RU2797117C2 - New quinoline-substituted compound - Google Patents

Abstract

FIELD: pharmaceuticals.

SUBSTANCE: invention relates to a method for producing a new compound (I-4), where R₁, R₂, R₉ are defined in the claims having inhibitory effects on EGFR and inhibitory effects on cell growth.

EFFECT: obtaining a new quinoline-substituted compound.

17 cl, 3 tbl, 29 ex

Description

The field of technology to which the invention belongs

Cross-reference to related applications

The present application claims priority of the description of Japanese Patent Application 2013-172746 (the entire disclosure is hereby incorporated by reference), filed August 22, 2013.

The present invention relates to quinoline-substituted compounds having epidermal growth factor receptor (EGFR) inhibitory activity and pharmaceutical compositions containing these compounds as an active ingredient.

Prerequisites for the creation of the invention

EGFR is a tyrosine kinase receptor type, exhibits its physiological function in normal tissue when bound to epidermal growth factor (EGF) which is a ligand, and promotes growth, inhibition of apoptosis, etc. in epithelial tissues (non-patent document (NPD) 1).

In addition, EGFR is one of the oncogenes, and amplification of the EGFR gene and high expression or mutation of its protein are found in various types of cancers, such as head and neck cancer, breast cancer, colorectal cancer, esophageal cancer, pancreatic cancer, lung cancer. , ovarian cancer, kidney cancer, bladder cancer, skin cancer and brain tumor (non-patent document (NPD) 2). In Japan and Western countries, approximately 170 to 375 people per 100,000 of the population die due to cancer each year, and cancer ranks high as the cause of death (Non-Patent Document (NPD) 3). Among the types of cancers, death due to lung cancer is approximately 1,400,000 per year worldwide, and since non-small cell lung cancer accounts for 80% or more of lung cancers, there is a need to develop an effective therapy for this disease (non-patent document (NPD) 4) .

In recent years, the genes responsible for these cancers have been identified, and a mutation in the EGFR gene is also one of them and results in an active mutated EGFR protein. An active mutated EGFR protein is, for example, a deletion of amino acids at positions 746-750 (EGFR (d746-750)), a mutation of the amino acid at position 858 from leucine to arginine (EGFR (L858R)) or the like. Such mutations have been reported, for example, in 20-40% of non-small cell lung cancers in Japan and 10-15% of non-small cell lung cancers in Western countries. Since non-small cell lung cancer with these mutations is highly sensitive to gefitinib (product name: Iressa (registered trademark)) and erlotinib (product name: Tarceva (registered trademark)), which are chemical agents (EGFR inhibitors) , inhibiting kinase activity of EGFR, these chemical agents have been used as therapeutic agents in Japan and Western countries. However, cancer develops resistance to gefitinib and erlotinib after 6 to 12 months of use and the therapeutic effect becomes weak. Therefore, this acquired resistance is a major problem for the treatment of non-small cell lung cancer having a highly sensitive mutated EGFR. Approximately 50% of acquired resistance has been found to be due to the appearance of a resistant mutated EGFR protein (EGFR (d746-750/T790M) or EGFR (T790M/L858R)), which has a second mutation in the EGFR gene, resulting in the replacement of the amino acid at position 790 of threonine with methionine. It is an important task to find a therapeutic agent that is effective against non-small cell lung cancer having this drug-resistant mutated EGFR (Non-Patent Document (NPD) 5).

On the other hand, skin abnormalities and gastrointestinal disorders have been reported as common side effects of EGFR inhibitors gefitinib and erlotinib, which are clinically used as therapeutic agents at present, and EGFR inhibitors such as BIBW2992, etc., which are used in clinical study. In all cases, these side effects are believed to be caused by EGFR inhibitors inhibiting not only the activity of mutated EGFR expressed in non-small cell lung cancer, but also the activity of wild-type EGFR (EGFR(WT)) expressed in the skin or gastrointestinal tract (non-patent document (NPD) 1). From the point of view of reducing the side effect, it is considered preferable to have a weak inhibitory activity against EGFR (WT) in normal tissues.

Thus, it is possible to suppress the growth of non-small cell lung cancer cells having drug-resistant mutated EGFR by administering a chemical agent that has weaker inhibitory activity against wild-type EGFR compared to inhibitory activity against drug-resistant mutated EGFR whose amino acid at position 790 is mutated to methionine at an administered dose where the side effect on the skin or in the digestive tract is not severe. This is predicted to promote cancer treatment and longevity and improve the QOL ratio of patients. In addition, if a chemical agent has weak inhibitory activity against wild-type EGFR, however, is strong in inhibitory activity not only against drug-resistant mutated EGFR, but also against highly sensitive mutated EGFRs, such as EGFR ( d746-750) and EGFR (L858R), etc., which are highly sensitive to gefitinib and erlotinib; there is a possibility of possible suppression of growth of non-small cell lung cancer cells expressing highly sensitive mutated EGFR or drug-resistant mutated EGFR at an administered dose where the side effect on the skin or in the digestive tract is not severe, or the possibility of a possible decrease in the incidence of drug-resistant EGFR , as acquired resistance, from non-small cell lung cancer cells expressing highly sensitive mutated EGFR. This is predicted to promote cancer treatment and longevity and improve the QOL ratio of patients. In addition, since the expressions of highly sensitive mutated EGFR and drug-resistant mutated EGFR can be used in the current field of therapy as indicators of stratification for possible selection of patients, they greatly contribute to this from an ethical point of view.

As a compound having a structure similar to that of the present invention, N-(3-(4-amino-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-5-yl)phenyl)benzamide derivative is known (patent document (PTD) 1). Although Patent Document 1 describes the use of an amide compound for the treatment of diseases characterized by B-RAF kinase, the document does not disclose specific tests and results confirming kinase inhibitory activity, and the activity is not confirmed either.

Link List

Patent Documents

PTD 1: International publication WO 2006/102079 brochure

Non-Patent Documents

NPD 1: Nature Rev. Cancer, vol. 6, pp. 803-811 (2006)

NPD 2: J. Clin. Oncol., vol. 19, pp. 32-40 (2001)

NPD 3: Ministry of Internal Affairs and Communications Statistics Bureau homehage/statistical data/world statistics “World Statistics 2011”

NPD 4: Lung Cancer, Volume 69, pp. 1-12 (2010)

NPD 5: Nature Rev. Cancer, vol. 10, pp. 760-774 (2010)

Brief summary of the invention

Technical problem

As described above, EGFR inhibitors, while believed to be effective in cancer therapy, are currently clinically ineffective.

Therefore, it is an object of the present invention to provide a novel compound that strongly inhibits EGFR, or a salt thereof. A further object of the present invention is to provide a novel compound that inhibits mutated EGFR, e.g. EGFR (d746-750), EGFR (L858R), EGFR (d746-750/T790M) and EGFR (T790M/L858R), but does not inhibit EGFR (WT ); or its salt.

Solution

The inventors of the present invention have conducted a comprehensive study to achieve the above object. As a result, the present inventors have found that a group of quinoline-substituted compounds of the present invention has an excellent EGFR inhibitory activity and exhibits a cancer cell growth inhibitory action, and these compounds are useful as a cancer treatment drug. The present inventors have thus arrived at the present invention.

Thus, the present invention provides the following items.

1. The compound represented by the following formula (I), or a salt thereof:

where group:

represents

(1) a group represented by formula A1:

(in the formula A1, B means the group represented by the formula:

[R ₁ means a hydrogen atom or C ₁ -C ₆ -alkyl group;

and R ₂ means a group represented by the formula:

where R ₃ , R ₄ and R ₅ are the same or different, and each represents a hydrogen atom, a halogen atom, a C ₁ -C ₆ alkyl group, a C ₆ -C ₁₂ aryl group, a C ₄ -C ₉ heteroaryl group, an aminomethyl group which may be substituted by a C ₁ -C ₆ alkyl group, or a 1-piperidinomethyl group,

or a group represented by the formula:

where R ₆ means a hydrogen atom or a C ₁ -C ₆ -alkyl group],

R ₇ and R ₈ are the same or different and each represents a hydrogen atom or a C ₁ -C ₆ alkyl group; m means 0 or 1; n means 1 or 2);

(2) a group represented by formula A2:

(in formula A2, B and n have the meanings as described in formula A1; and R ₉ is a hydrogen atom or a C ₁ -C ₆ alkyl group); or

(3) a group represented by formula A3:

(in formula A3, B, m and n have the meanings as defined in formula A1; and R ₁₀ is a C ₁ -C ₆ alkyl group).

2. The compound, or salt thereof, according to claim 1, where R ₂ is a group represented by the formula:

where R ₃ , R ₄ and R ₅ are the same or different and each represents a hydrogen atom, a halogen atom, a C ₁ -C ₆ alkyl group, an aminomethyl group which may be substituted with a C ₁ -C ₆ alkyl group, or 1 -piperidinomethyl group,

or a group represented by the formula:

where R ₆ means a hydrogen atom or a C ₁ -C ₆ -alkyl group.

3. The compound, or salt thereof, according to claim 1 or 2, where R ₂ is a group represented by the formula:

where R ₃ , R ₄ and R ₅ are the same or different, and each represents a hydrogen atom, a halogen atom, an aminomethyl group which may be substituted with a methyl group, or a 1-piperidinomethyl group.

4. A compound or salt thereof according to any one of claims 1-3, where the group is:

represents

(1) a group represented by formula A1:

(in the formula A1, B means the group represented by the formula:

where R ₁ means a hydrogen atom or a C ₁ -C ₆ -alkyl group; and R ₂ means a group represented by the formula:

where R ₃ , R ₄ and R ₅ are the same or different and each represents a hydrogen atom or a halogen atom,

R ₇ and R ₈ are the same or different and each represents a hydrogen atom or a C ₁ -C ₆ alkyl group; m means 0 or 1; n means 1); or

(2) a group represented by formula A2:

(in formula A2, B and n have the meanings as defined in formula A1; and R ₉ is a hydrogen atom or a C ₁ -C ₆ alkyl group).

5. A compound, or salt thereof, according to any one of claims 1-4, where the group is:

represents

(1) a group represented by formula A1:

(in the formula A1, B means the group represented by the formula:

where R ₁ means a hydrogen atom; and R ₂ means a group represented by the formula:

where R ₃ , R ₄ and R ₅ each represent a hydrogen atom,

R ₇ and R ₈ each represent a hydrogen atom; m means 0; and n means 1); or

(2) a group represented by formula A2:

(in formula A2, B and n have the meanings as defined in formula A1; and R ₉ is a C ₁ -C ₆ alkyl group).

6. A compound, or salt thereof, according to any one of claims 1 to 5, wherein the compound is selected from the following group of compounds consisting of:

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)acrylamide (Compound 1);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)methacrylamide (Compound 2);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)but-2 α-enamide (mixture of E and Z isomers) (Compound 3);

(S,E)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(dimethylamino)but-2-enamide (Compound 4);

(S,E)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 3-chloroacrylamide (Compound 5);

(S,Z)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 3-chloroacrylamide (Compound 6);

(S,E)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(piperidin-1-yl)but-2-enamide (Compound 7);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)propiolamide (Compound 8);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)but-2 -inamide (Compound 9);

(S,E)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(diethylamino)but-2-enamide (Compound 10);

(S,E)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(ethyl(methyl)amino)but-2-enamide (Compound 11);

(S,E)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(isopropyl(methyl)amino)but-2-enamide (Compound 12);

(R)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-7-yl)acrylamide (Compound 13);

(S)-N-(4-amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 14);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 15);

(R)-N-(4-amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 16);

(R)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 17);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8,9,10-tetrahydro-6H-pyrimido[5',4':4,5]pyrrolo [1,2-a]azepin-8-yl)acrylamide (Compound 18);

(S,E)-N-(4-amino-6-ethyliden-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 19);

(S)-N-(4-amino-6-isopropyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 20A);

(S)-N-(4-amino-6-(propan-2-ylidene)-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8 -yl)acrylamide (Compound 20B);

(R)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)-N- methylacrylamide (Compound 21);

(R)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)-N- methylacrylamide (Compound 22);

(R)-N-(4-Amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)-N-methylacrylamide (Compound 23 );

N-((7S)-4-amino-6-methyl-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)acrylamide (Compound 24);

(R)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)acrylamide (Compound 25);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)-N- methylacrylamide (Compound 26);

(S)-N-(4-amino-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 27);

(R)-N-(4-amino-5-(quinolin-3-yl)-9,10-dihydro-8H-pyrimido[5',4':4,5]pyrrolo[1,2-a]azepine -8-yl)acrylamide (Compound 28);

N-((6R*,8S)-4-amino-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl) acrylamide (Compound 29A); And

N-((6S*,8S)-4-amino-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl) acrylamide (Compound 29B).

7. A compound, or salt thereof, according to any one of claims 1 to 5, wherein the compound is selected from the following group of compounds consisting of:

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)acrylamide (Compound 1);

(S,E)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 3-chloroacrylamide (Compound 5);

(S,Z)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 3-chloroacrylamide (Compound 6);

(S)-N-(4-amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 14);

(S)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 15);

(S,E)-N-(4-amino-6-ethyliden-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 19);

(R)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)-N- methylacrylamide (Compound 21);

(R)-N-(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)-N- methylacrylamide (Compound 22);

(R)-N-(4-Amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)-N-methylacrylamide (Compound 23 ); And

(R)-N-(4-amino-5-(quinolin-3-yl)-9,10-dihydro-8H-pyrimido[5',4':4,5]pyrrolo[1,2-a]azepine -8-yl)acrylamide (Compound 28).

8. (S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)acrylamide (Compound 1) or its salt.

9. (S)-N-(4-Amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 14) or its salt.

10. An EGFR inhibitor containing a compound or salt thereof according to any one of claims 1 to 9 as an active ingredient.

11. A pharmaceutical composition containing a compound, or salt thereof, according to any one of claims 1-9.

12. An antitumor agent containing a compound, or salt thereof, according to any one of claims 1 to 9 as an active ingredient.

13. A method of treating or preventing a cancer, the method comprising the step of administering to a mammal a compound, or salt thereof, according to any one of claims 1 to 9, at a dose effective to treat or prevent the cancer.

14. The use of a compound or salt thereof, according to any one of claims 1 to 9, for the preparation of an antitumor agent.

15. A compound or salt thereof, according to any one of claims 1 to 9, for use in the prevention or treatment of cancer.

Advantageous Effects of the Invention

According to the present invention, a novel compound represented by the above formula (I) or a salt thereof useful as an EGFR inhibitor is obtained.

It has been found that the compound of the present invention, or a salt thereof, has an excellent EGFR inhibitory activity and an inhibitory effect on the growth of cancer cell strains. In addition, the compound of the present invention, or a salt thereof, advantageously has few side effects as a result of excellent selectivity against EGFR. Therefore, the compound of the present invention, or a salt thereof, is useful as an agent for the prevention and/or treatment of cancer.

Description of Embodiments

The compound represented by formula (I) according to the present invention is a quinoline-substituted compound having a quinoline structure and an unsaturated α,β-amide structure, and is a novel compound not described anywhere in any of the above prior art documents and etc.

Specifically, the compound specifically disclosed in PTD 1 is N-(3-(4-amino-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-5-yl)phenyl)benzamide derivative.

The compound of the present invention differs from the compound disclosed in PTD 1 because the compound of the present invention has a quinoline structure and an unsaturated α,β-amide structure.

In the description of the present invention, examples of "halogen atom" include fluorine, chlorine, bromine and iodine.

In the description of the present invention, the term "C ₁ -C ₆ -alkyl group" refers to a straight or branched alkyl group having 1-6 carbon atoms. Specific examples of the alkyl group include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, t-butyl, pentyl, and hexyl.

In the description of the present invention, the term "C ₆ -C ₁₂ -aryl group" refers to an aryl group having 6-12 carbon atoms. Specific examples of the aryl group include phenyl, naphthyl and biphenyl.

In the description of the present invention, the term "C ₄ -C ₉ -heteroaryl group" refers to a monocyclic or bicyclic C ₄ -C ₉ -heteroaryl group containing from 1 to 3 identical or different heteroatoms selected from nitrogen, oxygen and sulfur atoms; and preferably represents a monocyclic or bicyclic C ₄ -C ₉ -heteroaryl group containing 1-3 nitrogen atoms. Specific examples of the heteroaryl group include thienyl, furyl, pyrrolyl, triazolyl, imidazolyl, pyrazolyl, isothiazolyl, isoxazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, isobenzofuryl, indolizinyl, isoindolyl, indolyl, indazolyl, quinolyl, isoquinolyl, phthalazinyl, and naphthyridinyl.

In the description of the present invention, the term "aminomethyl group which may be substituted with a C ₁ -C ₆ alkyl group" refers to an aminomethyl group in which at least one hydrogen atom of the amino group may be substituted with a straight or branched alkyl group, having 1-6 carbon atoms. Specific examples include aminomethyl, N-methylaminomethyl, N,N-dimethylaminomethyl, N-ethylaminomethyl, N,N-diethylaminomethyl, N-methyl-N-ethylaminomethyl, N-methyl-N-isopropylaminomethyl, N-propylaminomethyl, N-butylaminomethyl, N -pentylaminomethyl and N-hexylaminomethyl.

In the above formula (I), the fragment represented by the formula:

is (1) a group represented by formula A1:

(in the formula A1, B, m, n, R ₇ and R ₈ have the meanings as defined above);

(2) a group represented by formula A2:

(in the formula A2, B, n and R ₉ have the meanings as defined above);

or

(3) a group represented by formula A3:

(in the formula A3, B, m, n, R ₇ and R ₈ have the meanings as defined above);

and preferably is (1) a group represented by formula A1:

(in the formula, B, m, n, R ₇ and R ₈ have the meanings as defined above); or

(2) a group represented by formula A2:

(in the formula, B, n and R ₉ have the meanings as defined above);

m, in formula (I), preferably means 0.

n, in formula (I), preferably means 1.

R ₁ in formula (I) preferably represents a hydrogen atom.

R ₂ in formula (I) preferably means a group represented by the formula:

(where R ₃ , R ₄ and R ₅ have the meanings as defined above).

R ₃ , R ₄ and R ₅ in formula (I) are preferably the same or different, and each represents a hydrogen atom, a halogen atom, a C ₁ -C ₆ alkyl group, an aminomethyl group which may be substituted with C ₁ - With a ₆ -alkyl group, or 1-piperidinomethyl group; more preferably, R ₃ , R ₄ and R ₅ are the same or different and each represents a hydrogen atom, a halogen atom, an aminomethyl group which may be substituted with a methyl group, or a 1-piperidinomethyl group; further preferably, R ₃ , R ₄ and R ₅ are the same or different, and each represents a hydrogen atom or a halogen atom, particularly preferably a hydrogen atom.

R ₇ and R ₈ in formula (I) are preferably the same or different, and each represents a hydrogen atom or a C ₁ -C ₆ alkyl group (preferably a C ₁ -C ₃ alkyl group, more preferably a methyl group ); further, preferably, at least one of R ₇ and R ₈ represents a hydrogen atom; especially preferably both R ₇ and R ₈ are hydrogen atoms.

R ₉ in formula (I) preferably represents a C ₁ -C ₆ alkyl group, more preferably a C ₁ -C ₃ alkyl group, further preferably a methyl group.

