Substituted quinolinonyl piperazine compounds useful as T cell activators

Abstract

Formula (I): TIFF2023508055000073.tif45150 [wherein R1, R2, R3, R4, R5, R6, and m are as defined herein] or a salt thereof. Also disclosed are methods of using such compounds, and pharmaceutical compositions containing such compounds, to inhibit the activity of one or both of diacylglycerol kinase alpha (DGKα) and diacylglycerol kinase zeta (DGKζ). These compounds are useful in the treatment of proliferative disorders such as viral infections and cancer.

Description

[Technical field]

[0001] (Related Applications) This application claims priority to Indian Provisional Application No. 201911053557, filed December 23, 2019, the entire contents of which are incorporated herein by reference.

[0002] Description of the Invention The present invention generally relates to substituted quinolinonyl compounds that activate T cells, promote T cell proliferation, and / or exhibit anti-tumor activity. Provided herein are substituted quinolinonyl compounds, compositions containing the compounds, and methods of using them. The present invention further relates to pharmaceutical compositions containing at least one compound of the present invention that are useful for treating proliferative disorders (e.g., cancer, and viral infections). [Background technology]

[0003] Human cancers contain numerous genetic and epigenetic alterations, producing neoantigens that can be recognized by the immune system (Sjoblom et al., (2006) Science 314:268-74). The adaptive immune system, consisting of T and B lymphocytes, has the potential to exert potent anticancer effects, and is endowed with a broad capacity and high specificity to respond to a variety of tumor antigens. Furthermore, this immune system has considerable plasticity and memory components. Successfully exploiting all these properties of the adaptive immune system makes immunotherapy unique among all cancer treatment approaches. However, although an endogenous immune response against cancer has been observed in preclinical models and patients, this response is ineffective and established cancers are considered as "self" and tolerated by the immune system. This state of tolerance allows tumors to utilize multiple different mechanisms to actively subvert antitumor immunity. These mechanisms include dysfunctional T cell signaling (Mizoguchi et al., (1992) Science 258:1795-98), suppressive regulatory cells (Facciabene et al., (2012) Cancer Res. 72:2162-71), and the use of endogenous "immune checkpoints" that function to reduce the strength of the adaptive immune response and protect normal tissues from collateral damage by the tumor and escape immune destruction (Topalian et al., (2012) Curr. Opin. Immunol. 24:1-6; Mellman et al., (2011) Nature 480:480-489).

[0004] Diacylglycerol kinase (DGK) is a lipid kinase that mediates the conversion of diacylglycerol to phosphatidic acid, thereby halting T cell function propagating through the TCR signaling pathway. Therefore, DGK functions as an intracellular checkpoint, and inhibition of DGK is expected to promote T cell signaling pathways and T cell activation. Supporting evidence is that knockout mouse models of either DGKα or DGKζ show hyperreactive T cell phenotypes and improved antitumor immune activity (Riese MJ et al., Journal of Biological Chemistry, (2011) 7: 5254-5265; Zha Y et al., Nature Immunology, (2006) 12:1343; Olenchock BA et al., (2006) 11: 1174-81). Furthermore, it has been confirmed that tumor-infiltrating lymphocytes isolated from human renal cell carcinoma patients overexpress DGKα and inhibit T cell function (Prinz, PU et al., J Immunology (2012) 12:5990-6000). Therefore, DGKα and DGKζ are considered as targets in cancer immunotherapy (Riese MJ et al., Front Cell Dev Biol. (2016) 4: 108; Chen, SS et al., Front Cell Dev Biol. (2016) 4: 130; Avila-Flores, A. et al., Immunology and Cell Biology (2017) 95: 549-563; Noessner, E., Front Cell Dev Biol. (2017) 5: 16; Krishna, S., et al., Front Immunology (2013) 4:178; Jing, W. et al., Cancer Research (2017) 77: 5676-5686).

[0005] There is a need for compounds useful as inhibitors of DGKα and / or DGKζ. Additionally, there is a need for compounds useful as inhibitors of DGKα and / or DGKζ that have selectivity over other diacylglycerol kinases, protein kinases, and / or other lipid kinases.

[0006] Thus, safe and effective agents to restore T cell activity, lower the antigen threshold, enhance antitumor function, and / or overcome the suppressive effects of one or more endogenous immune checkpoints (e.g., PD-1, LAG-3, and TGFβ) would be an important addition to the treatment of patients with proliferative disorders (e.g., cancer, as well as viral infections). Summary of the Invention

[0007] The present inventors have found compounds that have activity as inhibitors of one or both of DGKα and DGKζ. Furthermore, the present inventors have found compounds that have activity as inhibitors of one or both of DGKα and DGKζ and have selectivity over other diacylglycerol kinases, protein kinases, and / or other lipid kinases. These compounds provide useful pharmaceutical agents with desirable stability, bioavailability, therapeutic index, and toxicity values ​​important to their druggability.

[0008] The present invention provides substituted quinolinonyl compounds of formula (I), or salts and prodrugs thereof, that are useful as inhibitors of DGKα, DGKζ, or both DGKα and DGKζ.

[0009] The present invention also provides a pharmaceutical composition comprising a compound of formula (I) and / or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable carrier.

[0010] The present invention also provides a method for treating a disease or disorder associated with the activity of DGKα, DGKζ, or both DGKα and DGKζ, comprising administering to a mammalian patient a compound of formula (I) and / or a pharma- ceutical acceptable salt thereof.

[0011] The present invention also provides processes and intermediates for preparing the compounds of formula (I) and / or salts thereof.

[0012] The present invention also provides compounds of formula (I) and / or pharma- ceutically acceptable salts thereof for use in therapy.

[0013] The invention also provides the use of a compound of formula (I) and / or a pharma- ceutically acceptable salt thereof for the manufacture of a medicament for the treatment of proliferative disorders (eg cancer and viral infections).

[0014] The compounds of formula (I) and compositions containing the compounds of formula (I) can be used to treat, prevent, or cure viral infections and various proliferative disorders, such as cancer. Pharmaceutical compositions containing these compounds are useful for treating, preventing, or inhibiting the progression of diseases or disorders in various therapeutic areas, such as viral infections and cancer.

