HK1258833A1 - Heterocyclic compound used as fgfr inhibitor - Google Patents

Description

Heterocyclic compounds as FGFR inhibitors

Technical Field

The present invention relates to a class of heterocyclic compounds and a method for preparing a pharmaceutical composition containing the same, and the use of the heterocyclic compounds as fibroblast growth factor receptor (FGFR) inhibitors. The compounds of the present invention can be used to treat or prevent diseases mediated by FGFR, such as cancer.

Background Art

Fibroblast growth factors (FGFs) are a family of structurally related polypeptides with diverse biological activities, encoded by the FGF gene family. To date, 22 members of the FGF family have been identified. Fibroblast growth factor receptors (FGFRs) are transmembrane tyrosine kinase receptors that mediate FGF signaling into the cytoplasm. Currently, four FGFRs, encoded by independent genes, have been identified: FGFR1, FGFR2, FGFR3, and FGFR4. They are all single-chain glycoprotein molecules composed of an extracellular domain, a transmembrane domain, and an intracellular domain. These receptor-ligand interactions lead to receptor dimerization and autophosphorylation, followed by complex formation with membrane-bound proteins and cytoplasmic accessory proteins, mediating a variety of signaling pathways. The FGFR-FGF signaling system plays an important role in many biological processes, including cell proliferation, differentiation, migration, angiogenesis, and tissue repair.

FGFR4 is the major FGF receptor subtype in the liver. Of the more than 20 fibroblast growth factors (FGFs) discovered to date, 10 can bind to FGFR4, and only FGF19 specifically binds to FGFR4. Recent studies have shown that alterations in FGFR4, such as overexpression, mutation, translocation, and truncation, are associated with the progression of various human cancers, including rhabdomyosarcoma, renal cell carcinoma, myeloma, breast cancer, gastric cancer, colon cancer, bladder cancer, pancreatic cancer, and hepatocellular carcinoma.

Therefore, it can be predicted that selective inhibition of FGFR4 can be used to treat the above cancers, especially tumors with activating mutants of receptor tyrosine kinases or upregulation of receptor tyrosine kinases will be particularly sensitive to this type of inhibitor.

Summary of the Invention

The present invention aims to provide a compound as shown in formula I, its isomers, prodrugs, stable isotope derivatives or pharmaceutically acceptable salts

wherein ring A and R are each independently selected from substituted or unsubstituted aryl, heteroaryl, and when A or R is substituted, they may be substituted by one or more substituents independently selected from hydrogen, halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, aryl, heteroaryl, aldehyde, -C(O)R ¹ , carboxyl, alkenyl, alkynyl, -OR ¹ , -NR ² R ³ , wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more substituents selected from halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , or a substituent of -S(O)mNR ⁵ R ⁶ ;

X is selected from CR ⁷ R ⁸ , NR ⁷ , O, S;

Y is selected from -C(O)-, -C(=NR ⁹ )-, and -S(O)m-;

R ¹ , R ² , R ³ , R ⁴ , R ⁵ , and R ⁶ are each independently selected from hydrogen, a C1-C8 alkyl group, a C3-C8 cyclyl group, a 3-8 membered monocyclic heterocyclic group, a monocyclic heteroaryl group or a monocyclic aryl group, an alkenyl group, and an alkynyl group, wherein R ² and R ³ , R ⁵ and R ⁶ may form a 3-7 membered heterocyclic group together with the nitrogen atom to which they are attached; and R ⁷ and R ⁸ may form a 3-8 membered cyclyl group or a 3-8 membered monocyclic heterocyclic group together with the carbon atom to which they are attached;

R ⁷ and R ⁸ are each independently selected from hydrogen, halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, aryl, heteroaryl, aldehyde, -C(O)R ¹ , carboxyl, alkenyl, alkynyl, -OR ¹ , -NR ² R ³ , wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ is substituted by a substituent selected from S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , and -S(O)mNR ⁵ R ⁶ ;

R ⁹ is independently selected from hydrogen, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, aryl, heteroaryl, aldehyde, -C(O)R ¹ , alkenyl, alkynyl, wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more selected from halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , -S(O)mNR ⁵ R ⁶ ;

And m is 1 or 2.

In one embodiment of the present invention, a compound of general formula (I), its isomer, prodrug or pharmaceutically acceptable salt thereof, characterized in that the compound shown in formula I is shown in formula II:

Z ¹ , Z ² , and Z ³ are each independently selected from CR ^{Z 1} , CR ^{Z 2} , CR ^{Z 3} or N, and:

When Z ¹ is N, Z ² and Z ³ are not N at the same time;

When Z ² is N, Z ¹ and Z ³ are not N at the same time;

When Z ³ is N, Z ¹ and Z ² are not N at the same time;

R ^Z1 , R ^Z2 , and R ^Z3 are each independently selected from hydrogen, halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, aryl, heteroaryl, aldehyde, -C(O)R ¹ , carboxyl, alkenyl, alkynyl, -OR ¹ , -NR ² R ³ , wherein the alkyl, cyclyl, heterocyclyl, aryl, or heteroaryl is optionally substituted by one or more halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ is substituted by a substituent selected from S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , and -S(O)mNR ⁵ R ⁶ ;

X is selected from CR ⁷ R ⁸ , NR ⁷ , O, S;

Y is selected from -C(O)-, -C(=NR ⁹ )-, and -S(O)m-;

R ¹ , R ² , R ³ , R ⁴ , R ⁵ , and R ⁶ are each independently selected from hydrogen, a C1-C8 alkyl group, a C3-C8 cyclyl group, a 3-8 membered monocyclic heterocyclic group, a monocyclic heteroaryl group or a monocyclic aryl group, an alkenyl group, and an alkynyl group, wherein R ² and R ³ , R ⁵ and R ⁶ may form a 3-7 membered heterocyclic group together with the nitrogen atom to which they are attached; and R ⁷ and R ⁸ may form a 3-8 membered cyclyl group or a 3-8 membered monocyclic heterocyclic group together with the carbon atom to which they are attached;

R ⁷ and R ⁸ are each independently selected from hydrogen, halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, aryl, heteroaryl, aldehyde, -C(O)R ¹ , carboxyl, alkenyl, alkynyl, -OR ¹ , -NR ² R ³ , wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ is substituted by a substituent selected from S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , and -S(O)mNR ⁵ R ⁶ ;

R ⁹ is independently selected from hydrogen, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, aryl, heteroaryl, aldehyde, -C(O)R ¹ , alkenyl, alkynyl, wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more selected from halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , -S(O)mNR ⁵ R ⁶ ;

When Z ¹ is CCH ₂ OH, CCH ₂ COOH, or C-(4-piperidine), compounds (2-1), (2-2), and (2-3) may exist as isomers (2-1A), (2-2A), and (2-3A):

In another embodiment of the present invention, a compound represented by general formula (I), its isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt, characterized in that the compound represented by formula I is formula IIIa, IIIb, IIIc, IIId, IIIe, IIIf:

R ^Z1 , R ^Z2 , R ^Z3 , R ¹⁰ , R ¹¹ are each independently selected from hydrogen, halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, aryl, heteroaryl, aldehyde, -C(O)R ¹ , carboxyl, alkenyl, alkynyl, -OR ¹ , -NR ² R ³ , wherein said alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , or -S(O)mNR ⁵ R ⁶ ;

R ⁷ is independently selected from H, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered monocyclic heterocyclyl, monocyclic heteroaryl or monocyclic aryl, wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more selected from halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , -S(O)mNR ⁵ R substituted by a substituent of ⁶ ;

R ¹ , R ² , R ³ , R ⁴ , R ⁵ , and R ⁶ are each independently selected from hydrogen, a C1-C8 alkyl group, a C3-C8 cyclyl group, a 3-8 membered monocyclic heterocyclic group, a monocyclic heteroaryl group or a monocyclic aryl group, an alkenyl group, and an alkynyl group, wherein R ² and R ³ , R ⁵ and R ⁶ may form a 3-7 membered heterocyclic group together with the nitrogen atom to which they are attached; and R ⁷ and R ⁸ may form a 3-8 membered cyclyl group or a 3-8 membered monocyclic heterocyclic group together with the carbon atom to which they are attached;

The aforementioned R ² and R ³ , R ⁵ and R ⁶ together with the nitrogen atom to which they are attached may form a 3-8 membered heterocyclic group.

In another embodiment of the present invention, a compound represented by general formula (I), its isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt thereof, characterized in that the compound represented by formula I is formula IV;

R ^Z1 and R ^Z2 are each independently selected from hydrogen, halogen, C1-C4 alkyl, C3-C7 cyclyl, 4-6 membered monocyclic heterocyclyl, 5-6 membered monocyclic heteroaryl or monocyclic aryl, aldehyde, keto, carboxyl, cyano, OR ¹ , NR ² R ³ , wherein said alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted with one or more substituents selected from halogen, hydroxyl, C1-C4 alkyl, C3-C7 cyclyl, 4-6 membered heterocyclyl, aryl or heteroaryl;

R ⁷ is selected from H, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered monocyclic heterocyclyl, monocyclic heteroaryl or monocyclic aryl, wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more selected from halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁴ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁴ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁴ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁴ , -NR ⁴ C(O)NR ⁵ R ⁶ , -S(O)mR ⁴ , -NR ⁵ S(O)mR ⁴ , -SR ⁴ , -NR ⁴ S(O)mNR ⁵ R ⁶ , -S(O)mNR ⁵ R substituted by a substituent of ⁶ ;

R ¹⁰ is independently selected from hydrogen, halogen, halogenated C1-C4 alkyl, cyano;

R ¹¹ is independently selected from hydrogen, halogen, C1-C4 alkyl, halo-C1-C4 alkoxy, C1-C6 alkoxy, HO-C1-C4 alkoxy, cyano, NR ² R ³ , C1-C4 alkoxy C1-C4 alkoxy, C1-C4 alkoxy halo-C1-C4 alkoxy;

R ¹ , R ² , and R ³ are each independently selected from hydrogen, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered monocyclic heterocyclyl, monocyclic heteroaryl or monocyclic aryl, C1-C3 alkylthio, and halogenated alkoxy substituted with hydroxyl at any position;

R ⁴ , R ⁵ , and R ⁶ are each independently selected from hydrogen, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered monocyclic heterocyclic group, monocyclic heteroaryl or monocyclic aryl, alkenyl, and alkynyl, wherein R ² and R ³ , R ⁵ and R ⁶ can form a 3-7 membered heterocyclic group together with the N atom to which they are connected; and R ⁷ and R ⁸ can form a 3-8 membered cyclyl or a 3-8 membered monocyclic heterocyclic group together with the C atom to which they are connected.

In another embodiment of the present invention, a compound represented by general formula (I), its isomer, prodrug, stable isotope derivative or pharmaceutically acceptable salt, characterized in that the compound represented by formula I is formula V;

R ^Z1 and R ^Z2 are each independently selected from hydrogen, halogen, C1-C4 alkyl, C3-C7 cyclyl, 5-6 membered monocyclic heterocyclyl, 5-6 membered monocyclic heteroaryl or monocyclic aryl, aldehyde, carboxyl, wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted with one or more substituents selected from halogen, hydroxyl, C1-C4 alkyl, 5-6 membered heterocyclyl, aryl or heteroaryl;

R ⁷ is selected from hydrogen, C1-C4 alkyl, C3-6 cyclyl, wherein the alkyl or cyclyl is optionally substituted with one or more substituents selected from C1-C3 alkyl, 4-6 membered monocyclic heterocyclyl;

R ¹¹ is selected from NR ² R ³ , C1-C3 alkoxy, -O(CH ₂ ) _0-1 -R ⁴ ; wherein, R ⁴ is independently selected from hydrogen, C1-C8 alkyl, HO-C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered monocyclic heterocyclyl, monocyclic heteroaryl or monocyclic aryl, wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more selected from halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁵ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁵ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁵ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁶ , -NR ⁷ C(O)NR ⁵ R ⁶ , -S(O)mR ⁵ , -NR ⁵ S(O)mR ⁶ , -SR ⁵ , -NR ⁷ S(O)mNR ⁵ R ⁶ , -S(O)mNR ⁵ R ⁶ substituents; R ² , R ³ are independently selected from hydrogen, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered monocyclic heterocyclyl, monocyclic heteroaryl or monocyclic aryl, C1-C3 alkylthio, halogenated alkoxy substituted by hydroxy at any position, wherein the alkyl, cyclyl, heterocyclyl, aryl or heteroaryl is optionally substituted by one or more halogen, cyano, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered heterocyclyl, -OR ⁵ , -OC(O)NR ⁵ R ⁶ , -C(O)OR ⁵ , -C(O)NR ⁵ R ⁶ , -C(O)R ⁵ , -NR ⁵ R ⁶ , -NR ⁵ C(O)R ⁶ , -NR ⁷ C(O)NR ⁵ R ⁶ , -S(O)mR ⁶ , -NR ⁵ S(O)mR ⁶ , -SR ⁵ , -NR ⁷ S(O)mNR ⁵ R ⁶ , or -S(O)mNR ⁵ R ⁶ ;

R ⁴ , R ⁵ , and R ⁶ are each independently selected from hydrogen, C1-C8 alkyl, C3-C8 cyclyl, 3-8 membered monocyclic heterocyclic group, monocyclic heteroaryl or monocyclic aryl, alkenyl, and alkynyl, wherein R ² and R ³ , R ⁵ and R ⁶ can form a 3-7 membered heterocyclic group together with the N atom to which they are connected; and R ⁷ and R ⁸ can form a 3-8 membered cyclyl or a 3-8 membered monocyclic heterocyclic group together with the C atom to which they are connected.

Typical compounds of the present invention include, but are not limited to:

or its tautomers, meso racemates, racemates, enantiomers, diastereomers, mixtures thereof, and pharmaceutically acceptable salts thereof.

The compounds of this invention are effective FGFR inhibitors, particularly effective and selective inhibitors of FGFR4. Therefore, the compounds of this invention can be used to treat or prevent FGFR-mediated diseases, particularly FGFR4-mediated diseases, including but not limited to cancer and inflammatory diseases. The compounds of this invention can be used to treat or prevent cancers such as rhabdomyosarcoma, renal cell carcinoma, myeloma, breast cancer, gastric cancer, colon cancer, bladder cancer, pancreatic cancer, and hepatocellular carcinoma. The compounds of this invention are particularly useful in treating or preventing liver cancer, particularly hepatocellular carcinoma. In particular, tumors harboring activating mutants of receptor tyrosine kinases or upregulation of receptor tyrosine kinases are particularly sensitive to these inhibitors.

The compounds of the present invention, as FGFR4 selective inhibitors, have fewer side effects.

The present invention further relates to a pharmaceutical composition comprising the compound represented by general formula (I) or its isomers, prodrugs, stable isotope derivatives or pharmaceutically acceptable salts thereof, and pharmaceutically acceptable carriers, diluents and excipients.

The present invention also includes a method for preparing the pharmaceutical composition, for example, mixing the compound of the present invention with a pharmaceutically acceptable carrier, diluent, or excipient. The pharmaceutical composition of the present invention can be prepared by conventional methods in the art.

Another aspect of the present invention relates to the use of a compound represented by general formula (I) or its isomers, prodrugs, stable isotopic derivatives or pharmaceutically acceptable salts, and pharmaceutically acceptable carriers, diluents, and excipients in the preparation of a medicament, wherein the medicament is used to treat or prevent diseases mediated by FGFR, especially FGFR4, such as tumors or inflammatory diseases.