The compound of the present invention, or a salt thereof, preferably has strong enzyme-inhibiting activity against EGFR (T790M/L858R); more preferably, the concentration of the compound at which the enzyme can be inhibited by 50% is 2 nM or less. Further, the compound of the present invention or a salt thereof preferably has strong enzyme-inhibiting activity against EGFR (d746-750/T790M); the 50% inhibitory concentration of the compound is also preferably 2 nM or less. Further, the compound of the present invention or a salt thereof preferably has a strong cell growth inhibitory effect against EGFR tumor cells (T790M/L858R); more preferably, the 50% inhibitory concentration of the compound is 200 nM or less, further preferably 100 nM or less, and particularly preferably 40 nM or less.

Next, a method for preparing a compound of the present invention is disclosed.

The compound (I) according to the present invention can be produced, for example, by the following production method, the methods described in the Examples, and the like. However, the method for producing the compound of the present invention is not limited to these reaction examples.

Production method 1

where R ₁ represents a protective group of the amino group; L ₁ is a leaving group; and R ₂ , R ₇ , R ₈ , m and n are as defined above.

Stage a

In this step, a compound represented by formula (IV) is obtained by a Mitsunobu reaction using compounds represented by formulas (II) and (III).

Examples of the leaving group represented by L ₁ in the compound represented by formula (II) include a bromine atom or an iodine atom. The compound represented by formula (II) may be a commercially available product, or it may be obtained by a known method. Examples of the amino protecting group represented by P ₁ in the formula (III) include a t-butoxycarbonyl group and a benzoyl group. The compound represented by formula (III) may be a commercially available product, or it may be obtained by a known method. The compound represented by the formula (III) can be used in an amount of 1 mol to 10 mol, and preferably 1 to 5 mol, based on the mol of the compound represented by the formula (II).

The Mitsunobu reaction can be carried out according to a known method (eg, the method disclosed in Synthesis, p. 1, 1981) or using a similar method.

Examples of azodicarboxylic acid esters include diethyl azodicarboxylate and diisopropyl azodicarboxylate. Such an azodicarboxylic acid ester can be used in an amount of 1-10 mol, and preferably 1-5 mol, based on the mol of the compound represented by formula (II).

As the phosphine compound, triphenylphosphine, tributylphosphine or the like can be used. The phosphine compound can be used in an amount of 1-10 mol, and preferably 1-5 mol, based on the mol of the compound represented by formula (II).

As the solvent, tetrahydrofuran, 1,2-dimethoxyethane, 1,4-dioxane, toluene, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, N-methylpyrrolidin-2-one and the like can be used alone or in the form of a mixture. The reaction time is 0.1 hour to 100 hours, and preferably 0.1 to 24 hours. The reaction temperature is from 0°C to the boiling point of the solvent, and preferably from 0°C to 100°C.

The thus obtained compound represented by formula (IV) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation and chromatography.

Stage b

In this step, the compound represented by formula (IV) is reacted with ammonia, or a salt thereof, to obtain a compound represented by formula (V).

The amount of ammonia or its salt used in this step is usually from an equimolar amount to an excess molar amount per mol of the compound represented by formula (IV).

You can use any solvent for the reaction, which does not adversely affect the reaction. Examples of suitable reaction solvents include water, methanol, ethanol, isopropanol, tert-butyl alcohol, tetrahydrofuran, 1,2-dimethoxyethane, 1,4-dioxane, N,N-dimethylformamide, N-methylpyrrolidin-2-one, dimethyl sulfoxide and those derived from them mixed solvents.

The reaction temperature is usually from 0°C to 200°C, preferably from room temperature to a temperature of 150°C. The reaction time is usually 5 minutes to 7 days and preferably 30 minutes to 24 hours.

The compound represented by formula (V) thus obtained can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, concentration in vacuo, crystallization, solvent extraction, reprecipitation and chromatography.

Stage c

In this step, the compound represented by formula (V) is subjected to a coupling reaction with 3-quinolineboronic acid or 3-quinolineboronic acid ester to obtain the compound represented by formula (VI).

This step can be carried out in accordance with a well-known method (for example, Chemical Reviews, vol. 95, p. 2457, 1995). This step can be carried out in the presence of a transition metal and base catalyst, in a solvent that does not adversely affect the reaction.

The amount of 3-quinolineboronic acid or 3-quinolineboronic acid ester used may be 1-10 mol, and preferably 1-3 mol, based on the mol of the compound represented by formula (V).

Examples of transition metal catalysts include palladium catalysts (e.g., palladium acetate, palladium chloride, tetrakis(triphenylphosphine)palladium, 1,1'-bis(diphenylphosphino)ferrocenepalladium(II)dichloride, and tris(dibenzylideneacetone)dipalladium(0)) , nickel-based catalysts (eg nickel chloride), and the like. If necessary, a ligand (e.g., triphenylphosphine, tri-tert-butylphosphine, or 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl) can be added and, as a cocatalyst, a metal oxide (such as copper oxide or silver oxide) can be used. ). The amount of transition metal catalyst used may vary depending on the type of catalyst. The transition metal catalyst is generally used in an amount of 0.0001 mol to 1 mol, and preferably 0.01 mol to 0.5 mol, based on the mol of the compound represented by formula (V). The amount of ligand used is usually 0.0001 mol to 4 mol, and preferably 0.01 mol to 2 mol, based on the mol of the compound represented by formula (V). The amount of cocatalyst used is usually 0.0001 mol to 4 mol, and preferably 0.01 mol to 2 mol, based on the mol of the compound represented by formula (V).

Examples of suitable bases include organic amines (e.g., trimethylamine, triethylamine, diisopropylethylamine, N-methylmorpholine, 1,8-diazabicyclo[5.4.0]undec-7-ene, pyridine, and N,N-dimethylaniline), alkali metal salts ( e.g. sodium hydrogen carbonate, potassium hydrogen carbonate, sodium carbonate, potassium carbonate, cesium carbonate, sodium phosphate, potassium phosphate, sodium hydroxide and potassium hydroxide), metal hydrides (e.g. potassium hydride and sodium hydride), alkali metal alkoxides (e.g. sodium methoxide , sodium ethoxide, sodium t-butoxide, and potassium t-butoxide), alkali metal disilazides (eg, lithium disilazide, sodium disilazide, and potassium disilazide), and the like. Among them, alkali metal salts such as sodium carbonate, potassium carbonate, cesium carbonate, sodium phosphate and potassium phosphate are preferable; alkali metal alkoxides such as sodium t-butoxide and potassium t-butoxide; and organic amines such as triethylamine and diisopropylethylamine. The amount of base used is usually 0.1-10 mol, and preferably 1-5 mol, based on the mol of the compound represented by formula (V).

You can use any solvent that does not adversely affect the reaction. Examples of a suitable solvent include hydrocarbons (eg benzene, toluene and xylene), halogenated hydrocarbons (eg chloroform and 1,2-dichloroethane), nitriles (eg acetonitrile), ethers (eg 1,2-dimethoxyethane, tetrahydrofuran and 1 ,4-dioxane), alcohols (eg methanol and ethanol), aprotic polar solvents (eg N,N-dimethylformamide, dimethyl sulfoxide and hexamethylphosphorylamide), water and mixed solvents derived therefrom. The reaction time is from 0.1 hour to 100 hours and preferably from 0.5 hour to 24 hours. The reaction temperature is from 0°C to the boiling point of the solvent, and preferably from 20°C to 150°C.

The thus obtained compound represented by formula (VI) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation and chromatography.

Stage d

In this step, the compound represented by formula (VI) is brominated by reacting with N-bromosuccinimide to obtain the compound represented by formula (VII).

Halogenation can be carried out using the method disclosed in patent application number WO 2006/102079 or using a similar method.

The amount of N-bromosuccinimide used in this step is 0.5 mol to 2.0 mol, and preferably 0.9 mol to 1.2 mol, based on the mol of the compound represented by formula (VI).

You can use any solvent for the reaction, which does not adversely affect the reaction. For example, tetrahydrofuran, 1,4-dioxane, N,N-dimethylformamide, N-methylpyrrolidin-2-one, or mixed solvents derived therefrom can be preferably used.

The reaction temperature is usually -20°C to 50°C, and preferably from 0°C to room temperature. The reaction time is usually from 1 minute to 2 days and preferably from 5 minutes to 12 hours.

The thus obtained compound represented by formula (VI) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation and chromatography.

Stage e

In this step, the compound represented by formula (VII) is subjected to an intramolecular cyclization reaction to obtain a compound represented by formula (VIII).

This stage can be carried out in accordance with a well-known method (for example, the method disclosed in Chemical Reviews, volume 103, p. 2945, 2003).

Examples of transition metal catalysts include divalent palladium catalysts (e.g., palladium acetate, palladium chloride, 1,1'-bis(diphenylphosphino)ferrocenepalladium(II) dichloride, etc.) and zero valence palladium catalysts ( for example, tetrakis(triphenylphosphine)palladium, tris(dibenzylideneacetone)dipalladium, etc.). If necessary, a ligand (eg, triphenylphosphine, tri-tert-butylphosphine, etc.) can be added. The amount of transition metal catalyst used may vary depending on the type of catalyst. The transition metal catalyst is usually used in an amount of 0.0001 mol to 1 mol, and preferably 0.01 mol to 0.5 mol, based on the mol of the compound represented by formula (VII). The amount of ligand used is usually 0.0001 mol to 4 mol, and preferably 0.01 mol to 2 mol, based on the mol of the compound represented by formula (VII).

Examples of suitable bases include inorganic bases such as sodium hydrogen carbonate, sodium carbonate, potassium carbonate, cesium carbonate, sodium hydroxide, potassium hydroxide, cesium hydroxide, sodium hydride or potassium hydride. Such a base can be used in an amount of 1 mol to 100 mol, and preferably 2 mol to 20 mol, based on the mol of the compound represented by formula (VII).

You can use any solvent that does not adversely affect the reaction. Examples of suitable solvents include hydrocarbons (e.g. benzene, toluene and xylene), ethers (e.g. 1,2-dimethoxyethane, tetrahydrofuran and 1,4-dioxane), aprotic polar solvents (e.g. N,N-dimethylformamide, dimethyl sulfoxide and hexamethylphosphorylamide ), water and mixed solvents derived from them. The reaction time is from 0.1 hour to 100 hours and preferably from 0.5 hour to 24 hours. The reaction temperature is from -20°C to the boiling point of the solvent, and preferably from 0°C to 150°C.

The thus obtained compound represented by formula (VIII) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation and chromatography.

Stage f

In this step, the protected amino group of the compound represented by formula (VIII) is deprotected to obtain the compound represented by formula (IX).

Deprotection can be carried out using a known method, such as the method described in Protective Groups in Organic Synthesis, T.W. Greene, John Wiley & Sons (1981); or using a similar method.

When a t-butoxycarbonyl group is used as a protecting group, hydrochloric acid, sulfuric acid, methanesulfonic acid, trifluoroacetic acid, and the like are used. used as a deprotection agent. The amount of the reagent used is preferably 1 mol to 100 mol, based on the mol of the compound (VIII).

You can use any solvent that does not adversely affect the reaction. Examples of suitable solvents include water, methanol, ethanol, methylene chloride, chloroform, and the like, and mixed solvents derived therefrom. The reaction time is from 0.1 hour to 100 hours and preferably from 0.5 hour to 24 hours. The reaction temperature is from 0°C to the boiling point of the solvent, and preferably from 0°C to 50°C.

The thus obtained compound represented by formula (IX) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation and chromatography.

Stage g

In this step, the compound represented by formula (IX) is amidated with an α,β-unsaturated carboxylic acid or an α,β-unsaturated carboxylic acid chloride or acid bromide to obtain a compound represented by formula (I-1) according to the present invention.

When a carboxylic acid is used as the amidation reagent, the carboxylic acid can be used in an amount of 0.5 mol to 10 mol, preferably 1 mol to 3 mol, based on the mol of the compound represented by formula (IX), in the presence of a suitable condensing agent. agent. The carboxylic acid may be a commercially available product or may be prepared according to a known method.

You can use any solvent for the reaction, which does not adversely affect the reaction. Examples of a suitable solvent include toluene, benzene, methylene chloride, chloroform, tetrahydrofuran, 1,4-dioxane, N,N-dimethylformamide, dimethylacetamide, N-methylpyrrolidin-2-one, dimethyl sulfoxide, and mixed solvents derived therefrom. The reaction temperature is usually -78°C to 200°C and preferably 0°C to 50°C. The reaction time is usually 5 minutes to 3 days, and preferably 5 minutes to 10 hours.

Examples of condensing agents include diphenylphosphoryl azide, N,N'-dicyclohexylcarbodiimide, benzotriazol-1-yloxy-trisdimethylaminophosphonium salts, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride, 1- ethyl 3-(3-dimethylaminopropyl)carbodiimide, combination of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and 1-hydroxybenzotriazole, 2-chloro-1,3-dimethylimidazolinium chloride, O-(7-azabenzotriazo-1-yl) -N,N,N',N'-tetramethylhexauronium hexafluorophosphate and the like.

If necessary, a base may optionally be added during the reaction. Examples of suitable bases include organic bases such as triethylamine, diisopropylethylamine, pyridine, lutidine, collidine, 4-(N,N-dimethylamino)pyridine, potassium t-butyrate, sodium t-butyrate, sodium methoxide, sodium ethoxide, lithium hexamethyldisilazide, hexamethyldisilazide sodium, potassium hexamethyldisilazide and butyllithium; and inorganic bases such as sodium hydrogen carbonate, sodium carbonate, potassium carbonate, cesium carbonate, sodium hydroxide and sodium hydride. Such a base can be added in an amount of 1 mol to 100 mol, and preferably 1 mol to 10 mol, based on the mol of the compound represented by formula (IX).

When an acid chloride or an acid bromide is used as an amidation reagent, the acid halide is used in an amount of 0.5 mol to 5 mol, and preferably 0.9 mol to 1.1 mol, based on the mol of the compound represented by the formula ( IX). The acid halide may be a commercially available product or may be prepared according to a known method.

You can use any solvent for the reaction, which does not adversely affect the reaction. Examples of such solvents include toluene, benzene, methylene chloride, chloroform, tetrahydrofuran, 1,4-dioxane, N,N-dimethylformamide, dimethylacetamide, N-methylpyrrolidin-2-one, acetonitrile, water, and mixed solvents derived therefrom. The reaction temperature is usually -78°C to 200°C, preferably 0°C to 50°C. The reaction time is usually 5 minutes to 3 days, and preferably 5 minutes to 10 hours.

If necessary, a base may be added during the reaction. Examples of suitable bases include organic bases such as triethylamine, diisopropylethylamine, pyridine, lutidine, collidine, 4-(N,N-dimethylamino)pyridine, potassium t-butyrate, sodium t-butyrate, sodium methoxide, sodium ethoxide, lithium hexamethyldisilazide, hexamethyldisilazide sodium, potassium hexamethyldisilazide and butyllithium; and inorganic bases such as sodium hydrogen carbonate, sodium carbonate, potassium carbonate, cesium carbonate, sodium hydroxide and sodium hydride. Such a base can be added in an amount of 1 mol to 100 mol, and preferably 1 mol to 20 mol, based on the mol of the compound represented by formula (IX).

The thus obtained compound represented by formula (I-1) can be isolated and purified by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation, and chromatography.

Obtaining Method 2

where R ₁ represents a protective group for the amino group; and R ₂ , R ₇ , R ₈ , R ₉ and n are as defined above.

Stage h

At this step, the protected amino group of the compound represented by formula (X) is deprotected to obtain the compound represented by formula (XI).

Deprotection can be carried out using a known method, such as the method described in Protective Groups in Organic Synthesis, T.W. Greene, John Wiley & Sons (1981); or using a similar method.

When a t-butoxycarbonyl group is used as a protecting group, hydrochloric acid, sulfuric acid, methanesulfonic acid, trifluoroacetic acid, and the like are used. can be used as a deprotection agent. Preferably, the reagent is used in an amount of 1 mol to 100 mol, based on the mol of the compound (X).

You can use any solvent that does not adversely affect the reaction. Examples of suitable solvents include water, methanol, ethanol, methylene chloride, chloroform, and mixed solvents thereof. The reaction time is from 0.1 hour to 100 hours and preferably from 0.5 hour to 24 hours. The reaction temperature is from 0°C to the boiling point of the solvent, and preferably from 50°C to the boiling point of the solvent.

The thus obtained compound represented by formula (XXI) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation, and chromatography.

Stage i

This step can be carried out in the same manner as step g.

Obtaining method 3

where R ₁ represents a protective group for the amino group; L ₁ and L ₂ represent a leaving group; and R ₁ , R ₂ , R ₇ , R ₈ , m and n are as defined above.

Stage j

In this step, the compound represented by the formula (XIII) is obtained by an alkylation reaction using the compounds represented by the formulas (IV) and (XII) in the presence of a base.

In the compound represented by formula (XII), examples of the leaving group represented by L ₂ include bromine atom, iodine atom, methanesulfonic acid ester, p-toluenesulfonic acid ester, and the like. The compound represented by formula (XII) may be a commercially available product, or it may be obtained by a known method. The compound represented by formula (XII) can be used in an amount of 1 mol to 10 mol, and preferably 1 mol to 5 mol, based on mol of the compound represented by formula (IV). Examples of suitable bases include organic amines (e.g., trimethylamine, triethylamine, diisopropylethylamine, N-methylmorpholine, 1,8-diazabicyclo[5.4.0]undec-7-ene, pyridine, and N,N-dimethylaniline), alkali metal salts ( e.g. sodium hydrogen carbonate, potassium hydrogen carbonate, sodium carbonate, potassium carbonate, cesium carbonate, sodium phosphate, potassium phosphate, sodium hydroxide and potassium hydroxide), metal hydrides (e.g. potassium hydride and sodium hydride), alkali metal alkoxides (e.g. sodium methoxide , sodium ethoxide, sodium t-butoxide and potassium t-butoxide), alkali metal disilazides (for example, lithium disilazide, sodium disilazide and potassium disilazide), and the like. Among these, alkali metal salts such as sodium carbonate, potassium carbonate, cesium carbonate, sodium phosphate and potassium phosphate; and metal hydrides such as sodium hydride; alkali metal alkoxides such as sodium t-butoxide and potassium t-butoxide. The amount of base used is usually 0.1 mol to 10 mol, and preferably 1 mol to 5 mol, based on the mol of the compound represented by formula (V).

As the solvent, tetrahydrofuran, 1,2-dimethoxyethane, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, N-methylpyrrolidin-2-one and the like can be used, alone or as mixtures. The reaction time is from 0.1 hour to 100 hours and preferably from 0.5 hour to 24 hours. The reaction temperature is from 0°C to the boiling point of the solvent, and preferably from 20°C to 150°C.

The thus obtained compound represented by formula (XIII) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation and chromatography.

Stage k

This step can be carried out in the same manner as step b.

Stage l

This step can be carried out in the same manner as step c.

stage m

This step can be carried out in the same manner as step d.

Stage n

This step can be carried out in the same manner as step e.

Stage o

This step can be carried out in the same manner as step f.

Stage p

This step can be carried out in the same manner as step g.

Production method 4

where R ₁ represents a protective group for the amino group; and R ₁ , R ₂ , R ₇ , R ₈ , R ₉ and n are as defined above.

Stage q

This step can be carried out in the same manner as step h.