[0015] These and other features of the present invention are set forth in the broader disclosure. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0016] The first aspect of the present invention relates to a compound of formula (I): [ka] [In formula: R1 is H, F, Cl, Br, -CN, C 1-3 Alkyl (0 to 4 R 1a ), C 3-4 Cycloalkyl (0 to 4 R 1a ), C 1-3 Alkoxy (0 to 4 R 1asubstituted with -C(O)NR a R a , -NR a R a , -S(O) n R e , or -P(O)R e R e and; Each R 1a are independently F, Cl, -CN, -OH, -OCH3, or -NR a R a and; Each R a are independently H or C 1-3 is alkyl; Each R e is independently 3-4 Cycloalkyl or C 1-3 Alkyl (0 to 4 R 1a ) and R2 is H, C 1-3 Alkyl (0 to 4 R 2a ), C 2-3 Alkenyl (0 to 4 R 2a ), or C 3-4 Cycloalkyl (0 to 4 R 2a ) and Each R 2a are independently F, Cl, -CN, -OH, -O(C 1-2 Alkyl), C 3-4 Cycloalkyl, C 3-4 Alkenyl, or C 3-4 is alkynyl; R3 is H, F, Cl, Br, -CN, C 1-3 Alkyl, C 1-2 Fluoroalkyl, C 3-4 Cycloalkyl, C 3-4 Fluorocycloalkyl, -NO2, or pyridinyl (0 to 2 R 3a ) and Each R 3a , halo, -CN, C 1-3 Alkyl, or C 1-3 is alkoxy; R4 is -CH2R 4a, -CH2CH2R 4a , -CHCHR 4a R 4d , -CHR 4a R 4b , or -CR 4a R 4b R 4c and; R 4a and R 4b is, independently, (I C 1-6 Alkyl (F, Cl, -CN, -OH, -OCH3, -SCH3, C 1-3 Fluoroalkoxy, -NR a R a , -S(O)2R e , or -NR a S(O)2R e substituted with 0 to 4 substituents independently selected from the group consisting of: (ii) C 3-6 cycloalkyl, heterocyclyl, phenyl, or heteroaryl, each of which is F, Cl, Br, -CN, -OH, C 1-6 Alkyl, C 1-3 Fluoroalkyl, C 1-4 Hydroxyalkyl, -(CH2) 1-2 O(C 1-3 Alkyl), C 1-4 Alkoxy, -O(C 1-4 Hydroxyalkyl), -O(CH) 1-3 O(C 1-3 Alkyl), C 1-3 Fluoroalkoxy, -O(CH) 1-3 NR c R c , -OCH2CH=CH2, -OCH2C≡CH, -C(O)(C 1-4 alkyl), -C(O)OH, -C(O)O(C 1-4 alkyl), -NR c R c , -NR a S(O)2(C 1-3 alkyl), -NR a C(O)(C 1-3 alkyl), -NR a C(O)O(C 1-4 alkyl), -P(O)(C1-3 alkyl)2, -S(O)2(C 1-3 alkyl), -O(CH2) 1-2 (C 3-6 Cycloalkyl), -O(CH2) 1-2 (morpholinyl), cyclopropyl, cyanocyclopropyl, methylazetidinyl, acetylazetidinyl, (tert-butoxycarbonyl)azetidinyl, triazolyl, tetrahydropyranyl, morpholinyl, thiophenyl, methylpiperidinyl, and R d or is substituted with 0 to 4 substituents independently selected from (iii) C 1-4 Alkyl, C 3-6 cycloalkyl, heterocyclyl, aryl, and heteroaryl, and the ring group is selected from F, Cl, Br, -OH, -CN, C 1-6 Alkyl, C 1-3 Fluoroalkyl, C 1-3 Alkoxy, C 1-3 Fluoroalkoxy, -OCH2CH=CH2, -OCH2C≡CH, -NR c R c , -NR a S(O)2(C 1-3 alkyl), -NR a C(O)(C 1-3 alkyl), -NR a C(O)O(C 1-4 alkyl), and C 3-6 substituted with 0-3 substituents independently selected from cycloalkyl; Or R 4a and R 4b together with their attached carbon atoms, C 3-6 cycloalkyl or 3- to 6-membered heterocyclyl, each of which is 0 to 3 R f Replaced with; Each R f are independently F, Cl, Br, -OH, -CN, C 1-6 Alkyl, C 1-3 Fluoroalkyl, C 1-3 Alkoxy, C 1-3Fluoroalkoxy, -OCH2CH=CH2, -OCH2C≡CH, -NR c R c , or C 3-6 A ring group selected from cycloalkyl, 3- to 6-membered heterocyclyl, phenyl, monocyclic heteroaryl, and bicyclic heteroaryl, each of which is selected from F, Cl, Br, -OH, -CN, C 1-6 Alkyl, C 1-3 Fluoroalkyl, C 1-3 Alkoxy, C 1-3 Fluoroalkoxy, and -NR c R c substituted with 0 to 3 substituents independently selected from R 4c is C 1-6 Alkyl or C 3-6 cycloalkyl, each of which is F, Cl, -OH, C 1-2 Alkoxy, C 1-2 substituted with 0 to 4 substituents independently selected from: -fluoroalkoxy, -CN; R 4d is -OCH3; Each R c are independently H or C 1-2 is alkyl; R d is phenyl and is substituted with 0-1 substituents selected from F, Cl, -CN, -CH3, and -OCH3; Each R5 is independently F, Cl, -CN, -OH, C 1-6 Alkyl (0 to 4 R g ), C 1-3 Alkoxy (0 to 4 R g ), C 2-4 Alkenyl (0 to 4 R g ), C 2-4 Alkynyl (0 to 4 R g ), C 3-4 Cycloalkyl (0 to 4 R g phenyl (substituted with 0 to 4 R g substituted with 0 to 3 R gsubstituted with 0 to 4 R g substituted with -(CH2) 1-2 (0 to 4 R g heterocyclyl substituted with -(CH2) 1-2 NR c C(O)(C 1-4 alkyl), -(CH2) 1-2 NR c C(O)O(C 1-4 alkyl), -(CH2) 1-2 NR c S(O)2(C 1-4 alkyl), -C(O)(C 1-4 alkyl), -C(O)OH, -C(O)O(C 1-4 alkyl), -C(O)O(C 3-4 Cycloalkyl), -C(O)NR a R a or -C(O)NR a (C 3-4 cycloalkyl) or two R5 are attached to the same carbon atom to form =O; Each R g are independently F, Cl, -CN, -OH, C 1-3 Alkoxy, C 1-3 Fluoroalkoxy, -O(CH2) 1-2 O(C 1-2 Alkyl), C 3-5 Cycloalkyl, or -NR c R c and; Each R6 is H, F, Cl, -CN, -CH3, -CH2F, -CHF2, -CF3, or -OCH3; m is 0, 1, 2, or 3; and n is 0, 1, or 2] The present invention provides at least one compound represented by the formula:

[0017] In one embodiment, the compound of formula (I) or a salt thereof, wherein R1 is H, F, Cl, Br, -CN, C 1-3 Alkyl (0 to 4 R 1a substituted with cyclopropyl (substituted with 0 to 3 R 1a), C 1-3 Alkoxy (0 to 3 R 1a substituted with -C(O)NR a R a , -NR a R a , -S(O) n CH3, or -P(O)(CH3)2; each R 1a is independently F, Cl, or -CN; each R a are independently H or C 1-3 alkyl; R2 is H, C 1-2 Alkyl (0 to 2 R 2a ), or C 2-3 Alkenyl (0 to 2 R 2a (substituted with each R 2a are independently F, Cl, -CN, -OH, -O(C 1-2 Alkyl), cyclopropyl, C 3-4 Alkenyl, or C 3-4 Alkynyl; R3 is H, F, Cl, Br, -CN, C 1-2 Alkyl, C 1-2 Fluoroalkyl, C 3-4 Cycloalkyl, -NO2, or pyridinyl (0 to 2 R 3a substituted with R 4a and R 4b But independently, (i) C 1-4 Alkyl (F, Cl, -CN, -OH, -OCH3, -SCH3, C 1-3 Fluoroalkoxy, and -NR a R a (ii) C 3-6 cycloalkyl, heterocyclyl, phenyl, or heteroaryl, each of which is selected from F, Cl, Br, -CN, -OH, C 1-6 Alkyl, C 1-3 Fluoroalkyl, -CH2OH, -(CH2) 1-2 O(C 1-2 Alkyl), C 1-4 Alkoxy, -O(C 1-4 Hydroxyalkyl), -O(CH) 1-2 O(C1-2 Alkyl), C 1-3 Fluoroalkoxy, -O(CH) 1-2 NR c R c , -OCH2CH=CH2, -OCH2C≡CH, -C(O)(C 1-4 alkyl), -C(O)OH, -C(O)O(C 1-4 alkyl), -NR c R c , -NR a S(O)2(C 1-3 alkyl), -NR a C(O)(C 1-3 alkyl), -NR a C(O)O(C 1-4 alkyl), -P(O)(C 1-2 alkyl)2, -S(O)2(C 1-3 alkyl), -O(CH2) 1-2 (C 3-4 Cycloalkyl), -O(CH2) 1-2 (morpholinyl), cyclopropyl, cyanocyclopropyl, methylazetidinyl, acetylazetidinyl, (tert-butoxycarbonyl)azetidinyl, triazolyl, tetrahydropyranyl, morpholinyl, thiophenyl, methylpiperidinyl, and R d or (iii) C 1-3 Alkyl, C 3-6 Cycloalkyl, heterocyclyl, phenyl, and heteroaryl are substituted with one ring group selected from F, Cl, Br, -OH, -CN, C 1-3 Alkyl, C 1-2 Fluoroalkyl, C 1-3 Alkoxy, C 1-2 Fluoroalkoxy, -OCH2CH=CH2, -OCH2C≡CH, -NR c R c , -NR a S(O)2(C 1-3 alkyl), -NR a C(O)(C 1-3 alkyl), -NR a C(O)O(C 1-4 alkyl), and C 3-4cycloalkyl; or R 4a and R 4b together with the carbon atoms to which they are attached, C 3-6 cycloalkyl or 3- to 6-membered heterocyclyl, each of which is 0 to 3 R f Each R f are independently F, Cl, Br, -OH, -CN, C 1-4 Alkyl, C 1-2 Fluoroalkyl, C 1-3 Alkoxy, C 1-2 Fluoroalkoxy, -OCH2CH=CH2, -OCH2C≡CH, -NR c R c , or C 3-6 A ring group selected from cycloalkyl, 3- to 6-membered heterocyclyl, phenyl, monocyclic heteroaryl, and bicyclic heteroaryl, each of which is selected from F, Cl, Br, -OH, -CN, C 1-4 Alkyl, C 1-2 Fluoroalkyl, C 1-3 Alkoxy, C 1-2 Fluoroalkoxy, and -NR c R c substituted with 0 to 3 substituents independently selected from the group consisting of: 4c C 1-4 Alkyl or C 3-6 cycloalkyl, each of which is F, Cl, -OH, C 1-2 Alkoxy, C 1-2 substituted with 0 to 4 substituents independently selected from: -fluoroalkoxy, -CN; each R5 is independently selected from F, -CN, -OH, C 1-5 Alkyl (0 to 4 R g ), C 1-2 Alkoxy (0 to 3 R g ), C 2-3 Alkenyl (0 to 4 R g ), C 2-3 Alkynyl (0 to 4 R g ), C 3-4 Cycloalkyl (0 to 4 R gphenyl (substituted with 0 to 3 R g substituted with 0 to 3 R g substituted with 0 to 3 R g substituted with -(CH2) 1-2 (0 to 4 R g heterocyclyl substituted with -(CH2) 1-2 NR c C(O)(C 1-4 alkyl), -(CH2) 1-2 NR c C(O)O(C 1-4 alkyl), -(CH2) 1-2 NR c S(O)2(C 1-4 alkyl), -C(O)(C 1-4 alkyl), -C(O)OH, -C(O)O(C 1-4 alkyl), -C(O)O(C 3-4 Cycloalkyl), -C(O)NR a R a or -C(O)NR a (C 3-4 cycloalkyl) or two R5 are attached to the same carbon atom to form =O; each R6 is H, F, or -CH3; and m is 0, 1, 2, or 3, or a salt thereof.

