.14 - 1.77 (m, 4H), 1.04 (s, 9H). ESI-MS: m/z = 536.59 [M+H] ⁺ .

Example 56

Preparation of 1-(4-(3-(1-(3,5-di-tert-butylbenzoyl)piperidin-4-yl)-7-morpholinylisoxazole using GL-26 [4,5- d]pyrimidin-5-yl)phenyl)-3-methylurea (GL-57)

GL-57 is a white solid (66 mg, yield 67%) which was synthesized from 3,5-di-tert-butylbenzoic acid. Mp 158-159°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.75 (s, 1H), 8.27–8.21 (m, 2H), 7.53–7.48 (m, 3H), 7.23 (d, J= 1.8 Hz, 2H), 6.06 (q, J = 4.8 Hz, 1H), 4.77-4.24 (m, 1H), 4.07 - 4.04 (m, 4H), 3.82 - 3.59 (m, 5H), 3.56-3.46 (m , 1H), 3.41 - 3.02 (m, 2H), 2.66 (d, J = 4.5 Hz, 3H), 2.17 - 1.95 (m, 4H), 1.30 (s, 18H). ESI-MS: m/z = 654.93 [M+H] ⁺ .

Example 57

Preparation of 1-(4-(3-(1-(3,5-difluorobenzoyl)piperidin-4-yl)-7-morpholinylisoxazole [4,5-d] using GL-26 And pyrimidine-5-yl)phenyl)-3-methylurea (GL-58)

GL-58 was obtained as yellow solid (60 mg, yield 81%) from 3,5-difluorobenzoic acid. Mp 162-164 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.77 (s, 1H), 8.26-8.21 (m, 2H), 7.53-7.50 (m, 2H), 7.36 (tt, J = 9.6, 2.4 Hz, 1H), 7.22–7.19 (m, 2H), 6.07 (q, J=4.5 Hz, 1H), 4.61-4.39 (m, 1H), 4.07-4.04 (m, 4H), 3.82–3.79 (m, 4H), 3.71-3.59 (m, 1H), 3.58-3.47 (m, 1H), 3.40-3.30 (m, 1H), 2.66 (d, J = 4.8 Hz, 3H), 2.37-2.06 (m , 2H), 2.04 - 1.90 (m, 2H). ESI-MS: m/z = 578.74 [M+H] ⁺ .

Example 58

Preparation of 1-(4-(3-(1-(2-(4-fluorophenyl)acetyl)piperidin-4-yl)-7-morpholinylisoxazole using GL-26 [4,5- d]pyrimidin-5-yl)phenyl)-3-methylurea (GL-59)

GL-59 was obtained as a yellow solid (68 mg, yield 93%) from </ br></br> Mp 138-140 ° C. ¹ H NMR (300 MHz, DMSO-d ₆ ) δ 8.78 (s, 1H), 8.25 - 8.20 (m, 2H), 7.54 - 7.50 (m, 2H), 7.33 - 7.28 (m, 2H), 7.17–7.09 (m, 2H), 6.08 (q, J=4.8 Hz, 1H), 4.47-4.42 (m, 1H), 4.10-4.03 (m, 5H), 3.81–3.78 (m, 6H) , 3.45 (tt, J = 11.1, 3.6 Hz, 1H), 3.33 - 3.26 (m, 1H), 2.94 - 2.87 (m, 1H), 2.66 (d, J = 4.5 Hz, 3H), 2.15 - 2.09 (m , 2H), 1.85-1.72 (m, 2H). ESI-MS: m/z = 574.91 [M+H] ⁺ .