Another aspect of the present invention relates to the use of a compound represented by general formula (I) or its tautomers, meso-, racemic-, enantiomers, diastereoisomers, mixtures thereof, and pharmaceutically acceptable salts thereof, or the pharmaceutical composition thereof in the preparation of drugs for treating and/or preventing diseases such as tumors and inflammation.

According to the present invention, the medicine can be any pharmaceutical dosage form, including but not limited to tablets, capsules, solutions, lyophilized preparations, injections. The pharmaceutical preparation of the present invention can be administered in the dosage unit form comprising a predetermined amount of active ingredient per dosage unit. This unit can comprise, for example, 0.5 milligram to 1 gram, preferably 1 milligram to 700 milligrams, particularly preferably 5 milligrams to 300 milligrams of the compound of the present invention, or the pharmaceutical preparation can be administered in the dosage unit form comprising a predetermined amount of active ingredient per dosage unit according to the disease, administration method and patient's age, body weight and condition for treatment. It is preferred that the dosage unit formulation comprises the active ingredient of a predetermined amount of daily dose or divided dose or its corresponding fraction as indicated above. In addition, methods known in the pharmaceutical field can be used to prepare this type of pharmaceutical preparation.

The pharmaceutical formulations of the present invention may be suitable for administration by any desired suitable method, for example, by oral (including buccal or sublingual), rectal, nasal, topical (including buccal, sublingual or transdermal), vaginal or parenteral (including subcutaneous, intramuscular, intravenous or intradermal) administration. Such formulations may be prepared using all methods known in the pharmaceutical art, for example, by combining the active ingredient with one or more excipients or one or more adjuvants.

Pharmaceutical formulations suitable for oral administration can be administered as discrete units such as capsules or tablets; powders or granules; solutions or suspensions in aqueous or non-aqueous liquids; or oil-in-water liquid emulsions or water-in-oil liquid emulsions.

The present invention also relates to a method for treating or preventing FGFR, especially FGFR4-mediated diseases (such as tumors or inflammatory diseases), which comprises administering to a patient in need thereof a therapeutically effective amount of a compound represented by general formula (I) of the present invention or its isomers, prodrugs, stable isotope derivatives or pharmaceutically acceptable salts, and a pharmaceutically acceptable carrier, diluent, or excipient.

Another aspect of the present invention relates to a compound represented by general formula (I) or its isomers, prodrugs, stable isotopic derivatives, or pharmaceutically acceptable salts, and pharmaceutically acceptable carriers, diluents, and excipients, for use in treating or preventing diseases mediated by FGFR, particularly FGFR4, such as tumors or inflammatory diseases.

Another aspect of the present invention relates to a compound represented by general formula (I) or its tautomers, mesomers, racemates, enantiomers, diastereomers, mixtures thereof, and pharmaceutically acceptable salts thereof for treating and/or preventing diseases such as tumors.

Preparation process

The present invention also provides a method for preparing the compound.

Process 1

Step 1: R ^z2 and R ⁷ in the structure of compound (VI-1) are consistent with those in the structure of (VI). Compound (VI-1) is subjected to carbonyl insertion using palladium (palladium acetate) catalysis, and 1,1-bis(diphenylphosphino)ferrocene is used as a ligand and triethylamine is used as a base to synthesize compound (VI-2). R ^z2 and R ⁷ in compound (VI-2) are consistent with those in the structure of (VI).

Step 2: The ester in compound (VI-2) is reduced with a reducing agent (sodium borohydride) to synthesize compound (VI-3), and R ^z2 and R ⁷ in the structure of compound (VI-3) are consistent with those in the structure of (VI).

Step 3: The hydroxyl group in the structure (VI-3) is replaced, and phosphorus tribromide is used as a reaction reagent to synthesize compound (VI-4), and R ^z2 and R ⁷ in the structure of compound (VI-4) are consistent with those in the structure (VI).

Step 4: The bromine in the structure of (VI-4) is replaced by a nucleophilic reagent (morpholin-3-one, 4-methylpiperazine-2-one, etc.), and a base (sodium hydride) is used as a deprotonating agent to synthesize compound (VI-5). The R ^z2 and R ⁷ in the structure of compound (VI-5) are consistent with those in the structure of (VI).

Step 5: R ^z2 and R ⁷ in the structure of compound (VI-5) are consistent with those in the structure of (VI). The amino protecting group can be removed with acid (trifluoroacetic acid), basified with triethylamine, and purified to obtain compound (VI).

Process 2

Step 1: R ^z1 , R ^z2 and R ⁷ in the structure of compound (VI) are consistent with those in the structure of (IV). Compound (VI) is activated with an acylating agent (diphenyl carbonate or phenyl chloroformate) to synthesize compound (VII). Compound (VI) is acylated to synthesize compound (VII) using a base (lithium hexamethyldisilazide) as a deprotonating agent.

Step 2: R ¹⁰ and R ¹¹ on the 2-aminopyridine ring are consistent with those in structure (IV), and replace the phenol group in compound (VII). Using base (lithium hexamethyldisilazide) as a deprotonating agent, compound (VIII) is synthesized, and R ^z1 , R ^z2 and R ⁷ in compound (VIII) are consistent with those in structure (IV).

Step 3: R ^z1 , R ^z2 and R ⁷ in compound (VIII) are consistent with those in the structure of (IV). The acetal protecting group can be removed with acid, alkalized with sodium bicarbonate, and purified to obtain compound (IV).

Process 3

Step 1: R ^z2 and R ⁷ in the structure of compound (VI-1) are consistent with those in structure (IV). Compound (VI-1) is subjected to carbon-carbon coupling with a boronic acid compound using palladium catalysis to synthesize compound (VI-5). The catalyst ([1,1′-bis(diphenylphosphino)ferrocene]palladium dichloride) and a base (potassium carbonate) are used. R ^z1 , R ^z2 , and R ¹⁰ in the structure of compound (VI-5) are consistent with those in structure (IV). The operations of steps 2, 3, and 4 are described in Scheme 2.

definition

Unless stated otherwise, the following terms used in the specification and claims have the following meanings.

The notation "Cx-y" used herein represents a range of carbon atoms, wherein x and y are both integers, for example, a C3-8 cyclyl group represents a cyclyl group having 3-8 carbon atoms, and a -C0-2 alkyl group represents an alkyl group having 0-2 carbon atoms, wherein -C0 alkyl refers to a chemical single bond.

As used herein, the term "alkyl" refers to a saturated aliphatic hydrocarbon group, including straight and branched chain groups of 1 to 20 carbon atoms, for example, straight and branched chain groups of 1 to 18 carbon atoms, 1 to 12 carbon atoms, 1 to 8 carbon atoms, 1 to 6 carbon atoms, or 1 to 4 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, the tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, and various branched chain isomers thereof. The alkyl group may be substituted or unsubstituted.

In this article, term " cyclic radical " refers to saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon group, and it includes 3 to 12 ring carbon atoms, for example, can be 3 to 12, 3 to 10, 3 to 8 or 3 to 6 ring carbon atoms, or can be 3, 4, 5, 6, 7, 8 yuan ring.The non-limiting embodiment of monocyclic cyclic radical comprises cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl etc.Cyclic radical can be substituted or unsubstituted.

As used herein, the term "heterocyclyl" refers to a saturated or partially unsaturated monocyclic or polycyclic ring group containing 3 to 20 ring carbon atoms, for example, 3 to 16, 3 to 12, 3 to 10, 3 to 8 or 3 to 6 ring atoms, wherein one or more ring atoms are selected from nitrogen, oxygen or S(O)m (wherein m is an integer from 0 to 2) heteroatoms, but excluding the ring portion of -O-O-, -O-S- or -S-S-, and the remaining ring atoms are carbon. Preferably, it contains 3 to 12 ring atoms, of which 1 to 4 are heteroatoms, more preferably the heterocyclyl ring contains 3 to 10 ring atoms, most preferably a 5-membered ring or a 6-membered ring, of which 1 to 4 are heteroatoms, more preferably 1 to 3 are heteroatoms, and most preferably 1 to 2 are heteroatoms. Non-limiting examples of monocyclic heterocyclyls include pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, homopiperazinyl, and the like. Polycyclic heterocyclic groups include spiro, fused and bridged heterocyclic groups.

In this article, the term "spiro heterocyclic radical" refers to a polycyclic heterocyclic group of 5 to 20 yuan, one atom (called spiral atom) shared between the monocycles, wherein one or more ring atoms are selected from nitrogen, oxygen or S (O) m (wherein m is an integer 0 to 2) heteroatom, and the remaining ring atoms are carbon. These can contain one or more double bonds, but no ring has a completely conjugated π electron system. Preferably, it is 6 to 14 yuan, more preferably 7 to 10 yuan. According to the number of shared spiral atoms between the rings, the spiro heterocyclic radical is divided into a monospiro heterocyclic radical, a dispiro heterocyclic radical or a polyspiro heterocyclic radical, preferably a monospiro heterocyclic radical and a dispiro heterocyclic radical. More preferably, it is a 4 yuan/4 yuan, 4 yuan/5 yuan, 4 yuan/6 yuan, 5 yuan/5 yuan or 5 yuan/6 yuan monospiro heterocyclic radical. The non-limiting examples of spiro heterocyclic radicals include

In this article, the term "fused heterocyclic radical" refers to a polycyclic heterocyclic group of 5 to 20 members, each ring in the system shares a pair of atoms adjacent to the other rings in the system, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron system, wherein one or more ring atoms are selected from nitrogen, oxygen or S (O) m (wherein m is an integer 0 to 2) heteroatom, and the remaining ring atoms are carbon. Preferably it is 6 to 14 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic fused heterocyclic groups, preferably bicyclic or tricyclic, more preferably 5 yuan / 5 yuan or 5 yuan / 6 yuan bicyclic fused heterocyclic groups. Non-limiting examples of fused heterocyclic groups include

The heterocyclyl ring may be fused to an aryl, heteroaryl or cyclyl ring, wherein the ring attached to the parent structure is a heterocyclyl, non-limiting examples of which include:

etc. The heterocyclic group may be substituted or unsubstituted.

As used herein, the term "aryl" refers to a 6- to 14-membered all-carbon monocyclic or fused polycyclic (i.e., rings that share adjacent pairs of carbon atoms) group, a polycyclic (i.e., rings with adjacent pairs of carbon atoms) group having a conjugated π electron system, preferably 6- to 10-membered, such as phenyl and naphthyl, with phenyl being most preferred. The aryl ring may be fused to a heteroaryl, heterocyclyl, or cyclyl ring, wherein the ring attached to the parent structure is the aryl ring, non-limiting examples of which include:

Aryl groups can be substituted or unsubstituted.

As used herein, the term "heteroaryl" refers to a heteroaromatic system containing 1 to 4 heteroatoms and 5 to 14 ring atoms, wherein the heteroatoms include oxygen, sulfur, and nitrogen. Preferably, it is 5 to 10-membered. More preferably, the heteroaryl group is 5-membered or 6-membered, such as furyl, thienyl, pyridyl, pyrrolyl, N-alkylpyrrolyl, pyrimidinyl, pyrazinyl, imidazolyl, tetrazolyl, oxazolyl, isoxazolyl, etc. The heteroaryl ring may be fused to an aryl, heterocyclyl, or cyclyl ring, wherein the ring connected to the parent structure is a heteroaryl ring. Non-limiting examples include:

Heteroaryl groups can be substituted or unsubstituted.

As used herein, the term "halogen" refers to fluorine, chlorine, bromine or iodine.

As used herein, the term "cyano" refers to -CN.

As used herein, the term "alkenyl" refers to a linear, branched, or cyclic non-aromatic hydrocarbon group containing at least one carbon-carbon double bond. One to three carbon-carbon double bonds may be present, preferably one. Examples include ethenyl, propenyl, butenyl, 2-methylbutenyl, and cyclohexenyl. These alkenyl groups may be substituted. The term "C2-4 alkenyl" refers to an alkenyl group having 2 to 4 carbon atoms.

As used herein, the term "alkynyl" refers to a straight-chain, branched, or cyclic hydrocarbon group containing at least one carbon-carbon triple bond. There may be 1 to 3 carbon-carbon triple bonds, preferably 1. Examples include ethynyl, propynyl, butynyl, and 3-methylbutynyl. The term "C2-4 alkynyl" refers to an alkynyl group having 2 to 4 carbon atoms.

As used herein, the term "alkoxy" refers to a cyclic or acyclic alkyl group having the stated number of carbon atoms attached through an oxygen bridge, including alkyloxy, cycloalkyloxy, and heterocycloalkyloxy. Thus, "alkoxy" encompasses the aforementioned definitions of alkyl, heterocycloalkyl, and cycloalkyl. "Optional" or "optionally" means that the subsequently described event or circumstance may, but need not, occur, and the description includes instances where the event or circumstance occurs or does not occur. For example, "a heterocyclic group optionally substituted with an alkyl group" means that an alkyl group may, but need not, be present, and the description includes both instances where the heterocyclic group is substituted with an alkyl group and instances where the heterocyclic group is not substituted with an alkyl group.

As used herein, the term "substituted" refers to a group in which one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3, are independently replaced by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and a person skilled in the art will be able to determine (by experiment or theory) which substitutions are possible or impossible without undue effort. For example, an amino or hydroxyl group with free hydrogen may be unstable when combined with a carbon atom with an unsaturated (e.g., olefinic) bond.

As used herein, the term "pharmaceutical composition" refers to a mixture containing one or more compounds described herein, or their physiologically/pharmaceutically acceptable salts or prodrugs, together with other chemical components, as well as other components such as physiologically/pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, facilitating absorption of the active ingredient and thereby exerting its biological activity.

The "room temperature" mentioned in the present invention refers to 15-30°C.

As used herein, the term "stable isotope derivative" includes: an isotope substituted derivative in which any hydrogen atom in Formula I is replaced by 1-5 deuterium atoms, an isotope substituted derivative in which any carbon atom in Formula I is replaced by 1-3 carbon-14 atoms, or an isotope substituted derivative in which any oxygen atom in Formula I is replaced by 1-3 oxygen-18 atoms.

The "pharmaceutically acceptable salts" of the present invention are discussed in Berge, et al., "Pharmaceutically acceptable salts", J. Pharm. Sci., 66, 1-19 (1977) and are obvious to pharmaceutical chemists. The salts are substantially non-toxic and can provide the desired pharmacokinetic properties, palatability, absorption, distribution, metabolism or excretion.

The pharmaceutically acceptable salts of the present invention can be synthesized by general chemical methods.

Generally, salts can be prepared by reacting a free base or acid with an equal chemical equivalent or excess of an acid (inorganic or organic) or base in a suitable solvent or solvent combination.

As used herein, a "prodrug" refers to a compound that is converted into the original active compound after metabolism in the body. Typically, a prodrug is inactive or less active than the active parent compound, but may provide convenient handling, administration, or improved metabolic properties.

The term "isomers" as used herein refers to compounds of formula (I) of the present invention that may have asymmetric centers and racemates, racemic mixtures, and individual diastereomers, and all such isomers, including stereoisomers and geometric isomers, are encompassed by the present invention. The geometric isomers include cis-trans isomers.

As used herein, the term "tumor" includes benign tumors and malignant tumors, such as cancer.

As used herein, the term "cancer" includes various malignancies, especially malignancies in which FGFR, especially FGFR4, is involved, including but not limited to rhabdomyosarcoma, renal cell carcinoma, myeloma, breast cancer, gastric cancer, colon cancer, bladder cancer, pancreatic cancer and hepatocellular carcinoma.