[0018] In one embodiment, there is provided a compound of formula (I) or a salt thereof, wherein: R1 is H, F, -CN, or -OCH3; R2 is H, -CH3, -CH2CN, -CH2CH2F, or -CH2CH=CH2; R3 is H, -CN, -CH2OH, -C(O)OCH2CH3, -NO2, or pyridinyl; and R4 is -CH2R 4a or -CHR 4a R 4b ;R 4a is phenyl, naphthalenyl, or indolyl, each of which is substituted with 0 to 2 substituents independently selected from F, -CH3, -CH2CH3, and -OCH3; R 4bis phenyl or fluorophenyl; each R5 is -CH3 or two R5 are attached to the same carbon atom to form =O; and m is 0, 1, or 2, or a salt thereof.

[0019] In one embodiment, the compound of formula (I) or a salt thereof, wherein R1 is H, F, Cl, Br, -CN, C 1-3 Alkyl (0 to 4 R 1a substituted with cyclopropyl (substituted with 0 to 3 R 1a ), C 1-3 Alkoxy (0 to 3 R 1a substituted with -C(O)NR a R a , -NR a R a , -S(O) n The present invention provides a compound or a salt thereof, wherein R1 is H, F, Cl, Br, -CN, -CH3, -CHF2, -CF3, cyclopropyl, or -OCH3. The present invention also provides a compound or a salt thereof, wherein R1 is H, F, -CN, or -OCH3. The present invention also provides a compound or a salt thereof, wherein R1 is H, F, -CN, or -OCH3. The present invention also provides a compound or a salt thereof, wherein R1 is H, F, -CN, or -OCH3.

[0020] In one embodiment, the compound of formula (I) or a salt thereof, wherein R2 is H, C 1-2 Alkyl (0 to 2 R 2a ), or C 2-3 Alkenyl (0 to 2 R 2a or a salt thereof. Included in this embodiment are compounds where R2 is H, -CH3, -CH2CN, -CH2CH2F, or -CH2CH=CH2. Included in this embodiment are compounds where R2 is H, -CH3, -CH2CN, or -CH2CH2F.

[0021] In one embodiment, the compound of formula (I) or a salt thereof, wherein R3 is H, F, Cl, Br, -CN, C 1-2 Alkyl, C 1-2 Fluoroalkyl, C 3-4Cycloalkyl, -NO2, or pyridinyl (0 to 2 R 3a or a salt thereof. Included in this embodiment are compounds where R3 is H, -CN, -CH2OH, -C(O)OCH2CH3, -NO2, or pyridinyl. Also included in this embodiment are compounds where R3 is H, -CN, -CH2OH, or -NO2.

[0022] In one embodiment, the compound of formula (I) or a salt thereof, wherein R4 is -CH2R 4a or -CH2CH2R 4a In one embodiment, R4 is -CH2R 4a or -CD2R 4a Compounds are included in the present invention, In one embodiment, in a compound of formula (I) or a salt thereof, R4 is -CH2R 4a In another embodiment, R4 is -CD2R 4a Compounds are included in the present invention, In one embodiment, in a compound of formula (I) or a salt thereof, R4 is -CHR 4a R 4b or -CR 4a R 4b R 4c or a salt thereof. In one embodiment, in a compound of formula (I) or a salt thereof, R4 is -CHR 4a R 4b or a salt thereof. In one embodiment, in a compound of formula (I) or a salt thereof, R4 is -CH2R 4a or -CHR 4a R 4b or a salt thereof.

[0023] In one embodiment, a compound of formula (I) or a salt thereof, wherein: m is 1, 2, or 3; each R5 is independently F, -CN, -OH, C 1-5 Alkyl (0 to 4 Rg ), C 1-2 Alkoxy (0 to 3 R g ), C 2-3 Alkenyl (0 to 4 R g ), C 2-3 Alkynyl (0 to 4 R g ), C 3-4 Cycloalkyl (0 to 4 R g phenyl (substituted with 0 to 3 R g substituted with 0 to 3 R g substituted with 0 to 3 R g substituted with -(CH2) 1-2 (0 to 4 R g heterocyclyl substituted with -(CH2) 1-2 NR c C(O)(C 1-4 alkyl), -(CH2) 1-2 NR c C(O)O(C 1-4 alkyl), -(CH2) 1-2 NR c S(O)2(C 1-4 alkyl), -C(O)(C 1-4 alkyl), -C(O)OH, -C(O)O(C 1-4 alkyl), -C(O)O(C 3-4 Cycloalkyl), -C(O)NR a R a or -C(O)NR a (C 3-4 In one embodiment, each R is independently selected from the group consisting of F, -CN, -OH, -C, -Cycloalkyl ... 1-2 Alkyl (0 to 3 R g ), C 1-2 Alkoxy (0 to 3 R g ), C 3-4 Cycloalkyl (0 to 2 R g substituted with -C(O)(C 1-4 alkyl), -C(O)OH, -C(O)O(C 1-2alkyl), or -C(O)NR a R a or two R5 are attached to the same carbon atom to form =O. Also included in this embodiment are compounds where each R5 is -CH3 or two R5 are attached to the same carbon atom to form =O.

[0024] In one embodiment, there is provided a compound of formula (I) or a salt thereof, wherein m is 2 and two R5 are attached to the same carbon atom to form =O. In one embodiment, there is provided a compound of formula (I) or a salt thereof, wherein m is 0. In one embodiment, there is provided a compound of formula (I) or a salt thereof, wherein m is 1, 2, or 3. Within this embodiment are compounds where m is 1 or 2. Within this embodiment are also compounds where m is 1. In one embodiment, there is provided a compound of formula (I) or a salt thereof, wherein m is 2 or 3. Within this embodiment, there is included a compound wherein m is 2. In one embodiment, there is provided a compound of formula (I) or a salt thereof, wherein m is 3.

[0025] In one embodiment, a compound of formula (I) or a salt thereof has the structural formula (II): [ka] [In the formula: R 5a , R 5b , R 5c , and R 5d One, two, or three of 5a , R 5b , R 5c , and R 5d the remainder being hydrogen. or a salt thereof. This embodiment includes compounds where each R5 is independently -CN, -CH3, -CH2CH3, -CH(CH3)2, -CHC(CH3)2, -CH2F, -C(CH3)2F, -CF(CH3)CH(CH3)2, -CH2OH, -C(CH3)2OH, -C(CH3)(OH)CH(CH3)2, -CHOCH3, -C(O)C(CH3)2, -C(O)OH, -C(O)OCH3, -C(O)OC(CH3)2, -C(O)NH2, -C(O)NH(cyclopropyl), -C(O)O(cyclopropyl), cyclopropyl, phenyl, methyloxadiazolyl, or methylpyridinyl.

[0026] In one embodiment, a compound of formula (I) or a salt thereof has structural formula (III): [ka] The present invention provides a compound represented by the formula:

[0027] In one embodiment, there is provided a compound of formula (III) or a salt thereof, wherein R 5a is -CH3, and R 5c is -CH3, or a salt thereof. In one embodiment, there is provided a compound of formula (III) or a salt thereof, wherein R 5a is -CH3, and R 5c is -CH2CH3, or a salt thereof. In one embodiment, there is provided a compound of formula (III) or a salt thereof, wherein R 5a is -CH2CH3, and R 5c is -CH3, or a salt thereof. In one embodiment, there is provided a compound of formula (III) or a salt thereof, wherein R 5a is -CH2CH3, and R 5c is -CH2CH3, or a salt thereof. In one embodiment, there is provided a compound of formula (III) or a salt thereof, wherein R 5a is -CH3, and R5c is -CHOH, or a salt thereof. In one embodiment, there is provided a compound of formula (III) or a salt thereof, wherein R 5a is -CH3 and R 5c is -CH2OCH3, or a salt thereof.

[0028] In one embodiment, the compound of formula (I) or the compound of formula (III) or a salt thereof has the following structure: [ka] Available in.

[0029] In one embodiment, the compound of formula (I) or a salt thereof, wherein R4 is: (i) [ka] Is it; (ii) [ka] or (iii) [ka] or a salt thereof. Within this embodiment are compounds where R1 is H, Br, -CN, or -OCH3; and R2 is -CH3.

[0030] In one embodiment, the compound of formula (I) or a salt thereof, wherein R4 is: [ka] or a salt thereof. Included in this embodiment are compounds where R1 is H, Br, -CN, or -OCH3; and R2 is -CH3. Also included in this embodiment are compounds where R1 is -CN; and R2 is -CH3.