Stage r

This step can be carried out in the same manner as step g.

Production method 5

where R ₁ represents a protective group for the amino group; and R ₁ , R ₂ , R ₇ , R ₈ , R ₁₀ , m and n are as defined above.

Stage s

In this step, the compound represented by the formula (XX) is obtained by hydrogenating the compound represented by the formula (XVII) in the presence of a catalyst.

Examples of the catalyst include palladium-on-charcoal, palladium-on-charcoal hydroxide, and Raney nickel. The catalyst can be used in an amount of 0.01 mol to an excess amount, preferably 0.1 mol to 10 mol, based on the mol of the compound represented by the formula (XVII).

Hydrogenation can be carried out at a pressure of 1 atm. up to 100 atm., preferably from 1 to 10 atm. As a solvent, methanol, ethanol, ethyl acetate, tetrahydrofuran, 1,2-dimethoxyethane, 1,4-dioxane, N,N-dimethylformamide, N,N-dimethylacetamide, N-methylpyrrolidin-2-one, water, etc. can be used. ., individually or as a mixture. The reaction time is 0.1 hour to 100 hours, and preferably 0.5 hour to 48 hours. The reaction temperature is in the range from room temperature to the boiling point of the solvent, preferably from room temperature to 100°C.

The thus obtained compound represented by formula (XX) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation, and chromatography.

Stage t

This step can be carried out in the same manner as step f.

Stage u

This step can be carried out in the same manner as step g.

Production method 6

where R ₁ represents a protective group for the amino group; and R ₂ and n are as defined above.

stage v

In this step, an organic borane reagent is reacted with a compound represented by formula (XXII) to obtain an intermediate alkylborane compound in the system, and then, in the presence of a transition metal catalyst and a base, a compound represented by formula (XXIII) is obtained.

This step can be carried out according to a generally known method (eg WO 2006/102079).

Examples of organic borane reagents include 9-BBN(9-borabicyclo[3,3,1]-nonane), 9-BBN(9-borabicyclo[3,3,1]-nonane dimer), disyamilborane(bis(1,2- dimethylpropyl)borane), texylborane (1,1,2-trimethylpropyl)borane) and the like. The organic borane reagent is preferably 9-BBN(9-borabicyclo[3.3.1]-nonane or 9-BBN(9-borabicyclo[3.3.1]-nonane) dimer and particularly preferably 9- BBN(9-borabicyclo[3,3,1]-nonane) The amount of organic borane reagent used is not particularly limited, as long as an intermediate alkylborane compound can be obtained The organic borane reagent can be used in an amount of 1 mol to 20 mol , in terms of mol of the compound represented by the formula (XXII), the amount of the organic borane reagent is preferably 6 mol to 10 mol from the viewpoint of facilitating the progression of the reaction.

As the transition metal catalyst, for example, a divalent palladium catalyst (eg, palladium acetate, palladium chloride and 1,1'-bis(diphenylphosphino)ferrocenepalladium(II)dichloride) can be used. If necessary, a ligand (eg, triphenylphosphine and tri-tert-butylphosphine) can be used. The amount of transition metal catalyst used may vary depending on the type of catalyst. The transition metal catalyst is typically used in an amount of 0.0001 mol to 1 mol, and preferably 0.01 mol to 0.5 mol, based on the mol of the compound represented by formula (XXII). The ligand is typically used in an amount of 0.0001 mol to 4 mol, and preferably 0.01 mol to 2 mol, based on the mol of the compound represented by formula (XXII).

Alternatively, for example, a zero valent palladium catalyst can be used. Examples of zero valent palladium catalysts include tetrakis(triphenylphosphine)palladium, tris(dibenzylideneacetone)dipalladium, palladium-on-charcoal, and the like. Tetrakis(triphenylphosphine)palladium or tris(dibenzylideneacetone)dipalladium is preferred, and tetrakis(triphenylphosphine)palladium is particularly preferred. The amount of the zero-valent palladium catalyst used is not particularly limited, as long as the intramolecular cyclization reaction can be carried out, and may vary depending on the type of catalyst. The zero valence palladium catalyst can be used in an amount of 0.0001 mol to 1 mol, and preferably 0.01 mol to 0.5 mol, based on the mol of the compound represented by formula (XXII).

If necessary, a ligand can be added along with the zero valent palladium catalyst. Examples of such ligands include triphenylphosphine, 1,1'-bis(diphenylphosphino)ferrocene, tri-tert-butylphosphine, tricyclohexylphosphine, 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl, 2-dicyclohexylphosphino-2',4',6'- triisopropylbiphenyl, 2-(di-tert-butylphosphino)biphenyl, 2-dicyclohexylphosphino-2'-(N,N-dimethylamino)biphenyl, 4,5'-bis(diphenylphosphino)-9,9'-dimethylxanthene and the like. When tris(dibenzylideneacetone)dipalladium is used as the zero valent palladium catalyst, triphenylphosphine can be added as a ligand. The amount of ligand used is not particularly limited, as long as the intramolecular cyclization reaction can be carried out. The ligand can be used in an amount of 0.0001 mol to 4 mol, and preferably 0.01 mol to 2 mol, based on the mol of the compound represented by formula (XXII).

Examples of bases include inorganic bases such as sodium hydrogen carbonate, sodium carbonate, potassium carbonate, cesium carbonate and alkali metal hydroxides. Alkali metal hydroxides are preferred. Examples of alkali metal hydroxides include lithium hydroxide, sodium hydroxide, potassium hydroxide and cesium hydroxide. Lithium hydroxide, sodium hydroxide, potassium hydroxide or cesium hydroxide are preferably used. Especially preferred is lithium hydroxide or sodium hydroxide. The amount of base used is not particularly limited as long as the reaction proceeds. The base can be used in an amount of 1 mol to 100 mol, and preferably 2 mol to 20 mol, based on the mol of the compound represented by formula (XXII). The alkali metal hydroxide can be used in the form of an aqueous solution of the alkali metal hydroxide.

As a combination of an organic borane reactant, an alkali metal hydroxide and a zero valent palladium catalyst, a combination of a preferred organic borane reactant, a preferred alkali metal hydroxide and a preferred zero valence palladium catalyst is preferred. Particularly preferred is the combination of a particularly preferred organic borane reactant, a particularly preferred alkali metal hydroxide, and a particularly preferred zero valence palladium catalyst.

You can use any solvent that does not adversely affect the reaction. Examples of such a solvent include hydrocarbons (such as benzene, toluene, and xylene), ethers (such as 1,2-dimethoxyethane, tetrahydrofuran, and 1,4-dioxane), aprotic polar solvents (such as N,N-dimethylformamide, dimethyl sulfoxide, and hexamethylphosphorylamide ), water, and mixtures thereof. Preferably, 1,2-dimethoxyethane or tetrahydrofuran is used. Tetrahydrofuran is particularly preferred in terms of the stability of the organic borane reagent and the generated alkylborane intermediate. The amount of solvent used is not particularly limited as long as the reaction proceeds. The solvent can be used in an amount that is 1 to 300 times, and preferably 10 to 96 times, by weight of the compound represented by formula (XXII).

The reaction time is not particularly limited, as long as the compound of formula (XXIII) can be obtained. The reaction time may be from 0.1 hour to 100 hours, and preferably from 0.5 hour to 24 hours.

The reaction temperature is not particularly limited, as long as the compound represented by the formula (XXIII) can ultimately be obtained. The reaction temperature may be from -20°C to the boiling point of the solvent, and preferably from 0°C to 150°C. In the case of an intramolecular cyclization reaction of an alkylborane intermediate using a zero valence palladium catalyst and an aqueous alkali metal hydroxide solution, the low reaction temperature leads to side reactions, which in turn lead to low yields. Therefore, the temperature is preferably 61° C. or higher.

The thus obtained compound represented by formula (XXIII) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, concentration in vacuo, crystallization, solvent extraction, reprecipitation and chromatography.

At this stage, the formation of an alkylborane intermediate in the system can be confirmed. For example, LCMS spectra can be used as a confirmation method.

Stage w

In this step, the compound represented by formula (XXIV) is obtained by reacting the compound represented by formula (XXIII) with iodobenzene diacetate.

The amount of iodobenzene diacetate used in this step is 1 mol to 10 mol, preferably 1 mol to 2 mol, based on the mol of the compound represented by the formula (XXIII).

In the case of the above reaction, if necessary, tetrabutylammonium iodide can be added. Tetrabutylammonium iodide can be added in an amount of 0.01 mol to 10 mol, preferably 0.1 mol to 1 mol, based on the mol of the compound represented by formula (XXIII).

You can use any solvent for the reaction, which does not adversely affect the reaction. Examples of suitable reaction solvents include methylene chloride, chloroform, 1,2-dichloroethane, acetic acid, and mixed solvents derived therefrom.

The reaction temperature is usually from 0°C to the boiling point of the solvent, and preferably from 0°C to room temperature. The reaction time is from 0.1 hour to 100 hours, and preferably from 0.1 hour to 24 hours.

The thus obtained compound represented by formula (XXIV) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation, and chromatography.

Stage x

At this stage, the protected hydroxyl group of the compound represented by formula (XXIV) is deprotected to obtain a compound of formula (XXV).

Deprotection can be carried out using a known method, such as the method described in Protective Groups in Organic Synthesis, T.W. Greene, John Wiley & Sons (1981); or using a similar method.

When the acetyl group is deprotected, examples of the deprotection agent include sodium hydroxide, potassium hydroxide, and the like. The amount of the reagent used is preferably 1 mol to 100 mol, based on the mol of the compound represented by the formula (XXIV).

You can use any solvent that does not adversely affect the reaction. Examples of the solvent include water, methanol, ethanol, tetrahydrofuran, and mixed solvents derived therefrom. The reaction time is from 0.1 hour to 100 hours and preferably from 0.5 hour to 24 hours. The reaction temperature is from 0°C to the boiling point of the solvent, and preferably from 0°C to 50°C.

The thus obtained compound represented by formula (XXV) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation, and chromatography.

Stage y

In this step, a compound represented by formula (XXVI) is obtained by subjecting a compound represented by formula (XXV) to an elimination reaction.

In the case of the elimination reaction, acids such as p-toluenesulfonic acid monohydrate, 10-camphorsulfonic acid and the like are used. The amount of acid used is 0.1 mol to 100 mol, preferably 1 mol to 10 mol, based on the mol of the compound represented by formula (XXV).

You can use any solvent that does not adversely affect the reaction. Examples of the solvent include toluene, xylene, and mixed solvents derived therefrom. The reaction time is from 0.1 hour to 100 hours and preferably from 0.5 hour to 24 hours. The reaction temperature is in the range from room temperature to the boiling point of the solvent.

The thus obtained compound represented by formula (XXVI) can be subjected to a subsequent step, after or without isolation or purification by known isolation and purification methods such as concentration, vacuum concentration, crystallization, solvent extraction, reprecipitation and chromatography.

Stage z

This step can be carried out in the same manner as step g.

In the case of the above production methods 1 to 6, for functional groups having an active proton, such as amino group, imino group, hydroxyl group, carboxyl group, carbonyl group and amide group, and indole, protective reagents can be used or a protective group can be introduced into such a functional group in accordance with the usual method; subsequently, the protecting group can be removed at an appropriate step for each production method.

The "amino protecting group or imino protecting group" is not particularly limited, as long as the group has a protective function. Examples of such protecting groups include aralkyl groups such as benzyl, p-methoxybenzyl, 3,4-dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl, benzhydryl, trityl and cumyl; lower alkanoyl groups such as formyl, acetyl, propionyl, butyryl, pivaloyl, trifluoroacetyl and trichloroacetyl; benzoyl; arylalkanoyl groups such as phenylacetyl and phenoxyacetyl; lower alkoxycarbonyl groups such as methoxycarbonyl, ethoxycarbonyl, propyloxycarbonyl and t-butoxycarbonyl; aralkyloxycarbonyl groups such as p-nitrobenzyloxycarbonyl and phenethyloxycarbonyl; lower alkylsilyl groups such as trimethylsilyl and t-butyldimethylsilyl; tetrahydropyranyl; trimethylsilylethoxymethyl; lower alkylsulfonyl groups such as methylsulfonyl, ethylsulfonyl and t-butylsulfonyl; lower alkylsulfinyl groups such as t-butylsulfinyl; arylsulfonyl groups such as benzenesulfonyl and toluenesulfonyl; and imido groups such as phthalimido. Particularly preferred are trifluoroacetyl, acetyl, t-butoxycarbonyl, benzyloxycarbonyl, trimethylsilylethoxymethyl, cumyl, and the like.

The "hydroxyl protecting group" is not particularly limited, as long as the group has a protective function. Examples of such protecting groups include lower alkyl groups such as methyl, ethyl, propyl, isopropyl and t-butyl; lower alkylsilyl groups such as trimethylsilyl and t-butyldimethylsilyl; lower alkoxymethyl groups such as methoxymethyl and 2-methoxyethoxymethyl; tetrahydropyranyl; trimethylsilylethoxymethyl; aralkyl groups such as benzyl, p-methoxybenzyl, 2,3-dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl and trityl; and acyl groups such as formyl, acetyl and trifluoroacetyl. Particularly preferred are methyl, methoxymethyl, tetrahydropyranyl, trimethylsilylethoxymethyl, t-butyldimethylsilyl and acetyl.

The "carboxyl group protecting group" is not particularly limited, as long as the group has a protective function. Examples of such protecting groups include lower alkyl groups such as methyl, ethyl, propyl, isopropyl and t-butyl; lower haloalkyl groups such as 2,2,2-trichloroethyl; lower alkenyl groups such as allyl; trimethylsilylethoxymethyl; and aralkyl groups such as benzyl, p-methoxybenzyl, p-nitrobenzyl, benzhydryl and trityl. Particularly, methyl, ethyl, t-butyl, allyl, benzyl, p-methoxybenzyl, trimethylsilylethoxymethyl and the like are preferred.

The "carbonyl protecting group" is not particularly limited, as long as the group has a protective function. Examples of such protecting groups include ethylene ketal, trimethylene ketal, dimethyl ketal, and the like ketals and acetals.

The method for removing such a protecting group may vary depending on the type of protecting group, the stability of the desired compound (I), and so on. For example, the following methods can be used: solvolysis using acid or base, in accordance with the method disclosed in the publication (Protective Groups in Organic Synthesis, third edition, T.W. Greene, John Wiley & Sons (1999)), or a similar method, then that is, a method comprising reacting with 0.01 mol or more of an acid, preferably trifluoroacetic acid, formic acid, or hydrochloric acid, or with an equimolar to a large excess molar amount of a base, preferably potassium hydroxide or calcium hydroxide; chemical reduction using a metal hydride complex, etc.; or catalytic reduction using a palladium-on-charcoal catalyst, a Raney nickel catalyst, etc.

The compound of the present invention can be isolated and purified using conventional isolation and purification methods. Examples of such methods include solvent extraction, recrystallization, preparative reverse phase high performance liquid chromatography, column chromatography, preparative thin layer chromatography, and the like.

Where the compound of the present invention has isomers such as optical isomers, stereoisomers, regioisomers and rotational isomers, any isomers and mixtures thereof are included within the scope of the compound of the present invention. For example, if a compound has optical isomers, then the optical isomer isolated from the racemic mixture is also included within the scope of the compound of the present invention. Each of these isomers can be obtained as an individual compound using known synthesis and separation methods (eg, concentration, solvent extraction, column chromatography, recrystallization, etc.).

According to the present invention, the carbon atom associated with substituent B in formula (I) is an asymmetric carbon atom; therefore, the compound includes isomers. As indicated above, except as otherwise indicated, the compound of the present invention includes all enantiomers and mixtures thereof. The compound of the present invention may be a mixture of R and S enantiomers. Such a mixture may be a mixture containing 90% or more, 95% or more, or 99% or more of the R-enantiomer; or a mixture containing 90% or more, 95% or more, or 99% or more of the S-enantiomer.

Chiral separation methods include, for example: a diastereomeric method based on reacting a chiral resolving agent with a compound of the present invention to form a salt and isolating one of the enantiomers using difference in solubility, etc., from the resulting salt; the preferred method of crystallization by adding one of the enantiomers to a supersaturated solution of the racemate, as a seed, for crystallization; and column chromatography such as HPLC using a chiral column. The chiral resolving agent that can be used in the diastereomeric process can be suitably selected from, for example, the group consisting of acidic resolving agents such as tartaric acid, malic acid, lactic acid, mandelic acid, 10-camphorsulfonic acid and derivatives thereof; and basic separating agents such as brucine, strychnine, quinine and the like alkaloid compounds, amino acid derivatives, cinchonidine and α-methylbenzylamine. In addition, one of the enantiomers of the compound of the present invention can be obtained not only by obtaining the compound of the present invention as a mixture of each of the enantiomers and then performing the above chiral separation methods, but also by obtaining, by chiral separation using the above methods, etc. .d., and the use of one enantiomer of the compound according to the present invention as a synthetic raw material. In addition, methods for producing one of the enantiomers of the compound of the present invention or the compound as a raw material thereof include a method for preferentially producing one of the enantiomers by adjusting the reaction conditions for the catalyst or the like in the asymmetric carbon atom production reaction step.

The compound of the present invention, or a salt thereof, may be in the form of crystals. Individual crystals and polymorphic mixtures are included within the scope of the compound of the present invention or a salt thereof. Such crystals can be obtained by crystallization in accordance with the method of crystallization itself known in this field. A compound of the present invention, or a salt thereof, may be in solvate (eg, hydrate) or unsolvated form. Any such forms are included within the scope of the compound of the present invention, or a salt thereof. Isotopically labeled compounds (eg, 3H, 14C, 35S, and 125I) are also included within the scope of a compound of the present invention or a salt thereof.

A salt of a compound of the present invention or an intermediate thereof refers to a common salt used in the field of organic chemistry. Examples of such salts include base addition salts at the carboxyl group when the compound has a carboxyl group, and acid addition salts at the amino group or a basic heterocyclic group when the compound has an amino group or a basic heterocyclic group.

Examples of base addition salts include alkali metal salts such as sodium salts and potassium salts; alkaline earth metal salts such as calcium salts and magnesium salts; ammonium salts; and organic amine salts such as trimethylamine salts, triethylamine salts, dicyclohexylamine salts, ethanolamine salts, diethanolamine salts, triethanolamine salts, procaine salts and N,N'-dibenzylethylenediamine salts.

Examples of acid addition salts include inorganic acid salts such as hydrochlorides, sulfates, nitrates, phosphates and perchlorates; organic acid salts such as acetates, formates, maleates, fumarates, tartrates, citrates, ascorbates and trifluoroacetates; and sulfonates such as methanesulfonates, isethionates, benzenesulfonates and p-toluenesulfonates.