As used herein, the term "inflammatory disease" refers to any inflammatory disease in which FGFR, particularly FGFR4, is involved in the occurrence of inflammation.

Example

The present invention is further illustrated by way of examples below, but the present invention is not limited to the scope of the examples. Experimental methods in the following examples where specific conditions are not specified were performed according to conventional methods and conditions, or selected according to the product specifications.

The structures of all compounds of the present invention can be identified by nuclear magnetic resonance (1H NMR) and/or mass spectrometry (MS).

¹ H NMR chemical shifts (δ) are reported in PPM (unit: 10 ⁻⁶ PPM). NMR was performed on a Bruker AVANCE-400 spectrometer. Suitable solvents were deuterated chloroform (CDCl ₃ ), deuterated methanol (CD ₃ OD), deuterated dimethyl sulfoxide (DMSO-d ⁶ ), and tetramethylsilane as an internal standard (TMS).

Low-resolution mass spectrometry (MS) was performed on an Agilent 1260 HPLC/6120 mass spectrometer using an Agilent ZORBAXX DB-C18, 4.6 × 50 mm, 3.5 μm column. Gradient elution conditions were: 0: 95% solvent A1 and 5% solvent B1; 1-2: 5% solvent A1 and 95% solvent B1; 2.01-2.50: 95% solvent A1 and 5% solvent B1. Percentages represent the volume percentage of a particular solvent relative to the total solvent volume. Solvent A1: 0.01% formic acid in water; Solvent B1: 0.01% formic acid in acetonitrile. Percentages represent the volume percentage of the solute relative to the total solvent volume.

Thin layer silica gel plates are Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plates. Column chromatography generally uses Yantai Huanghai 100-200 or 200-300 mesh silica gel as a carrier.

The known starting materials of the present invention can be synthesized by methods known in the art, or can be purchased from companies such as Acros Organics, Aldrich Chemical Company, Accela ChemBio Inc, Shanghai Bid Pharmaceutical, Shanghai Aladdin Chemical, Shanghai Myrel Chemical, and J&K Chemical.

Unless otherwise specified in the examples, all solvents used in the reactions were anhydrous solvents, with commercially available anhydrous tetrahydrofuran being used. Sodium block was used as a desiccant, and benzophenone was used as an indicator. The reaction was refluxed under nitrogen until the solution turned blue-purple, then collected by distillation and stored at room temperature under nitrogen. Other anhydrous solvents were purchased from Aladdin Chemical and J&K Chemical. The transfer and use of all anhydrous solvents were carried out under nitrogen unless otherwise specified.

Unless otherwise specified in the examples, all reactions were carried out under an argon or nitrogen atmosphere.

Argon atmosphere or nitrogen atmosphere means that the reaction bottle is connected to an argon or nitrogen balloon with a capacity of about 1 L.

Hydrogen atmosphere means that the reaction bottle is connected to a hydrogen balloon with a capacity of about 1L.

Carbon monoxide atmosphere means that the reaction bottle is connected to a carbon monoxide balloon with a volume of about 1L;

The hydrogenation reaction is usually carried out by evacuating the chamber and filling it with hydrogen, and the operation is repeated three times.

Unless otherwise specified in the examples, the reaction temperature is room temperature, which ranges from 15°C to 30°C.

The reaction progress in the examples was monitored by thin layer chromatography (TLC). The developing solvent systems used in the reactions were A: dichloromethane and methanol system; B: petroleum ether and ethyl acetate system. The volume ratio of the solvents was adjusted according to the polarity of the compounds.

The eluent system for column chromatography and the developing solvent system for thin-layer chromatography used for purifying compounds include A: dichloromethane and methanol system; B: petroleum ether and ethyl acetate system. The volume ratio of the solvents is adjusted according to the polarity of the compound, and a small amount of triethylamine and acidic or alkaline reagents can also be added for adjustment.

Example 1

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

first step

6-Chloro-2-hydroxymethylpyridine

Compound 6-chloro-2-pyridinecarboxylic acid methyl ester 1a (1.00 g, 5.85 mmol), sodium borohydride (0.38 g, 9.95 mmol), and ethanol (15 mL) were mixed and stirred at room temperature for 6 hours. The mixture was quenched with 30 mL of water and extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated brine (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolventized under reduced pressure to obtain the desired product, 6-chloro-2-hydroxymethylpyridine 1b (0.70 g, yellow oil). Yield: 84%. The product was used directly in the next reaction without purification.

¹ H NMR (400MHz, CDCl3) δ7.68 (dd, J=8.0, 7.6Hz, 1H), 7.27 (d, J=8.0Hz, 1H), 7.26 (d, J=7.6Hz, 1H), 4.76 (d, J=5.2Hz, 2H), 3.07 (t, J=5.6Hz, 1H).

Step 2

(6-(Methylamino)pyridin-2-yl)methanol

Compound 6-chloro-2-hydroxymethylpyridine 1b (1.50 g, 10.5 mmol) and methylamine (15 mL, 30% ethanol solution) were mixed and stirred at 100°C for 48 hours. The mixture was cooled to room temperature and desolventized under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 10:1-1:2) to obtain the desired product (6-(methylamino)pyridin-2-yl)methanol 1c (0.70 g, yellow oil) in a 48% yield.

MS m/z (ESI): 139 [M+1].

Step 3

6-(Methylamino)methylpyridine aldehyde

Compound (6-(methylamino)pyridin-2-yl)methanol 1c (0.60 g, 4.35 mmol), manganese dioxide (3.78 g, 43.5 mmol), and dichloromethane (15 mL) were mixed and stirred at 40°C for 16 hours. The mixture was cooled to room temperature, filtered, and the filtrate was desolvated under reduced pressure to obtain the desired product, 6-(methylamino)methylpyridine aldehyde 1d (0.50 g, yellow solid), in a 72% yield.

MS m/z (ESI): 137 [M+1].

Step 4

6-(1,3-dioxolane-2-yl)-N-methylpyridin-2-amine

Compound 6-(methylamino)methylpyridine aldehyde 1d (0.80 g, 5.95 mmol), ethylene glycol (1.80 g, 29.7 mmol), p-toluenesulfonic acid (0.10 g, 0.60 mmol), 4A molecular sieves (0.2 g), and toluene (15 mL) were mixed and stirred at 120°C for 5 hours. After cooling to room temperature, the mixture was diluted with 30 mL of water and extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated brine (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 6:1-2:1) to obtain the desired product, 6-(1,3-dioxolane-2-yl)-N-methylpyridin-2-amine (0.60 g, yellow solid) in a 57% yield.

MS m/z(ESI):181[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ7.51 (t, J=8.0Hz, 1H), 6.83 (d, J=7.2Hz, 1H), 6.39 (d, J=8.0Hz, 1H), 5.72 (s, 1 H), 4.66 (brs, 1H), 4.20-4.14 (m, 2H), 4.09-4.03 (m, 2H), 2.93 (d, J=4.8Hz, 3H).

Step 5

Phenyl(6-(1,3-dioxolane-2-yl)pyridin-2-yl)(methyl)aminocarboxylate

Compound 6-(1,3-dioxolane-2-yl)-N-methylpyridin-2-amine 1e (54 mg, 0.30 mmol), diphenyl carbonate (1.28 g, 0.60 mmol), lithium hexamethyldisilazide (0.41 mL, 0.41 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (3 mL) were mixed and stirred at 0°C for 2 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate 4:1) to obtain the target product (6-(1,3-dioxolane-2-yl)pyridin-2-yl)(methyl)aminocarboxylate (60 mg, white solid) in a 67% yield.

MS m/z(ESI):301[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ7.96-7.94(m, 1H), 7.74-7.70(m, 1H), 7.45-7.32(m, 2H), 7.40-7.38(m, 1H), 7.28-7.25( m, 1H), 7.20-7.17 (m, 2H), 5.87 (s, 1H), 4.24-4.21 (m, 2H), 4.13-4.09 (m, 2H), 3.67 (s, 3H).

Step 6

1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(4-chloro-5-cyanopyridin-2-yl)-1-methylurea

Phenyl(6-(1,3-dioxolane-2-yl)pyridin-2-yl)(methyl)aminocarboxylate 1f (60 mg, 0.20 mmol), 6-amino-4-chloronicotinonitrile (76 mg, 0.50 mmol), lithium hexamethyldisilazide (0.4 mL, 0.4 mmol, 1 M tetrahydrofuran solution), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 2 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (dichloromethane/methanol 30:1) to give the target product 1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(4-chloro-5-cyanopyridin-2-yl)-1-methylurea (22 mg, white solid) in a yield of 31%.

MS m/z(ESI):360&362[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ13.44(s, 1H), 8.87(s, 1H), 8.33(s, 1H), 8.04-8.00(m, 1H), 7.42(d, J=8.4Hz, 1H), 7 .36 (d, J=7.6Hz, 1H), 5.82 (s, 1H), 4.24-4.21 (m, 2H), 4.04-4.01 (m, 2H), 3.44 (s, 3H).

Step 7

1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea

1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(4-chloro-5-cyanopyridin-2-yl)-1-methylurea 1 g (22 mg, 0.06 mmol), 2-methoxyethylamine (14 mg, 0.18 mmol), diisopropylethylamine (24 mg, 0.18 mmol), and N,N-dimethylacetamide (0.4 mL) were mixed and stirred at 70°C for 16 hours. The mixture was diluted with 10 mL of water and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 2:1) to give the target product 1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea (10 mg, white solid) in a yield of 41%.

MS m/z (ESI): 399 [M+1].

Step 8

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

Compound 1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea 1h (10 mg, 0.025 mmol), hydrochloric acid (0.5 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 6 hours. The mixture was quenched with saturated sodium carbonate solution and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (dichloromethane/methanol 20:1) to give the target product 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea (8 mg, white solid) in a yield of 90%.

MS m/z(ESI):355[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.03 (s, 1H), 10.18 (s, 1H), 8.21 (s, 1H), 8.02-7.98 (m, 1H), 7.75 (d, J=7.6Hz, 1H), 7.59 (s, 1H), 7. 36 (d, J=8.4Hz, 1H), 5.82 (s, 1H), 3.66 (t, J=4.8Hz, 2H), 3.56 (s, 3H), 3.52-3.50 (m, 2H), 3.44 (s, 3H).

Example 2

3-(6-chloropyrimidin-4-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

Example 2 was synthesized by referring to the operating procedures of Example 1, except that 6-chloropyrimidin-4-amine was used instead of 6-amino-4-chloronicotinonitrile in the sixth step.

MS m/z(ESI):292&294[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.51 (s, 1H), 10.18 (s, 1H), 8.67 (s, 1H), 8.25 (s, 1H), 8.02 (dd, J=8.4, 7.6Hz, 1H), 7.79 (d, J=7.6Hz, 1H), 7.40 (d, J=8.4Hz, 1H), 3.59 (s, 3H).

Example 3

3-(5-cyanopyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

Example 3 was synthesized by referring to the operating procedures of Example 1, except that 6-aminonicotinonitrile was substituted for 6-amino-4-chloronicotinonitrile in the sixth step.

MS m/z(ESI):282[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.49 (s, 1H), 10.19 (s, 1H), 8.60 (s, 1H), 8.34 (d, J = 7.6Hz, 1H), 8.03 (dd, J = 8.8, 7. 6Hz, 1H), 7.95-7.93 (m, 1H), 7.77 (d, J=7.6Hz, 1H), 7.39 (d, J=8.8Hz, 1H), 3.59 (s, 3H).

Example 4

3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

Example 4 was synthesized by referring to the operating steps of Example 1, except that 4-chloropyridin-2-amine was used instead of 6-amino-4-chloronicotinonitrile in the sixth step.

MS m/z(ESI):291&293[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.09 (s, 1H), 10.19 (s, 1H), 8.29 (d, J = 2.0Hz, 1H), 8.22 (d, J = 5.2Hz, 1H), 8.00 (dd, J = 8.4, 7. 2Hz, 1H), 7.74 (d, J=7.2Hz, 1H), 7.37 (d, J=8.4Hz, 1H), 7.03 (dd, J=5.2, 2.0Hz, 1H), 3.58 (s, 3H).

Example 5

3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

Example 5 was synthesized by referring to the procedure of Example 1, except that isopropylamine was used instead of 2-methoxyethylamine in the seventh step.

MS m/z(ESI):339[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.00(s, 1H), 10.18(s, 1H), 8.20(s, 1H), 8.02-7.97(m, 1H), 7.76-7.74(m, 1H), 7.60(s, 1H), 7.57-7.54 (m, 1H), 7.36 (d, J=8.4Hz, 1H), 3.92-3.89 (m, 1H), 3.57 (s, 3H), 1.34 (d, J=6.4Hz, 6H).

Example 6

3-(5-cyano-4-isopropoxypyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

first step

6-Amino-4-isopropoxynicotinonitrile

Compound 6-amino-4-chloronicotinonitrile 6a (46 mg, 0.30 mmol), isopropyl alcohol (90 mg, 1.50 mmol), sodium hydride (72 mg, 1.80 mmol, in a 60% mineral oil mixture), and N-methylpyrrolidone (1.5 mL) were mixed and stirred at 70°C for 24 hours. After cooling to room temperature, the mixture was quenched with 20 mL of water and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolventized under reduced pressure. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate 1:1) to obtain the desired product, 6-amino-4-isopropoxynicotinonitrile 6b (16 mg, yellow solid). Yield: 30%.

MS m/z (ESI): 178 [M+1].

Step 2

1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(5-cyano-4-isopropoxypyridin-2-yl)-1-methylurea

Example 6c was synthesized by following the procedures of Step 6 of Example 1, substituting 6-amino-4-isopropoxynicotinonitrile for 6-amino-4-chloronicotinonitrile. The target product, 1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(5-cyano-4-isopropoxypyridin-2-yl)-1-methylurea 6c, was obtained (8 mg, white solid). Yield: 46%.

MS m/z (ESI): 384 [M+1].

Step 3

3-(5-cyano-4-isopropoxypyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

Example 6 was synthesized by referring to the procedure of Step 8 of Example 1. 1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(5-cyano-4-isopropoxypyridin-2-yl)-1-methylurea was substituted with 1-(6-(1,3-dioxolane-2-yl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea. The target product, 3-(5-cyano-4-isopropoxypyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea 7, was obtained (5 mg, white solid). Yield: 71%.

MS m/z(ESI):340[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.33(s, 1H), 10.19(s, 1H), 8.38(s, 1H), 8.02(dd, J=8.0, 7.6Hz, 1H), 7.96(s, 1H), 7.77(d , J=7.6Hz, 1H), 7.38 (d, J=8.0Hz, 1H), 4.88-4.86 (m, 1H), 3.58 (s, 3H), 1.48 (d, J=6.0Hz, 6H).

Example 7

3-(5-cyano-4-(2-methoxyethoxy)pyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea

Example 7 was synthesized by following the procedure of Example 6, except that 2-methoxyethylene glycolamine was used instead of isopropyl alcohol in the first step.

MS m/z(ESI):356[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.38 (s, 1H), 10.18 (s, 1H), 8.40 (s, 1H), 8.02 (dd, J=8.4, 7.2Hz, 1H), 7.99 (s, 1H), 7.77 (d, J=7.2H z, 1H), 7.38 (d, J=8.4Hz, 1H), 4.38 (t, J=4.8Hz, 2H), 3.86 (t, J=4.8Hz, 2H), 3.58 (s, 3H), 3.51 (s, 3H).