[0031] In one embodiment, the compound of formula (I) or a salt thereof is 6-Fluoro-4-{4-[(3-fluorophenyl)methyl]piperazin-1-yl}-1-methyl-3-nitro-1,2-dihydroquinolin-2-one (1); 6-Fluoro-4-{4-[(3-fluoro-5-methylphenyl)methyl]piperazin-1-yl}-1-methyl-3-(pyridin-4-yl)-1,2-dihydroquinolin-2-one (2); 4-(4-((1H-indol-4-yl)methyl)piperazin-1-yl)-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one (3); 4-{4-[(1-ethyl-1H-indol-4-yl)methyl]piperazin-1-yl}-6-fluoro-1-methyl-3-nitro-1,2-dihydroquinolin-2-one (4); 4-(4-benzyl-3-oxopiperazin-1-yl)-6-fluoro-1-methyl-3-nitro-1,2-dihydroquinolin-2-one (5); 4-[(2S,5R)-4-[bis(4-fluorophenyl)methyl]-2,5-dimethylpiperazin-1-yl]-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbonitrile (6); 4-[(2S,5R)-4-[bis(4-fluorophenyl)methyl]-2,5-dimethylpiperazin-1-yl]-1-methyl-2-oxo-1,2-dihydroquinoline-6-carbonitrile (7); 6-Methoxy-1-methyl-4-{4-[(naphthalen-1-yl)methyl]piperazin-1-yl}-3-nitro-1,2-dihydroquinolin-2-one (8); 4-(4-benzhydrylpiperazin-1-yl)-1-methylquinolin-2(1H)-one (9); 4-[4-(diphenylmethyl)piperazin-1-yl]-3-(hydroxymethyl)-1-methyl-1,2-dihydroquinolin-2-one (11); Ethyl 4-{4-[(2-methoxynaphthalen-1-yl)methyl]piperazin-1-yl}-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (12); 4-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-3-nitroquinolin-2(1H)-one (13); 4-{4-[bis(4-fluorophenyl)methyl]piperazin-1-yl}-1-(2-fluoroethyl)-3-nitro-1,2-dihydroquinolin-2-one (14); 2-(4-{4-[bis(4-fluorophenyl)methyl]piperazin-1-yl}-3-nitro-2-oxo-1,2-dihydroquinolin-1-yl)acetonitrile (15); or 4-{4-[bis(4-fluorophenyl)methyl]piperazin-1-yl}-3-nitro-1-(prop-2-en-1-yl)-1,2-dihydroquinolin-2-one (16) or a salt thereof.

[0032] The present invention may be embodied in other specific forms without departing from its essence or essential characteristics. The present invention includes any and all combinations of the aspects and / or embodiments of the present invention described herein. It is understood that any and all embodiments of the present invention may be combined with any and all other embodiments to describe further embodiments. It is also understood that individual elements of the embodiments are meant to be combined with any and all other elements from any of the embodiments to describe further embodiments.

[0033] definition The features and advantages of the present invention may be more readily understood by those skilled in the art upon reading the following detailed description. It is understood that, for clarity, certain features of the invention that are described in the context of another embodiment may be combined to form a single embodiment. Conversely, various features of the invention that are described in the context of a single embodiment for brevity may also be combined to form subcombinations thereof. The embodiments identified herein as exemplary or preferred are intended to be illustrative and not limiting.

[0034] Unless otherwise stated herein, references in the singular may also include the plural, for example, "a" and "an" may refer to either one, or one or more.

[0035] As used herein, the phrase "compound and / or salt thereof" refers to at least one compound, at least one salt of a compound, or a combination thereof. For example, a compound of formula (I) and / or salt thereof includes a compound of formula (I); two compounds of formula (I); a salt of a compound of formula (I); a compound of formula (I) and one or more salts of a compound of formula (I); and a salt of two or more compounds of formula (I).

[0036] Unless otherwise specified, any atom with unsatisfied valences is assumed to include enough hydrogen atoms to satisfy the valences.

[0037] The definitions set forth herein take precedence over definitions set forth in any patents, patent applications, and / or published patent applications incorporated herein by reference.

[0038] Listed below are definitions of various terms used to describe this invention. These definitions apply to the terms as they are used throughout the specification (unless otherwise limited in specific instances) either individually or as part of a larger group.

[0039] Throughout the specification, groups and substituents may be chosen by one skilled in the art to provide stable moieties and compounds.

[0040] According to the practice used in the art, [ka] is used in structural formulas herein to represent the bond that is the point of attachment of a moiety or substituent to a core or backbone structure.

[0041] The terms "halo" and "halogen" as used herein refer to F, Cl, Br, and I.

[0042] The term "cyano" refers to the group --CN.

[0043] The term "amino" refers to the group -NH2. The term "oxo" refers to the group =O.

[0044] The term "alkyl" as used herein refers to both branched and straight chain saturated aliphatic hydrocarbon groups, for example, having 1 to 12 carbon atoms, 1 to 6 carbon atoms, and 1 to 4 carbon atoms. Examples of alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (e.g., n-propyl and i-propyl), butyl (e.g., n-butyl, i-butyl, sec-butyl, and t-butyl), and pentyl (e.g., n-pentyl, isopentyl, neopentyl), n-hexyl, 2-methylpentyl, 2-ethylbutyl, 3-methylpentyl, and 4-methylpentyl. When a number appears in subscript after the symbol "C," the subscript more specifically qualifies the number of carbon atoms that a particular group may contain. For example, "C 1-4 "Alkyl" means straight and branched chain alkyl groups having 1 to 4 carbon atoms.

[0045] As used herein, the term "fluoroalkyl" is meant to include both branched and straight-chain saturated aliphatic hydrocarbon groups substituted with one or more fluorine atoms. For example, 1-4 "Fluoroalkyl" is meant to include C1, C2, C3, and C4 alkyl groups substituted with one or more fluorine atoms. Representative examples of fluoroalkyl groups include, but are not limited to, -CF3 and -CH2CF3.

[0046] The term "hydroxyalkyl" includes both branched and straight chain saturated alkyl groups substituted with one or more hydroxyl groups. For example, "hydroxyalkyl" includes -CHOH, -CHCHOH, and -CH 1-4 Hydroxyalkyl is an example.

[0047] The term "alkenyl" refers to a straight or branched chain hydrocarbon group containing 2 to 12 carbon atoms and at least one carbon-carbon double bond. Examples of such groups include ethenyl or allyl. For example, "C 2-6 "Alkenyl" refers to a straight or branched chain alkenyl group having 2 to 6 carbon atoms.

[0048] The term "alkynyl" refers to a straight or branched chain hydrocarbon group containing from 2 to 12 carbon atoms and at least one carbon-carbon triple bond. Examples of such groups include ethynyl. For example, "C 2-6 "Alkynyl" refers to a straight or branched chain alkynyl group having 2 to 6 carbon atoms.

[0049] The term "cycloalkyl" as used herein refers to a group derived from a non-aromatic monocyclic or polycyclic hydrocarbon molecule by removing one hydrogen atom from a saturated ring carbon atom. Representative examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclopentyl, and cyclohexyl. When a number appears as a subscript after the symbol "C," the subscript more specifically qualifies the number of carbon atoms that a particular cycloalkyl group may contain. For example, "C3-6 "Cycloalkyl" means a cycloalkyl group having 3 to 6 carbon atoms.

[0050] As used herein, the term "fluorocycloalkyl" is meant to include cycloalkyl groups substituted with one or more fluorine atoms.

[0051] The term "alkoxy" as used herein refers to an alkyl group attached to the parent molecular moiety through an oxygen atom, such as a methoxy group (-OCH). 1-3 "Alkoxy" means an alkoxy group having 1 to 3 carbon atoms.

[0052] The terms "fluoroalkoxy" and "-O(fluoroalkyl)" refer to a fluoroalkyl group as described above attached via an oxygen linkage (-O-). For example, 1-4 "Fluoroalkoxy" is intended to include C1, C2, C3, and C4 fluoroalkoxy groups.

[0053] The terms "carbocyclo", "carbocycle" or "carbocyclyl" may be used interchangeably and refer to a cyclic group having at least one saturated or partially saturated non-aromatic ring in which all atoms of all rings are carbon. A carbocyclyl ring can be unsubstituted or can have one or more substituents up to the valence range. Thus, the term includes non-aromatic rings (e.g., cycloalkyl, cycloalkenyl, and cycloalkynyl rings). Examples of bicyclic carbocyclyl groups include indanyl, indenyl, dihydronaphthalenyl, tetrahydronaphthalenyl, hexahydronaphthalenyl, octahydronaphthalenyl, decahydronaphthalenyl, bicycloheptanyl, bicyclooctanyl, and bicyclononanyl.

[0054] The term "aryl" as used herein refers to a group of atoms obtained by removing one hydrogen attached to an aromatic ring from a molecule containing an aromatic carbocyclic ring. Representative examples of aryl groups include, but are not limited to, phenyl and naphthyl. The aryl ring can be unsubstituted or can have one or more substituents, as valences permit.