The compound of the present invention, or a salt thereof, has excellent EGFR inhibitory activity and is useful as an antitumor agent. Further, the compound of the present invention, or a salt thereof, has excellent selectivity for EGFR and advantageously fewer side effects caused by other kinases. The type of cancer to be treated with the compound of the present invention or a salt thereof is not particularly limited. Examples of malignancies include epithelial cancers (eg, cancers of the respiratory system, cancers of the digestive system, cancers of the reproductive system, cancers of the secretion system, and the like), sarcomas, hematopoietic tumors, tumors of the central nervous system, and tumors of the peripheral nervous system. . Preferred examples include epithelial cancers. More preferred examples include cancers of the respiratory system. Further, the organ from which the tumor develops is not particularly limited. Examples include head and neck cancers, esophageal cancer, gastric cancer, colon cancer, rectal cancer, liver cancer, gallbladder cancer, cholangiocarcinoma, bile duct cancer, pancreatic cancer, lung cancer, breast cancer, ovarian cancer, cervical cancer, intrauterine cancer, kidney cancer, bladder cancer, prostate cancer, testicular tumor, osteosarcoma, soft tissue sarcoma, blood cancer, multiple myeloma, skin cancer, brain tumor and mesothelioma. Preferably, the target cancer is head and neck cancer, gastric cancer, colon cancer, rectal cancer, liver cancer, pancreatic cancer, lung cancer, breast cancer, ovarian cancer, kidney cancer, or prostate cancer, in especially, lung cancer is preferred.

Further, the compound of the present invention, or a salt thereof, has excellent inhibitory activity against mutated EGFR. Examples of such mutated EGFR include drug-tolerant mutated EGFR and hypersensitive mutated EGFR. Therefore, the compound of the present invention, or a salt thereof, is useful as an antitumor agent for the treatment of the above-mentioned EGFR-mutated cancers.

When the compound of the present invention, or a salt thereof, is used as a pharmaceutical preparation, a pharmaceutical carrier may be added if necessary, thereby obtaining a suitable dosage form according to prevention and treatment purposes. Exemplary dosage forms include oral preparations, injections, suppositories, ointments, patches, and the like. Of these, oral preparations are preferred. Such dosage forms may be prepared by methods generally known to those skilled in the art.

As a pharmaceutical carrier, various standard organic or inorganic carriers used as substances for the preparation of a pharmaceutical preparation can be used, as an excipient, binder, disintegrant, lubricant or coloring agent, in the case of solid preparations; or as a solvent, solubilizing agent, suspending agent, isotonizing agent, buffer or sedative, in the case of liquid preparations. In addition, pharmaceutical additives such as antiseptics, antioxidants, colorants, sweeteners, and stabilizers can also be used, if necessary.

Oral solid preparations are prepared as follows.

After the excipient, a binder, a disintegrating agent, a lubricant, a coloring agent, a taste-masking agent or a flavoring agent, etc. are optionally added to the compound of the present invention, the resulting mixture is adjusted to the finished dosage form in the form of tablets, coated tablets, granules , powders, capsules or the like, by standard methods.

Examples of excipients include lactose, sucrose, D-mannitol, glucose, starch, calcium carbonate, kaolin, microcrystalline cellulose and silicic anhydride. Examples of binders include water, ethanol, 1-propanol, 2-propanol, sugar syrup, glucose solution, liquid α-starch, liquid gelatin, D-mannitol, carboxymethylcellulose, hydroxypropylcellulose, hydroxypropylstarch, methylcellulose, ethylcellulose, shellac, calcium phosphate, polyvinylpyrrolidone and so on. Examples of disintegrating agents include dry starch, sodium alginate, agar powder, sodium hydrogen carbonate, calcium carbonate, sodium lauryl sulfate, stearic acid monoglyceride, lactose, and the like. Examples of lubricants include purified talc, sodium stearic acid, magnesium stearate, borax, polyethylene glycol, and the like. Examples of dyes include titanium oxide, iron oxide, and the like. Examples of taste masking agents or flavors include sucrose, bitter orange peel, citric acid, tartaric acid, and the like.

When a liquid preparation for oral administration is prepared, a taste masking agent, a buffer, a stabilizer, a flavoring agent, and the like can be added to the compound of the present invention; and the resulting mixture can be adjusted to the finished dosage form in the form of an oral liquid preparation, syrup, elixir, etc., in accordance with the standard method.

In this case, the same taste-masking agent or flavoring agent as mentioned above can be used. An example of a buffer is sodium citrate and examples of stabilizers include tragacanth, gum arabic and gelatin. If desired, these oral formulations may be coated according to methods known in the art as an enteric coating or other coating for the purpose of, for example, persistence of effects. Examples of such coating agents include hydroxypropyl methylcellulose, ethylcellulose, hydroxymethylcellulose, hydroxypropylcellulose, polyoxyethylene glycol, and Tween 80 (registered trademark).

When an injectable agent is prepared, a pH adjuster, a buffer, a stabilizer, an isotonic agent, a local anesthetic, and the like can be added to the compound of the present invention; and the mixture can be brought to the finished dosage form as a subcutaneous, intramuscular or intravenous injection, in accordance with the standard method.

Examples of a pH adjuster and buffer used in this context include sodium citrate, sodium acetate and sodium phosphate. Examples of the stabilizer include sodium pyrosulfite, EDTA, thioglycolic acid and thiolactic acid. Examples of a local anesthetic include procaine hydrochloride and lidocaine hydrochloride. Examples of the tonic agent include sodium chloride, dextrose, D-mannitol and glycerin.

When a suppository is prepared, pharmaceutically acceptable carriers known in the art such as polyethylene glycol, lanolin, cocoa butter and fatty acid triglyceride can be added to the compound of the present invention; and, if necessary, surfactants such as Tween 80 (registered trademark), and the resulting mixture can be adjusted to the finished dosage form in the form of a suppository, in accordance with the standard method.

When an ointment is prepared, a commonly used base, stabilizer, wetting agent, preservative, and the like. can be mixed with the compound according to the present invention, when necessary; and the resulting mixture can be mixed and adjusted to the finished dosage form in the form of an ointment, in accordance with the standard method.

Examples of the base include liquid paraffin, white petrolatum, white wax, octyldodecyl alcohol, and paraffin.

Examples of preservatives include methyl paraoxybenzoate, ethyl paraoxybenzoate, and propyl paraoxybenzoate.

When a patch is prepared, the above ointment, cream, gel, paste or the like can be added to the standard substrate according to the standard method.

Suitable substrates are woven or non-woven materials, including cotton, staple fibers or man-made fibers; and films or sheet foams of plastic PVC, polyethylene, polyurethane, etc.

The amount of a compound of the present invention included in each of such unit dosage forms depends on the condition of the patient to whom the compound is administered, the dosage form of the compound, and so forth. In general, in the case of an oral agent, the amount of the compound is from 0.05 mg to 1000 mg per unit dosage form. In the case of injection, the amount of the compound is from 0.01 mg to 500 mg per unit dosage form; and, in the case of a suppository, the amount of the compound is from 1 mg to 1000 mg per unit dosage form.

The daily dose of the drug in such a dosage form depends on the condition, body weight, age, anatomical sex, etc. patient and cannot be generalized. For example, the daily dose for an adult patient (body weight: 50 kg) in most cases may be 0.05-5000 mg and preferably 0.1-1000 mg; and preferably this dose is administered as a single dose or as two to three divided doses per day.

Examples of mammals administered with the compound of the present invention include humans, monkeys, mice, rats, rabbits, dogs, cats, cows, horses, pigs and sheep.

Examples

The present invention is more specifically explained below with reference to examples; however, the present invention is not limited to these examples.

In the examples, unless otherwise indicated, commercially available reagents were used. Silica gel column chromatography used Purif-Pack® SI available from Moritex Corp. (manufactured by Shoko Scientific Co., Ltd.); pre-filled KP-Sil® Silica column available from Biotage; a pre-filled HP-Si® Silica column available from Biotage; or a pre-filled HP-Sphere® Silica column available from Biotage. For column chromatography on basic silica gel, Purif-Pack® NH, available from Moritex Corp., was used. (manufactured by Shoko Scientific Co., Ltd.); or a pre-filled KP-NH® column available from Biotage. For preparative thin layer chromatography, Kieselgel TM 60F254, Art. 5744 from Merk or NH2 Silica Gel 60F254 from Wako. The NMR spectrum was taken using an AL400 (400 MHz; manufactured by JEOL) or Mercury 400 (400 MHz; manufactured by Agilent Technologies, Inc.) spectrometer. When the deuterated solvent contains tetramethylsilane, tetramethylsilane is used as an internal standard. Alternatively, the NMR solvent was used as an internal standard. All δ values were presented in ppm. The reaction under the influence of microwave radiation was carried out using an initiator manufactured by Biotage.

Next, an LCMS spectrum was obtained using an Acquity SQD (quadrupole) instrument manufactured by Waters Corporation under the following conditions.

Column: Acquity UPLC® BEH C18, 2.1×50 mm, 1.7 µm, available from Waters Corporation;

MS-detection: ESI with the formation of positive ions;

UV detection: 254 and 210 nm;

Volumetric flow rate through the column: 0.5 ml/min;

Mobile phase: water/acetonitrile (0.1% formic acid);

Injected sample volume: 1 µl;

(Table)
Gradient Time (min) Water Acetonitrile 0 95 5 0.1 95 5 2.1 5 95 3.0 termination

Purification by reverse phase preparative HPLC was carried out using a preparative separation system available from Gilson.

Column: CombiPrep Pro C18, 50×30 mm (id), S-5 µm (manufactured by YMC);

UV detection: 254 nm;

Volumetric flow rate through the column: 40 ml/min;

Mobile phase: water/acetonitrile (0.1% trifluoroacetic acid);

The volume of the injected sample: from 0.1 to 1 ml.

Each symbol represents the following.

s (s): singlet

d(d): doublet

t (t): triplet

q (square): quartet

dd (dd): doublet of doublets

dt (dt): doublet of triplets

ddd (ddd): doublet of doublets of doublets

m (m): multiplet

brs (br. s): broadened singlet

DMSO-d ₆ (DMSO-d ₆ ): deuterated dimethyl sulfoxide

CDCl ₃ : deuterated chloroform

CD ₃ OD: deuterated methanol

THF (THF): tetrahydrofuran

DMF (DMF): N,N-dimethylformamide

DME: 1,2-dimethoxyethane

DMSO (DMSO): dimethyl sulfoxide

HATU: O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate

Example 1

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-1-yl)acrylamide (Compound 1)

Step 1: Synthesis of (S)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but-3-en-2-yl)carbamate

Diisopropyl azodicarboxylate (2.44 ml) was slowly added to a solution of triphenylphosphine (13.1 g) in THF (70 ml), under ice-cooling. The reaction mixture was stirred, under ice-cooling, for 1 hour, and then a solution of (S)-tert-butyl(1-hydroxybut-3-en-2-yl)carbamate (7.0 g) synthesized according to with the method described in NPD Org. Lett., 2005, vol. 7, no. 5, pp. 847-849, and 4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (6.97 g), in THF (35 ml) . After the reaction mixture was stirred at room temperature for 2 hours, the solvent was distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluting solvent: hexane/ethyl acetate), thereby obtaining the title compound (20.84 g) as a light yellow oily substance.

ESI-MS m/z: 448, 450 [M+H] ⁺ .

Step 2: Synthesis of (S)-tert-butyl(1-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but-3-en-2-yl)carbamate

An 8 N solution of ammonia in methanol (89.4 ml) was added to (S)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but -3-en-2-yl)carbamate (20.84 g) obtained in Step 1, and the mixture was stirred in an autoclave at 120° C. for 6 hours. The reaction mixture is cooled with ice, and the solvent is distilled off under reduced pressure. Then the resulting residue was diluted with a small amount of methanol, the precipitated precipitate was collected by filtration, washed with cold methanol (11 ml), and then dried under reduced pressure, thus obtaining the title compound (8.28 g) as a milky white solid .

ESI-MS m/z: 430 [M+H] ⁺ .

Step 3: Synthesis of (S)-tert-butyl(1-(4-amino-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but-3-ene -2-yl) carbamate

A mixture of (S)-tert-butyl (1- (4-amino-5-iodo-7H-pyrrolo[2,3-d] pyrimidin-7-yl) but-3-en-2-yl) carbamate (8, 26 g) obtained in Step 2, 3-quinolineboronic acid (4.99 g), cesium carbonate (12.54 g), 1,1'-bis(diphenylphosphino)ferrocene-palladium(II)dichloride(II) (785 .6 mg), DME (66 ml) and water (33 ml) were stirred under nitrogen at 100° C. for 2 hours. After the reaction mixture was cooled, water and ethyl acetate were added thereto to separate the organic layer. The aqueous layer was then extracted twice with ethyl acetate. The resulting organic layer was dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: hexane/ethyl acetate, ethyl acetate/methanol), thereby obtaining the title compound (8.0 g) as a light orange solid.

ESI-MS m/z: 431 [M+H] ⁺ .

Step 4: Synthesis of (S)-tert-butyl(1-(4-amino-6-bromo-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but -3-en-2-yl) carbamate

N-Bromosuccinimide (3.63 g) was added to a solution of (S)-tert-butyl(1-(4-amino-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7 -yl)but-3-en-2-yl)carbamate (7.98 g) obtained in step 3 in DMF (64 ml) at -15°C, and the mixture is stirred at -15°C, within 1 hour. A 10% aqueous sodium thiosulfate solution and ethyl acetate were added to the reaction mixture, and stirred at room temperature for 10 minutes. The organic layer is separated and the aqueous layer is extracted twice with ethyl acetate. The resulting organic layer was washed twice with saturated sodium chloride solution and dried over anhydrous magnesium sulfate. The solvent is distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (6.30 g) as a light brown solid.

ESI-MS m/z: 509, 511 [M+H] ⁺ .

Step 5: Synthesis of (S)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7- yl) carbamate

4 N Aqueous sodium hydroxide solution (28.8 ml) was added to a solution of (S)-tert-butyl(1-(4-amino-6-bromo-5-(quinolin-3-yl)-7H-pyrrolo[2, 3-d]pyrimidin-7-yl)but-3-en-2-yl)carbamate (5.00 g) obtained in step 4 in THF (100 ml) and the mixture was degassed under reduced pressure, then purged with nitrogen . Tetrakis(triphenylphosphine)palladium (1.13 g) was then added, and the mixture was stirred overnight at reflux. Then, the reaction mixture was cooled to room temperature, extraction was carried out using ethyl acetate, and the resulting organic layer was washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration and concentration, the resulting residue was purified by silica gel chromatography (elution solvent: ethyl acetate/methanol), thus obtaining the title compound as an orange solid (5.01 g).

ESI-MS m/z: 429 [M+H] ⁺ .

Step 6: Synthesis of Compound 1

A 4 N solution of hydrogen chloride in dioxane (1 ml) and methanol (1 ml) were added to a solution of (S)-tert-butyl (4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro -6H-pyrimido[5,4-b]pyrrolisin-7-yl)carbamate (100 mg) obtained in Step 5 in chloroform (1 ml) and the mixture was stirred for 5 hours at room temperature. The reaction mixture was concentrated under reduced pressure, thus obtaining (S)-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-4,7- diamine hydrochloride.

Diisopropylethylamine (0.187 ml) was added to a solution of (S)-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisine-4,7-diamine hydrochloride in chloroform (4 ml) and, under ice-cooling, add 100 mg/ml solution of acryloyl chloride in chloroform (0.19 ml). The mixture is stirred for 40 minutes. The reaction mixture is purified by silica gel chromatography (elution solvent: ethyl acetate/methanol); after concentration, the resulting residue is suspended and washed with ethyl acetate/hexane. The resulting solid was collected by filtration and dried under reduced pressure, thus obtaining the title compound (49 mg) as a light green solid.

¹ H-NMR (DMSO-d ₆ ) δ: 3.88-3.93 (1H, m), 4.57-4.63 (1H, m), 5.03 (1H, d, J=2, 4 Hz), 5.24 (1H, d, J=2.4 Hz), 5.55-5.62 (1H, m), 5.68 (1H, dd, J=10.0, 2.4 Hz), 6.12-6.38 (4H, m), 7.65 (1H, dd, J=7.8, 7.8 Hz), 7.77-7.83 (1H, m), 8 .04-8.11 (2H, m), 8.15 (1H, s), 8.41 (1H, d, J=2.2 Hz), 8.82 (1H, d, J=7.8 Hz), 8.98 (1H, d, J=2.2 Hz).

ESI-MS m/z: 383 [M+H] ⁺ .

Example 2

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)methacrylamide (Compound 2)

Step 1: Synthesis of (S)-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-4,7-diamine

5 N Hydrochloric acid (12 ml) was added to a solution of (S)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5.4 -b]pyrrolisin-7-yl)carbamate (5.01 g) obtained in step 5 of Example 1 in ethanol (25 ml) and the resulting mixture was stirred for 1 hour, at an external temperature of 70°C to 80° WITH. Then, the reaction mixture was cooled to room temperature, washed with chloroform, and the pH of the aqueous layer was adjusted to about 10 using 5 N sodium hydroxide aqueous solution, followed by extraction with chloroform. The resulting organic layer was dried over anhydrous sodium sulfate, then filtered and concentrated. The resulting residue was purified by basic silica gel chromatography (elution solvent: chloroform/methanol), thereby obtaining the title compound (2.10 g) as a yellow solid.

ESI-MS m/z: 329 [M+H] ⁺ .

Step 2: Synthesis of Compound 2

Diisopropylethylamine (0.0318 ml) and methacryloyl chloride (0.0148 ml) were added successively to a solution of (S)-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5.4 -b]pyrrolysine-4,7-diamine (50.0 mg) obtained in step 1 in acetonitrile (2.0 ml) and water (2.0 ml) at 0°C, and the resulting mixture was stirred in for 45 minutes at the same temperature. The reaction mixture was poured into a saturated aqueous sodium bicarbonate solution, then extracted with chloroform. After the organic layer was dried over anhydrous sodium sulfate, the solvent was distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (38.0 mg) as a light yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 2.02 (3H, s), 4.03 (1H, dd, J=11.6, 4.9 Hz), 4.70 (1H, dd, J=11 .6, 8.2 Hz), 5.20-5.22 (3H, m), 5.44 (2H, d, J=1.7 Hz), 5.73-5.75 (1H, m) , 5.82 (1H, s), 6.89 (1H, d, J=7.6 Hz), 7.61-7.65 (1H, m), 7.76-7.80 (1H, m ), 7.87 (1H, d, J=8.3 Hz), 8.15 (1H, d, J=8.5 Hz), 8.21-8.25 (2H, m), 9.06 (1H, d, J=2.2 Hz).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 3

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)but-2 -enamide (mixture of E and Z isomers) (Compound 3)

The same synthesis as in Step 2 of Example 2 was carried out using crotonoyl chloride instead of the methacryloyl chloride used in Step 2 of Example 2, thus obtaining the title mixture (8.8 mg) as a light yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 1.83-1.90 (3H, m), 4.01 (1H, dd, J=11.6, 5.2 Hz), 4.73 (1H, dd , J=11.6, 8.2 Hz), 5.00 (2H, s), 5.19 (1H, d, J=1.5 Hz), 5.44 (1H, d, J=2, 0 Hz), 5.75-5.77 (1H, m), 5.87-5.91 (1H, m), 6.27 (1H, d, J=8.3 Hz), 6.92- 7.01 (1H, m), 7.62-7.66 (1H, m), 7.78-7.82 (1H, m), 7.88 (1H, d, J=7.3 Hz) , 8.18 (1H, d, J=8.0 Hz), 8.28-8.29 (2H, m), 9.08 (1H, d, J=2.2 Hz).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 4

(S,E)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(dimethylamino)but-2-enamide (Compound 4)

(E)-4-(dimethylamino)but-2-enoic acid hydrochloride (60.6 mg), HATU (139 mg), diisopropylethylamine (0.106 ml) and DMF (1.0 ml) are added successively to the suspension (S)- 6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisine-4,7-diamine (100 mg) obtained in step 1 of Example 2, in methylene chloride (3.0 ml), at room temperature, and the mixture is stirred for 1.5 hours at the same temperature. The reaction mixture was poured into a saturated aqueous sodium bicarbonate solution, then extracted with ethyl acetate. After the organic layer was dried over anhydrous sodium sulfate, the solvent was distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (114 mg) as a light yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 2.23 (6H, s), 3.07 (2H, dd, J=6.0, 1.3 Hz), 4.01 (1H, dd, J=11 .6, 5.0 Hz), 4.70 (1H, dd, J=11.6, 8.2 Hz), 5.09 (2H, br.s), 5.20 (1H, d, J= 2.0 Hz), 5.43 (1H, d, J=2.0 Hz), 5.73-5.79 (1H, m), 6.07 (1H, dt, J=15.4, 1 .3 Hz), 6.72 (1H, br.s), 6.94 (1H, dt, J=15.4, 6.0 Hz), 7.61-7.65 (1H, m), 7 .77-7.81 (1H, m), 7.87 (1H, d, J=8.0 Hz), 8.16 (1H, d, J=8.5 Hz), 8.25-8, 26 (2H, m), 9.05 (1H, d, J=2.0 Hz).