Example 59

Preparation of 1-methyl-3-(4-(7-morpholine-3-(1-(4-(trifluoromethyl)benzoyl)piperidin-4-yl)isoxazole from GL-26 [ 4,5-d]pyrimidin-5-yl)phenyl)urea (GL-60)

GL-60 was a yellow-white solid (70 mg, yield 90%) from </RTI></RTI></RTI></RTI></RTI> Mp 208-210°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.76 (s, 1H), 8.26 - 8.23 (m, 2H), 7.84 (d, J = 8.1 Hz, 2H), 7.67 (d , J=8.1 Hz, 2H), 7.54–7.49 (m, 2H), 6.07 (q, J=4.5 Hz, 1H), 4.66-4.44 (m, 1H), 4.06–4.07 (m, 4H), 3.82– 3.79 (m, 4H), 3.70-3.58 (m, 1H), 3.58-3.49 (m, 1H), 3.43-3.27 (m, 1H), 3.25-3.07 (m, 1H), 2.66 (d, J = 4.8) Hz, 3H), 2.38-2.08 (m, 2H), 2.08-1.81 (m, 2H). ESI-MS: m/z = 610.87 [M+H] ⁺ .

Example 60

Preparation of (3-(3-(1-benzylpiperidin-4-yl)-7-morpholinylisoxazole[4,5-d]pyrimidin-5-yl)phenyl)methanol using Compound 1 GL-61)

1 (207 mg, 0.5 mmol), 3-hydroxyethylbenzeneboronic acid (114 mg, 0.75 mmol), Pd(Pcy ₃ ) ₂ Cl ₂ (19 mg, 0.025 mmol) and CsF (228 mg, 1.5 mmol) were placed in a three-neck bottle. The vacuum was exhausted and then Ar gas was introduced, and the exhaust gas was repeatedly taken three times, and 4 mL of solvent (NMP/water=9/1) was injected, and reacted at 100 ° C for 48 hours under the protection of Ar gas. Water and ethyl acetate were added to the reaction mixture, and the organic layer was washed with EtOAc (EtOAc) Mp 87-89°C. ¹ H NMR (300MHz, DMSO-d ₆ ) δ 8.32 (s, 1H), 8.26 - 8.23 (m, 1H), 7.48-7.43 (m, 2H), 7.35 - 7.23 (m, 5H), 5.28 (t, J = 5.7 Hz, 1H), 4.59 (d, J = 5.7 Hz, 1H), 4.08 - 4.05 (m, 4H), 3.82 - 3.79 (m, 4H), 3.54 (s, 2H) ), 3.22-3.12 (m, 1H), 2.95-2.91 (m, 2H), 2.23-1.95 (m, 6H). ESI-MS: m/z = 486.55 [M+H] ⁺ .

Example 61

Preparation of 3-(3-(1-benzylpiperidin-4-yl)-7-morpholinylisoxazole [4,5-d]pyrimidin-5-yl)phenol (GL-62) from Compound 1

1 (207 mg, 0.5 mmol), 3-hydroxybenzeneboronic acid (103 mg, 0.75 mmol), Pd(Pcy ₃ ) ₂ Cl ₂ (19 mg, 0.025 mmol) and CsF (228 mg, 1.5 mmol) were placed in a three-necked flask. The exhaust gas was further introduced into the Ar gas, and the exhaust gas was repeatedly injected three times, and 4 mL of the solvent (NMP/water = 9/1) was injected, and reacted at 100 ° C for 48 hours under the protection of Ar gas. Water and ethyl acetate were added to the reaction mixture, and the organic layer was washed with EtOAc EtOAc. Mp 86-88 ° C. ¹ H NMR (300 MHz, CDCl ₃ ) δ 7.93 (d, J = 7.8 Hz, 1H), 7.83 (s, 1H), 7.39 - 7.25 (m, 5H), 6.92 (dd, J = 7.8, 1.8 Hz, 1H), 4.13 - 4.10 (m, 4H), 3.87 - 3.84 (m, 4H), 3.66 (s, 2H), 3.19 - 3.11 (m, 3H), 2.33 - 2.11 (m, 7H) ). ESI-MS: m/z = 472.31 [M+H] ⁺ .