[0055] The term "benzyl" as used herein refers to a methyl group in which one of the hydrogen atoms has been replaced by a phenyl group. The phenyl ring can be unsubstituted or can have one or more substituents, valences permitting.

[0056] The term "heteroatom" refers to oxygen (O), sulfur (S), and nitrogen (N).

[0057] The terms "heterocyclo", "heterocycle", or "heterocyclyl", which may be used interchangeably, refer to a cyclic group having at least one saturated or partially saturated non-aromatic ring, where one or more of the rings have at least one heteroatom (O, S, or N), and the heteroatom-containing rings preferably have 1 to 3 heteroatoms independently selected from O, S, and / or N. The rings of such groups having heteroatoms may contain 1 or 2 oxygen or sulfur atoms, and / or 1 to 4 nitrogen atoms, provided that the total number of heteroatoms in each ring is 4 or less, and further provided that the ring contains at least one carbon atom. The nitrogen and sulfur atoms may be optionally oxidized, and the nitrogen atom may be optionally quaternized. The heterocyclo group may be attached at any available nitrogen or carbon atom. The heterocyclo ring may be unsubstituted or may have one or more substituents, as valences permit.

[0058] Examples of single-ring heterocyclyl groups include pyrrolidinyl, imidazolinyl, oxazolidinyl, isoxazolinyl, thiazolidinyl, isothiazolidinyl, tetrahydrofuranyl, piperidinyl, piperazinyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolodinyl, 2-oxoazepinyl, azepinyl, 4-piperidonyl, tetrahydropyranyl, morpholinyl, thiamorpholinyl, thiamorpholinyl sulfoxide, thiamorpholinyl sulfone, 1,3-dioxolane, tetrahydro-1,1-dioxothienyl, dihydroisoindolyl, and tetrahydroquinolinyl.

[0059] The term "heteroaryl" refers to substituted and unsubstituted aromatic 5- or 6-membered monocyclic and 9- or 10-membered bicyclic groups having at least one heteroatom (O, S, or N) in at least one ring, the heteroatom-containing ring preferably having 1, 2, or 3 heteroatoms independently selected from O, S, and / or N. Heteroaryl groups in each ring containing a heteroatom may contain 1 or 2 oxygen or sulfur atoms, and / or 1 to 4 nitrogen atoms, provided that the total number of heteroatoms in each ring is 4 or less, and each ring contains at least one carbon atom. The fused rings forming the bicyclic group may be aromatic and contain only carbon atoms. The nitrogen and sulfur atoms may be optionally oxidized, and the nitrogen atoms may be optionally quaternized. Bicyclic heteroaryl groups must contain only aromatic rings. The heteroaryl group may be attached at any available nitrogen or carbon atom. The heteroaryl ring system may be unsubstituted or may have one or more substituents.

[0060] Examples of single-ring heteroaryl groups include pyrrolyl, pyrazolyl, pyrazolinyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, thiadiazolyl, isothiazolyl, furanyl, thiophenyl, oxadiazolyl, pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, and triazinyl.

[0061] Examples of bicyclic heteroaryl groups include indolyl, benzothiazolyl, benzodioxolyl, benzoxazolyl, benzothienyl, quinolinyl, tetrahydroisoquinolinyl, isoquinolinyl, benzimidazolyl, benzopyranyl, indolizinyl, benzofuranyl, chromonyl, coumarinyl, benzopyranyl, cinnolinyl, quinoxalinyl, indazolyl, and pyrrolopyridyl.

[0062] As used herein, the term "pharmaceutical acceptable" refers to compounds, substances, compositions, and / or dosage forms that are suitable, within the scope of ordinary medical judgment, for contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response, or other problem or complication and that afford a reasonable benefit / risk ratio.

[0063] The compounds of formula (I) may form salts, which are also within the scope of the present invention. Unless otherwise specified, a reference to a compound of the invention is understood to include a reference to one or more salts thereof. The term "salt" refers to acid and / or base salts formed with inorganic and / or organic acids and bases. In addition, the term "salt" may include zwitterions (internal salts), for example, when the compound of formula (I) has both a basic moiety (e.g., an amine or a pyridine or imidazole ring) and an acidic moiety (e.g., a carboxylic acid). Pharmaceutically acceptable (i.e., non-toxic and physiologically acceptable) salts are preferred, for example, acceptable metal salts and amine salts, in which the cation does not significantly contribute to the toxicity or biological activity of the salt. However, other salts may also be useful, for example, in isolation or purification steps that may be used in the preparation process, and therefore other salts are considered to be within the scope of the present invention. Salts of compounds of formula (I) may be formed, for example, by reacting a compound of formula (I) with a certain amount of acid or base (e.g., one equivalent) and precipitating the salt in a solvent, for example, or from an aqueous solution followed by lyophilization.

[0064] Examples of acid addition salts include acetates (e.g., acetates prepared from acetic acid or trihaloacetic acids (e.g., trifluoroacetic acid)), adipates, alginates, ascorbates, aspartates, benzoates, benzenesulfonates, hydrogensulfates, borates, butyrates, citrates, camphorates, camphorsulfonates, cyclopentanepropionates, digluconates, dodecylsulfates, ethanesulfonates, fumarates, glucoheptanoates, glycerophosphates, hemisulfates, heptanoates, hexanoates, hydrochlorides (prepared from hydrochloric acid), hydrobromides (prepared from hydrobromide), and the like. salts such as those described herein), hydroiodides, maleates (prepared from maleic acid), 2-hydroxyethanesulfonates, lactates, methanesulfonates (prepared from methanesulfonic acid), 2-naphthalenesulfonates, nicotinates, nitrates, oxalates, pectinates, persulfates, 3-phenylpropionates, phosphates, picrates, pivalates, propionates, salicylates, succinates, sulfates (e.g., prepared from sulfuric acid), sulfonates (e.g., those described herein), tartrates, thiocyanates, toluenesulfonates (e.g., tosylates), undecanoates, and the like.

[0065] Examples of base salts include ammonium salts, alkali metal salts (e.g., sodium, lithium, and potassium salts), alkaline earth metal salts (e.g., calcium and magnesium salts), barium, zinc, and aluminum salts, organic base salts such as organic amines (e.g., trialkylamines (e.g., triethylamine), procaine, dibenzylamine, N-benzyl-β-phenethylamine, 1-ephenamine, N,N'-dibenzylethylene-diamine, dehydroabietylamine, N-ethylpiperidine, benzylamine, dicyclohexylamine), or similar pharma- ceutically acceptable amines, and amino acid salts (e.g., arginine, lysine), and the like. Basic nitrogen-containing groups may be quaternized with reagents such as lower alkyl halides (e.g., methyl, ethyl, propyl, and butyl chlorides, bromides and iodides), dialkyl sulfates (e.g., dimethyl, diethyl, dibutyl, and diamyl sulfates), longer chain halides (e.g., decyl, lauryl, myristyl, and stearyl chlorides, bromides and iodides), aralkyl halides (e.g., benzyl and phenethyl bromide), and other groups. Preferred salts include monohydrochlorides, hydrogen sulfates, methanesulfonates, phosphates, or nitrates.

[0066] The compound of formula (I) may be provided as an amorphous solid or a crystalline solid. The compound of formula (I) may be provided as a solid by lyophilization.

[0067] In addition, solvates (e.g., hydrates) of the compounds of formula (I) should be considered within the scope of the present invention. The term "solvate" refers to a physical association of a compound of formula (I) with one or more organic or inorganic solvent molecules. This physical association includes hydrogen bonding. In some cases, it is possible to isolate the solvate, for example when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid. "Solvate" includes both solution-phase and isolable solvates. Examples of solvates include hydrates, ethanolates, methanolates, isopropanolates, acetonitrile solvates, and ethyl acetate solvates. Methods of solvation are known in the art.

[0068] Various forms of prodrugs are well known in the art and are described in Rautio, J. et al., Nature Review Drug Discovery, 17, 559-587 (2018).

[0069] In addition, the compounds of formula (I) may be isolated and purified following their preparation to obtain compositions containing 99% or greater of the compounds of formula (I) ("substantially pure"), which are then used or formulated as described herein. Such "substantially pure" compounds of formula (I) are also considered herein to be part of the invention.

[0070] By "stable compound" and "stable structure" is intended a compound that is sufficiently robust that it will not decompose upon isolation to a useful degree of purity from a reaction mixture, or upon formulation into an efficacious therapeutic agent. The present invention embodies stable compounds.

[0071] A "therapeutically effective amount" is intended to include an amount of a compound of the invention alone, or in combination with a compound of the claimed invention, or in combination with other active ingredients, that is effective to act as an inhibitor of DGKα and / or DGKζ, or to treat or prevent viral infections and proliferative disorders (e.g., cancer).

[0072] The term "treating" or "treatment" as used herein extends to the treatment of a disease state in a mammal, particularly a human, and includes (a) preventing the mammal from acquiring the disease state, particularly where the mammal is susceptible to, but has not yet been diagnosed as having, the disease state; (b) inhibiting the disease state, i.e., arresting the progression of the disease state; and / or (c) relieving the disease state, i.e., causing regression of the disease state.