Example 8

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(4-formylpyrimidin-2-yl)-1-methylurea,

first step

4-(Dimethoxymethyl)pyrimidin-2-amine

Compound 1,1-dimethoxy-N,N-dimethylmethanamine 8a (4.90 g, 41.36 mmol) and 1,1-dimethoxypropan-2-one (4.90 g, 41.36 mmol) were mixed and stirred at 100°C for 16 hours. The mixture was then desolvated under reduced pressure. The residue was mixed with guanidine hydrochloride (4.30 g, 45.00 mmol), sodium hydroxide (1.80 g, 45.00 mmol), and water (15 mL) and stirred at room temperature for 48 hours. Filtration afforded the desired product, 4-(dimethoxymethyl)pyrimidin-2-amine 8b (2.00 g, white solid). Yield: 27%.

MS m/z(ESI):170[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ 8.36 (d, J=4.8Hz, 1H), 6.87 (d, J=5.2Hz, 1H), 5.16 (s, 1H), 5.15 (brs, 2H), 3.42 (s, 6H).

Step 2

4-(Dimethoxymethyl)-N-methylpyrimidin-2-amine

Compound 4-(dimethoxymethyl)pyrimidin-2-amine 8b (1.00 g, 5.65 mmol), iodomethane (2.80 g, 19.77 mmol), and acetone (30 mL) were mixed and stirred at 70°C for 16 hours, cooled to room temperature, and filtered. The solid was then mixed with 10% sodium hydroxide (8 mL), stirred at 80°C for 0.5 hours, and cooled to room temperature. The mixture was quenched with 250 mL of ice water and extracted with dichloromethane (50 mL x 2). The organic phase was washed with saturated brine (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolvated under reduced pressure to obtain the desired product, 4-(dimethoxymethyl)-N-methylpyrimidin-2-amine 8c (0.70 g, yellow oil). Yield: 67%.

MS m/z(ESI):184[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ 8.37 (d, J=4.8Hz, 1H), 6.77 (d, J=5.2Hz, 1H), 5.18 (brs, 1H), 5.13 (s, 1H), 3.42 (s, 6H), 3.03 (d, J=5.2Hz, 3H).

Step 3

Phenyl (4-(dimethoxymethyl) pyrimidin-2-yl)(methyl) aminocarboxylate

Compound 4-(dimethoxymethyl)-N-methylpyrimidin-2-amine 8c (0.20 g, 1.09 mmol), diphenyl carbonate (0.47 g, 2.19 mmol), lithium hexamethyldisilazide (1.5 mL, 1.51 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (5 mL) were mixed and stirred at 0°C for 2 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified on preparative silica gel (dichloromethane/methanol 50:1) to obtain the desired product, phenyl (4-(dimethoxymethyl)pyrimidin-2-yl)(methyl)aminocarboxylate 8d (40 mg, white solid). Yield: 12%.

MS m/z (ESI): 304 [M+1].

Step 4

3-(4-chloro-5-cyanopyridin-2-yl)-1-(4-(dimethoxymethyl)pyrimidin-2-yl)-1-methylurea

Phenyl(4-(dimethoxymethyl)pyrimidin-2-yl)(methyl)aminocarboxylate 8d (40 mg, 0.13 mmol), 6-amino-4-chloronicotinonitrile (21 mg, 0.13 mmol), lithium hexamethyldisilazide (0.26 mL, 0.26 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 2 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with dichloromethane (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified on preparative silica gel (dichloromethane/methanol 50:1) to obtain the desired product, 3-(4-chloro-5-cyanopyridin-2-yl)-1-(4-(dimethoxymethyl)pyrimidin-2-yl)-1-methylurea 8e (25 mg, white solid). Yield: 52%.

MS m/z(ESI):363&365[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.59 (s, 1H), 8.74 (d, J=5.2Hz, 1H), 8.56 (s, 1H), 8.55 (s, 1H), 7.30 (d, J=5.2Hz, 1H), 5.34 (s, 1H), 3.68 (s, 3H), 3.51 (s, 6H).

Step 5

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(4-(dimethoxymethyl)pyrimidin-2-yl)-1-methylurea

Compound 3-(4-chloro-5-cyanopyridin-2-yl)-1-(4-(dimethoxymethyl)pyrimidin-2-yl)-1-methylurea 8e (18 mg, 0.05 mmol), 2-methoxyethylamine (15 mg, 0.20 mmol), diisopropylethylamine (13 mg, 0.10 mmol), and N,N-dimethylacetamide (0.4 mL) were mixed and stirred at 70°C for 16 hours. The mixture was diluted with 10 mL of water and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (dichloromethane/methanol 50:1) to afford the desired product, 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(4-(dimethoxymethyl)pyrimidin-2-yl)-1-methylurea 8f (15 mg, yellow solid). Yield: 75%.

MS m/z(ESI):402[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.11 (s, 1H), 8.72 (d, J = 5.2Hz, 1H), 8.23 (s, 1H), 7.65 (s, 1H), 7.25 (d, J = 5.2Hz, 1H), 5.31 (s, 1H), 5.30(brs, 1H), 3.67-3.65(m, 2H), 3.66(s, 3H), 3.54-3.52(m, 2H), 3.51(s, 6H), 3.44(s, 3H).

Step 6

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(4-formylpyrimidin-2-yl)-1-methylurea

Compound 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(4-(dimethoxymethyl)pyrimidin-2-yl)-1-methylurea 8f (15 mg, 0.04 mmol), hydrochloric acid (0.8 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 3 hours. The mixture was quenched with saturated sodium carbonate solution and extracted with dichloromethane (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified on preparative silica gel (dichloromethane/methanol 50:1) to obtain the desired product, 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(4-formylpyrimidin-2-yl)-1-methylurea 8f (6 mg, white solid). Yield: 27%.

MS m/z(ESI):356[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.88 (s, 1H), 10.06 (s, 1H), 8.95 (d, J = 4.8Hz, 1H), 8.23 (s, 1H), 7.65 (s, 1H), 7.52 (d, J = 4.8Hz, 1H), 5.35(brs, 1H), 3.73(s, 3H), 3.69-3.65(m, 2H), 3.55-3.52(m, 2H), 3.45(s, 3H).

Example 9

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

first step

Di-tert-butyl (5-bromo-6-methylpyridin-2-yl)imido dicarbonate

Compound 5-bromo-6-methylpyridin-2-amine 9a (9.30 g, 50.00 mmol), 2-tert-butyl 2-carbonate (27.00 g, 125 mmol), N,N-dimethylpyridin-4-amine (0.31 g, 2.50 mmol), and tetrahydrofuran (300 mL) were mixed and stirred at room temperature for 16 hours. The mixture was quenched with 300 mL of water and extracted with ethyl acetate (200 mL x 2). The organic phase was washed with saturated brine (200 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolventized under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 100:0-95:5) to obtain the target product, di-tert-butyl (5-bromo-6-methylpyridin-2-yl)imido dicarbonate 9b (15.00 g, white solid). Yield: 78%.

¹ H NMR (400MHz, CDCl ₃ ) δ 7.80 (d, J=8.4Hz, 1H), 7.00 (d, J=8.4Hz, 1H), 2.61 (s, 3H), 1.46 (s, 18H).

Step 2

Di-tert-butyl (5-bromo-6-(dibromomethyl)pyridin-2-yl)imido dicarbonate

Di-tert-butyl (5-bromo-6-methylpyridin-2-yl)imido dicarbonate 9b (3.86 g, 10.00 mmol), 1-bromopyrrolidine-2,5-dione (4.45 g, 25.00 mmol), benzoperoxyanhydride (0.24 g, 1.00 mmol), and carbon tetrachloride (100 mL) were mixed and stirred at 90°C for 16 hours. The mixture was cooled to room temperature and desolvated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 94:6) to obtain the target product, di-tert-butyl (5-bromo-6-(dibromomethyl)pyridin-2-yl)imido dicarbonate 9c (4.00 g, yellow solid) in a yield of 74%.

¹ H NMR (400MHz, CDCl ₃ ) δ 7.82 (d, J=8.0Hz, 1H), 7.26 (d, J=8.0Hz, 1H), 7.08 (s, 1H), 1.50 (s, 18H).

Step 3

tert-Butyl (5-bromo-6-(dimethoxy)pyridin-2-yl)aminocarboxylate

Compound di-tert-butyl (5-bromo-6-(dibromomethyl)pyridin-2-yl)imidodicarbonate 9c (5.00 g, 96.00 mmol), potassium hydroxide (2.23 g, 0.38 mol), and methanol (30 mL) were mixed and stirred at 70°C for 16 hours. The mixture was cooled to room temperature and desolventized under reduced pressure. The residue was dissolved in 50 mL of water and extracted with ethyl acetate (50 mL x 3). The organic phase was washed with saturated brine (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was filtered to remove the desiccant, and desolventized under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 100:0-12:1) to obtain the target product, tert-butyl (5-bromo-6-(dimethoxy)pyridin-2-yl)aminocarboxylate 9d (0.50 g, yellow solid) in a yield of 16%.

MS m/z(ESI):347&349[M+1].

Step 4

tert-Butyl (5-bromo-6-(dimethoxy)pyridin-2-yl)(methyl)aminocarboxylate

Compound tert-butyl (5-bromo-6-(dimethoxy)pyridin-2-yl)aminocarboxylate 9d (1.20 g, 3.47 mmol), sodium hydride (0.18 g, 4.51 mmol, 60% mineral oil mixture), iodomethane (0.59 g, 4.16 mmol), and N,N-dimethylformamide (8 mL) were mixed and stirred at room temperature for 16 hours. The mixture was quenched with 30 mL of water and extracted with ethyl acetate (50 mL x 3). The organic phase was washed with saturated brine (50 mL x 3). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolventized under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 100:0-96:4) to obtain the target product, tert-butyl (5-bromo-6-(dimethoxy)pyridin-2-yl)(methyl)aminocarboxylate 9e (0.40 g, yellow oil) in a 32% yield.

MS m/z(ESI):361&363[M+1].

Step 5

6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl) nicotinate

Compound tert-butyl(5-bromo-6-(dimethoxy)pyridin-2-yl)(methyl)carbamic acid 9e (0.45 g, 1.25 mmol), palladium acetate (28 mg, 0.13 mmol), 1,1-bis(diphenylphosphino)ferrocene (0.14 g, 0.25 mmol), triethylamine (0.25 g, 2.50 mmol), methanol (3 mL), and N,N-dimethylformamide (20 mL) were mixed and stirred at 100°C under a carbon monoxide atmosphere (1 atm) for 16 hours. The mixture was quenched with 100 mL of water and extracted with ethyl acetate (50 mL x 3). The organic phase was washed with saturated brine (50 mL x 3). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 94:6) to obtain the target product, methyl-6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)nicotinate 9f (0.25 g, yellow oil) in a yield of 59%.

MS m/z(ESI):341[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ 8.07 (d, J=8.8Hz, 1H), 7.84 (d, J=8.8Hz, 1H), 6.08 (s, 1H), 3.91 (s, 3H), 3.52 (s, 6H), 3.41 (s, 3H), 1.53 (s, 9H).

Step 6

tert-Butyl (6-(dimethoxymethyl)-5-(hydroxymethyl)pyridin-2-yl)(methyl)aminocarboxylate

Methyl 6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)nicotinate 9f (0.30 g, 0.88 mmol), sodium borohydride (0.67 g, 17.65 mmol), anhydrous calcium chloride (0.19 g, 1.77 mmol), and methanol (10 mL) were mixed and stirred at 65°C for 8 hours. The mixture was quenched with 10 mL of water and extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated brine (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolvated under reduced pressure to obtain the target product, tert-butyl (6-(dimethoxymethyl)-5-(hydroxymethyl)pyridin-2-yl)(methyl)aminocarboxylate 9g (0.20 g, white solid) in a yield of 73%.

MS m/z (ESI): 313 [M+1].

Step 7

tert-Butyl (5-(bromomethyl)-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate

The compound tert-butyl (6-(dimethoxymethyl)-5-(hydroxymethyl)pyridin-2-yl)(methyl)aminocarboxylate 9g (0.20g, 0.64mmol), phosphorus tribromide (0.21g, 0.77mmol), and dichloromethane (5mL) were mixed and stirred at 0°C for 1 hour. The mixture was quenched with 10mL of aqueous sodium bicarbonate solution and extracted with ethyl acetate (50mL×2). The organic phase was washed with saturated brine (50mL×2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 100:0-93:7) to obtain the target product, tert-butyl (5-(bromomethyl)-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate 9h (0.15g, colorless solid) in a yield of 63%.

MS m/z(ESI):375&377[M+1].

Step 8

tert-Butyl-(6-(2-methoxyethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate

Compound tert-butyl (5-(bromomethyl)-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate 9h (70 mg, 0.19 mmol), morpholin-3-one (38 mg, 0.38 mmol), sodium hydride (19 mg, 0.47 mmol, 60% mineral oil mixture) and N,N-dimethylformamide (3 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with water and extracted with ethyl acetate (20 mL×2). The organic phase was washed with saturated brine (20 mL×2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate 1.5:1) to obtain the target product tert-butyl-(6-(2-methoxyethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate 9i (70 mg, white solid) in a yield of 95%.

MS m/z(ESI):396[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ7.63-7.61(m, 2H), 5.22(s, 1H), 4.91(s, 2H), 4.26(s, 2H), 3.82-3.81(m, 2H), 3.45(s, 6H), 3.40(s, 3H), 3.27-3.26(m, 2H), 1.52(s, 9H).

Step 9

4-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)morpholin-3-one

Compound tert-butyl-(6-(2-methoxyethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate 9i (70 mg, 0.18 mmol), trifluoroacetic acid (1 mL) and dichloromethane (4 mL) were mixed and stirred at room temperature for 6 hours. The mixture was basified with triethylamine and desolvated under reduced pressure. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate 1:1) to give the target product, 4-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)morpholin-3-one 9j (46 mg, colorless solid) in an 86% yield.

MS m/z (ESI): 296 [M+1].

Step 10

Phenyl-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate

Compound 4-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)morpholin-3-one 9j (60 mg, 0.20 mmol), diphenyl carbonate (87 mg, 0.40 mmol), lithium hexamethyldisilazide (1.0 mL, 1.01 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (5 mL) were mixed and stirred at 0°C for 0.5 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate 2:1) to obtain the target product, phenyl (6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate 9k (45 mg, colorless oil) in a 54% yield.

MS m/z (ESI): 416 [M+1].

Step 11

3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea

Phenyl(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate 9k (45 mg, 0.11 mmol), 6-amino-4-chloronicotinonitrile (33 mg, 0.22 mmol), lithium hexamethyldisilazide (0.3 mL, 0.33 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (3 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 1:1) to give the target product 3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea 91 (40 mg, white solid) in a yield of 78%.

MS m/z (ESI): 475 & 477 [M+1].

Step 12

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

Compound 3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea 91 (20 mg, 0.04 mmol), 2-methoxyethylamine (13 mg, 0.17 mmol), diisopropylethylamine (11 mg, 0.08 mmol) and N,N-dimethylacetamide (0.4 mL) were mixed and stirred at 50°C for 16 hours. The mixture was diluted with 10 mL of water and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 1:1) to give the target product 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea 9m (15 mg, white solid) in a yield of 69%.

MS m/z (ESI): 514 [M+1].

Step 13

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

Compound 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea 9m (15 mg, 0.03 mmol), hydrochloric acid (0.8 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with saturated sodium carbonate solution and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was washed with ethyl acetate to give the target product 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea 9 (7 mg, white solid) in a yield of 52%.