Example 62

Preparation of 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinolinylisoxazole [4,5-d]pyrimidin-5-yl)phenyl)-3- Methylurea hydrochloride

GL-2 (528 mg, 1 mmol) was dissolved in 10 mL of anhydrous dioxane, and 0.52 mL of a self-made 2.3 mol/L hydrogen chloride anhydrous dioxane solution was added thereto, stirred at room temperature overnight, and evaporated to dryness under reduced pressure. The hexacyclohexane was ultrasonically washed by adding 10 mL of anhydrous dichloromethane, and filtered to give a white solid (535 mg, yield 95%). Mp 255-257 ° C. ESI-MS: m/z = 528.38 [M+H] ⁺ .

Test Example 63

The compound of the present invention having the structure of the formula (I) and a pharmaceutically acceptable salt thereof, in the antitumor side

The surface has obvious utility and is now illustrated by the following pharmacological experiments:

The half-inhibitory concentration IC ₅₀ value of the compound against the three tumor cell lines (U-87MG, PC-3 and BT-474) was measured by a CCK-8 test kit.

1. Materials and methods

U-87MG human malignant glioblastoma cell line (ordered at the Shanghai Cell Resource Center of the Chinese Academy of Sciences)

PC-3 human prostate cancer cell line (ordered at the Shanghai Cell Resource Center of the Chinese Academy of Sciences)

BT-474 human breast cancer cell line (ordered at the Shanghai Cell Resource Center of the Chinese Academy of Sciences)

Cell Counting Kit-8 (Cat#CK04-13, Dojindo)

96-well culture plate (Cat# 3599, Corning Costar)

Fetal bovine serum (Cat#10099-141, GIBCO)

Medium (Invitrogen)

Benchtop Microplate Reader SpectraMax M5Microplate Reader (Molecular Devices)

2, the experimental steps

2.1 reagent preparation

Cell line	Medium
		U-87MG	EMEM+1mM sodium pyruvate+1.5g/L NaHCO3+10%FBS
PC-3	F12K+10%FBS
		BT-474	RPMI1640+10%FBS

Preparation of the compound: The compound was diluted with DMSO to a final concentration of 10 mM.

2.2IC ₅₀ experiment (CCK-8 test)

The logarithmic growth phase cells were collected, counted, and the cells were resuspended in complete medium, and the cell concentration was adjusted to an appropriate concentration (determined according to the cell density optimization test results), and 96-well plates were seeded, and 100 μL of the cell suspension was added to each well. The cells were incubated for 24 hours at 37 ° C in a 100% relative humidity, 5% CO ₂ incubator. The test compound was diluted with the medium to the corresponding concentration (5X) set, and added to the cells at 25 μL/well. The final concentration of the compound was from 100 μM to 0 μM, a 4-fold gradient dilution, a total of 10 concentration points; or from 10 μM to 0 μM, a 4-fold gradient dilution, a total of 10 concentration points. The cells were then incubated for 72 hours at 37 ° C in 100% relative humidity, 5% CO 2 incubator. The medium was aspirated, and the complete medium containing 10% CCK-8 was added and incubated in a 37 ° C incubator for 2-4 hours. The absorbance at a wavelength of 450 nm was measured on a SpectraMax M5 Microplate Reader with gentle shaking, and the absorbance at 650 nm was used as a reference to calculate the inhibition rate.

3, data processing

The inhibition rate of the drug on tumor cell growth was calculated as follows: tumor cell growth inhibition rate % = [(Ac-As) / (Ac-Ab)] × 100%

As: OA of the sample (cell + CCK-8 + test compound)

Ac: negative control OA (cell + CCK-8 + DMSO)

Ab: blank control OA (medium + CCK-8 + DMSO)

The IC ₅₀ curve was fitted and the IC ₅₀ value was calculated using the software Graphpad Prism 5 and using the calculation formula log(inhibitor) vs. normalized response-Variable slope.