[0073] The compounds of the present invention are intended to contain all isotopes of atoms contained in the compounds of the present invention. Isotopes include atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of hydrogen include deuterium (D) and tritium (T). Isotopes of carbon include 13 C and 14 C. Isotopically labeled compounds of the invention can generally be prepared by conventional techniques known to those of skill in the art, or by methods analogous to those described herein, substituting the appropriate isotopically labeled reagent for the non-labeled reagent otherwise used.

[0074] The compounds according to formula (I) and / or pharma- ceutically acceptable salts thereof may be administered by any means appropriate to the condition to be treated, which may depend on the need for site-specific treatment or the amount of compound of formula (I) to be delivered.

[0075] The present invention also includes a class of pharmaceutical compositions comprising a compound of formula (I) and / or a pharma- ceutically acceptable salt thereof; and one or more non-toxic, pharma- cetically acceptable carriers and / or diluents and / or adjuvants (substances collectively referred to herein as "carriers"), and optionally other active ingredients. The compound of formula (I) may be administered by any suitable route, preferably in the form of a pharmaceutical composition adapted for such route, and in a dosage effective for the intended treatment. The compounds and compositions of the present invention may be administered, for example, orally, transmucosally, or parenterally, including intravascularly, intravenously, intraperitoneally, subcutaneously, intramuscularly, and intrasternally, in dosage unit formulations containing pharma- cetically acceptable conventional carriers, adjuvants, and vehicles. For example, the pharmaceutical carrier may include a mixture of mannitol or lactose and microcrystalline cellulose. The mixture may include additional components, such as lubricants (e.g., magnesium stearate) and disintegrants (e.g., crospovidone). The carrier mixture may be filled into gelatin capsules or compressed into tablets.The pharmaceutical composition may be administered, for example, as an oral dosage form or by infusion.

[0076] For oral administration, the pharmaceutical composition may be in the form of, for example, a tablet, capsule, liquid capsule, suspension, or liquid. The pharmaceutical composition is preferably formulated in a dosage unit form with a specific amount of active ingredient. For example, the pharmaceutical composition may be provided as a tablet or capsule containing an amount of active ingredient ranging from about 0.1 to 1000 mg, preferably from about 0.25 to 250 mg, more preferably from about 0.5 to 100 mg. The appropriate daily dose to be administered to a human or other mammal may vary widely depending on the patient's condition and other factors, but can be determined using routine methods.

[0077] Any pharmaceutical composition discussed herein can be delivered orally, for example, via any acceptable and suitable oral formulation. Examples of oral formulations include, but are not limited to, tablets, troches, lozenges, aqueous and oily suspensions, dispersible powders or granules, emulsions, hard and soft capsules, liquid capsules, syrups, and elixirs. Pharmaceutical compositions for oral administration can be prepared according to any method known in the art for preparing pharmaceutical compositions for oral administration. In order to provide a medicament that is easy to swallow, the pharmaceutical compositions described in the present invention can include at least one substance selected from sweeteners, flavoring agents, coloring agents, demulcents, antioxidants, and preservatives.

[0078] Tablets can be prepared, for example, by mixing at least one compound of formula (I) and / or at least one pharma- ceutically acceptable salt with at least one non-toxic, pharma-ceutically acceptable additive suitable for tablet preparation. Examples of additives include, but are not limited to, inert diluents (e.g., calcium carbonate, sodium carbonate, lactose, calcium phosphate, and sodium phosphate), granulating and disintegrating agents (e.g., microcrystalline cellulose, croscarmellose sodium, corn starch, and alginic acid), binding agents (e.g., starch, gelatin, polyvinylpyrrolidone, and gum arabic), and lubricants (e.g., magnesium stearate, stearic acid, and talc). In addition, tablets can be uncoated or coated by known techniques to mask the unpleasant taste of unpleasant drugs or to delay disintegration and absorption of the active ingredient in the digestive tract, thus prolonging the effect of the active ingredient for a longer period of time. Examples of water-soluble taste-masking materials include, but are not limited to, hydroxypropyl methylcellulose and hydroxypropyl cellulose. Examples of time delay materials include, but are not limited to, ethyl cellulose and cellulose acetate butyrate.

[0079] Hard gelatin capsules can be prepared, for example, by mixing at least one compound of formula (I) and / or at least one salt thereof with at least one inert solid diluent (e.g., calcium carbonate, calcium phosphate, and kaolin).

[0080] Soft gelatin capsules can be prepared, for example, by mixing at least one compound of formula (I) and / or at least one pharma- ceutically acceptable salt thereof with at least one water-soluble carrier (e.g., polyethylene glycol) and at least one oil medium (e.g., peanut oil, liquid paraffin, and olive oil).

[0081] Aqueous suspensions can be prepared, for example, by mixing at least one compound of formula (I) and / or at least one pharma- ceutically acceptable salt thereof with at least one excipient suitable for the preparation of aqueous suspensions. Examples of excipients suitable for the preparation of aqueous suspensions include, but are not limited to, suspending agents (e.g., sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, alginic acid, polyvinylpyrrolidone, gum tragacanth, and gum arabic), dispersing or wetting agents (e.g., naturally occurring phosphatides (e.g., lecithin), condensation products of alkylene oxides with fatty acids (e.g., polyoxyethylene stearate), condensation products of ethylene oxide with long chain aliphatic alcohols (e.g., heptadecaethyleneoxycetanol), condensation products of ethylene oxide with fatty acids and Examples of suitable condensation products include condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides (e.g., polyoxyethylene sorbitol monooleate), and condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides (e.g., polyethylene sorbitan monooleate). The aqueous suspension may also contain at least one preservative (e.g., ethyl p-hydroxybenzoate and n-propyl p-hydroxybenzoate), at least one coloring agent, at least one flavoring agent, and / or at least one sweetening agent (e.g., but not limited to, sucrose, saccharin, and aspartame).

[0082] Oil suspensions can be prepared, for example, by suspending at least one compound of formula (I) and / or at least one pharma- ceutically acceptable salt thereof in either vegetable oil (e.g., peanut oil, olive oil, sesame oil, and coconut oil) or mineral oil (e.g., liquid paraffin). Oil suspensions can also include at least one thickening agent (e.g., beeswax, hard paraffin, and cetyl alcohol). To provide a palatable oil suspension, at least one sweetener already described above and / or at least one flavoring agent can be added to the oil suspension. Oil suspensions can further include at least one preservative, including but not limited to, for example, an antioxidant (e.g., butylated hydroxyanisole, and alpha-tocopherol).

[0083] Dispersible powders and granules can be prepared, for example, by mixing at least one compound of formula (I) and / or at least one pharma- ceutically acceptable salt thereof with at least one dispersing agent and / or wetting agent, at least one suspending agent, and / or at least one preserving agent. Suitable dispersing agents, wetting agents, and suspending agents have already been described above. Examples of preservatives include, but are not limited to, antioxidants (e.g., ascorbic acid). In addition, dispersible powders and granules can also include at least one excipient (e.g., but are not limited to, sweeteners, flavoring agents, and coloring agents).

[0084] Emulsions of at least one compound of formula (I) and / or at least one pharma- ceutically acceptable salt may be prepared, for example, as oil-in-water emulsions. The oil phase of the emulsions containing the compounds of formula (I) may be composed of known ingredients in a known manner. The oil phase may be provided, for example, but not limited to, vegetable oils (e.g., olive oil and peanut oil), mineral oils (e.g., liquid paraffin), and mixtures thereof. The oil phase may include only an emulsifier, but may also include a mixture of at least one emulsifier and a fat or oil, or both a fat and an oil. Suitable emulsifiers include, but are not limited to, naturally occurring phosphatides (e.g., soybean lecithin), esters or partial esters derived from fatty acids and hexitol anhydrides (e.g., sorbitan monooleate), and condensation products of partial esters with ethylene oxide (e.g., polyoxyethylene sorbitan monooleate). Preferably, a hydrophilic emulsifier is included together with a lipophilic emulsifier acting as a stabilizer. It is also preferred to include both oil and fat. Together, the emulsifier, with or without stabilizer, makes up the so-called emulsifying wax, and the wax, together with the oil and fat, makes up the so-called emulsifying ointment base, which forms the oily dispersed phase of the cream formulation. The emulsion may also include sweeteners, flavorings, preservatives, and / or antioxidants. Emulsifiers and emulsion stabilizers suitable for use in the formulation of the present invention include Tween 60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl monostearate, sodium lauryl sulfate, glyceryl distearate, alone or with wax; or other materials known in the art.

[0085] Also, the compound of formula (I) and / or at least one of its pharmaceutically acceptable salts can be delivered, for example, intravenously, subcutaneously, and / or intramuscularly via any pharmaceutically acceptable and suitable injection form. Examples of injection forms include, but are not limited to, sterile aqueous solutions containing acceptable vehicles and solvents (e.g., water, Ringer's solution, and isotonic sodium chloride solution), sterile oil-in-water microemulsions, and aqueous or oily suspensions.