ESI-MS m/z: 440 [M+H] ⁺ .

Example 5

(S,E)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 3-chloroacrylamide (Compound 5)

The same synthesis as in Example 4 was carried out using trans-3-chloroacrylic acid instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride used in Example 4, thus obtaining the title compound (49 .2 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 4.04 (1H, dd, J=11.7, 4.6 Hz), 4.69 (1H, dd, J=11.7, 8.0 Hz), 5.03 (2H, s), 5.22 (1H, d, J=1.7 Hz), 5.44 (1H, d, J=1.7 Hz), 5.72-5.75 (1H , m), 6.34 (1Н, d, J=13.0 Hz), 6.89 (1Н, br.s), 7.41 (1Н, d, J=13.0 Hz), 7.63 -7.65 (1H, m), 7.79-7.81 (1H, m), 7.87 (1H, d, J=8.0 Hz), 8.16 (1H, d, J=8 .3 Hz), 8.25-8.26 (2H, m), 9.02 (1H, d, J=2.0 Hz).

ESI-MS m/z: 417, 419 [M+H] ⁺ .

Example 6

(S,Z)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 3-chloroacrylamide (Compound 6)

The same synthesis as in Example 4 was carried out using cis-3-chloroacrylic acid instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride used in Example 4, thus obtaining the title compound (74 .7 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 4.07 (1H, dd, J=11.6, 5.4 Hz), 4.82 (1H, dd, J=11.6, 8.2 Hz), 4.96 (2H, s), 5.26 (1H, d, J=2.0 Hz), 5.48 (1H, d, J=2.0 Hz), 5.75-5.78 (1H , m), 6.28 (1H, d, J=8.3 Hz), 6.61 (1H, d, J=8.3 Hz), 6.81 (1H, d, J=7.3 Hz ), 7.64-7.67 (1H, m), 7.79-7.83 (1H, m), 7.90 (1H, d, J=7.1 Hz), 8.19 (1H, d, J=8.5 Hz), 8.31 (1H, d, J=2.0 Hz), 8.33 (1H, s), 9.12 (1H, d, J=2.2 Hz) .

ESI-MS m/z: 417, 419 [M+H] ⁺ .

Example 7

(S,E)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(piperidin-1-yl)but-2-enamide (Compound 7)

The same synthesis as in Example 4 is carried out using (E)-4-(piperidin-1-yl)but-2-enoic acid hydrochloride instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride, used in Example 4, thus obtaining the title compound (122 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 1.42-1.59 (5H, m), 2.14 (1H, s), 2.39 (4H, br.s), 3.10 (2H, dd , J=6.0, 1.4 Hz), 4.01 (1H, dd, J=11.6, 5.0 Hz), 4.69 (1H, dd, J=11.5, 8.0 Hz), 5.10 (2H, br.s), 5.20 (1H, d, J=2.1 Hz), 5.43 (1H, d, J=2.1 Hz), 5.73- 5.79 (1H, m), 6.06 (1H, dt, J=15.3, 1.4 Hz), 6.76 (1H, br.s), 6.96 (1H, dt, J= 15.3, 6.0 Hz), 7.62-7.64 (1H, m), 7.77-7.80 (1H, m), 7.87 (1H, d, J=8.0 Hz ), 8.16 (1H, d, J=8.3 Hz), 8.24-8.25 (2H, m), 9.04 (1H, d, J=2.0 Hz).

ESI-MS m/z: 480 [M+H] ⁺ .

Example 8

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)propiolamide (Compound 8)

The same synthesis as in Example 4 was carried out using propiolic acid instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride used in Example 4, thus obtaining the title compound (30.0 mg) , as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 2.91 (1H, s), 4.08 (1H, dd, J=11.7, 4.9 Hz), 4.76 (1H, dd, J=11 .7, 8.0 Hz), 4.91 (2H, s), 5.24 (1H, d, J=1.2 Hz), 5.49 (1H, d, J=1.7 Hz), 5.68-5.69 (1H, m), 6.34-6.37 (1H, m), 7.65-7.67 (1H, m), 7.81-7.83 (1H, m ), 7.90 (1H, d, J=8.3 Hz), 8.20 (1H, d, J=8.5 Hz), 8.30 (1H, d, J=2.0 Hz), 8.34 (1H, s), 9.11 (1H, d, J=2.2 Hz).

ESI-MS m/z: 381 [M+H] ⁺ .

Example 9

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)but-2 -inamide (Compound 9)

The same synthesis as in Example 4 was carried out using but-2-ic acid instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride used in Example 4, thus obtaining the title compound (98 0 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 1.96 (3H, s), 4.02 (1H, dd, J=11.6, 5.0 Hz), 4.70 (1H, dd, J=11 .6, 8.3 Hz), 5.05 (2H, s), 5.24 (1H, d, J=1.6 Hz), 5.45 (1H, d, J=1.6 Hz), 5.70-5.72 (1H, m), 6.95-7.01 (1H, br.s), 7.63-7.67 (1H, m), 7.78-7.83 (1H , m), 7.88 (1Н, d, J=8.3 Hz), 8.21 (1Н, d, J=8.5 Hz), 8.26-8.28 (2Н, m), 9 .02 (1H, d, J=2.0 Hz).

ESI-MS m/z: 395 [M+H] ⁺ .

Example 10

(S,E)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(diethylamino)but-2-enamide (Compound 10)

The same synthesis as in Example 4 was carried out using (E)-4-(diethylamino)but-2-enoic acid hydrochloride instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride used in Example 4 , thus obtaining the title compound (38.2 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 1.00 (6H, t, J=7.2 Hz), 2.51 (4H, q, J=7.2 Hz), 3.22 (2H, dd , J=5.9, 1.5 Hz), 4.00 (1H, dd, J=11.5, 4.9 Hz), 4.62 (1H, dd, J=11.5, 8.2 Hz), 5.22-5.24 (3H, m), 5.40 (1H, d, J=2.0 Hz), 5.74-5.80 (1H, m), 6.12 (1H , dt, J=15.4, 1.5 Hz), 6.98 (1Н, dt, J=15.4, 5.9 Hz), 7.50 (1Н, br.s), 7.58- 7.62 (1H, m), 7.74-7.78 (1H, m), 7.84 (1H, d, J=7.6 Hz), 8.12-8.14 (2H, m) , 8.22 (1H, d, J=1.7 Hz), 8.99 (1H, d, J=2.0 Hz).

ESI-MS m/z: 468 [M+H] ⁺ .

Example 11

(S,E)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(ethyl(methyl)amino)but-2-enamide (Compound 11)

The same synthesis as in Example 4 is carried out using (E)-4-(ethyl(methyl)amino)but-2-enoic acid hydrochloride instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride, used in Example 4, thus obtaining the title compound (13.5 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 1.06 (3H, t, J=7.2 Hz), 2.24 (3H, s), 2.44 (2H, q, J=7.2 Hz ), 3.15 (2Н, dd, J=6.0, 1.6 Hz), 4.03 (1Н, dd, J=11.6, 5.1 Hz), 4.73 (1Н, dd, J=11.6, 8.2 Hz), 5.01(2H, s), 5.20(1H, d, J=1.7 Hz), 5.44(1H, d, J=1.7 Hz), 5.75-5.77 (1H, m), 6.05 (1H, dt, J=15.4, 1.6 Hz), 6.42 (1H, d, J=6.8 Hz ), 6.96 (1H, dt, J=15.4, 6.0 Hz), 7.62-7.66 (1H, m), 7.78-7.82 (1H, m), 7, 88 (1H, d, J=8.0 Hz), 8.18 (1H, d, J=8.5 Hz), 8.29 (2H, s), 9.08 (1H, d, J=2 .2 Hz).

ESI-MS m/z: 454 [M+H] ⁺ .

Example 12

(S,E)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)- 4-(isopropyl(methyl)amino)but-2-enamide (Compound 12)

The same synthesis as in Example 4 is carried out using (E)-4-(isopropyl(methyl)amino)but-2-enoic acid hydrochloride instead of (E)-4-(dimethylamino)but-2-enoic acid hydrochloride, used in Example 4, thus obtaining the title compound (25.8 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 1.00 (6H, d, J=6.6 Hz), 2.15-2.36 (3H, m), 2.79-2.88 (1H, m ), 3.18 (2Н, dd, J=5.9, 1.5 Hz), 4.01 (1Н, dd, J=11.5, 5.0 Hz), 4.70 (1Н, dd, J=11.5, 8.0 Hz), 5.11(2H, s), 5.21(1H, d, J=1.8 Hz), 5.43(1H, d, J=1.8 Hz), 5.73-5.79 (1H, m), 6.10 (1H, dt, J=15.4, 1.5 Hz), 6.74 (1H, br.s), 6.94 (1Н, dt, J=15.4, 5.9 Hz), 7.61-7.65 (1Н, m), 7.77-7.81 (1Н, m), 7.87 (1Н, d , J=8.5 Hz), 8.16 (1H, d, J=8.8 Hz), 8.23-8.26 (2H, m), 9.05 (1H, d, J=2, 2 Hz).

ESI-MS m/z: 468 [M+H] ⁺ .

Example 13

(R)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-7-yl)acrylamide (Compound 13)

Step 1: Synthesis of (R)-tert-butyl(1-hydroxy-5-(methylthio)pentan-3-yl)carbamate

N-methylmorpholine (3.63 ml) and ethyl chloroformate (3.01 ml) are added to a solution of (R)-3-((tert-butoxycarbonyl)amino)-5-(methylthio)pentanoic acid (7.92 g), in THF (79.2 ml), at -10°C. After stirring at -10° C. for 15 minutes, the resulting insoluble matter is filtered off. An aqueous solution of sodium borohydride (1.55 g) (15 ml) was added to the filtrate at -10°C, and the mixture was stirred at -10°C for 1 hour. A saturated ammonium chloride aqueous solution was added to the reaction mixture, and the mixture was stirred at room temperature for 30 minutes. Ethyl acetate was added to the reaction mixture to separate the organic layer. The organic layer was washed with a 0.5 N aqueous potassium hydrogen sulfate solution, water, a 0.5 N aqueous sodium hydroxide solution, and a saturated sodium chloride solution, and dried over anhydrous sodium sulfate. The solvent is then distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: chloroform/ethyl acetate), thereby obtaining the title compound as a light yellow oily substance (7.18 g).

Stage 2: Synthesis of tert-butyl ((3R)-1-hydroxy-5-(methylsulfinyl)pentan-3-yl)carbamate

A suspension of sodium periodate (7.0 g), in water (32 ml), was added to a solution of (R)-tert-butyl (1-hydroxy-5-(methylthio)pentan-3-yl)carbamate (8.16 g) obtained in step 1, in methanol (98 ml), at a temperature of 10°C or below, and the mixture is stirred at room temperature for 2 hours.

The resulting insoluble matter was filtered off, and the filtrate was subjected to distillation under reduced pressure. The residue obtained is taken up in saturated sodium chloride solution and then extracted 3 times with chloroform. The organic layer was dried over anhydrous sodium sulfate, and the solvent was distilled off under reduced pressure, thereby obtaining the title compound (9.38 g) as a light yellow solid.

Step 3: Synthesis of (R)-tert-butyl(5-hydroxypent-1-en-3-yl)carbamate

Sodium acetate (13.45 g) is added to a solution of tert-butyl ((3R)-1-hydroxy-5-(methylsulfinyl)pentan-3-yl)carbamate (9.38 g) obtained in step 2, in 1, 2-dichlorobenzene (140 ml), at room temperature. The mixture is stirred at an internal temperature of 166°C for 18 hours. After the reaction mixture was cooled, the insoluble matter was filtered off, and 1,2-dichlorobenzene was distilled off under reduced pressure. The resulting residue was dissolved in ethyl acetate, washed with aqueous sodium hypochlorite solution, water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. The solvent is then distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluting solvent: hexane/ethyl acetate), thereby obtaining the title compound (2.50 g) as a light yellow oily substance.

Step 4: Synthesis of (R)-tert-butyl(5-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-1-en-3-yl)carbamate

Triphenylphosphine (3.25 g) is added and dissolved in a solution of (R)-tert-butyl (5-hydroxypent-1-en-3-yl) carbamate (2.5 g) obtained in step 3 and 4-chloro- 5-iodo-7H-pyrrolo[2,3-d]pyrimidine (2.31 g), in DME (23 ml), under ice-cooling. Thereafter, diisopropyl azodicarboxylate (2.44 ml) was gradually added. The reaction mixture was stirred under ice-cooling for 30 minutes and at room temperature for 1 hour, and the solvent was then distilled off under reduced pressure. The resulting residue was dissolved in ethyl acetate, washed with water and dried over anhydrous sodium sulfate. The solvent is then distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: hexane/ethyl acetate), thereby obtaining the title compound (3.49 g) as a light yellow solid.

ESI-MS m/z: 463, 465 [M+H] ⁺ .

Step 5: Synthesis of (R)-tert-butyl(5-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-1-en-3-yl)carbamate

28% aqueous ammonia solution (17.5 ml) was added to a solution of (R)-tert-butyl(5-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl )pent-1-en-3-yl)carbamate (3.49 g) obtained in step 4 in DME (17.5 ml) and the mixture was stirred in an autoclave at an internal temperature of 105° C. for 8 hours. After the reaction mixture was cooled, water (70 ml) was added and the mixture was stirred at room temperature for 4 hours. The resulting precipitate was collected by filtration, washed with water and dried, thus obtaining the title compound (3.20 g) as a light yellow solid.

ESI-MS m/z: 444 [M+H] ⁺ .

Step 6: Synthesis of (R)-tert-butyl(5-(4-amino-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-1-ene -3-yl) carbamate

A mixture of (R)-tert-butyl (5-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-1-en-3-yl)carbamate (3, 2 g) obtained in step 5, 3-quinolineboronic acid (1.37 g), sodium carbonate (843 mg), tetrakis(triphenylphosphine)palladium (250 mg), DME (32 ml) and water (32 ml) are stirred in nitrogen atmosphere, at a temperature of 100°C, for 6 hours. After cooling the reaction mixture, saturated aqueous sodium bicarbonate and ethyl acetate were added. The resulting mixture was stirred at room temperature for 30 minutes. After filtering off the insoluble matter, the organic layer was separated and dried over anhydrous sodium sulfate. The solvent is then distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: hexane/ethyl acetate, ethyl acetate/methanol), thereby obtaining the title compound (3.21 g) as a light orange solid.

ESI-MS m/z: 445 [M+H] ⁺ .

Step 7: Synthesis of (R)-tert-butyl(5-(4-amino-6-bromo-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent -1-en-3-yl) carbamate

A solution of N-bromosuccinimide (1.35 g), in THF (23 ml), was added to a solution of (R)-tert-butyl(5-(4-amino-5-(quinolin-3-yl)-7H-pyrrolo[ 2,3-d]pyrimidin-7-yl)pent-1-en-3-yl)carbamate (3.21 g) obtained in step 6 in THF (26 ml) with ice-cooling for 30 min . The mixture was stirred under ice-cooling for 30 minutes. After a 5% sodium thiosulfate aqueous solution was added to the reaction mixture, the mixture was poured into a saturated sodium bicarbonate aqueous solution, followed by extraction with ethyl acetate. The organic layer was washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. The solvent is then distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: hexane/ethyl acetate, ethyl acetate/methanol), thereby obtaining the title compound (3.15 g) as a light brown solid.

ESI-MS m/z: 523, 525 [M+H] ⁺ .

Step 8: Synthesis of (R)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-7- yl) carbamate

4 N Aqueous sodium hydroxide solution (0.454 ml) was added to a solution of (R)-tert-butyl(5-(4-amino-6-bromo-5-(quinolin-3-yl)-7H-pyrrolo[2,3- d]pyrimidin-7-yl)pent-1-en-3-yl)carbamate (475 mg) obtained in step 7 in THF (5 ml). Thereafter, the mixture is degassed under reduced pressure, then purged with nitrogen. Then add tetrakis(triphenylphosphine)palladium (41.6 mg), the mixture is stirred over night at the boil under reflux. The reaction mixture is then cooled to room temperature, poured into water and extracted with ethyl acetate. The resulting organic layer was washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration and concentration, the resulting residue was purified by silica gel chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (293 mg) as a yellow solid.

ESI-MS m/z: 443 [M+H] ⁺ .

Step 9: Synthesis of (R)-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-4,7-diamine

5 N Hydrochloric acid (1 ml) was added to a solution of (R)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4 -b]indolizin-7-yl)carbamate (290 mg) obtained in step 8 in ethanol (4 ml). After that, the mixture is stirred for 6 hours at a temperature of 70°C. After the reaction mixture was cooled to room temperature, the pH of the reaction mixture was adjusted to about 10 using 5N aqueous sodium hydroxide, then extracted with chloroform. The resulting organic layer was dried over anhydrous sodium sulfate, then filtered and concentrated. The resulting residue was purified by basic silica gel chromatography (elution solvent: chloroform/methanol), thereby obtaining the title compound (218 mg) as a light brown solid.

ESI-MS m/z: 343 [M+H] ⁺ .

Step 10: Synthesis of Compound 13

Diisopropylethylamine (0.129 ml) and a solution of acryloyl chloride (56.1 mg), in chloroform (0.5 ml), are added to a solution of (R)-6-methylene-5-(quinolin-3-yl)-6.7.8 ,9-tetrahydropyrimido[5,4-b]indolizine-4,7-diamine (215 mg) obtained in step 9 in chloroform (4 ml) under ice-cooling, and the resulting mixture was stirred for 30 minutes. After the reaction mixture was concentrated, the residue was purified by silica gel chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (117 mg) as a yellow solid.