4. Experimental results

Test Example 64

The action of the compound of formula (I) as a PIK3 inhibitor is illustrated by the following test

Application of ^{PI3-Kinase (human) HTRF TM} Assay kit detection test compound inhibitory concentrations (IC ₅₀₎ of the inhibition of PI3K delta and half of the enzyme.

1, materials and instruments

2104En

Multilabel Reader (Cat: 2104-0010, PerkinElmer)

384well opaque balck plate (Cat.6007270, PerkinElmer)

PI 3-Kinase (human) HTRF ^TM Assay kit (Cat.33-016, Millipore)

4×Reaction Buffer (Cat.33-002, Millipore)

PIP21mM (Cat.33-004, Millipore)

Stop A (Cat.33-006, Millipore)

Stop B (Cat.33-008, Millipore)

DM A (Cat.33-010, Millipore)

DM B (Cat.33-012, Millipore)

DM C (Cat.33-014, Millipore)

PI3K delta (Cat. 14-604, Millipore)

ATP 10mM (cat PV3227, Invitrogen)

DTT 1M (cat D5545, Sigma)

2, reagent preparation

2.11×Reaction Buffer

4×Reaction Buffer (Cat. 33-002, Millipore) was diluted to 1× with ddH ₂ O, and 1 M DTT was added to give a final concentration of 5 mM. Freshly prepared before each use. For example, 10 mL of 1×Reaction Buffer is prepared, and 2.5 mL of 4×Reaction Buffer, 50 μL of 1 M concentration of DTT and ddH ₂ O 7.45 mL are added. Throughout the experiment, the ATP working solution, the substrate and the enzyme mixed working solution were prepared by using freshly prepared 1×Reaction Buffer.

2.24× compound working fluid

Primary screening: The test compound was dissolved in DMSO to 50 μM as a stock solution, and 2 μL of each was added to 48 μL of ddH ₂ O to obtain 2 μM of a compound solution containing 4% DMSO. After mixing, 2 μL of each was added to add 18 μL of 4% DMSO (in ddH _{2 ).} O) A 0.2 μM compound solution was obtained. 5 μL of each diluted solution was added to a 384-well plate such that the final concentration of the compound in the final 20 μL of the kinase reaction system was 500 nM and 50 nM, respectively, and contained 1% DMSO.

IC ₅₀ : The test compound was dissolved in DMSO to 10 mM as a stock solution, and 2 μL of each was added to 48 μL of 1×Reaction Buffer to obtain 400,000 nM of a compound solution containing 4% DMSO. After mixing, 5 μL of each was added to the next 15 μL of 4% DMSO ( In the wells of 1×Reaction Buffer), it was diluted sequentially to obtain 10 concentration gradients. 5 μL of each diluted solution was added to the 384-well plate, so that the final concentration of the compound in each well in the final 20 μL kinase reaction system was 100000 nM, 25000 nM, 6250 nM, 1562.5 nM, 390.63 nM, 97.65 nM, 24.42 nM, 6.10 nM. 1.53 nM, 0.38 nM and containing 1% DMSO. Each of the 10 assay concentrations was a duplicate well. At the same time, CAL-101 was selected as a reference compound, and the compound was diluted 4 times from 2 μM.

2.32×PIP2 working fluid

2×PIP2 working solution was prepared with 1×reaction buffer to a final concentration of 20 μM, and the final concentration of PIP2 was 10 μM. For example, 1 mL of 1× reaction buffer/PIP2 working solution was prepared, and 20 μL of PIP2 was added to 980 μL of 1×reaction buffer. This working fluid should be mixed with 0.1-0.2 mL to meet the control use and dead volume.

2.42×PIP2/Kinase working fluid

The kinase was diluted with 2 x PIP2 working solution at a concentration of 80 ng/well of kinase working solution. No kinase control (can be considered 100% inhibition) ie 2 x PIP2 working solution.