MS m/z(ESI):468[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ12.99 (s, 1H), 10.26 (s, 1H), 8.17 (s, 1H), 7.98 (d, J = 8.4Hz, 1H), 7.56 (s, 1H), 7.27 (d, J = 8.4Hz, 1H), 5.31 (brs, 1H), 5.13 (s, 2H) , 4.26 (s, 2H), 3.89 (t, J = 4.4Hz, 2H), 3.61 (t, J = 4.0Hz, 2H), 3.53 (s, 3H), 3.51 (t, J = 4.4Hz, 2H), 3.42 (s, 3H), 3.41 (d, J = 4.0Hz, 2H).

Example 10

3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

first step

Di-tert-butyl (5-bromo-6-methylpyridin-2-yl)imido dicarbonate

Compound 5-bromo-6-methylpyridin-2-amine 10a (50 g, 0.27 mol), 2-tert-butyl 2-carbonate (145.16 g, 0.67 mol), N,N-dimethylpyridin-4-amine (3.20 g, 27.00 mmol), and tetrahydrofuran (300 mL) were mixed and stirred at room temperature for 16 hours. The mixture was quenched with 300 mL of water and extracted with ethyl acetate (200 mL x 2). The organic phase was washed with saturated brine (200 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was filtered to remove the desiccant, and the solvent was removed under reduced pressure. The residue was washed with petroleum ether to obtain the target product, di-tert-butyl (5-bromo-6-methylpyridin-2-yl)imido dicarbonate 10b (80.00 g, white solid) in a yield of 77%.

¹ H NMR (400MHz, CDCl ₃ ) δ 7.80 (d, J=8.4, 1H), 7.00 (d, J=8.4Hz, 1H), 2.61 (s, 3H), 1.46 (s, 18H).

Step 2

Di-tert-butyl (5-bromo-6-(dibromomethyl)pyridin-2-yl)imido dicarbonate

Di-tert-butyl (5-bromo-6-methylpyridin-2-yl)imido dicarbonate 10b (80 g, 0.21 mol), 1-bromopyrrolidine-2,5-dione (110.00 g, 0.63 mol), benzoperoxyanhydride (0.24 g, 0.06 mol), and carbon tetrachloride (600 mL) were mixed and stirred at 90°C for 16 hours. The mixture was cooled to room temperature and desolventized under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 94:6) to obtain the target product, di-tert-butyl (5-bromo-6-(dibromomethyl)pyridin-2-yl)imido dicarbonate 10c (90.00 g, yellow solid) in an 80% yield.

¹ H NMR (400MHz, CDCl ₃ ) δ 7.82 (d, J=8.0Hz, 1H), 7.26 (d, J=8.0Hz, 1H), 7.08 (s, 1H), 1.50 (s, 18H).

Step 3

5-Bromo-6-(dimethoxymethyl)pyridin-2-amine

Compound di-tert-butyl (5-bromo-6-(dibromomethyl)pyridin-2-yl)imido dicarbonate 10c (90.00 g, 0.17 mol), potassium hydroxide (38.52 g, 0.66 mol), and methanol (300 mL) were mixed and stirred at 70°C for 72 hours. The mixture was cooled to room temperature and desolventized under reduced pressure. The residue was dissolved in 500 mL of water and extracted with ethyl acetate (500 mL x 3). The organic phase was washed with saturated brine (500 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was filtered off, and desolventized under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 100:0-12:1) to obtain the desired product, 5-bromo-6-(dimethoxymethyl)pyridin-2-amine 10d (22 g, yellow solid) in a 54% yield.

MS m/z(ESI):247&249[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ 7.54 (d, J=8.4Hz, 1H), 6.38 (d, J=8.4Hz, 1H), 5.61 (s, 1H), 4.63 (brs, 2H), 3.48 (s, 6H).

Step 4

5-Bromo-6-(dimethoxymethyl)-N-methylpyridin-2-amine

Compound 5-bromo-6-(dimethoxymethyl)pyridin-2-amine 10d (22.00 g, 89.43 mmol), sodium methoxide (24.15 g, 447 mmol), paraformaldehyde (10.74 g, 358 mmol), and methanol (300 mL) were mixed and stirred at 80°C for 16 hours. After cooling, sodium borohydride (13.59 g, 358 mmol) was added and stirred at 80°C for 1 hour. The mixture was quenched with 300 mL of water and extracted with ethyl acetate (500 mL x 3). The organic phase was washed with saturated brine (500 mL x 3). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 100:0-96:4) to obtain the target product 5-bromo-6-(dimethoxymethyl)-N-methylpyridin-2-amine 10e (8.20 g, yellow solid) in a yield of 34%.

MS m/z(ESI):261&263[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ7.58 (d, J=8.8Hz, 1H), 6.27 (d, J=8.8Hz, 1H), 5.59 (s, 1H), 4.87 (brs, 1H), 3.48 (s, 6H), 2.87 (d, J=5.2Hz, 3H).

Step 5

2-(dimethoxymethyl)-6-(methylamino) nicotinate

Compound 5-bromo-6-(dimethoxymethyl)-N-methylpyridin-2-amine 10e (8.00 g, 30.77 mmol), palladium acetate (0.69 g, 3.08 mmol), 1,1-bis(diphenylphosphino)ferrocene (3.41 g, 6.16 mmol), triethylamine (6.22 g, 61.54 mmol), methanol (30 mL), and N,N-dimethylformamide (400 mL) were mixed and stirred at 100°C under a carbon monoxide atmosphere for 16 hours. The mixture was quenched with 700 mL of water and extracted with ethyl acetate (500 mL × 3). The organic phase was washed with saturated brine (500 mL × 3). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 94:6) to obtain the desired product, methyl 2-(dimethoxymethyl)-6-(methylamino) nicotinate 10f (2.30 g, yellow solid) in a 30% yield.

MS m/z(ESI):241[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ 8.05 (d, J=8.8Hz, 1H), 6.32 (d, J=8.8Hz, 1H), 6.24 (s, 1H), 5.30 (brs, 1H), 3.86 (s, 3H), 3.51 (s, 6H), 2.95 (d, J=5.2Hz, 3H).

Step 6

6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl) nicotinate

Methyl 2-(dimethoxymethyl)-6-(methylamino)nicotinate 10f (2.00 g, 8.33 mmol), 2-tert-butyl 2-carbonate (3.60 g, 16.67 mol), N,N-dimethylpyridin-4-amine (0.10 g, 0.83 mmol), and tetrahydrofuran (50 mL) were mixed and stirred at room temperature for 2 hours. The mixture was quenched with 100 mL of water and extracted with ethyl acetate (100 mL x 3). The organic phase was washed with saturated brine (100 mL x 3). The organic phase was dried over anhydrous sodium sulfate, the desiccant was filtered to remove the solvent, and the solvent was removed under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 94:6) to obtain the target product, methyl 6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)nicotinate 10 g (2.20 g, yellow oil), in a yield of 78%.

MS m/z(ESI):341[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ 8.07 (d, J=8.8Hz, 1H), 7.84 (d, J=8.8Hz, 1H), 6.08 (s, 1H), 3.91 (s, 3H), 3.52 (s, 6H), 3.49 (s, 3H), 1.53 (s, 9H).

Step 7

tert-Butyl (6-(dimethoxymethyl)-5-(hydroxymethyl)pyridin-2-yl)(methyl)aminocarboxylate

Methyl 6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)nicotinate (10 g, 2.20 g, 6.47 mmol), sodium borohydride (2.46 g, 60.47 mmol), anhydrous calcium chloride (1.42 g, 12.90 mmol), and methanol (20 mL) were mixed and stirred at 65°C for 2 hours. The mixture was quenched with 100 mL of water and extracted with ethyl acetate (100 mL x 2). The organic phase was washed with saturated brine (100 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolvated under reduced pressure to obtain the target product, tert-butyl-(6-(dimethoxymethyl)-5-(hydroxymethyl)pyridin-2-yl)(methyl)aminocarboxylate 10h (1.70 g, white solid) in an 84% yield.

MS m/z (ESI): 313 [M+1].

Step 8

tert-Butyl (5-(bromomethyl)-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate

The compound tert-butyl (6-(dimethoxymethyl)-5-(hydroxymethyl)pyridin-2-yl)(methyl)aminocarboxylate 10 g (1.70 g, 5.45 mmol), phosphorus tribromide (1.75 g, 6.54 mmol), and dichloromethane (50 mL) were mixed and stirred at 0°C for 0.5 hours. The mixture was quenched with 10 mL of aqueous sodium bicarbonate solution and extracted with ethyl acetate (100 mL x 2). The organic phase was washed with saturated brine (100 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate 100:0-94:6) to obtain the target product, tert-butyl-(5-(bromomethyl)-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate 10i (0.80 g, colorless solid) in a 40% yield.

MS m/z(ESI):375&377[M+1].

Step 9

tert-Butyl-(6-(2-methoxyethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate

Compound tert-butyl-(5-(bromomethyl)-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate 10i (0.28 g, 0.76 mmol), morpholin-3-one (0.15 g, 1.52 mmol), sodium hydride (76 mg, 1.88 mmol, 60% mineral oil mixture) and N,N-dimethylformamide (10 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with water and washed with ethyl acetate (5 mL). The product was extracted with 1% ethyl acetate (50 mL x 3), and the organic phase was washed with saturated brine (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate = 100:0-7:3) to obtain the target product, tert-butyl-(6-(2-methoxyethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate 10j (0.27 g, white solid) in a yield of 91%.

MS m/z (ESI): 396 [M+1].

Step 10

4-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)morpholin-3-one

The compound tert-butyl-(6-(2-methoxyethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate 10j (0.27 g, 0.18 mmol), trifluoroacetic acid (1 mL), and dichloromethane (4 mL) were mixed and stirred at room temperature for 6 hours. The solvent was removed under reduced pressure to obtain the target product, 4-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)morpholin-3-one trifluoroacetate 10k (0.27 g, yellow solid).

MS m/z (ESI): 296 [M+1].

Step 11

Phenyl-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate

Compound 4-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)morpholin-3-one trifluoroacetate 10k (0.25 g, 0.60 mmol), diphenyl carbonate (0.26 g, 1.20 mmol), lithium hexamethyldisilazide (1.8 mL, 1.80 mmol, 1 M tetrahydrofuran solution), and tetrahydrofuran (8 mL) were mixed and stirred at 0°C for 0.5 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (50 mL × 2). The organic phase was washed with saturated brine (50 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 2:1) to give the target product phenyl-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)(methyl)aminocarboxylate 101 (0.13 g, colorless oil) in a yield of 51%.

MS m/z (ESI): 416 [M+1].

Step 12

3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea

Phenyl(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)(methyl)aminocarboxylate 101 (45 mg, 0.11 mmol), 6-amino-4-chloronicotinonitrile (33 mg, 0.22 mmol), lithium hexamethyldisilazide (0.3 mL, 0.33 mmol, 1 M tetrahydrofuran solution), and tetrahydrofuran (3 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 1:1) to give the target product 3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea 10m (38 mg, white solid) in a yield of 74%.

MS m/z (ESI): 475 & 477 [M+1].

Step 13

3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

The compound 3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea 10 mg (10 mg, 0.02 mmol), isopropylamine (5 mg, 0.08 mmol), diisopropylethylamine (6 mg, 0.04 mmol) and N,N-dimethylacetamide (0.4 mL) were mixed and stirred at 50°C for 16 hours. The mixture was diluted with 10 mL of water and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 1:1) to give the target product 3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea 10n (5 mg, white solid) in a yield of 48%.

MS m/z (ESI): 498 [M+1].

Step 14

3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

Compound 3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea 10n (5 mg, 0.01 mmol), hydrochloric acid (0.8 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with saturated sodium carbonate solution and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 1:1) to give the target product 3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea 10 (3 mg, white solid) in a yield of 66%.

MS m/z(ESI):452[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ12.95 (s, 1H), 10.26 (s, 1H), 8.15 (s, 1H), 7.98 (d, J = 8.8Hz, 1H), 7.56 (s, 1H), 7.30 (d, J = 8.8Hz, 1H), 5.13 (s, 2H), 4.78- 4.76 (m, 1H), 4.26 (s, 2H), 3.89 (t, J=4.4Hz, 2H), 3.88 (brs, 1H), 3.53 (s, 3H), 3.43 (t, J=4.4Hz, 2H), 1.31 (d, J=4.8Hz, 6H).

Example 11

1-(4-chloro-5-cyanopyridin-2-yl)-3-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)urea

Example 11 was synthesized by following the procedures of Example 9, except that in the thirteenth step, 1-(4-chloro-5-cyanopyridin-2-yl)-3-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)urea was used to replace 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea.

MS m/z(ESI):429&431[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.60 (s, 1H), 10.26 (s, 1H), 8.51 (s, 1H), 8.46 (s, 1H), 8.01 (d, J=8.8Hz, 1H), 7.28 (d, J=8. 8Hz, 1H), 5.13 (s, 2H), 4.26 (s, 2H), 3.91 (t, J=3.2Hz, 2H), 3.55 (s, 3H), 3.44 (t, J=3.2Hz, 2H).

Example 12

1-(5-cyano-4-(2-methoxyethoxy)pyridin-2-yl)-3-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)urea

first step

6-Amino-4-(2-methoxyethoxy)nicotinenitrile

Compound 6-amino-4-chloronicotinonitrile 12a (60 mg, 0.39 mmol), 2-methoxyethanol (60 mg, 0.78 mmol), sodium hydride (34 mg, 0.86 mmol, in a 60% mineral oil mixture), and N-methylpyrrolidone (1.5 mL) were mixed and stirred at 70°C for 16 hours. After cooling to room temperature, the mixture was quenched with 20 mL of water and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolventized under reduced pressure. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate = 1:1) to obtain the desired product, 6-amino-4-(2-methoxyethoxy)nicotinonitrile 12b (30 mg, white solid), in a 40% yield.

MS m/z (ESI): 194 [M+1].

Step 2

3-(5-cyano-4-(2-methoxyethoxy)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

Example 12c was synthesized by following the procedure of Step 11 of Example 9, substituting 6-amino-4-(2-methoxyethoxy)nicotinonitrile for 6-amino-4-chloronicotinonitrile. The target product, 3-(5-cyano-4-(2-methoxyethoxy)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea 12c (8 mg, white solid), was obtained in a 70% yield.

MS m/z (ESI): 515 [M+1].

Step 3

3-(5-cyano-4-(2-methoxyethoxy)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea

Example 12 was synthesized by referring to the procedure of Step 13 of Example 9. 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea was substituted with 3-(5-cyano-4-(2-methoxyethoxy)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((3-carbonylmorpholine)methyl)pyridin-2-yl)-1-methylurea. The target product, 3-(5-cyano-4-isopropoxypyridin-2-yl)-1-(6-formylpyridin-2-yl)-1-methylurea 12, was obtained (6 mg, white solid). Yield: 82%.

MS m/z(ESI):469[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.34 (s, 1H), 10.27 (s, 1H), 8.36 (s, 1H), 8.00 (d, J = 8.8Hz, 1H), 7.96 (s, 1H), 7.18 (d, J = 7.6Hz, 1H), 5.13 (s, 2H), 4.35 (s , 2H), 4.26 (d, J=3.2Hz, 2H), 3.90 (t, J=3.2Hz, 2H), 3.83 (t, J=4.0Hz, 2H), 3.54 (s, 3H), 3.38 (s, 3H), 3.43 (t, J=4.0Hz, 2H).