¹ H-NMR (CDCl ₃ ) δ: 2.28-2.41 (2H, m), 4.32-4.50 (2H, m), 4.64 (2H, br.s), 4.97 (1H, s), 5.07 (1H, s), 5.05-5.12 (1H, m), 5.72 (1H, dd, J=10.2, 1.2 Hz), 5, 75-5.85 (1H, m), 6.14 (1H, dd, J=16.8, 10.2 Hz), 6.36 (1H, dd, J=16.8, 1.2 Hz) , 7.61-7.68 (1H, m), 7.77-7.84 (1H, m), 7.87 (1H, d, J=8.4 Hz), 8.18 (1H, d , J=8.4 Hz), 8.27 (1H, d, J=2.0 Hz), 8.33 (1H, s), 8.98 (1H, d, J=2.0 Hz).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 14

(S)-N-(4-Amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 14)

Step 1: Synthesis of (S)-methyl-2-((tert-butoxycarbonyl)amino)pent-4-enoate

5N aqueous sodium hydroxide solution (2.1 ml) and di-tert-butyl dicarbonate (2.128 ml) were added to a suspension of (S)-2-amino-4-pentenoic acid (1.016 g), in methanol (20 ml), at room temperature, and the resulting mixture was stirred for 8 hours at the same temperature. Di-tert-butyl dicarbonate (0.304 ml) was added at room temperature and the mixture was stirred for 1 hour at the same temperature. 4-Hydroxy-1H-benzotriazole (1.796 g) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (3.882 g) were added to the reaction mixture at room temperature, and the mixture was stirred overnight at the same temperature. The reaction mixture was concentrated under reduced pressure and then poured into a saturated sodium bicarbonate aqueous solution, followed by extraction with ethyl acetate. The resulting organic layer was dried over anhydrous sodium sulfate, then filtered and concentrated. The resulting residue was purified by silica gel chromatography (eluting solvent: chloroform/methanol), thereby obtaining the title compound (2.0311 g) as a light yellow oily substance.

Step 2: Synthesis of (S)-tert-butyl(1-hydroxypent-4-en-2-yl)carbamate

Lithium aluminum hydride (668.2 mg) was added to a solution of (S)-methyl-2-((tert-butoxycarbonyl)amino)pent-4-enoate (1.983 g) obtained in Step 1 in THF (50 ml) with cooling ice, and the mixture is stirred for 1.5 hours at the same temperature. Sodium sulfate decahydrate (1.1375 g) and THF (10 ml) were added to the reaction mixture at room temperature, and the mixture was stirred overnight at the same temperature. The insoluble matter is filtered off. After washing with THF and ethyl acetate, the filtrate is concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography (eluting solvent: hexane/ethyl acetate), thereby obtaining the title compound (1.2601 g) as a light yellow oily substance.

Step 3: Synthesis of (S)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-4-en-2-yl)carbamate

The same synthesis as in step 4 of Example 13 was carried out using (S)-tert-butyl(1-hydroxypent-4-en-2-yl)carbamate obtained in step 2 instead of (R)-tert-butyl( 5-hydroxypent-1-en-3-yl)carbamate used in step 4 of Example 13, thus obtaining the title compound (2.7679 g) as a light yellow solid.

ESI-MS m/z: 463, 465 [M+H] ⁺ .

Step 4: Synthesis of (S)-tert-butyl(1-(4-amino-6-bromo-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent -4-en-2-yl) carbamate

The same synthesis as in steps 5-7 of Example 13 was carried out using (S)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl )pent-4-en-2-yl)carbamate obtained in step 3 instead of (R)-tert-butyl(5-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine- 7-yl)pent-1-en-3-yl)carbamate used in step 5 of Example 13, thus obtaining the title compound (2.669 g) as a light brown solid.

ESI-MS m/z: 523, 525 [M+H] ⁺ .

Step 5: Synthesis of (S)-tert-butyl(4-amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)carbamate and (S)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)carbamate

Aqueous 4 N sodium hydroxide solution (5.1 ml) and tetrakis(triphenylphosphine)palladium (235.4 mg) were added to a solution of (S)-tert-butyl(1-(4-amino-6-bromo-5-(quinoline -3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-4-en-2-yl)carbamate (2.669 g) obtained in step 4 in THF (50 ml), and the mixture is degassed under reduced pressure, then purged with nitrogen. The reaction mixture is stirred over night at the boil under reflux. The reaction mixture was then cooled to room temperature, poured into a saturated sodium bicarbonate aqueous solution and extracted with ethyl acetate. The resulting organic layer was dried over anhydrous sodium sulfate. After filtration and concentration, the resulting residue was purified by silica gel chromatography (elution solvent: chloroform/methanol), thereby obtaining a mixture of the title compounds (2.427 g) as a yellow solid.

ESI-MS m/z: 443 [M+H] ⁺ .

Step 6: Synthesis of (S)-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizine-4,8-diamine and (S)-6-methylene- 5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-4,8-diamine

5 N Hydrochloric acid (5.1 ml) was added to a mixture solution (2.427 g) of (S)-tert-butyl(4-amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[ 5,4-b]indolizin-8-yl)carbamate and (S)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[ 5,4-b]indolizin-8-yl)carbamate obtained in step 5 in ethanol (20 ml) and the mixture was stirred overnight at 60°C. Then add an additional amount of 5 N hydrochloric acid (5.1 ml), the mixture is stirred for 10 hours at the boil under reflux. The reaction mixture is then cooled to room temperature, diluted with water and washed with chloroform. A 5N sodium hydroxide aqueous solution (10 ml) was added to the aqueous layer, followed by extraction with chloroform. The resulting organic layer was dried over anhydrous sodium sulfate, then filtered and concentrated. The resulting residue was purified by basic silica gel chromatography (elution solvent: ethyl acetate/methanol), thus obtaining a mixture of the title compounds (1.4098 g) as a light yellow solid.

ESI-MS m/z: 343 [M+H] ⁺ .

Step 7: Synthesis of Compound 14

Diisopropylethylamine (0.8452 ml) and a solution of acryloyl chloride (0.35 ml) in acetonitrile (3.5 ml) were added successively to a mixture solution (1.407 g) of (S)-6-methyl-5-(quinoline-3- yl)-8,9-dihydropyrimido[5,4-b]indolizin-4,8-diamine and (S)-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido [5,4-b]indolizine-4,8-diamine prepared in step 6 in acetonitrile (10 ml) and water (10 ml) at 0° C. and the mixture is stirred for 45 minutes at the same temperature. Then the reaction mixture was diluted with water, poured into a saturated aqueous sodium bicarbonate solution, then extracted with ethyl acetate. After the organic layer was dried over anhydrous sodium sulfate, the solvent was distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (897.1 mg) as a light yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 1.51 (3H, s), 4.17 (1H, dd, J=13.2, 5.1 Hz), 4.25 (1H, dd, J =13.2, 5.1 Hz), 4.73-4.83 (1H, m), 5.61 (1H, dd, J=9.8, 2.7 Hz), 5.65-6, 00 (2H, br.s), 5.81 (1H, dd, J=5.1, 1.2 Hz), 6.14 (1H, dd, J=17.1, 2.4 Hz), 6 .24 (1H, dd, J=17.1, 9.8 Hz), 7.67 (1H, ddd, J=8.1, 7.1, 1.0 Hz), 7.81 (1H, ddd , J=8.3, 7.1, 1.5 Hz), 8.05 (1H, d, J=8.1 Hz), 8.09 (1H, d, J=8.3 Hz), 8 .13 (1H, s), 8.39 (1H, br.s), 8.46 (1H, d, J=7.3 Hz), 8.94 (1H, s).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 15

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 15)

The mixed fraction obtained by purification by silica gel column chromatography in step 7 of Example 14 was purified by reverse phase preparative HPLC, thereby obtaining the title compound (32.2 mg) as a light yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 2.56-2.70 (1H, m), 2.72-2.82 (1H, m), 4.19 (1H, dd, J=14, 1, 8.3 Hz), 4.25-4.36 (1H, m), 4.72 (1H, d, J=13.7 Hz), 5.62 (1H, dd, J=10.0 , 2.2 Hz), 5.80 (1H, dt, J=12.2, 4.6 Hz), 6.00-6.20 (3H, m), 6.20-6.31 (2H, m), 7.65 (1H, t, J=8.1 Hz), 7.80 (1H, t, J=8.3 Hz), 8.05 (1H, d, J=8.5 Hz) , 8.08 (1H, d, J=8.3 Hz), 8.15 (1H, s), 8.28-8.39 (2H, m), 8.87 (1H, d, J=2 .2 Hz).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 16

(R)-N-(4-Amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 16)

Step 1: Synthesis of (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-4-en-2-yl)carbamate

The same synthesis as in steps 1-3 of Example 14 is carried out using (R)-2-amino-4-pentenoic acid instead of (S)-2-amino-4-pentenoic acid used in step 1 of Example 14, such thus obtaining the title compound (1.4217 g) as a light yellow solid.

ESI-MS m/z: 463, 465 [M+H] ⁺ .

Step 2: Synthesis of Compound 16

The same synthesis as in steps 4-7 of Example 14 was carried out using (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl )pent-4-en-2-yl)carbamate obtained in step 1, thus obtaining the title compound (21.0 mg) as a light yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 1.51 (3H, s), 4.17 (1H, dd, J=13.2, 5.1 Hz), 4.25 (1H, dd, J =13.2, 5.1 Hz), 4.73-4.83 (1H, m), 5.61 (1H, dd, J=9.8, 2.7 Hz), 5.65-6, 00 (2H, br.s), 5.81 (1H, dd, J=5.1, 1.2 Hz), 6.14 (1H, dd, J=17.1, 2.4 Hz), 6 .24 (1H, dd, J=17.1, 9.8 Hz), 7.67 (1H, t, J=7.1 Hz), 7.81 (1H, t, J=7.6 Hz) , 8.05 (1H, d, J=8.1 Hz), 8.09 (1H, d, J=8.6 Hz), 8.13 (1H, s), 8.39 (1H, br. s), 8.46 (1H, d, J=7.1 Hz), 8.94 (1H, s).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 17

(R)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 17)

The mixed fraction obtained by purification by silica gel column chromatography in Example 16 was purified by the same procedure as in Example 15, thereby obtaining the title compound (8.4 mg) as a light yellow solid .

¹ H-NMR (DMSO-d ₆ ) δ: 2.57-2.70 (1H, m), 2.71-2.82 (1H, m), 4.19 (1H, dd, J=13, 2, 8.3 Hz), 4.25-4.36 (1H, m), 4.72 (1H, d, J=13.2 Hz), 5.62 (1H, dd, J=10.0 , 2.2 Hz), 5.80 (1H, dt, J=12.2, 4.6 Hz), 6.00-6.20 (3H, m), 6.20-6.31 (2H, m), 7.65 (1H, t, J=7.6 Hz), 7.79 (1H, ddd, J=8.3, 7.1, 1.5 Hz), 8.05 (1H, d , J=8.5 Hz), 8.08 (1H, d, J=8.5 Hz), 8.15 (1H, s), 8.33 (1H, d, J=2.2 Hz), 8.36 (1H, d, J=7.6 Hz), 8.87 (1H, d, J=2.2 Hz).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 18

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8,9,10-tetrahydro-6H-pyrimido[5',4':4,5]pyrrolo [1,2-a]azepin-8-yl)acrylamide (Compound 18)

Step 1: Synthesis of (S)-tert-butyl(1-hydroxyhex-5-en-3-yl)carbamate

N-methylmorpholine (2.1 g) was added to a solution of (S)-3-((tert-butoxycarbonyl)amino)hex-5-enoic acid (4.0 g) in THF (40 ml) and ethyl chloroformate was gradually added (1.75 ml), at -10°C. Thereafter, the mixture was stirred for 20 minutes at the same temperature, and the resulting insoluble matter was filtered off through celite. To the resulting filtrate was gradually added a solution of sodium tetrahydroborate (904 mg) in water (8 ml) at -10°C. After the mixture was stirred for 30 minutes, at the same temperature, a saturated aqueous ammonium chloride solution was added. After extraction with ethyl acetate, washed with 0.5 N potassium hydrogen sulfate aqueous solution, saturated sodium bicarbonate aqueous solution and saturated sodium chloride solution in that order. The resulting organic layer was dried over anhydrous sodium sulfate and then filtered, then concentrated. The resulting residue was purified by silica gel chromatography (eluting solvent: hexane/ethyl acetate), thereby obtaining the title compound (3.54 g) as an oily substance.

Step 2: Synthesis of Compound 18

The same synthesis as in steps 4-10 of Example 13 is carried out using (S)-tert-butyl(1-hydroxyhex-5-en-3-yl)carbamate obtained in step 1 instead of (R)-tert- butyl(5-hydroxypent-1-en-3-yl)carbamate used in step 4 of Example 13, thus obtaining the title compound (147 mg) as a colorless solid.

¹ H-NMR (DMSO-d ₆ ) δ: 1.68-1.82 (1H, m), 2.17-2.25 (1H, m), 2.42-2.53 (1H, m) , 2.75-2.83(1H, m), 3.86-4.16(2H, m), 4.64-4.78(1H, m), 4.84(1H, d, J= 1.7 Hz), 5.26 (1H, s), 5.60 (1H, dd, J=10.0, 2.4 Hz), 6.00 (2H, br.s), 6.11 ( 1Н, dd, J=17.1, 2.4 Hz), 6.27 (1Н, dd, J=17.1, 10.0 Hz), 7.60-7.64 (1Н, m), 7 .73-7.77 (1H, m), 7.99-8.04 (2H, m), 8.17 (1H, s), 8.22 (1H, d, J=8.0 Hz), 8.30 (1H, d, J=2.0 Hz), 8.78 (1H, d, J=2.0 Hz).

ESI-MS m/z: 411 [M+H] ⁺ .

Example 19

(S,E)-N-(4-Amino-6-ethyliden-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 19)

Step 1: Synthesis of (S)-methyl-3-((tert-butoxycarbonyl)amino)-4-hydroxybutanoate

N-methylmorpholine (9.78 ml) and ethyl chloroformate (8.09 ml) were added to a solution of (S)-2-((tert-butoxycarbonyl)amino)-4-methoxy-4-oxobutanoic acid (20.0 g), in THF (200 ml), under ice-cooling, and the mixture is stirred for 1 hour, under ice-cooling. The resulting insoluble matter is filtered off. A solution of sodium borohydride (4.14 g) in water (41.4 ml) was added to the filtrate under ice-cooling, and the mixture was stirred for 30 minutes under ice-cooling. A 0.5 N aqueous potassium hydrogen sulfate solution and ethyl acetate were added to the reaction mixture. The organic layer was separated, washed with saturated sodium bicarbonate aqueous solution and saturated sodium chloride solution, and dried over anhydrous magnesium sulfate. The solvent was then distilled off under reduced pressure, thus obtaining the title compound (10.82 g).

Stage 2: Synthesis of (S)-tert-butyl-4-(2-methoxy-2-oxoethyl)-2,2-dimethyloxazolidin-3-carboxylate

2,2-Dimethoxypropane (28.52 ml) and boron trifluoride-diethyl ether complex (0.294 ml) were added to a solution of (S)-methyl-3-((tert-butoxycarbonyl)amino)-4-hydroxybutanoate (10.82 g ) obtained in step 1 in acetone (108.2 ml) at room temperature, and the mixture is stirred for 4 hours at the same temperature. The reaction mixture was poured into saturated aqueous sodium bicarbonate and extracted with ethyl acetate. The resulting organic layer was dried over anhydrous magnesium sulfate and then filtered, then concentrated. The resulting residue was purified by silica gel chromatography (elution solvent: hexane/ethyl acetate), thereby obtaining the title compound (8.72 g).

Stage 3: Synthesis of (S)-tert-butyl-4-(2-hydroxyethyl)-2,2-dimethyloxazolidine-3-carboxylate

A 1 M solution of diisobutylaluminum hydride in toluene (65.7 ml) was added to a solution of (S)-tert-butyl-4-(2-methoxy-2-oxoethyl)-2,2-dimethyloxazolidin-3-carboxylate (8.71 g) obtained in step 2, in methylene chloride (87.1 ml), under ice-cooling, and the mixture is stirred for 2 hours, under ice-cooling. A 5% mixed potassium sodium tartrate aqueous solution and ethyl acetate were added to the reaction mixture, and the mixture was stirred overnight at room temperature. The resulting organic layer was dried over anhydrous magnesium sulfate and then filtered, then concentrated. The resulting residue was purified by silica gel chromatography (elution solvent: hexane/ethyl acetate), thereby obtaining the title compound (5.94 g).

Step 4: Synthesis of (S)-tert-butyl-2,2-dimethyl-4-(2-oxoethyl)oxazolidine-3-carboxylate

Triethylamine (16.9 ml) and sulfur trioxide-pyridine complex (12.56 g) were added to a solution of (S)-tert-butyl-4-(2-hydroxyethyl)-2,2-dimethyloxazolidine-3-carboxylate (5, 94 g) obtained in step 3 in DMSO (59.4 ml) at room temperature, and the mixture was stirred for 1 hour at the same temperature. The reaction mixture was poured into saturated aqueous sodium bicarbonate and extracted with ethyl acetate. The resulting organic layer was dried over anhydrous magnesium sulfate and then filtered, then concentrated. The resulting residue was purified by silica gel chromatography (elution solvent: hexane/ethyl acetate), thereby obtaining the title compound (6.06 g).

Step 5: Synthesis of (S)-tert-butyl-4-(but-2-en-1-yl)-2,2-dimethyloxazolidin-3-carboxylate

A 2.69 M solution of n-butyl lithium in hexane (7.0 ml) was added to a suspension of ethyltriphenylphosphonium bromide (5.60 g), in THF (25.2 ml), with ice-cooling, and the mixture was stirred for 30 minutes, with cooling ice. To the reaction mixture was added a solution of (S)-tert-butyl-2,2-dimethyl-4-(2-oxoethyl)oxazolidine-3-carboxylate (3.06 g) obtained in Step 4 in THF (3.06 ml ), under ice-cooling, and the mixture was stirred for 14 hours at room temperature. Hexane is added to the reaction mixture and the insoluble matter is filtered off, then washed with a mixture of THF-hexane = 2/1. The filtrate is concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography (eluting solvent: hexane/ethyl acetate), thereby obtaining the title compound (1.81 g).

Step 6: Synthesis of (S)-tert-butyl(1-hydroxyhex-4-en-2-yl)carbamate

A 4 N solution of hydrogen chloride in dioxane (18.1 ml) was added to (S)-tert-butyl-4-(but-2-en-1-yl)-2,2-dimethyloxazolidin-3-carboxylate (1.81 g ) obtained in step 5 at room temperature, and the mixture is stirred for 2 hours at 70°C. The reaction mixture is cooled and then concentrated under reduced pressure. The obtained residue was dissolved in THF (18.1 ml), and a saturated sodium bicarbonate aqueous solution (18.1 ml) and di-t-butyl dicarbonate (1.53 g) were added thereto at room temperature. The mixture is stirred overnight at the same temperature. The reaction mixture was poured into water and extracted with ethyl acetate. The resulting organic layer was dried over anhydrous magnesium sulfate and then filtered, then concentrated. The resulting residue was purified by silica gel chromatography (eluting solvent: hexane/ethyl acetate), thereby obtaining the title compound (1.57 g).

Step 7: Synthesis of Compound 19

The same synthesis as in steps 4-10 of Example 13 is carried out using (S)-tert-butyl(1-hydroxyhex-4-en-2-yl)carbamate obtained in step 6 instead of (R)-tert- butyl(5-hydroxypent-1-en-3-yl)carbamate used in step 4 of Example 13, thus obtaining the title compound (327 mg).