2.54×ATP working fluid

10 mM ATP was diluted to 40 μM with 1×reaction buffer. In a 20 μL kinase reaction system, the concentration of ATP was 10 μM. For example, 2 mL of ATP working solution was prepared, and 8 μL of 10 mM ATP was added to a 1992 μL 1×reaction buffer.

2.6 stop solution

Stop A and Stop B are mixed in a ratio of 3:1 and allowed to stand at room temperature for at least 2 hours before use. The stop solution can be stabilized at room temperature for 12 hours.

2.7 test solution

DM C, DM A and DM B were mixed in a ratio of 18:1:1 and allowed to stand at room temperature for at least 2 hours before use. The test solution was stable at room temperature for 12 hours.

3, the experimental process

4, data analysis

Calculate the Emission Ratio (ER) of each hole

Emission Ratio (ER)=665nm Emission signal/620nm Emission signal

The average Emission Ratio of the 100% inhibition control is recorded as: ER _100%

The average Emission Ratio of the 0% inhibition control is recorded as: ER _0%

The inhibition rate is calculated by the following formula: inhibition rate = (ER _{sample -} ER _0% ) / (ER _{100% -} ER _0% ) × 100%

Using Graphpad Prism 5 software for the IC ₅₀ curve ₅₀ value _was calculated to be merged IC.

5, the experimental results

^a NT= no test

Claims (10)

1. a pyrimidine derivative of the formula (I) and a plurality of crystal forms thereof or a pharmaceutically acceptable salt thereof,

   

   among them,

   R ₁ is hydrogen, optionally substituted aryl having C ₁ -C ₆ as a substituent, optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl, acylamino, halogen, in optionally substituted C ₁ -C ₆ linear, branched, cyclic alkyl group, a substituted aryl group, optionally substituted at C ₁ -C ₆ linear, branched, or cyclic alkyl group substituted a pyridyl group of the group, an optionally substituted C ₁ -C ₆ linear, branched, or cyclic alkyl group as a substituent of a sulfone group, optionally substituted C ₁ -C ₆ straight chain, branched or cyclic a sulfonyl group in which an alkyl group is a substituent, and an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group, pyridyl group, halogenated phenyl group, amine group, nitrile group or halogenated alkyl group as a substituent Carbonyl group, optionally substituted C ₁ -C ₆ linear, branched or cyclic alcohol group, ester group, ether group or carboxyl group;

   R ₂ is hydrogen,

   

   NHR ₂ ' or NHCONHR ₃ ;

   Wherein R ₁₀ is H or an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group; said R ₂ 'is H or a C ₁ -C ₆ linear, branched alkyl group An optionally substituted amide group having a C ₃ -C ₆ cycloalkyl group as a substituent, a trisubstituted amino group having an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group as a substituent An optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group having one, two or three halogen atoms as a substituent, a C ₁ -C ₆ straight or branched alkoxy group, optionally substituted a C ₁ -C ₆ linear, branched or cyclic alcohol group, piperazinyl pyridyl, aminopyridyl, piperazinylaryl or halogen aryl; wherein piperazinyl and amine are a substituted piperazinyl and a substituted amine group optionally substituted with a C ₁ -C ₆ linear, branched or cyclic alkyl group; said R ₃ = optionally substituted C ₁ -C ₆ straight chain, a branched or cyclic alkyl group and an optionally substituted aryl group;

   R ₀ is hydrogen, hydroxy or an optionally substituted C1-C6 linear, branched or cyclic alcohol group.
2. The pyrimidine derivative represented by the formula (I) according to Claim 1 or a plurality of crystal forms thereof, or a pharmaceutically acceptable salt thereof, characterized in that

   (1) When R ₁ = an optionally substituted aryl group, R ₂ = H, R ₀ = -OH or R ₀ 'OH, wherein R ₀ ' is an optionally substituted C ₁ - C ₆ straight chain, Chain or cyclic alkyl group;

   When R ₁ = an optionally substituted aryl group, R ₀ = H,

   