Example 13

1-(5-cyano-4-isopropoxypyridin-2-yl)-3-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)urea

Example 13 was synthesized by following the same procedure as Example 12, except that isopropanol was used in place of 2-methoxyethanol in the first step.

MS m/z(ESI):453[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.26 (s, 1H), 10.27 (s, 1H), 8.34 (s, 1H), 8.00 (d, J = 8.8Hz, 1H), 7.93 (s, 1H), 7.32 (d, J = 8.8Hz, 1H), 5.13 (s, 2H) , 4.87-4.85 (m, 1H), 4.26 (s, 2H), 3.54 (s, 3H), 3.47 (t, J=4.4Hz, 2H), 3.43 (t, J=4.4Hz, 2H), 1.45 (d, J=3.2Hz, 6H).

Example 14

(R)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((3-carbonylmorpholino)methyl)pyridin-2-yl)-1-methylurea

Example 14 was synthesized by following the same procedure as Example 12, except that (R)-1-methoxypropan-2-ol was used in place of 2-methoxyethanol in the first step.

MS m/z(ESI):483[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.27 (s, 1H), 10.26 (s, 1H), 8.34 (s, 1H), 7.99 (s, 1H), 7.98 (d, J = 8.8Hz, 1H), 7.32 (d, J = 8.8Hz, 1H), 5.13 (s, 2H), 4.87-4.86 (m, 1H), 4.26 (s, 2H), 3.90 (t, J=4.4Hz, 2H), 3.64-3.60 (m, 1H), 3.56-3.55 (m, 1H), 3.54 (s, 3H), 3.49 (t, J=4.4Hz, 2H), 3.43 (s, 3H), 1.43 (d, J=3.2Hz, 3H).

Example 15

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea hydrochloride

first step

tert-Butyl-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)(methyl)aminocarboxylate

Compound tert-butyl-(5-(bromomethyl)-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate 15a (70 mg, 0.19 mmol), 4-methylpiperazin-2-one (43 mg, 0.38 mmol), sodium hydride (19 mg, 0.47 mmol, 60% mineral oil mixture) and N,N-dimethylformamide (3 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with water and washed with dichloromethane (2 mL). The organic phase was extracted with 20 mL × 3 (0 mL × 3), and the organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and the residue was purified on a preparative silica gel plate (petroleum ether/ethyl acetate 1.5:1) to obtain the target product, tert-butyl-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2yl)(methyl)aminocarboxylate 15b (60 mg, colorless solid) in an 83% yield.

MS m/z (ESI): 409 [M+1].

Step 2

1-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)-4-methylpiperazin-2-one

The compound tert-butyl-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)(methyl)aminocarboxylate 15b (60 mg, 0.15 mmol) was mixed with trifluoroacetic acid (1 mL) and dichloromethane (4 mL) and stirred at room temperature for 6 hours. The solvent was removed under reduced pressure to obtain the target product, 1-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)-4-methylpiperazin-2-one trifluoroacetate 15c (60 mg, light yellow solid) as a crude product.

MS m/z (ESI): 309 [M+1].

Step 3

Phenyl-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)(methyl)aminocarboxylate

Compound 1-((2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methyl)-4-methylpiperazin-2-one trifluoroacetate 15c (60 mg, 0.14 mmol), diphenyl carbonate (60 mg, 0.28 mmol), lithium hexamethyldisilazide (0.56 mL, 0.56 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (5 mL) were mixed and stirred at 0°C for 0.5 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with dichloromethane (20 mL × 3). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (dichloromethane/methanol 30:1) to give the target product, phenyl-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)(methyl)aminocarboxylate 15d (30 mg, colorless solid) in a yield of 36%.

MS m/z (ESI): 429 [M+1].

Step 4

3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Phenyl-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2yl)(methyl)aminocarboxylate 15d (30 mg, 0.07 mmol), 6-amino-4-chloronicotinonitrile (21 mg, 0.14 mmol), lithium hexamethyldisilazide (0.21 mL, 0.21 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (3 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with dichloromethane (20 mL × 3). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (dichloromethane/methanol 30:1) to give the desired product, 3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 15e (17 mg, white solid). Yield: 50%.

MS m/z(ESI):488&490[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.82 (s, 1H), 8.52 (s, 1H), 8.46 (s, 1H), 7.77 (d, J = 8.4Hz, 1H), 7.04 (d, J = 8.4Hz, 1H), 5.47 (s, 1H), 4.86 (s, 2H), 3.47 (s, 6H), 3.46 (s, 3H), 3.27 (s, 2H), 3.20 (t, J=4.4Hz, 2H), 2.62 (t, J=4.4Hz, 2H), 2.35 (s, 3H).

Step 5

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Compound 3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 15e (4 mg, 0.008 mmol), 2-methoxyethylamine (2 mg, 0.024 mmol), diisopropylethylamine (2 mg, 0.016 mmol), and N,N-dimethylacetamide (0.4 mL) were mixed and stirred at 50°C for 16 hours. The mixture was diluted with 10 mL of water and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (dichloromethane/methanol 25:1) to give the target product 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 15f (2 mg, white solid) in a yield of 46%.

MS m/z (ESI): 527 [M+1].

Step 6

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea hydrochloride

Compound 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 15f (2 mg, 0.004 mmol), hydrochloric acid (0.8 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was desolvated under reduced pressure to give the desired product, 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea hydrochloride 15 (1.5 mg, white solid). Yield: 67%.

MS m/z(ESI):481[M+1]

¹ H NMR (400MHz, CD ₃ OD) δ8.40 (s, 1H), 7.98 (d, J=8.4Hz, 1H), 7.40 (d, J=8.4Hz, 1H), 6.94 (s, 1H), 6.92 (s, 1H), 4.91 (s, 2H), 3.75 (t, J=6.8Hz, 2H), 3.69-3.60 (m, 2H), 3.45-3.42 (m, 4H), 3.37 (s, 3H), 3.36 (s, 3H), 3.26-3.21 (m, 2H), 3.03 (s, 3H).

Example 16

3-(5-cyano-4-(isopropylamino)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Example 16 was synthesized by following the procedure of Example 15, except that isopropylamine was used instead of 2-methoxyethylamine in the fifth step.

MS m/z(ESI):465[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ12.95 (s, 1H), 10.25 (s, 1H), 8.16 (s, 1H), 7.95 (d, J=8.0Hz, 1H), 7.56 (s, 1H), 7.28 (d, J=8.0Hz, 1H), 5.10 (s, 2H), 4.76 (brs, 1H ), 3.89-3.87 (m, 1H), 3.52 (s, 3H), 3.36 (s, 2H), 3.20 (t, J=4.0Hz, 2H), 2.66 (t, J=4.0Hz, 2H), 2.35 (s, 3H), 1.32 (d, J=5.2Hz, 6H).

Example 17

3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Example 17 was synthesized by following the procedures of Example 15, except that in the sixth step, 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea was replaced with 3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea.

MS m/z(ESI):442&444[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.59 (s, 1H), 10.26 (s, 1H), 8.51 (s, 1H), 8.46 (s, 1H), 7.95 (d, J = 8.4Hz, 1H), 7.31 (d, J = 8.4Hz, 1H ), 5.10 (s, 2H), 3.54 (s, 3H), 3.38 (s, 2H), 3.20 (t, J = 4.4Hz, 2H), 2.68 (t, J = 4.4Hz, 2H), 2.36 (s, 3H).

Example 18

(R)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

first step

(R)-6-Amino-4-((1-methoxypropan-2-yl)oxy)nicotinenitrile

Compound 6-amino-4-chloronicotinonitrile 18a (60 mg, 0.39 mmol), (R)-1-methoxypropan-2-ol (70 mg, 0.78 mmol), sodium hydride (34 mg, 0.86 mmol, in a 60% mineral oil mixture), and N-methylpyrrolidone (1.5 mL) were mixed and stirred at 70°C for 16 hours. After cooling to room temperature, the mixture was quenched with 20 mL of water and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolventized under reduced pressure. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate 1:1) to obtain the desired product, (R)-6-amino-4-((1-methoxypropan-2-yl)oxy)nicotinonitrile 18b (17 mg, white solid) in a 21% yield.

MS m/z(ESI):208[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ8.17 (s, 1H), 6.00 (s, 1H), 4.94 (brs, 2H), 4.63-4.60 (m, 1H), 3.63-3.62 (m, 1H), 3.54-3.51 (m, 1H), 3.41 (s, 3H), 1.38-1.36 (m, 3H).

Step 2

(R)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Phenyl-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2yl)(methyl)aminocarboxylate (10 mg, 0.02 mmol), (R)-6-amino-4-((1-methoxypropan-2-yl)oxy)nicotinonitrile 18b (8 mg, 0.04 mmol), lithium hexamethyldisilazide (0.06 mL, 0.06 mmol, 1 M tetrahydrofuran solution), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with dichloromethane (20 mL × 3). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (dichloromethane/methanol 30:1) to give the desired product, (R)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 18c (6 mg, white solid). Yield: 47%.

MS m/z (ESI): 542 [M+1].

Step 3

(R)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Compound (R)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-(dimethoxymethyl)-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 18c (6 mg, 0.01 mmol), hydrochloric acid (0.8 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with saturated sodium carbonate solution and extracted with dichloromethane (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (dichloromethane/methanol 25:1) to give the target product (R)-3-(5-cyano-4-((1-methoxypropane-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 18 (4 mg, white solid) in a yield of 73%.

MS m/z(ESI):496[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.28 (s, 1H), 10.27 (s, 1H), 8.35 (s, 1H), 7.99 (s, 1H), 7.94 (d, J = 7.6Hz, 1H), 7.27 (d, J = 7.6Hz, 1H), 5.11 (s, 2H), 4.88-4.86 (m, 1H), 3.66-3.59 (m, 2H), 3.57 (s, 3H), 3.49 (s, 3H), 3.43 (s, 2H), 3.20 (t, J=4.4Hz, 2H), 2.67 (t, J=4.4Hz, 2H), 2.36 (s, 3H), 1.42-1.40 (m, 3H).

Example 19

3-(5-cyano-4-isopropoxypyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Example 19 was synthesized following the procedure of Example 18, except that isopropanol was substituted for (R)-1-methoxypropan-2-ol in the first step.

MS m/z(ESI):466[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.28 (s, 1H), 10.26 (s, 1H), 8.34 (s, 1H), 7.96 (d, J = 8.4Hz, 1H), 7.93 (s, 1H), 7.26 (d, J = 8.4Hz, 1H), 5.10 (s, 2H), 4.85 -4.83 (m, 1H), 3.49 (s, 3H), 3.38 (s, 2H), 3.21 (t, J=4.4Hz, 2H), 2.69 (t, J=4.4Hz, 2H), 2.36 (s, 3H), 1.45 (d, J=4.0Hz, 6H).

Example 20

3-(5-cyano-4-(2-methoxyethoxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Example 20 was synthesized following the procedure of Example 18, except that 2-methoxyethanol was substituted for (R)-1-methoxypropan-2-ol in the first step.

MS m/z(ESI):482[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.34 (s, 1H), 10.27 (s, 1H), 8.36 (s, 1H), 7.96 (s, 1H), 7.95 (d, J = 8.4Hz, 1H), 7.31 (d, J = 8.4Hz, 1H), 5.11 (s, 2H), 4.35 (t, J = 4. 0Hz, 2H), 3.83 (t, J=4.0Hz, 2H), 3.53 (s, 3H), 3.48 (s, 3H), 3.37 (s, 2H), 3.20 (t, J=4.4Hz, 2H), 2.67 (t, J=4.4Hz, 2H), 2.36 (s, 3H).

Example 21

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-(hydroxymethyl)pyridin-2-yl)-1-methylurea

first step

5-(((tert-Butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)-N-methylpyridin-2-amine

Compound (2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)methanol 21a (60 mg, 0.28 mmol), tert-butylchlorodimethylsilane (64 mg, 0.42 mmol), N-ethyl-N-isopropylpropane-2-amine (72 mg, 0.56 mmol), N,N-dimethylpyridin-4-amine (7 mg, 0.06 mmol), and dichloromethane (4 mL) were mixed and stirred at room temperature for 6 hours. The mixture was quenched with 20 mL of water and extracted with ethyl acetate (20 mL × 2). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the solvent was removed under reduced pressure. The residue was purified on a preparative silica gel plate (petroleum ether/ethyl acetate = 15:1) to give the target product 5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)-N-methylpyridin-2-amine 21b (60 mg, colorless oil) in a yield of 65%.

MS m/z(ESI):327[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ7.57 (d, J=8.0Hz, 1H), 6.28 (d, J=8.0Hz, 1H), 5.21 (s, 1H), 4.69 (s, 2H), 4.57 (brs, 1H), 3.32 (s, 6H), 2.80 (s, 3H), 0.84 (s, 9H), 0.08 (s, 6H).

Step 2

Phenyl-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)(methyl)aminocarboxylate

Compound 5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)-N-methylpyridin-2-amine 21b (22 mg, 0.07 mmol), diphenyl carbonate (15 mg, 0.14 mmol), lithium hexamethyldisilazide (0.21 mL, 0.21 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (3 mL) were mixed and stirred at 0°C for 0.5 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 3). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (petroleum ether/ethyl acetate 5:1). The target product, phenyl-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)(methyl)aminocarboxylate 21c (18 mg, white solid), was obtained in a 60% yield.

MS m/z(ESI):447[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ7.86 (d, J=8.4Hz, 1H), 7.58 (d, J=7.2Hz, 1H), 7.27-7.25 (m, 2H), 7.15-7.04 (m, 3 H), 5.19 (s, 1H), 4.83 (s, 2H), 3.49 (s, 3H), 3.34 (s, 6H), 0.85 (s, 9H), 0.08 (s, 6H).

Step 3

1-(5-(((tert-Butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(4-chloro-5-cyanopyridin-2-yl)-1-methylurea

Phenyl-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)(methyl)aminocarboxylate 21c (18 mg, 0.04 mmol), 6-amino-4-chloronicotinonitrile (12 mg, 0.08 mmol), lithium hexamethyldisilazide (0.12 mL, 0.12 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL × 3). The organic phase was washed with saturated brine (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 3:1) to give the target product 1-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(4-chloro-5-cyanopyridin-2-yl)-1-methylurea 21d (16 mg, white solid) in a yield of 79%.

MS m/z(ESI):506&508[M+1]

¹ H NMR (400MHz, CDCl ₃ ) δ13.78 (s, 1H), 8.40 (s, 1H), 8.34 (s, 1H), 7.95 (d, J = 8.8Hz, 1H), 6.97 (d, J = 8.8Hz, 1H), 5.35(s, 1H), 4.78(s, 2H), 3.37(s, 3H), 3.33(s, 6H), 0.83(s, 9H), 0.02(s, 6H).

Step 4

1-(5-(((tert-Butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea

Compound 1-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(4-chloro-5-cyanopyridin-2-yl)-1-methylurea 21d (10 mg, 0.02 mmol), 2-methoxyethylamine (3 mg, 0.06 mmol), diisopropylethylamine (5 mg, 0.06 mmol), and N,N-dimethylacetamide (0.4 mL) were mixed and stirred at 50°C for 16 hours. The mixture was diluted with 10 mL of water and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (petroleum ether/ethyl acetate 3:1). The target product, 1-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea 21e (9 mg, white solid), was obtained in a yield of 76%.

MS m/z (ESI): 545 [M+1].