¹ H-NMR (DMSO-d ₆ ) δ: 1.47 (3H, d, J=7.1 Hz), 2.64-2.70 (1H, m), 2.81-2.85 (1H , m), 4.00-4.09 (1H, m), 4.32-4.43 (2H, m), 5.41-5.46 (1H, m), 5.60-5.63 (1H, m), 5.82 (2H, br.s), 6.11-6.16 (1H, m), 6.21-6.31 (1H, m), 8.05 (1H, d , J=7.3 Hz), 8.08 (1H, d, J=8.5 Hz), 8.13 (1H, s), 8.35-8.39 (2H, m), 8.87 -8.92 (1H, m).

ESI-MS m/z: 411 [M+H] ⁺ .

Example 20

A mixture of (S)-N-(4-amino-6-isopropyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 20A) and (S)-N-(4-amino-6-(propan-2-ylidene)-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8 -yl) acrylamide (Compound 20B)

The same synthesis as in Example 19 was carried out using isopropyltriphenylphosphonium iodide instead of ethyltriphenylphosphonium bromide used in step 5 of Example 19, thus obtaining a mixture (18.9 mg) of the title compounds.

¹ H-NMR (DMSO-d ₆ ) δ: 0.85-4.71 (11H, m), 5.63-5.68 (1H, m), 5.88 (1H, br.s), 6 .03 (1H, br.s), 6.12-6.19 (1H, m), 6.24-6.31 (1H, m), 7.59-7.64 (1H, m), 7 .72-7.78(1H, m), 7.96-7.99(1H, m), 8.01-8.05(1H, m), 8.16(1H, d, J=3, 4 Hz), 8.24-8.28 (1H, m), 8.39-8.44 (1H, m), 8.90 (1H, d, J=2.0 Hz).

ESI-MS m/z: 425 [M+H] ⁺ .

Example 21

(R)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)-N- methylacrylamide (Compound 21)

Step 1: Synthesis of (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but-3-en-2-yl)carbamate

The same synthesis as in step 1 of Example 1 was carried out using (R)-tert-butyl(1-hydroxybut-3-en-2-yl)carbamate instead of (S)-tert-butyl(1-hydroxybut-3- en-2-yl)carbamate used in step 1 of Example 1, thus obtaining the title compound (1.083 g) as a light yellow solid.

ESI-MS m/z: 448, 450 [M+H] ⁺ .

Step 2: Synthesis of (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but-3-en-2-yl)( methyl) carbamate

Methyl iodide (1.58 ml) and sodium hydride (224 mg) dispersed in liquid paraffin are added to a solution of (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3- d]pyrimidin-7-yl)but-3-en-2-yl)carbamate (2.28 g) obtained in Step 1 in DMF (11.4 ml) at room temperature and the mixture was stirred for 1 hours at the same temperature. The resulting mixture was poured into water and extracted with ethyl acetate. The resulting organic layer was washed with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate; thereafter, the solvent is distilled off under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluting solvent: hexane/ethyl acetate), thereby obtaining the title compound (2.41 g) as a light yellow solid.

ESI-MS m/z: 462, 464 [M+H] ⁺ .

Step 3: Synthesis of Compound 21

The same synthesis as in steps 5-10 of Example 13 was carried out using (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl )but-3-en-2-yl)(methyl)carbamate obtained in step 2 instead of (R)-tert-butyl(5-(4-chloro-5-iodo-7H-pyrrolo[2,3-d ]pyrimidin-7-yl)pent-1-en-3-yl)carbamate used in step 5 of Example 13, thus obtaining the title compound (256 mg) as a light yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 2.75 and 2.94 (total 3H, each s), 4.06-4.20 (1H, m), 4.46-4.72 (1H, m), 4.81-4.89 (1H, m), 5.26-5.35 (1H, m), 5.70-5.80 (1H, m), 5.98-6.37 ( 4H, m), 6.77-7.01 (1H, m), 7.62-7.68 (1H, m), 7.77-7.83 (1H, m), 8.03-8, 10(2H, m), 8.14-8.18(1H, m), 8.41-8.45(1H, m), 8.96-9.01(1H, m).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 22

(R)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)-N- methylacrylamide (Compound 22)

Step 1: Synthesis of (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent-4-en-2-yl)( methyl) carbamate

The same synthesis as in step 2 of Example 21 was carried out using (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pent -4-en-2-yl)carbamate obtained in step 1 of Examples 16 and 17 instead of (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d] pyrimidin-7-yl)but-3-en-2-yl)carbamate used in step 2 of Example 21, thus obtaining the title compound (2.743 g) as a light yellow solid.

ESI-MS m/z: 477, 479 [M+H] ⁺ .

Step 2: Synthesis of (R)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8- yl)(methyl)carbamate

The same synthesis as in steps 5-8 of Example 13 was carried out using (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl )pent-4-en-2-yl)(methyl)carbamate obtained in step 1 instead of (R)-tert-butyl(5-(4-chloro-5-iodo-7H-pyrrolo[2,3-d ]pyrimidin-7-yl)pent-1-en-3-yl)carbamate used in step 5 of Example 13, thus obtaining the title compound (1.366 g) as a light brown solid.

ESI-MS m/z: 457 [M+H] ⁺ .

Stage 3: Synthesis of (R)-N ⁸ -methyl-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-4,8-diamine

5 N Hydrochloric acid (0.5 ml) was added to a solution of (R)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5 ,4-b]indolizin-8-yl)(methyl)carbamate (228 mg) obtained in step 2 in ethanol (3 ml) and the mixture was stirred for 4 days at 50°C. After cooling, the reaction mixture was made basic with a 5N aqueous sodium hydroxide solution, then extracted with chloroform. After the resulting organic layer was dried over anhydrous sodium sulfate, the solvent was distilled off under reduced pressure. The resulting residue was purified by basic silica gel column chromatography (elution solvent: chloroform/methanol), thereby obtaining the title compound (197 mg) as a light brown solid.

ESI-MS m/z: 357 [M+H] ⁺ .

Step 4: Synthesis of Compound 22

The same synthesis as in step 10 of Example 13 was carried out using (R)-N ⁸ -methyl-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4 -b]indolizin-4,8-diamine obtained in step 3 instead of ((R)-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4- b]indolizine-4,7-diamine used in step 10 of Example 13, thus obtaining the title compound (68.5 mg) as a light yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 2.53-2.68 (1H, m), 2.88-3.11 (4H, m), 4.04-4.20 (1H, m) , 4.34-4.96(4H, m), 5.58-6.21(4H, m), 6.72-7.01(1H, m), 7.62-7.68(1H, m), 7.78-7.83 (1H, m), 8.02-8.10 (2H, m), 8.15 (1H, br.s), 8.37-8.44 (1H, m), 8.86-8.90 (1H, m).

ESI-MS m/z: 411 [M+H] ⁺ .

Example 23

(R)-N-(4-Amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)-N-methylacrylamide (Compound 23 )

Stage 1: Synthesis of (R)-N ⁸ ,6-dimethyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-4,8-diamine

5 N Hydrochloric acid (1 ml) was added to a solution of (R)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4 -b]indolizin-8-yl)(methyl)carbamate (228 mg) obtained in step 2 of Example 22 in ethanol (4 ml) and the mixture was stirred for 24 hours at reflux. After cooling, the reaction mixture was made basic with a 5N aqueous sodium hydroxide solution, then extracted with chloroform. Then, the resulting organic layer was dried over anhydrous sodium sulfate, and the solvent was distilled off under reduced pressure. The resulting residue was purified by basic silica gel column chromatography (elution solvent: chloroform/methanol), thereby obtaining the title compound (190.8 mg) as a light brown solid.

ESI-MS m/z: 357 [M+H] ⁺ .

Step 2: Synthesis of Compound 23

The same synthesis as in step 10 of Example 13 was carried out using (R)-N ⁸ ,6-dimethyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-4 ,8-diamine obtained in step 1 instead of ((R)-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-4, 7-diamine used in step 10 of Example 13, thus obtaining the title compound (150.9 mg) as a yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 1.52 (3H, s), 2.74 and 2.88 (total 3H, each br.s), 4.14-4.42 (2H, m) , 5.21-5.98(5H, m), 6.09-6.22(1H, m), 6.71-7.03(1H, m), 7.64-7.69(1H, m), 7.79-7.85 (1H, m), 8.02-8.07 (1H, m), 8.08-8.12 (1H, m), 8.14 (1H, s) , 8.43-8.49(1H, m), 8.89-8.96(1H, m).

ESI-MS m/z: 411 [M+H] ⁺ .

Example 24

N-((7S)-4-Amino-6-methyl-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)acrylamide (Compound 24)

Step 1: Synthesis of tert-butyl((7S)-4-amino-6-methyl-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7- yl) carbamate

10% Palladium-on-charcoal (50 wt%, 15.0 mg) was added to a solution of (S)-tert-butyl (4-amino-6-methylene-5-(quinolin-3-yl)-7 ,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)carbamate (15.0 mg) obtained in step 5 of Example 1 in ethyl acetate (2 ml)-ethanol (1 ml), and the mixture is stirred for 12 hours at room temperature under a hydrogen atmosphere. The reaction mixture is filtered through Celite, and the filtrate is concentrated under reduced pressure. The resulting residue was purified by preparative thin layer chromatography (elution solvent: chloroform/methanol), thereby obtaining the title compound (10.0 mg).

ESI-MS m/z: 357 [M+H] ⁺ .

Step 2: Synthesis of Compound 24

The same synthesis as in steps 9 and 10 of Example 13 was carried out using tert-butyl((7S)-4-amino-6-methyl-5-(quinolin-3-yl)-7,8-dihydro-6H- pyrimido[5,4-b]pyrrolisin-7-yl)carbamate obtained in step 1 instead of (R)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6, 7,8,9-tetrahydropyrimido[5,4-b]indolizin-7-yl)carbamate used in step 9 of Example 13, thus obtaining the title compound (3.10 mg) as a colorless solid.

¹ H-NMR (DMSO-d ₆ ) δ: 0.86 (2.7H, d, J=7.6 Hz), 1.08 (0.3H, d, J=7.6 Hz), 3, 43-4.02 (2H, m), 4.34-4.62 (1H, m), 5.08-5.19 (1H, m), 5.59-5.68 (1H, m), 5.99-6.17 (3H, m), 6.18-6.33 (1H, m), 7.62 (1H, dd, J=7.6, 7.6 Hz), 7.75 ( 1Н, dd, J=7.6, 7.6 Hz), 7.99-8.07 (2Н, m), 8.12 (1Н, s), 8.29-8.36 (1Н, m) , 8.53-8.76 (1H, m), 8.98 (1H, d, J=2.2 Hz).

ESI-MS m/z: 385 [M+H] ⁺ .

Example 25

(R)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)acrylamide (Compound 25)

The same synthesis as in steps 2-6 of Example 1 was carried out using (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl )but-3-en-2-yl)carbamate obtained in step 1 of Example 21 instead of (S)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d] pyrimidin-7-yl)but-3-en-2-yl)carbamate used in step 2 of Example 1, thus obtaining the title compound (44.6 mg) as a light yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 3.88-3.93 (1H, m), 4.57-4.63 (1H, m), 5.03 (1H, d, J=2, 4 Hz), 5.24 (1H, d, J=2.4 Hz), 5.55-5.62 (1H, m), 5.68 (1H, dd, J=10.0, 2.4 Hz), 6.12-6.38 (4H, m), 7.65 (1H, dd, J=7.8, 7.8 Hz), 7.77-7.83 (1H, m), 8 .04-8.11 (2H, m), 8.15 (1H, s), 8.41 (1H, d, J=2.2 Hz), 8.82 (1H, d, J=7.8 Hz), 8.98 (1H, d, J=2.2 Hz).

ESI-MS m/z: 383 [M+H] ⁺ .

Example 26

(S)-N-(4-Amino-6-methylene-5-(quinolin-3-yl)-7,8-dihydro-6H-pyrimido[5,4-b]pyrrolisin-7-yl)-N- methylacrylamide (Compound 26)

The same synthesis as in steps 2 and 3 of Example 21 was carried out using (S)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl )but-3-en-2-yl)carbamate obtained in step 1 of Example 1 instead of (R)-tert-butyl(1-(4-chloro-5-iodo-7H-pyrrolo[2,3-d] pyrimidin-7-yl)but-3-en-2-yl)carbamate used in step 2 of Example 21, thus obtaining the title compound (143 mg) as a light yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 2.75 and 2.94 (total 3H, each s), 4.06-4.20 (1H, m), 4.46-4.72 (1H, m), 4.81-4.89 (1H, m), 5.26-5.35 (1H, m), 5.70-5.80 (1H, m), 5.98-6.37 ( 4H, m), 6.77-7.01 (1H, m), 7.62-7.68 (1H, m), 7.77-7.83 (1H, m), 8.03-8, 10(2H, m), 8.14-8.18(1H, m), 8.41-8.45(1H, m), 8.96-9.01(1H, m).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 27

(S)-N-(4-Amino-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)acrylamide (Compound 27)

Step 1: Synthesis of (S)-tert-butyl(4-amino-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)carbamate

A 0.5 M solution of 9-borabicyclo[3,3,1]nonane in tetrahydrofuran (141.3 ml) was added to a solution of (S)-tert-butyl(1-(4-amino-6-bromo-5-(quinoline -3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but-3-en-2-yl)carbamate (6.0 g) obtained in step 4 of Example 1 in tetrahydrofuran ( 42 ml), under a nitrogen atmosphere. The mixture is stirred at room temperature for 2 hours. A 2N aqueous sodium hydroxide solution (84.8 ml) was gradually added to the reaction mixture at room temperature, and degassed under reduced pressure. Under a nitrogen atmosphere, (tetrakistriphenylphosphine)palladium(0) (1.70 g) was added and the mixture was stirred at 66°C for 12 hours. After the reaction mixture was cooled, the organic layer was separated and washed with 20% ammonium chloride aqueous solution (60 ml). Then, silica gel SH (6.0 g) was added to the organic layer, and the mixture was stirred at 50° C., under nitrogen atmosphere, for 14 hours, and then filtered. Silica gel SH (manufactured by Fuji Silysia Chemical Ltd.) (6.0 g) was again added to the filtrate, and the mixture was stirred under nitrogen at 50° C. for 14 hours, and then filtered. The solvent is distilled off from the filtrate under reduced pressure. The resulting residue was purified by silica gel column chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (4.46 g; yield = 88%) as a light yellow solid.

ESI-MS m/z: 431 [M+H] ⁺ .

Step 2: Synthesis of (8S)-4-amino-8-((tert-butoxycarbonyl)amino)-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizine -6-yl acetate

Tetrabutylammonium iodide (37 mg) and iodobenzene diacetate (241 mg) are added to a solution of (S)-tert-butyl (4-amino-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4- b]indolizin-8-yl)carbamate (215 mg) obtained in step 1 in acetic acid (2 ml) and methylene chloride (2 ml) under ice-cooling. The mixture is stirred for 2 hours under ice-cooling and for 2 hours at room temperature. After concentration under reduced pressure, the reaction mixture was made basic with a saturated sodium bicarbonate aqueous solution, then extracted with ethyl acetate. The resulting organic layer was dried over anhydrous sodium sulfate, then filtered. Thereafter, the solvent is distilled off under reduced pressure. The resulting residue was purified by silica gel chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (216 mg) as a light brown solid.

ESI-MS m/z: 489 [M+H] ⁺ .

Step 3: Synthesis of tert-butyl((8S)-4-amino-6-hydroxy-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8- yl) carbamate

2 N Aqueous sodium hydroxide solution (1 ml) was added to a solution of (8S)-4-amino-8-((tert-butoxycarbonyl)amino)-5-(quinolin-3-yl)-6,7,8,9- tetrahydropyrimido[5,4-b]indolizin-6-ylacetate (215 mg) obtained in step 2 in methanol (3 ml) at room temperature, and the mixture was stirred for 1 hour at room temperature. The reaction mixture was poured into water and extracted with chloroform. The resulting organic layer was dried over anhydrous sodium sulfate, then filtered. Thereafter, the solvent is distilled off under reduced pressure. The resulting residue was purified by silica gel chromatography (elution solvent: ethyl acetate/methanol), thereby obtaining the title compound (166.7 mg) as a light yellow solid.

ESI-MS m/z: 447 [M+H] ⁺ .

Step 4: Synthesis of (S)-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizine-4,8-diamine

A mixture of tert-butyl ((8S)-4-amino-6-hydroxy-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)carbamate (44.6 mg) obtained in step 3, p-toluenesulfonic acid monohydrate (95 mg) and toluene (3 ml) were stirred for 4 hours at a temperature of 100°C. After the reaction mixture was cooled, the solvent was distilled off under reduced pressure. The resulting residue was purified by silica gel chromatography (eluting solvent: chloroform/methanol), thereby obtaining the title compound (39 mg) as a light brown oily substance.

ESI-MS m/z: 329 [M+H] ⁺ .

Step 5: Synthesis of Compound 27

The same synthesis as in step 10 of Example 13 was carried out using (S)-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizine-4,8-diamine obtained from step 4, instead of (R)-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-4,7-diamine used in step 10 Example 13, thus obtaining the title compound (44.1 mg) as a yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 4.19-4.30 (2H, m), 4.86-4.94 (1H, m), 5.63 (1H, dd, J=9, 8, 2.7 Hz), 6.08-6.32 (5H, m), 6.70 (1H, dd, J=9.8, 1.1 Hz), 7.62-7.69 (1H , m), 7.75-7.82 (1H, m), 8.04-8.09 (2H, m), 8.17 (1H, s), 8.34 (1H, d, J=2 .0 Hz), 8.55 (1H, d, J=7.3 Hz), 8.91 (1H, d, J=2.4 Hz).

ESI-MS m/z: 383 [M+H] ⁺ .

Example 28

(R)-N-(4-Amino-5-(quinolin-3-yl)-9,10-dihydro-8H-pyrimido[5',4':4,5]pyrrolo[1,2-a]azepine -8-yl)acrylamide (Compound 28)

The same synthesis as in Example 27 was carried out using (R)-tert-butyl(5-(4-amino-6-bromo-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d ]pyrimidin-7-yl)pent-1-en-3-yl)carbamate obtained in step 7 of Example 13 instead of (S)-tert-butyl(1-(4-amino-6-bromo-5-(quinoline -3-yl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)but-3-en-2-yl)carbamate used in Step 1 of Example 27, thus obtaining the title compound (101 .8 mg) as a yellow solid.

¹ H-NMR (DMSO-d ₆ ) δ: 2.01-2.12 (1H, m), 2.23-2.33 (1H, m), 4.28-4.38 (1H, m) , 4.55-4.63 (1H, m), 4.84-4.92 (1H, m), 5.62 (1H, dd, J=10.0, 2.4 Hz), 5.68 (1Н, dd, J=12.6, 3.8 Hz), 6.08-6.31 (5Н, m), 7.63-7.69 (1Н, m), 7.78-7.83 (1H, m), 8.03-8.10 (2H, m), 8.18 (1H, s), 8.31 (1H, d, J=2.0 Hz), 8.51 (1H, d, J=8.2 Hz), 8.84 (1H, d, J=2.2 Hz).

ESI-MS m/z: 397 [M+H] ⁺ .