   Or NHR ₂ ', wherein R ₁₀ is H or an optionally substituted C ₁ -C ₆ straight chain, branched chain, or cyclic alkyl group; R ₂ '=H or CONHR ₃ ; wherein R ₃ =H,C _1- C ₆ linear, branched alkyl, C ₃ -C ₆ cycloalkyl, trisubstituted amino group substituted by C ₁ -C ₆ straight or branched alkyl, monohalogenated C ₁ -C ₆ straight-chain, branched or cyclic alkyl, dihalo C ₁ -C ₆ straight-chain, branched or cyclic alkyl, trihalo C ₁ -C ₆ straight-chain, branched or cyclic alkyl,

   

   -OR ₈ , R ₉ OH or

   

   among them,

   When X=N, the R ₄ =H,

   

   Or NHR ₆ , wherein R ₅ and R _{6 are} independently a C ₁ -C ₆ linear or branched alkyl group;

   When X=C, the R ₄ =H,

   

   Halogen, wherein R7=H, C ₁ -C ₆ alkyl, BOC or

   

   The R ₈ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

   The R ₉ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

   (2) When R ₀ =H, R ₁ =NHCONHR ₃ ', R ₂ =CONHR ₁₁ , the R ₃ ', R ₁₁ are each independently H, an optionally substituted C ₁ -C ₆ straight chain, Chain or cyclic alkyl group;

   (3) When R ₀ = H, R ₁ = H or C ₁ - C ₆ linear, branched alkyl; R ₂ = NHCONHR ₃ , R ₃ is an optionally substituted C ₁ - C ₆ straight chain, branch Chain or cyclic alkyl group;

   (4) When R ₀ =H, R ₂ =NHCONHR ₃ , and R ₃ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group,

   

   

   

   -R ₂₅ -CY ₃ , -R ₂₇ -OH,

   

   or

   

   among them,

   The R ₁₁ , R ₂₀ , R ₂₁ are each independently H or an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

   R ₁₂ , R ₁₃ , R _14a , R ₁₆ , R _18a , R ₂₃ , R ₂₄ , R ₂₆ , R ₂₇ , R ₂₈ , R ₂₉ , R ₃₀ , R ₃₁ , R ₃₂ , R ₃₃ , R ₃₄ , Individually independently an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

   The R ₁₄ is an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl or trihalo C ₁ -C ₆ linear, branched, or cyclic alkyl group;

   The R ₁₅ is an optionally substituted C ₁ -C ₁₂ straight chain, branched chain, cyclic alkyl group or an optional substitution with a halogen or a C ₁ -C ₆ linear, branched or cyclic alkyl group as a substituent Aromatic hydrocarbon

   The R ₁₈ and R _{19 are} independently H, Boc or an optionally substituted C ₁ -C ₆ linear, branched or cyclic alkyl group;

   The Y = F, Cl, Br or I;

   The Z is F, Cl, Br, I or hydrogen;
3. The pyrimidine derivative according to claim 1 or a plurality of crystal forms thereof, or a pharmaceutically acceptable salt thereof, characterized in that

   (1) When R ₁ = benzyl, R ₂ = H, R ₀ is a hydroxyl group, methanol, ethanol, propanol, isopropanol or butanol;

   When R ₁ = benzyl, R ₀ = H,

   

   NHR ₂ ' or NHCONHR ₃ ;

   among them,

   The R ₁₀ is H, methyl, ethyl, propyl or butyl;

   The R ₂ '=H or CONHR ₃ ;

   The R ₃ =H, methyl, ethyl, propyl, isopropyl, butyl, cyclopropyl, dimethylethylamine, fluoroethyl, difluoroethyl, trifluoroethyl ,

   

   -OR ₈ or R ₉ OH;

   Wherein, when X=N, the R ₄ =H,

   

   Or NHR ₆ , wherein R ₅ and R _{6 are} independently methyl, ethyl, propyl or butyl;

   When X=C, the R ₄ =H,

   

   Or F, where R7=H, BOC or

   