Step 5

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-(hydroxymethyl)pyridin-2-yl)-1-methylurea

Compound 1-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea 21e (9 mg, 0.02 mmol), hydrochloric acid (0.8 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with saturated sodium carbonate solution and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (petroleum ether/ethyl acetate 1:1). The target product, 1-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylurea 21 (3 mg, white solid), was obtained in a yield of 47%.

MS m/z(ESI):385[M+1]

¹ H NMR (400MHz, CDCl ₃ )δmajor(21'): 12.60 (s, 1H), 8.08 (s, 1H), 7.70 (d, J=7.6Hz, 1H), 7.58 (s, 1H), 7.07 (d, J=7.6Hz, 1H), 6.53 (s, 1H), 5. 31(s, 1H), 5.25-5.23(m, 1H), 5.05-5.02(m, 1H), 3.63-3.61(m, 2H), 3.51-3.50(m, 2H), 3.49(s, 3H), 3.41(s, 3H).Mino r(21): 12.94 (s, 1H), 10.25 (s, 1H), 8.17 (s, 1H), 8.06 (d, J=8.4Hz, 1H), 7.57 (s, 1H), 7.27 (d, J=8.4Hz, 1H), 6.53 (s, 1H ), 5.31 (s, 1H), 5.25-5.23 (m, 1H), 5.05-5.02 (m, 1H), 3.63-3.61 (m, 2H), 3.51-3.50 (m, 2H), 3.49 (s, 3H), 3.41 (s, 3H).

Example 22

3-(4-chloro-5-cyanopyridin-2-yl)-1-(6-formyl-5-(hydroxymethyl)pyridin-2-yl)-1-methylurea

Example 22 was synthesized by following the procedures of Example 21, except that in the fifth step, 1-(5-(((tert-butyldimethylsilyl)oxy)methyl)-6-(dimethoxymethyl)pyridin-2-yl)-3-(4-chloro-5-cyanopyridin-2-yl)-1-methylurea was used to replace 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-(hydroxymethyl)pyridin-2-yl)-1-methylurea.

MS m/z(ESI):346&348[M+1]

¹ H NMR (400MHz, CDCl ₃ )δmajor(22'): 13.10(s, 1H), 8.46(s, 1H), 8.45(s, 1H), 7.77(d, J=8.4Hz, 1H), 7.13 (d, J=8.4Hz, 1H), 6.51 (s, 1H), 5.29 (s, 1H), 5.11-5.08 (m, 2H), 3.53 (s, 3H) .Minor(22): 13.55(s, 1H), 10.25(s, 1H), 8.48(s, 1H), 8.47(s, 1H), 8.10(d, J＝ 8.4Hz, 1H), 7.35 (d, J=8.4Hz, 1H), 4.97 (s, 2H), 4.08-4.06 (m, 1H), 3.57 (s, 3H).

Example 23

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-(piperidin-4-yl)pyridin-2-yl)-1-methylurea

first step

tert-Butyl-6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)-5′,6′-dihydro-[3,4′-bipyridine]-1′(2′H)-carboxylate

Compound tert-butyl (5-bromo-6-(dimethoxymethylpyridin-2-yl)(methyl)aminocarboxylate 23a (0.20 g, 0.56 mmol), [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium (90 mg, 0.11 mmol), tert-butyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-5,6-dihydropyridine-1-(2H)-carboxylate (0.34 g, 1.11 mmol), potassium carbonate (0.23 g, 1.67 mmol), water (1 mL) and dioxane (5 mL) were stirred at 4 °C for 1 hr. L) and stirred at 90°C under nitrogen for 16 hours. The mixture was diluted with water and extracted with ethyl acetate (50 mL x 3). The organic phase was washed with saturated brine (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified by silica gel chromatography (petroleum ether/ethyl acetate 84:16) to obtain the target tert-butyl-6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)-5′,6′-dihydro-[3,4′-bipyridine]-1′(2′H)-carboxylate 23b (0.13 g, colorless oil) in a yield of 51%.

MS m/z(ESI):464[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ7.64 (d, J=8.4Hz, 1H), 7.41 (d, J=8.4Hz, 1H), 5.67-5.65 (m, 1H), 5.54 (s, 1H), 4.07 (s, 2H), 3.65 (t, J=5.2Hz, 2H), 3.46 (s, 6H), 3.45 (s, 3H), 2.40-2.38 (m, 2H), 1.53 (s, 9H), 1.52 (s, 9H).

Step 2

tert-Butyl-4-(6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate

The compound tert-butyl-6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)-5′,6′-dihydro-[3,4′-bipyridine]-1′(2′H)-carboxylate 23b (0.13 g, 0.28 mmol), palladium on carbon (26 mg, 20%), methanol (3 mL), and ethyl acetate (2 mL) were mixed and stirred at room temperature for 16 hours. The mixture was filtered and desolvated under reduced pressure to obtain the target product, tert-butyl-4-(6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate 23c (0.13 g, colorless oil) as a crude product.

MS m/z (ESI): 466 [M+1].

Step 3

6-(Dimethoxymethyl)-N-methyl-5-(piperidin-4-yl)pyridin-2-amine

Compound tert-butyl-4-(6-((tert-butoxycarbonyl)(methyl)amino)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate 23c (0.13 g, 0.28 mmol), trifluoroacetic acid (1 mL), and dichloromethane (4 mL) were mixed and stirred at room temperature for 6 hours. The solvent was removed under reduced pressure to obtain the target product, 6-(dimethoxymethyl)-N-methyl-5-(piperidin-4-yl)pyridin-2-amine trifluoroacetate 23d (95 mg, light yellow solid), as a crude product.

MS m/z (ESI): 266 [M+1].

Step 4

tert-Butyl-4-(2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)piperidine-1-carboxylate

Compound 6-(dimethoxymethyl)-N-methyl-5-(piperidin-4-yl)pyridin-2-amine trifluoroacetate 23d (95 mg, 0.25 mmol), 2-tert-butyl 2-carbonate (82 mg, 0.38 mmol), triethylamine (76 mg, 0.75 mmol), and dichloromethane (4 mL) were mixed and stirred at room temperature for 6 hours. The mixture was quenched with 20 mL of water and extracted with ethyl acetate (20 mL x 3). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate and filtered to remove the desiccant. The residue was purified on preparative silica gel (petroleum ether/ethyl acetate 3:1) to obtain the desired product, tert-butyl-4-(2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)piperidine-1-carboxylate 23e (80 mg, white solid) in a 65% yield.

MS m/z (ESI): 366 [M+1].

Step 5

tert-Butyl-4-(2-(dimethoxymethyl)-6-(methyl(phenoxycarbonyl)amino)pyridin-3-yl)piperidine-1-carboxylate

Compound tert-butyl-4-(2-(dimethoxymethyl)-6-(methylamino)pyridin-3-yl)piperidine-1-carboxylate 23e (40 mg, 0.11 mmol), diphenyl carbonate (47 mg, 0.22 mmol), lithium hexamethyldisilazide (0.33 mL, 0.33 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (6 mL) were mixed and stirred at 0°C for 0.5 hours. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 3). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (petroleum ether/ethyl acetate 2:1). The target product, tert-butyl-4-(2-(dimethoxymethyl)-6-(methyl(phenoxycarbonyl)amino)pyridin-3-yl)piperidine-1-carboxylate 23f (20 mg, white solid), was obtained in a yield of 38%.

MS m/z (ESI): 486 [M+1].

Step 6

tert-Butyl-4-(6-(3-(4-chloro-5-cyanopyridin-2-yl)-1-methylureido)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate

Compound tert-butyl-4-(2-(dimethoxymethyl)-6-(methyl(phenoxycarbonyl)amino)pyridin-3-yl)piperidine-1-carboxylate 23f (20 mg, 0.04 mmol), 6-amino-4-chloronicotinonitrile (12 mg, 0.08 mmol), lithium hexamethyldisilazide (0.12 mL, 0.12 mmol, 1 M solution in tetrahydrofuran), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The mixture was quenched with 10 mL of saturated ammonium chloride solution and extracted with ethyl acetate (20 mL x 3). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified by preparative silica gel plate (petroleum ether/ethyl acetate 1.5:1) to give the target product tert-butyl-4-(6-(3-(4-chloro-5-cyanopyridin-2-yl)-1-methylureido)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate 23 g (10 mg, white solid) in a yield of 45%.

MS m/z (ESI): 545 & 547 [M+1].

Step 7

tert-Butyl-4-(6-(3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylureido)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate

23 g (10 mg, 0.02 mmol) of the compound tert-butyl-4-(6-(3-(4-chloro-5-cyanopyridin-2-yl)-1-methylureido)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate, 2-methoxyethylamine (3 mg, 0.06 mmol), diisopropylethylamine (5 mg, 0.06 mmol), and N,N-dimethylacetamide (0.4 mL) were mixed and stirred at 50°C for 16 hours. The mixture was diluted with 10 mL of water and extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (20 mL x 2). The organic phase was dried over anhydrous sodium sulfate, the desiccant was removed by filtration, and the residue was purified on a preparative silica gel plate (petroleum ether/ethyl acetate 1.5:1). The target product, tert-butyl-4-(6-(3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylureido)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate 23h (6 mg, white solid), was obtained in a yield of 56%.

MS m/z (ESI): 584 [M+1].

Step 8

3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-(piperidin-4-yl)pyridin-2-yl)-1-methylurea

The compound tert-butyl-4-(6-(3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-methylureido)-2-(dimethoxymethyl)pyridin-3-yl)piperidine-1-carboxylate 23h (6 mg, 0.01 mmol), hydrochloric acid (0.8 mL, 37%), water (1 mL), and tetrahydrofuran (2 mL) were mixed and stirred at room temperature for 1 hour. The solution was removed by decompression to obtain the target product, 3-(5-cyano-4-((2-methoxyethyl)amino)pyridin-2-yl)-1-(6-formyl-5-(piperidin-4-yl)pyridin-2-yl)-1-methylurea hydrochloride 23 (4 mg, white solid) in a yield of 89%.

MS m/z(ESI):438[M+1]

¹ H NMR (400MHz, CD ₃ OD) δ 8.40 (s, 1H), 8.01 (d, J = 8.4Hz, 1H), 7.37 (d, J = 7.6Hz, 1H), 6.92 (s, 1H), 6.07 (s, 1H), 3.71- 3.58 (m, 6H), 3.55 (s, 3H), 3.43 (s, 3H), 3.25-3.20 (m, 2H), 3.19-3.15 (m, 1H), 2.12-1.95 (m, 4H).

Example 24

(S)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Example 24 was synthesized following the procedure of Example 18, except that (S)-1-methoxypropan-2-ol was substituted for (R)-1-methoxypropan-2-ol in the first step.

MS m/z(ESI):496[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.31 (s, 1H), 10.26 (s, 1H), 8.34 (s, 1H), 7.99 (s, 1H), 7.94 (d, J = 8.7Hz, 1H), 7.29 (d, J = 8.7Hz, 1H), 5.10 (s, 2H), 4.99-4.74 (m, 1H), 3. 73-3.55 (m, 2H), 3.52 (s, 3H), 3.43 (s, 3H), 3.37 (t, J=5.3Hz, 2H), 3.20 (s, 2H), 2.67 (t, J=5.3Hz, 2H), 2.36 (s, 3H), 1.41 (d, J=6.3Hz, 3H).

Example 25

(R)-3-(5-cyano-4-((1-hydroxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Compound (R)-3-(5-cyano-4-((1-methoxypropane-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 18 (4.0 g, 8 mmol) and dichloromethane (80 mL) were mixed, and boron tribromide (20.2 g, 81 mmol) was added dropwise under an ice-salt bath. After the addition was complete, the ice-salt bath was removed and stirring was continued at room temperature for 30 minutes. The mixture was slowly poured into ice water (300 mL) for quenching. The pH was adjusted to 8-9 with saturated aqueous sodium carbonate solution (200 mL), and the mixture was extracted with dichloromethane (150 mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove the desiccant, and desolvated under reduced pressure to obtain a crude product. The crude product was purified by flash column chromatography (dichloromethane: methanol = 20: 1) to give the target product (R)-3-(5-cyano-4-((1-hydroxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea 25 (2.3 g, 4.8 mmol, white solid) in a yield of 60%.

MS m/z(ESI):482[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.36 (s, 1H), 10.26 (s, 1H), 8.35 (s, 1H), 7.99 (s, 1H), 7.94 (d, J = 8.5Hz, 1H), 7.30 (d, J = 8.5Hz, 1H), 5.10 (s, 2H), 4.88-4.76 (m, 1H), 3.91-3.78 (m, 2H), 3.53 (s, 3H), 3.42-3.38 (s, 2H), 3.20 (s, 2H), 2.75-2.63 (m, 2H), 2.36 (s, 3H), 1.41 (d, J=6.0Hz, 3H).

Example 26

(S)-3-(5-cyano-4-((1-hydroxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea

Example 26 was synthesized by following the procedures of Example 25, except that in the first step, (S)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea was used to replace (R)-3-(5-cyano-4-((1-methoxypropan-2-yl)oxy)pyridin-2-yl)-1-(6-formyl-5-((4-methyl-2-carbonylpiperazin-1-yl)methyl)pyridin-2-yl)-1-methylurea.

MS m/z(ESI):482[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ13.37 (s, 1H), 10.26 (s, 1H), 8.34 (s, 1H), 7.97 (s, 1H), 7.95 (d, J = 8.8Hz, 1H), 7.32 (d, J = 8.8Hz, 1H), 5.11 (s, 2H), 4.91-4.74 (m, 1H), 3.96-3.76 (m, 2H), 3.54 (s, 3H), 3.46-3.33 (m, 2H), 3.22 (s, 2H), 2.79-2.62 (m, 2H), 2.38 (s, 3H), 1.42 (d, J=6.0Hz, 3H).

Biological experiments

FGFR4 activity inhibition test

In vitro kinase assays were used to evaluate the effects of the compounds of the present invention on fibroblast growth factor receptor 4 (FGFR4) activity.

The experimental methods are outlined as follows:

FGFR4 in vitro activity was determined using the HTRF kinase assay kit by measuring substrate phosphorylation levels in the kinase reaction. The reaction buffer contained the following components: 5x diluted Enzymatic buffer/kinase 5X (Cisbio, catalog number 62EZBFDD) (mainly composed of 50 mM HEPES, pH 7.0), 5 mM MgCl2, _and 1 mM DTT; human recombinant FGFR4 catalytic domain protein (amino acids 460-802) purchased from Tsinghua Protein Research Technology Center was diluted in reaction buffer to a 0.5 ng/uL kinase solution; the substrate reaction solution contained 500 nM biotin-labeled tyrosine kinase substrate (Cisbio, catalog number 62TK0PEC) and 90 uM ATP diluted in reaction buffer; and the detection solution contained 0.125 ng/uL Eu2O3 (Cisbio, catalog number 62SDBRDF) diluted in detection buffer. ³⁺ -labeled caged antibody (Cisbio, Cat. No. 61T66KLB) and 31.25 nM streptavidin-labeled XL665 (Cisbio, Cat. No. 610SAXLB).

Dissolve the compound in 100% DMSO and dilute it to 100uM. Then, serially dilute it 4-fold with DMSO to a minimum concentration of 0.0061uM. Each concentration point is diluted 40-fold with reaction buffer. If the compound _IC50 value is very low, the starting concentration of the compound can be reduced.