Example 29

N-((6R*,8S)-4-Amino-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl) acrylamide (Compound 29A) and N-((6S*,8S)-4-amino-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b] indolizin-8-yl)acrylamide (Compound 29B)

Step 1: Synthesis of (S)-tert-butyl(4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8- yl) carbamate

Mixture of (S)-tert-butyl(4-amino-6-methyl-5-(quinolin-3-yl)-8,9-dihydropyrimido[5,4-b]indolizin-8-yl)carbamate and (S) -tert-butyl (4-amino-6-methylene-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)carbamate obtained by such the same synthesis as in step 5 of Example 14 using (S)-tert-butyl(1-(4-amino-6-bromo-5-(quinolin-3-yl)-7H-pyrrolo[2,3-d ]pyrimidin-7-yl)pent-4-en-2-yl)carbamate (1.263 g) obtained in step 4 of Example 14 was purified by silica gel chromatography (elution solvent: ethyl acetate/methanol), thus obtaining the title compound (0.805 g), as a light yellow solid.

ESI-MS m/z: 443 [M+H] ⁺ .

Step 2: Synthesis of tert-butyl((8S)-4-amino-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8- yl) carbamate

10% Palladium-on-charcoal (50 wt%, 100.6 mg) was added to a solution of (S)-tert-butyl (4-amino-6-methylene-5-(quinolin-3-yl)-6 ,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl)carbamate (100.0 mg) obtained in step 1 in methanol (10 ml) and the mixture was stirred for 12 hours, at room temperature, in a hydrogen atmosphere. The reaction mixture is filtered through a glass fiber filter and the filtrate is concentrated under reduced pressure. The resulting residue was purified by basic silica gel column chromatography (elution solvent: chloroform/methanol), thereby obtaining the title compound (122.1 mg).

ESI-MS m/z: 445 [M+H] ⁺ .

Step 3: Synthesis of (8S)-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-4,8-diamine

Trifluoroacetic acid (1 ml) is added to a solution of tert-butyl ((8S)-4-amino-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b ]indolizin-8-yl)carbamate (122.1 mg) obtained in Step 2 in chloroform (3 ml) and the mixture was stirred for 30 minutes at room temperature. The reaction mixture is concentrated under reduced pressure, and the resulting residue is dissolved in methanol. Then desalted using a basic solid phase extraction column (VARIAN BondElut), the mixture was concentrated under reduced pressure, thus obtaining the title compound (58.9 mg) as a light yellow solid.

ESI-MS m/z: 345 [M+H] ⁺ .

Step 4: Synthesis of N-((8S)-4-amino-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-8-yl ) acrylamide

The same synthesis as in step 7 of Example 14 was carried out using (8S)-6-methyl-5-(quinolin-3-yl)-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin- 4,8-diamine (58.9 mg) obtained in step 3, thus obtaining the title compound (30.3 mg) as a light yellow solid.

ESI-MS m/z: 399 [M+H] ⁺ .

Step 5: Synthesis of Compound 29A and Compound 29B

The mixture of diastereoisomers containing the (6R*,8S) isomer and the (6S*,8S) isomer obtained in Step 4 was separated by preparative chiral column chromatography (column: CHIRALCEL OZ-H (20 mm×250 mm×5 µm ) manufactured by Daicel Chemical Industries, Ltd., eluting solvent: hexane/ethanol/triethylamine = 60/40/0.01), thus obtaining Compound 29A and Compound 29B as a first fraction (15.8 mg) and a second fractions (6.1 mg), respectively, as colorless solids.

Compound 29A

¹ H-NMR (CDCl ₃ ) δ: 0.96 (3H, d, J=6.6 Hz), 1.68 (1H, m), 2.29-2.39 (1H, m), 3, 49-3.63 (1H, m), 3.81 (1H, dd, J=11.7, 10.2 Hz), 4.56-4.69 (1H, m), 4.76 (1H, dd, J=12.2, 5.1 Hz), 4.81 (2H, br.s), 5.70 (1H, dd, J=10.5, 1.2 Hz), 6.17 (1H , dd, J=16.6, 10.2 Hz), 6.15-6.25 (1H, m), 6.36 (1H, dd, J=17.1, 1.2 Hz), 7, 64 (1H, t, J=7.6 Hz), 7.75-7.82 (1H, m), 7.88 (1H, d, J=8.0 Hz), 8.18 (1H, d , J=8.5 Hz), 8.21-8.29 (2H, m), 9.05 (1H, d, J=2.0 Hz).

ESI-MS m/z: 399 [M+H] ⁺ .

Connection 29V

¹ H-NMR (CDCl ₃ ) δ: 1.06 (3H, d, J=6.8 Hz), 1.83-2.08 (1H, m), 2.18-2.31 (1H, m ), 3.46-3.59 (1H, m), 4.00 (1H, dd, J=12.4, 7.6 Hz), 4.59 (1H, dd, J=12.4, 4 .6 Hz), 4.73-4.93 (3H, m), 5.70 (1H, d, J=10.5 Hz), 6.18 (1H, dd, J=16.8, 10, 2 Hz), 6.26-6.42 (2H, m), 7.63 (1H, t, J=7.6 Hz), 7.78 (1H, t, J=8.3 Hz), 7 .88 (1H, d, J=8.0 Hz), 8.17 (1H, d, J=8.0 Hz), 8.20-8.27 (2H, m), 9.02 (1H, e, J=1.7 Hz).

ESI-MS m/z: 399 [M+H] ⁺ .

Comparative Example 1

N-(3-(4-Amino-6,7,8,9-tetrahydropyrimido[5,4-b]indolizin-5-yl)phenyl)benzamide (Comparative Compound 1)

The compound is synthesized according to the procedure disclosed in International Publication Article WO 2006/102079.

ESI-MS m/z: 384 [M+H] ⁺ .

Test cases

Compounds of the present invention were evaluated using the following test methods.

Test Example 1: Measurement of inhibitory action against various EGFR kinase activities ( in vitro )

1) Measurement of EGFR (T790M/L858R)-kinase inhibitory activity

The inhibitory activities of the compounds of the present invention against EGFR (T790M/L858R) kinase activity were measured.

Among substances for measuring inhibitory activity, a substrate peptide and a kinase protein were prepared as follows. A biotinylated N-terminal peptide (Biotin-EEPLYWSFPAKKK) was synthesized relative to FL-Peptide 22, a PerkinElmer, Inc. LabChip® reagent. For the kinase protein, purified recombinant human EGFR (T790M/L858R) protein was purchased from Carna Biosciences, Inc..

Measurement of inhibitory activity was carried out as follows. Compounds of the present invention were individually dissolved in dimethyl sulfoxide (DMSO) and serial dilutions of these compounds were prepared using DMSO. Then, a solution containing substrate peptide (final concentration: 250 nM), magnesium chloride (final concentration: 10 mM), manganese chloride (final concentration: 10 mM) and ATP (final concentration: 1 μM) in kinase reaction buffer (Carna Biosciences, Inc.) were mixed with compound dilutions (final concentration of DMSO after kinase reaction: 2.5%) or DMSO (final concentration: 2.5%). Thereafter, EGFR (T790M/L858R) protein was added and the mixtures were incubated at 25° C. for 120 minutes to carry out the kinase reaction. EDTA was then added thereto to a final concentration of 24 mM to thereby complete the reaction. Phosphorylated tyrosine detection solution containing europium (Eu) labeled PT66 antibody against phosphorylated tyrosine (PerkinElmer, Inc.) and SureLight APC-SA (PerkinElmer, Inc.) was added to the reaction mixtures, and the mixtures were allowed to stand at room temperature for two hours or more. For background, a sample to which EDTA was added prior to the addition of EGFR (T790M/L858R) protein was incubated at 25° C. for 120 minutes using DMSO instead of a DMSO solution of the compound. Also, the detection solution was added to this sample, and the mixture was left to stand for two hours or more. Finally, for all test samples, the fluorescence intensity was measured during irradiation with excitation light having a wavelength of 337 nm at two wavelengths of 620 nm and 665 nm using a PHERAstar FS instrument (BMG LABTECH) and obtained as data ratio of fluorescence intensities at two wavelengths.

When analyzing the measured data, the ratio of the fluorescence intensities of the sample in which the kinase reaction was carried out with the addition of DMSO at a final concentration of 2.5% was set as the degree of phosphorylation inhibition of 0%, and the ratio of the background fluorescence intensities was set as the degree of phosphorylation inhibition of 100%. Based on the above degrees, the degrees of inhibition of phosphorylation (%) were calculated for the samples to which were added various concentrations of compound solutions. Finally, for each compound, curves were plotted for the resulting degrees of reaction inhibition (%), at the respective concentrations, and the IC ₅₀ value (nM) was determined, which is the concentration of the compound at which phosphorylation by EGFR (T790M/L858R) is inhibited by 50 % using Xlfit curve regression analysis software (IDBS).

2) Measurement of EGFR (d746-750/T790M)-kinase inhibitory activity

The inhibitory activities of the compounds of the present invention against EGFR (d746-750/T790M) kinase activity were measured.

The substances, measurement method, and data analysis method were actually the same as those mentioned in the above description regarding "EGFR (T790M/L858R)-kinase inhibitory activity measurement", except that the purified recombinant human EGFR (d746-750/ T790M) protein purchased from Carna Biosciences, Inc. was used as the kinase protein and measured with a final ATP concentration of 1.5 μM. Finally, the IC ₅₀ value (nM) of each compound with respect to EGFR (d746-750/T790M) was determined by data analysis.

3) Measurement of EGFR (L858R)-kinase inhibitory activity

The inhibitory activities of the compounds of the present invention against EGFR (L858R) kinase activity were measured.

Substances, measurement method and data analysis method were actually the same as those mentioned in the above description regarding "Measurement of EGFR (T790M/L858R)-kinase inhibitory activity", except that the purified recombinant human EGFR (L858R)-protein , purchased from Carna Biosciences, Inc., was used as the kinase protein and measured with a final ATP concentration of 4 μM. Finally, the IC ₅₀ value (nM) of each compound, with respect to EGFR (L858R), was determined by data analysis.

4) Measurement of EGFR (d746-750)-kinase inhibitory activity

The inhibitory activities of the compounds of the present invention against EGFR (d746-750) kinase activity were measured.

The substances, measurement method and data analysis method were actually the same as those mentioned in the above description regarding "EGFR (T790M/L858R)-kinase inhibitory activity measurement", except that the purified recombinant human EGFR (b746-750) α-protein purchased from Carna Biosciences, Inc. was used as the kinase protein, measurement was performed with a final ATP concentration of 5 μM, and incubation for the kinase reaction was performed for 90 minutes. Finally, the IC ₅₀ value (nM) of each compound with respect to EGFR (d746-750) was determined by data analysis.

5) Measurement of EGFR (WT)-kinase inhibitory activity

The inhibitory activities of the compounds of the present invention against EGFR (WT) kinase activity were measured.

The substances, measurement method, and data analysis method were actually the same as those indicated in the above description regarding "EGFR (T790M/L858R)-kinase inhibitory activity measurement", except that the kinase protein obtained by expression of the cytoplasmic domain of human EGFR (WT) with its N-terminus FLAG fused to Sf9 insect cells using a baculovirus expression system and then purified with an anti-FLAG agarose antibody (Sigma Aldrich) was used as the kinase protein, the final the substrate protein concentration was 500 nM and the final ATP concentration was 4.7 μM. Finally, the IC ₅₀ value (nM) of each compound, with respect to EGFR (WT), was determined by data analysis.

Table 1 presents test results using 30 different EGFRs.

The compounds of the present invention were confirmed to have strong inhibitory activities not only against EGFR (L858R) and EGFR (d746-750), but also against EGFR (T790M/L858R) and EGFR (d746-750/T790M). It is also confirmed that their inhibitory activities against EGFR (WT) are lower than those against the above EGFR proteins.

In contrast, the compound of Comparative Example 1, which is a compound having a structure similar to that of the compound of the present invention, was confirmed to show almost no inhibitory activity against these EGFR kinases.

Table 1 Type EGFR EGFR (T790M/
L858R) EGFR (d746-750/
T790M) EGFR (L858R) EGFR (d746-750) EGFR (WT) Example 1 <0.50 <0.15 <0.50 <0.15 0.70 2 77 thirty 230 78 1400 3 6.1 1.8 15 7.6 110 4 0.40 <0.50 0.63 0.45 6.0 5 <0.15 <0.15 <0.15 <0.15 0.78 6 0.24 <0.15 0.45 0.24 4.9 7 1.0 0.53 1.7 1.2 12 8 <0.50 <0.15 <0.50 <0.15 1.0 9 2.1 0.45 5.9 2.3 52 10 1.5 0.61 2.6 1.8 22 eleven 0.76 0.32 1.4 0.88 12 12 0.66 0.29 1.2 0.75 eleven 13 2.1 1.1 2.8 2.1 24 14 0.24 <0.50 0.32 0.19 2.9 15 1.6 1.2 1.3 1.4 7.9 16 eleven 3.0 39 15 480 17 23 eleven 50 39 580 18 4.7 1.3 12 4.5 68 19 0.26 0.28 0.27 0.42 2.2 20 3.7 2.4 4.1 2.9 28 21 0.60 0.34 1.1 0.72 7.8 22 0.35 0.23 0.37 0.33 5.5 23 0.44 0.30 0.63 0.29 7.3 24 0.83 0.31 1.1 0.85 8.6 25 36 eleven 100 59 26 1.0 0.31 1.7 1.1 27 0.34 0.25 0.35 0.38 28 0.30 <0.50 0.49 0.27 29 14 8.4 19 21 Comparative Example 1 >5000 >5000 >5000 1500 >5000

Test Example 2: Test for growth inhibitory activity against cell lines expressing wild-type and mutated EGFR ( in vitro )

Human small cell lung cancer cell line NCI-H1975 expressing EGFR (T790M/L858R), (2) human non-small cell lung cancer cell line HCC827 expressing EGFR (d746-750), and (3) human epithelioid cancer cell line A431 expressing EGFR (WT) were each suspended in ATCC recommended complete medium. Cell suspensions were seeded in each well of a 384-well flat microplate or a 96-well flat plate and cultured in an incubator containing 5% carbon dioxide gas at 37° C. for one day. Each compound of the present invention was dissolved in DMSO and serial dilutions of test compounds were prepared using DMSO (to a concentration 500 times the final concentration). The DMSO solution of the test compound or DMSO was diluted with complete culture medium and the resulting solution was added to each well of the cell culture plate so that the final concentration of DMSO was 0.2%. The cells were then cultured in an incubator containing 5% carbon dioxide gas at 37°C for three days. The number of cells was measured before the addition of the DMSO solution of the test compound, and after the addition and culture using the CellTiter-Glo Assay® (manufactured by Promega) according to the protocol recommended by Promega.

For each cell, the degree of inhibition of cell growth in the wells to which the test compounds were added at various concentrations was calculated by the following formula. Then, the obtained degrees of inhibition (%) at the respective concentrations were plotted for each test compound, and the GI ₅₀ value (nM), which is the concentration of the test compound at which cell growth can be inhibited by 50%, was determined using the software for XLfit Curve Regression Analysis (IDBS).

Degree of cell growth inhibition (%)=(C-T)/(C-C0)×100

T: luminescence intensity in a well cultured for three days after the addition of the test compound solution

C: luminescence intensity in a well cultured for three days after the addition of DMSO

C0: luminescence intensity in the well before addition of the test compound solution

The results are presented in table 2.

The compounds of the present invention were confirmed to have strong growth inhibitory effects not only on cells expressing EGFR (d746-750), but also on cells expressing EGFR (T790M/L858R). It is also confirmed that their growth inhibitory effects on EGFR expressing cells (WT) are lower than those of the above EGFR proteins.

table 2 Type EGFR EGFR (T790M/L858R) EGFR (d746-750) EGFR (WT) cell name NCI-H1975 HCC827 A431 Example 1 9.0 1.9 370 2 4200 780 >10000 3 560 16 >10000 4 140 5.5 2400 5 51 8.1 2200 6 54 3.3 520 7 150 5.6 2600 8 800 66 1100 9 770 7.5 4200 10 310 8.1 4400 eleven 260 9.7 3900 12 240 7.9 3500 13 150 13 1200 14 22 2.5 1900 15 85 22 2000 16 600 27 >10000 17 2900 160 >10000 18 160 8.9 4000 19 49 5.6 990 20 310 23 3900 21 56 8.1 2600 22 56 5.7 1500 23 47 3.3 5100 24 110 4.9 3400 25 2500 260 >10000 26 150 18.4 4700 27 110 15 1000 28 48 14 1200 29 1600 89 >10000

Claims (36)

1. The method of obtaining the compound (I-4), including the interaction of the compound (XIX) with a carboxylic acid, an acid chloride or an acid bromide to form a compound (I-4);

Where n means 1; R ₁ means hydrogen or C ₁ -C ₆ alkyl group; R ₉ means CH ₃ ; R ₂ means a group represented by the formula

where R ₃ , R ₄ and R ₅ are the same or different, and each represents a hydrogen atom, a halogen atom, a C ₁ -C ₆ alkyl group, an aminomethyl group which may be substituted by a C ₁ -C ₆ alkyl group, or 1- piperidinomethyl group. 2. The method according to p. 1, in which the compound (XIX) is obtained from compound (VII) by cyclization in the presence of a palladium catalyst, followed by removal of the protective group P ₁ ;

Where P ₁ means the protective group of the amino group; R ₇ and R ₈ each represent a hydrogen atom; And m means 1. 3. The method according to p. 2, where the compound (VII) is obtained from the compound (VI) by bromination

. 4. The method according to p. 3, where the compound (VI) is obtained from the compound (V) by reacting it with quinoline-3-boronic acid in the presence of a palladium-based catalyst

where L ₁ means the leaving group. 5. The method according to p. 4, where the compound (V) is obtained from the compound (IV) by reacting it with ammonia

. 6. The method according to any one of paragraphs. 1-5, where R ₃ , R ₄ and R ₅ each represents a hydrogen atom. 7. The method according to claim 6, wherein the compound (XIX) is reacted with acryloyl chloride to obtain compound (I-4). 8. The method according to any one of paragraphs. 1-7, where R ₁ means a hydrogen atom. 9. The method according to any one of paragraphs. 2-8, where P ₁ means tert-butoxycarbonyl and deprotection of the P ₁ group includes a reaction using an acid as a catalyst. 10. The method according to any one of paragraphs. 2-9, wherein the cyclization in the presence of a palladium-based catalyst involves the use of tetrakis(triphenylphosphine)palladium(0). 11. The method according to any one of paragraphs. 3-10, where the bromination includes the use of N-bromosuccinimide. 12. The method according to any one of paragraphs. 4-11, where interactions with quinoline-3-boronic acid in the presence of a palladium-based catalyst include the use of tetrakis(triphenylphosphine)palladium(0) as the palladium-based catalyst. 13. The method according to any one of paragraphs. 4-12, where L ₁ means a bromine atom or an iodine atom. 14. The method according to p. 13, where L ₁ means an iodine atom. 15. The method according to any one of paragraphs. 1-14, where compound I-4 is obtained, which is 90% or more one of the enantiomers. 16. The method of claim 15 wherein one of the enantiomers is obtained by separating the enantiomers using chiral chromatography. 17. The method of claim 15 or 16 wherein one of the enantiomers is the R -enantiomer. 18. The method of claim 15 or 16 wherein one of the enantiomers is the S -enantiomer.