   The R8 is a methyl group, an ethyl group, a propyl group or a butyl group;

   The R9 is a methyl group, an ethyl group, a propyl group or a butyl group;

   (2) When R ₀ = H, R ₁ =NHCONHR ₃ '; R ₂ =CONHR ₁₁ , each of R ₃ ', R _{11 is} independently methyl, ethyl, propyl, isopropyl or butyl ;

   (3) When R ₀ = H, when R ₁ = H, methyl, ethyl, propyl, isopropyl or butyl; R ₂ = NHCONHR ₃ , R ₃ = methyl, ethyl, propyl, Isopropyl or butyl;

   (4) When R ₀ = H, R ₂ = NHCONHR ₃ , R ₃ = methyl, ethyl, propyl, isopropyl or butyl,

   

   

   

   -R ₂₅ -CY ₃ , -R ₂₇ -OH,

   

   -R ₃₀ -OR ₃₁ , -R ₃₂ COOH,

   

   or

   

   The R ₁₀ , R ₁₁ , R ₂₀ , R ₂₁ are each independently H or methyl, ethyl, propyl, isopropyl or butyl;

   The R ₁₂ , R ₁₃ , R ₁₆ , R _18a , R ₂₃ , R ₂₄ , R ₂₆ , R ₂₇ , R ₂₈ , R ₂₉ , R ₃₀ , R ₃₁ , R ₃₂ , R ₃₃ , R ₃₄ are each independently Methyl, ethyl, propyl, isopropyl, butyl, isobutyl, cyclopentyl or cyclohexyl;

   The R ₁₄ is methyl, ethyl, propyl, isopropyl, butyl, monofluoro C ₁ -C ₄ straight or branched alkyl, difluoro C ₁ -C ₄ straight or branched Alkyl, trifluoromethyl, trifluoroethyl, trifluoropropyl, trifluoroisopropyl, trifluorobutyl or trifluoroisobutyl;

   The R ₁₅ is methyl, ethyl, propyl, isopropyl, butyl, isobutyl, cyclopentyl, cyclohexyl, monofluoro C ₁ -C ₄ linear, branched, cyclic alkyl , difluoro C ₁ -C ₄ straight chain, branched chain, cyclic alkyl group, trifluoro C ₁ -C ₄ straight chain, branched chain, cyclic alkyl group, with fluorine or C ₁ -C ₄ straight chain, a branched or cyclic alkyl group is a phenyl group in which a substituent is substituted, ortho, meta or unsubstituted;

   The R ₁₈ , R _{19 are} independently H, methyl, ethyl, propyl, isopropyl, butyl, isobutyl or Boc;

   The Y=F;

   The Z = F.
4. The pyrimidine derivative according to claim 1 or a plurality of crystal forms thereof or a pharmaceutically acceptable salt thereof, which is one of the following compounds:

   

   

   
5. The method for producing a pyrimidine derivative according to claim 1 or a plurality of crystal forms thereof or a pharmaceutically acceptable salt thereof, which is prepared from the compound 1, which is prepared by:

   
6. A cytotoxic agent comprising the pyrimidine derivative according to any one of claims 1 to 5, and a plurality of crystal forms thereof, or a pharmaceutically acceptable salt thereof.
7. A pharmaceutical composition comprising a therapeutically effective amount of the pyrimidine derivative according to any one of claims 1 to 5, and a plurality of crystal forms thereof, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier .
8. Use of the pyrimidine derivative according to any one of claims 1 to 5, and various crystal forms thereof, or a pharmaceutically acceptable salt thereof, for the preparation of a cytotoxic agent and an antitumor drug.
9. Use of the cytotoxic agent of claim 6 for the preparation of a medicament for abnormal alteration of PI3K kinase and an antitumor drug.
10. Use of the pharmaceutical composition of claim 7 in the preparation of a medicament for the abnormal alteration of PI3K kinase and an antitumor medicament.