[0086] Preparations for parenteral administration may be in the form of aqueous or non-aqueous isotonic sterile injection solutions or suspensions. These solutions and suspensions may be prepared from sterile powders or granules by using one or more carriers or diluents described for use in preparations for oral administration, or by using other suitable dispersing or wetting agents and suspending agents. The compound may be dissolved in water, polyethylene glycol, propylene glycol, ethanol, corn oil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium chloride, tragacanth gum, and / or various buffer solutions. Other adjuvants and methods of administration are known and widely known in the pharmaceutical field. The active ingredient may also be administered by injection in a composition with a suitable carrier (e.g., saline, dextrose, or water), or cyclodextrin (i.e., Captisol), solubilizing cosolvent (i.e., propylene glycol), or solubilizing micelle (i.e., Tween 80).

[0087] A sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent (e.g., a solution in 1,3-butanediol). Among the acceptable vehicles and solvents that may be used are water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally used as a solvent or suspending medium. For this purpose, any sterile fixed oil may be used, including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used as injectable preparations.

[0088] Sterile injectable oil-in-water microemulsions can be prepared, for example, by: 1) dissolving at least one compound of formula (I) in an oil phase (e.g., a mixture of soybean oil and lecithin), 2) combining the formula (I) containing oil phase with a mixture of water and glycerol, and 3) treating the combination to form a microemulsion.

[0089] Sterile aqueous suspension or sterile oily suspension can be prepared according to a method known to those skilled in the art. For example, sterile aqueous solution or sterile aqueous suspension can be prepared using a non-toxic, parenterally acceptable diluent or solvent (e.g., 1,3-butanediol), and sterile oily suspension can be prepared using a non-toxic, acceptable solvent or suspension medium (e.g., sterile fixed oil (e.g., synthetic monoglyceride or diglyceride), and fatty acid (e.g., oleic acid).

[0090] Pharmaceutically acceptable carriers, adjuvants, and vehicles that may be used in the pharmaceutical compositions of the present invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, self-emulsifying drug delivery systems (SEDDS) (e.g., d-α-tocopherol polyethylene glycol 1000 succinate), surfactants used in pharmaceutical dosage forms (e.g., Tween, polyethoxylated castor oil (e.g., CREMOPHOR surfactants (BASF), or other similar polymeric delivery matrices), serum proteins (e.g., human serum albumin), buffer substances (e.g., phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts, or electrolytes (e.g., protamine sulfate, glyceryl stearate, sorbitol ... Examples of suitable cyclodextrins include cyclodextrins such as α-, β-, and γ-cyclodextrin, or chemically modified derivatives such as hydroxyalkylcyclodextrins, including 2- and 3-hydroxypropylcyclodextrin, or other solubilizing derivatives.

[0091] The pharma- ceutical active compounds of the present invention can be processed according to conventional pharmaceutical methods to prepare medicaments for administration to patients (e.g., humans and other mammals). The pharmaceutical compositions can be subjected to conventional pharmaceutical operations (e.g., sterilization) and / or can contain conventional adjuvants (e.g., preservatives, stabilizers, wetting agents, emulsifiers, buffers, etc.). Tablets and pills can additionally be prepared with enteric coatings. Such compositions can also contain adjuvants (e.g., wetting agents, sweeteners, flavoring agents, and aromatic agents).

[0092] The amount of compound administered to treat a condition using the compounds and / or compositions of the present invention, and the dosing schedule will depend on a variety of factors, such as age, weight, sex, the condition of the patient, the type of disease, the severity of the disease, the route and frequency of administration, and the specific compound utilized. Thus, the dosing schedule may vary widely, but can be routinely determined using standard methods. A daily dose of about 0.001 to 100 mg / kg body weight, preferably between about 0.0025 to about 50 mg / kg body weight, and most preferably between about 0.005 to 10 mg / kg body weight, may be appropriate. The daily dose may be administered 1 to 4 times a day. Other dosing schedules include weekly and biday cycles.

[0093] For therapeutic purposes, the active compound of the present invention is usually combined with one or more adjuvants suitable for the intended route of administration.When administered orally, the compound can be mixed with lactose, sucrose, starch powder, cellulose esters of alkanoic acid, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric acid and sulfate, gelatin, gum arabic, sodium alginate, polyvinylpyrrolidone, and / or polyvinyl alcohol, and then tableted or encapsulated for convenient administration.Such capsules or tablets can include controlled release formulations, and can be provided with the active compound dispersed in hydroxypropylmethylcellulose.

[0094] The pharmaceutical compositions of the present invention include at least one compound of formula (I) and / or at least one pharma- ceutically acceptable salt thereof, and optionally additives selected from any pharma- ceutically acceptable carrier, adjuvant, and vehicle. Another composition of the present invention includes a compound of formula (I) as described herein, or a prodrug thereof, and a pharma- ceutically acceptable carrier, adjuvant, or vehicle.

[0095] usefulness The compounds of formula (I) are useful in the treatment of cancer.

[0096] In another embodiment, the present invention provides a pharmaceutical combination of a compound of formula (I) and / or a pharma- ceutical acceptable salt thereof, a stereoisomer thereof or a tautomer thereof with another therapeutic agent for simultaneous, separate or sequential use in the treatment and / or prevention of multiple diseases or disorders associated with the targeted inhibition of DGK in T cells.

[0097] In another aspect, the present invention provides a method for treating a patient suffering from or susceptible to a disease condition associated with targeted inhibition of DGK in T cells. Numerous disease conditions may be treated. The method comprises administering to the patient a composition comprising a therapeutically effective amount of a compound of formula (I) and / or its pharma- ceutical acceptable salt, its stereoisomer or its tautomer. For example, the compounds described herein may be used to treat or prevent viral infections and proliferative diseases (e.g., cancer).

[0098] The compounds of formula (I) and pharmaceutical compositions comprising at least one compound of formula (I) are useful for treating or preventing any disease or condition associated with targeted inhibition of DGK in T cells. Such diseases or conditions include viral and other infectious diseases (e.g., skin infections, GI infections, urinary tract infections, urogenital infections, systemic infections), and proliferative diseases (e.g., cancer). The compounds of formula (I) and pharmaceutical compositions comprising at least one compound of formula (I) can be administered to animals, preferably mammals (e.g., livestock animals, cats, dogs, mice, rats), and more preferably humans. Any method of administration may be used to deliver the compound or pharmaceutical composition to the patient. In some embodiments, the compounds of formula (I) or pharmaceutical compositions comprising at least one compound of formula (I) are administered orally. In other embodiments, the pharmaceutical compositions comprising formula (I) or at least one compound of formula (I) are administered parenterally.

[0099] The compound of formula (I) can inhibit the activity of diacylglycerol kinase α and ζ (DGKα / ζ). For example, the compound of formula (I) can be used to inhibit the activity of DGKα and DGKζ by administering an inhibitory amount of the compound of formula (I) or a salt thereof to a cell or individual in need of regulation of DGKα and DGKζ.

[0100] The present invention further provides a method for treating a disease associated with activity or expression (e.g., abnormal activity and / or overexpression of DGKα and DGKζ in an individual (e.g., a patient)) by administering a therapeutically effective amount or dose of a compound of formula (I) or a pharmaceutical composition thereof to an individual in need of such treatment. Examples of diseases may include any disease, disorder, or condition that is directly or indirectly associated with expression or activity (e.g., overexpression or abnormal activity) of DGKα and DGKζ enzymes. DGKα and DGKζ-related diseases may also include any disease, disorder, or condition that may be prevented, ameliorated, or ameliorated by modulating DGKα and DGKζ enzyme activity. Examples of DGKα and DGKζ-related diseases include cancer and viral infections (e.g., HIV infection, hepatitis B, and hepatitis C).

[0101] In some embodiments, the compound of formula (I) is administered sequentially prior to administration of the immuno-oncology agent. In other embodiments, the compound of formula (I) is administered simultaneously with the immuno-oncology agent. In yet other embodiments, the compound of formula (I) is administered sequentially following administration of the immuno-oncology agent.

[0102] In another embodiment, the compound of formula (I) may be formulated together with an immuno-oncology agent.

[0103] Immuno-oncology drugs include, for example, small molecule drugs, antibodies, or other biological molecules or small molecules. Examples of biological immuno-oncology drugs include, but are not limited to, cancer vaccines, antibodies, and cytokines. In one embodiment, the antibody is a monoclonal antibody. In another embodiment, the monoclonal antibody is a humanized antibody or a human antibody.

[0104] In some embodiments, the immuno-oncology agent is either (i) an agonist of a stimulatory receptor (including costimulatory), or (ii) an antagonist of an inhibitory signal (including co-inhibitory) on a T cell, both of which result in amplification of an antigen-specific T cell response (often referred to as an immune checkpoint regulator).