To a 384-well assay plate (Thermo, Cat. No. 264706), add 4 μL of compound solution and 2 μL of FGFR4 kinase solution, mix thoroughly, and incubate at room temperature for 15 minutes. Then, add 4 μL of substrate reaction solution, and incubate the reaction mixture at room temperature for 60 minutes. Then, add 10 μL of detection solution (equal to the reaction volume), mix thoroughly, and let it sit at room temperature. After 60 minutes, the enzyme reaction is terminated by the EDTA in the detection solution. The phosphorylated product is recognized simultaneously by the Eu ³⁺ -labeled caged antibody (donor) and the streptavidin-labeled XL665 antibody (acceptor). Upon laser excitation, the adjacent donor and acceptor undergo energy resonance transfer. The energy transferred from the donor (620 nm) to the acceptor (665 nm) can be detected using Envision. The 665/620 ratio positively correlates with the degree of substrate phosphorylation, thus measuring FGFR4 kinase activity. In this experiment, the group without protein addition served as the negative control (100% inhibition), and the group with protein addition but no compound addition served as the positive control (0% inhibition). The percentage of FGFR4 activity inhibition by the compound can be calculated using the following formula:

Percent inhibition = 100 - 100 * (signal _compound - signal _{negative control} ) / (signal _{positive control} - signal _{negative control} )

The _IC50 values of the compounds were calculated from 10 concentration points using XLfit (ID Business Solutions Ltd., UK) software using the following formula:

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)*slope factor))

Wherein Y is the inhibition percentage, Bottom is the bottom plateau of the curve (the bottom platform value of the S-shaped curve), Top is the top plateau of the curve (the top platform value of the S-shaped curve), and X is the logarithm of the concentration of the test compound.

FGFR1 activity inhibition test

In vitro kinase assays were used to evaluate the effects of the compounds of the present invention on the activity of fibroblast growth factor receptor 1 (FGFR1).

The experimental methods are outlined as follows:

FGFR1 activity in vitro was determined using the HTRF Kinase Assay Kit by measuring substrate phosphorylation in a kinase reaction. The reaction buffer contained the following components: 5x diluted Enzymatic buffer/kinase 5X (Cisbio, catalog number 62EZBFDD) (50 mM HEPES, pH 7.0), ₅ mM MgCl₂, and 1 mM DTT; human recombinant FGFR1 catalytic domain protein (amino acids 308-731), purified in-house, diluted in reaction buffer to a 0.6 ng/µL kinase solution; the substrate reaction solution contained 400 nM biotinylated tyrosine kinase substrate (Cisbio, catalog number 62TK0PEC) and 40 µM ATP diluted in reaction buffer; and the detection solution contained 0.125 ng/µL Eu₂O₄ (Cisbio, catalog number 62SDBRDF) diluted in detection buffer. ³⁺ -labeled caged antibody (Cisbio, Cat. No. 61T66KLB) and 25 nM streptavidin-labeled XL665 (Cisbio, Cat. No. 610SAXLB).

Dissolve the compound in 100% DMSO and dilute it to 1 mM. Then, perform a 4-fold serial dilution in DMSO to a minimum concentration of 0.061 uM. Each concentration point is diluted 40-fold in reaction buffer. If the compound _IC50 value is very low, the starting concentration of the compound can be reduced.

To a 384-well assay plate (Thermo, Cat. No. 264706), add 4 μL of compound solution and 2 μL of FGFR1 kinase solution, mix thoroughly, and incubate at room temperature for 15 minutes. Then, add 4 μL of substrate reaction solution, and incubate the reaction mixture at room temperature for 60 minutes. Then, add 10 μL of detection solution (equal to the reaction volume), mix thoroughly, and let it stand at room temperature. After 60 minutes, the enzyme reaction is terminated by the EDTA in the detection solution. The phosphorylated product is recognized simultaneously by the Eu ³⁺ -labeled caged antibody (donor) and the streptavidin-labeled XL665 antibody (acceptor). Upon laser excitation, the adjacent donor and acceptor undergo energy resonance transfer. The energy transferred from the donor (620 nm) to the acceptor (665 nm) can be detected using Envision. The 665/620 ratio positively correlates with the degree of substrate phosphorylation, thus measuring FGFR1 kinase activity. In this experiment, the group without protein addition served as the negative control (100% inhibition), and the group with protein addition but no compound addition served as the positive control (0% inhibition). The percentage of FGFR1 activity inhibition by the compound can be calculated using the following formula:

Percent inhibition = 100 - 100 * (signal _compound - signal _{negative control} ) / (signal _{positive control} - signal _{negative control} )

The _IC50 values of the compounds were calculated from 10 concentration points using XLfit (ID Business Solutions Ltd., UK) software using the following formula:

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)*slope factor))

Wherein Y is the inhibition percentage, Bottom is the bottom plateau of the curve (the bottom platform value of the S-shaped curve), Top is the top plateau of the curve (the top platform value of the S-shaped curve), and X is the logarithm of the concentration of the test compound.

FGFR2 activity inhibition test

In vitro kinase assays were used to evaluate the effects of the compounds of the present invention on fibroblast growth factor receptor 2 (FGFR2) activity.

The experimental methods are outlined as follows:

FGFR2 activity in vitro was determined using the HTRF kinase assay kit by measuring substrate phosphorylation in the kinase reaction. The reaction buffer contained the following components: 5x diluted Enzymatic buffer/kinase 5X (Cisbio, catalog number 62EZBFDD) (mainly composed of 50 mM HEPES, pH 7.0), 5 mM MgCl₂, _and 1 mM DTT; human recombinant FGFR2 catalytic domain protein (amino acids 400-821) purchased from Sino Biological Biotechnology Co., Ltd., diluted in reaction buffer to a 0.045 ng/µL kinase solution; the substrate reaction solution contained 800 nM biotin-labeled tyrosine kinase substrate (Cisbio, catalog number 62TK0PEC) and 50 µM ATP diluted in reaction buffer; and the detection solution contained 0.125 ng/µL Eu₂O₄ (Cisbio, catalog number 62SDBRDF) diluted in detection buffer. ³⁺ -labeled caged antibody (Cisbio, cat. no. 61T66KLB) and 50 nM streptavidin-labeled XL665 (Cisbio, cat. no. 610SAXLB).

Dissolve the compound in 100% DMSO and dilute it to 100uM. Then, serially dilute it 4-fold with DMSO to a minimum concentration of 0.0061uM. Each concentration point is diluted 40-fold with reaction buffer. If the compound _IC50 value is very low, the starting concentration of the compound can be reduced.

To a 384-well assay plate (Thermo, Cat. No. 264706), add 4 μL of compound solution and 2 μL of FGFR2 kinase solution, mix thoroughly, and incubate at room temperature for 15 minutes. Then, add 4 μL of substrate reaction solution, and incubate the reaction mixture at room temperature for 60 minutes. Then, add 10 μL of detection solution (equal to the reaction volume), mix thoroughly, and let it sit at room temperature. After 60 minutes, the enzyme reaction is terminated by the EDTA in the detection solution. The phosphorylated product is recognized simultaneously by the Eu ³⁺ -labeled caged antibody (donor) and the streptavidin-labeled XL665 antibody (acceptor). Upon laser excitation, the adjacent donor and acceptor undergo energy resonance transfer. The energy transferred from the donor (620 nm) to the acceptor (665 nm) can be detected using Envision. The 665/620 ratio positively correlates with the degree of substrate phosphorylation, thus measuring FGFR2 kinase activity. In this experiment, the group without protein addition served as the negative control (100% inhibition), and the group with protein addition but no compound addition served as the positive control (0% inhibition). The percentage of FGFR2 activity inhibition by the compound can be calculated using the following formula:

Percent inhibition = 100 - 100 * (signal _compound - signal _{negative control} ) / (signal _{positive control} - signal _{negative control} )

The _IC50 values of the compounds were calculated from 10 concentration points using XLfit (ID Business Solutions Ltd., UK) software using the following formula:

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)*slope factor))

Wherein Y is the inhibition percentage, Bottom is the bottom plateau of the curve (the bottom platform value of the S-shaped curve), Top is the top plateau of the curve (the top platform value of the S-shaped curve), and X is the logarithm of the concentration of the test compound.

FGFR3 activity inhibition test

The effects of the compounds of the present invention on fibroblast growth factor receptor 3 (FGFR3) activity were evaluated using an in vitro kinase assay.

The experimental methods are outlined as follows:

FGFR3 activity in vitro was determined using the HTRF kinase assay kit by measuring substrate phosphorylation levels in the kinase reaction. The reaction buffer contained the following components: 5x diluted Enzymatic buffer/kinase 5X (Cisbio, catalog number 62EZBFDD) (mainly composed of 50 mM HEPES, pH 7.0), 5 mM MgCl2, _and 1 mM DTT; human recombinant FGFR3 catalytic domain protein (amino acids 399-806) purchased from Sino Biological Biotechnology Co., Ltd. was diluted in reaction buffer to a 0.3 ng/uL kinase solution; the substrate reaction solution contained 1000 nM biotin-labeled tyrosine kinase substrate (Cisbio, catalog number 62TK0PEC) and 90 uM ATP diluted in reaction buffer; and the detection solution contained 0.125 ng/uL Eu2O3 (Cisbio, catalog number 62SDBRDF) diluted in detection buffer. ³⁺ -labeled caged antibody (Cisbio, Cat. No. 61T66KLB) and 62.5 nM streptavidin-labeled XL665 (Cisbio, Cat. No. 610SAXLB).

Dissolve the compound in 100% DMSO and dilute it to 100uM. Then, serially dilute it 4-fold with DMSO to a minimum concentration of 0.0061uM. Each concentration point is diluted 40-fold with reaction buffer. If the compound _IC50 value is very low, the starting concentration of the compound can be reduced.

To a 384-well assay plate (Thermo, Cat. No. 264706), add 4 μL of compound solution and 2 μL of FGFR3 kinase solution, mix thoroughly, and incubate at room temperature for 15 minutes. Then, add 4 μL of substrate reaction solution, and incubate the reaction mixture at room temperature for 60 minutes. Then, add 10 μL of detection solution (equal to the reaction volume), mix thoroughly, and let it sit at room temperature. After 60 minutes, the enzyme reaction is terminated by the EDTA in the detection solution. The phosphorylated product is recognized simultaneously by the Eu ³⁺ -labeled caged antibody (donor) and the streptavidin-labeled XL665 antibody (acceptor). Upon laser excitation, the adjacent donor and acceptor undergo energy resonance transfer. The energy transferred from the donor (620 nm) to the acceptor (665 nm) can be detected using Envision. The 665/620 ratio positively correlates with the degree of substrate phosphorylation, thus measuring FGFR3 kinase activity. In this experiment, the group without protein addition served as the negative control (100% inhibition), and the group with protein addition but no compound addition served as the positive control (0% inhibition). The percentage of FGFR2 activity inhibition by the compound can be calculated using the following formula:

Percent inhibition = 100 - 100 * (signal _compound - signal _{negative control} ) / (signal _{positive control} - signal _{negative control} )

The _IC50 values of the compounds were calculated from 10 concentration points using XLfit (ID Business Solutions Ltd., UK) software using the following formula:

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)*slope factor))

Wherein Y is the inhibition percentage, Bottom is the bottom plateau of the curve (the bottom platform value of the S-shaped curve), Top is the top plateau of the curve (the top platform value of the S-shaped curve), and X is the logarithm of the concentration of the test compound.

Biological test example: A: <10nM, B: 10-100nM, C: 100-1000nM, D: >1000nM, ND: not tested

The compounds of the present invention have a selective inhibitory effect on FGFR4.

Determination of Hep3B cell proliferation inhibition

The effects of the compounds of the present invention on the proliferation of Hep3B cells were evaluated using a luminescent cell viability assay.

The experimental methods are outlined as follows:

The CellTilter-Glo (CTG) assay kit uses a unique, stable luciferase to detect ATP, an indicator of viable cell metabolism. The luminescent signal generated in the assay is proportional to the number of viable cells in the culture medium, thereby detecting the cell proliferation status of Hep3B cells.

CellTilter-Glo reagent (Promega, G7572) consists of CellTilter-Glo lyophilized powder and CellTilter-Glo buffer. When used, dissolve the lyophilized powder in the buffer.

Hep3B cells (ATCC, HB-8064) (cell source: Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences) were cultured in DMEM complete medium (Thermofisher, 11995073) containing 10% FBS (GBICO, 10099-141) and 100 units/ml penicillin-streptomycin mixture (Thermofisher, 15140122). When the cell coverage in the culture vessel reached 80-90%, the cells were digested and blown off with 0.25% trypsin (containing EDTA) (Thermofisher, 25200056) and then seeded into white 384-well plates (Thermofisher, 164610), with 1000 cells per well (27 μl DMEM complete medium), and then the 384-well plates were placed in a 37°C, 5% CO2 incubator and cultured overnight (18-20 hours). The compound was dissolved and diluted to 5 mM in 100% DMSO, then serially diluted 4-fold with DMSO to a minimum concentration of 0.061 μM. Each concentration point was further diluted 50-fold using FBS-free DMEM medium. If the compound IC50 value is very low, the starting concentration of the compound can be reduced. After overnight, 3 μl of the diluted DMEM compound was added to each well and gently centrifuged to mix. The group with 10 μM BLU9931 served as a negative control (100% inhibition), and the group with 0.2% DMSO served as a positive control (0% inhibition). The 384-well plate was placed in an incubator at 37°C and 5% CO2 and continued to be incubated. After 72 hours, the plate was removed and allowed to stand at room temperature for 30 minutes. The CTG reagent was also removed and equilibrated to room temperature. 15 μl of CTG reagent was added to each well and gently shaken on a shaker for 3 minutes to ensure sufficient cell lysis. The luminescence signal was allowed to stabilize for 10 minutes, and then the luminescence signal was read using EnVision (Perkin Elmer).

The percentage of Hep3B cell proliferation inhibition by the compound can be calculated using the following formula:

Percent inhibition = 100 - 100 * (signal _compound - signal _{negative control} ) / (signal _{positive control} - signal _{negative control} )

The IC50 values of the compounds were calculated from 8 concentration points using XLfit (ID Business Solutions Ltd., UK) software using the following formula:

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC50-X)*slope factor))

Wherein Y is the inhibition percentage, Bottom is the bottom plateau of the curve (the bottom platform value of the S-shaped curve), Top is the top plateau of the curve (the top platform value of the S-shaped curve), and X is the logarithm of the concentration of the test compound.

Biological test example: A: <50nM, B: 50-100nM,

As can be seen from the above table, the compounds of the present invention have a good inhibitory effect on the proliferation of Hep3B cells.

Claims (6)

1. A compound according to formula (IV) or a pharmaceutically acceptable salt thereof, characterized in that: R ^Z1 is selected from hydrogen, C1-C4 alkyl, wherein the C1-C4 alkyl is optionally substituted with one or more hydroxyl groups; R ^Z2 is hydrogen; ^R7 is selected from C1-C8 alkyl; R ¹⁰ is CN; R ¹¹ is selected from hydrogen, halogen, C1-C4 alkyl, C1-C6 alkoxy, HO-C1-C4 alkoxy, NR ² R ³ , C1-C4 alkoxy C1-C4 alkoxy; R ² and R ³ are each independently selected from hydrogen and C1-C8 alkyl. 2. A compound selected from the group consisting of: . 3. A pharmaceutical composition comprising the compound according to claim 1 or 2 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. 4. Use of the compound according to claim 1 or 2 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 3 in the preparation of a medicament, wherein the medicament is used to treat or prevent FGFR4-mediated diseases. 5. Use according to claim 4, wherein the disease is cancer. 6. Use according to claim 5, wherein the cancer is hepatocellular carcinoma.