[0105] Certain stimulatory and inhibitory molecules belong to the immunoglobulin superfamily (IgSF). One important family of membrane-bound ligands that bind to costimulatory or co-inhibitory receptors is the B7 family, which includes B7-1, B7-2, B7-H1 (PD-L1), B7-DC (PD-L2), B7-H2 (ICOS-L), B7-H3, B7-H4, B7-H5 (VISTA), and B7-H6. Another family of membrane-bound ligands that bind to costimulatory or co-inhibitory receptors are the TNF family molecules that bind to the cognate TNF receptor family, including CD40 and CD40L, OX-40, OX-40L, CD70, CD27L, CD30, CD30L, 4-1BBL, CD137 (4-1BB), TRAIL / Apo2-L, TRAILR1 / DR4, TRAILR2 / DR5, TRAILR3, TRAILR4, OPG, and R. ANK, RANKL, TWEAKR / Fn14, TWEAK, BAFFR, EDAR, XEDAR, TACI, APRIL, BCMA, LTβR, LIGHT, DcR3, HVEM, VEGI / TL1A, TRAMP / DR3, Includes EDAR, EDA1, XEDAR, EDA2, TNFR1, lymphotoxin α / TNFβ, TNFR2, TNFα, LTβR, lymphotoxin α1β2, FAS, FASL, RELT, DR6, TROY, and NGFR.

[0106] In certain embodiments, T cell responses may be stimulated by a combination of a compound of Formula (I) and one or more of: (i) protein antagonists that inhibit T cell activation (e.g., immune checkpoint inhibitors), such as CTLA-4, PD-1, PD-L1, PD-L2, LAG-3, TIM-3, Galectin 9, CEACAM-1, BTLA, CD69, Galectin 1, TIGIT, CD113, GPR56, VISTA, 2B4, CD48, GARP, PD1H, LAIR1, TIM-1, and TIM-4, and (ii) agonists of proteins that stimulate T cell activation, such as B7-1, B7-2, CD28, 4-1BB (CD137), 4-1BBL, ICOS, ICOS-L, OX40, OX40L, GITR, GITRL, CD70, CD27, CD40, DR3, and CD28H.

[0107] For cancer treatment, other drugs that can be combined with the compound of formula (I) can include antagonists of inhibitory receptors on NK cells, or agonists of activating receptors on NK cells. For example, the compound of formula (I) can be combined with antagonists of KIR, such as lirilumab.

[0108] Further agents for use in combination therapy include agents that inhibit or deplete macrophages or monocytes, including, but are not limited to, CSF-1R antagonists, such as RG7155 (WO11 / 70024, WO11 / 107553, WO11 / 131407, WO13 / 87699, WO13 / 119716, WO13 / 132044) or CSF-1R antagonist antibodies, including FPA-008 (WO11 / 140249; WO13169264; WO14 / 036357).

[0109] In another embodiment, the compounds of formula (I) may be used with agonistic agents that bind positive co-stimulatory receptors, blocking agents that attenuate signaling through inhibitory receptors, antagonists, and one or more agents that increase the frequency of anti-tumor T cells systemically, agents that overcome different immune suppressive pathways in the tumor microenvironment (e.g., blocking inhibitory receptor engagement (e.g., PD-L1 / PD-1 interaction), depleting or inhibiting Treg cells (e.g., using anti-CD25 monoclonal antibodies (e.g., daclizumab) or by depletion with anti-CD25 beads in vitro), inhibiting metabolic enzymes such as IDO, or restoring / preventing T cell anergy or T cell depletion), and one or more agents that cause innate immune activation and / or inflammation in the tumor area.

[0110] In some embodiments, the immuno-oncology agent is a CTLA-4 antagonist, such as an antagonistic CTLA-4 antibody. Suitable CTLA-4 antibodies include, for example, Yervoy (ipilimumab) or tremelimumab.

[0111] In another embodiment, the immuno-oncology agent is a PD-1 antagonist, for example an antagonistic PD-1 antibody. Suitable PD-1 antibodies include, for example, Opdivo (nivolumab), Keytruda (pembrolizumab), or MEDI-0680 (AMP-514; WO2012 / 145493). The immuno-oncology agent may also include pidilizumab (CT-011), although its specificity for PD-1 binding has been questioned. Another approach to target the PD-1 receptor is a recombinant protein consisting of the extracellular domain of PD-L2 (B7-DC) fused to the Fc portion of IgG1, called AMP-224.

[0112] In another embodiment, the immuno-oncology agent is a PD-L1 antagonist, such as an antagonistic PD-L1 antibody. Suitable PD-L1 antibodies include, for example, MPDL3280A (RG7446; WO2010 / 077634), durvalumab (MEDI4736), BMS-936559 (WO2007 / 005874), and MSB0010718C (WO2013 / 79174).

[0113] In another embodiment, the immuno-oncology agent is a LAG-3 antagonist, for example an antagonistic LAG-3 antibody.Suitable LAG3 antibodies include, for example, BMS-986016 (WO10 / 19570, WO14 / 08218), or IMP-731 or IMP-321 (WO08 / 132601, WO09 / 44273).

[0114] In another embodiment, the immuno-oncology agent is a CD137 (4-1BB) agonist, such as an agonistic CD137 antibody. Suitable CD137 antibodies include, for example, urelumab and PF-05082566 (WO12 / 32433).

[0115] In another embodiment, the immuno-oncology agent is a GITR agonist, for example, an agonistic GITR antibody.Suitable GITR antibodies include, for example, BMS-986153, BMS-986156, TRX-518 (WO06 / 105021, WO09 / 009116) and MK-4166 (WO11 / 028683).

[0116] In another embodiment, the immuno-oncology agent is an IDO antagonist.Suitable IDO antagonists include, for example, INCB-024360 (WO2006 / 122150, WO07 / 75598, WO08 / 36653, WO08 / 36642), indoximod, BMS-986205 or NLG-919 (WO09 / 73620, WO09 / 1156652, WO11 / 56652, WO12 / 142237).

[0117] In another embodiment, the immuno-oncology agent is an OX40 agonist, such as an agonistic OX40 antibody. Suitable OX40 antibodies include, for example, MEDI-6383 or MEDI-6469.

[0118] In another embodiment, the immuno-oncology agent is an OX40L antagonist, such as an antagonistic OX40 antibody. Suitable OX40L antagonists include, for example, RG-7888 (WO06 / 029879).

[0119] In another embodiment, the immuno-oncology agent is a CD40 agonist, for example an agonistic CD40 antibody.In yet another embodiment, the immuno-oncology agent is a CD40 antagonist, for example an antagonistic CD40 antibody.Suitable CD40 antibodies include, for example, lucatumumab or dacetuzumab.

[0120] In another embodiment, the immuno-oncology agent is a CD27 agonist, such as an antagonistic CD27 antibody. Suitable CD27 antibodies include, for example, varlilumab.

[0121] In another embodiment, the immuno-oncology agent against B7H3 is MGA271 (WO11 / 109400).

[0122] Combination therapy is intended to include administration of these therapeutic agents in a sequential manner, i.e., each therapeutic agent is administered at various different times, as well as the therapeutic agents or at least two therapeutic agents are administered in a substantially simultaneous manner. Substantially simultaneous administration can be achieved, for example, by administering to the patient a single dosage form with a fixed ratio of each therapeutic agent, or multiple single dosage forms for each therapeutic agent. Sequential or substantially simultaneous administration of each therapeutic agent can be performed by any suitable route, including, but not limited to, oral, intravenous, intramuscular, and direct absorption through mucosal membranes. The therapeutic agents can be administered by the same route or by different routes. For example, a first therapeutic agent in a selected combination can be administered by intravenous injection, while another therapeutic agent in the combination can be administered orally. Alternatively, for example, all therapeutic agents can be administered orally, or all therapeutic agents can be administered by intravenous injection. The administration of the therapeutic agents described above can also be combined with other biologically active ingredients and non-pharmaceutical therapies (e.g., surgery or radiation therapy) to perform combination therapy. When the combination therapy further includes a non-drug treatment, the non-drug treatment can be administered at any suitable time, so long as the beneficial effect resulting from the synergistic action of the combination of the therapeutic agent and the non-drug treatment is achieved, e.g., in suitable cases, the beneficial effect is achieved even when the non-drug treatment is temporarily suspended, perhaps for days or weeks, from administration of the therapeutic agent.

[0123] The term "cell" as used herein is intended to refer to in vitro, ex vivo or in vivo cells. In some embodiments, ex vivo cells can be part of a tissue sample removed from an organism (e.g., a mammal). In some embodiments, in vitro cells can be cells in cell culture. In some embodiments, in vivo cells are living cells in an organism (e.g., a mammal).

[0124] As used herein, the term "contact" refers to the joining of the indicated moieties in an in vitro system or an in vivo system. For example, "contacting" a compound of formula (I) with DGKα and DGKζ enzymes includes administering a compound of the present invention to an individual or patient (e.g., a human) having DGKα and DGKζ, as well as introducing a compound of formula (I) into, for example, a sample containing cells or a purified product containing DGKα and DGKζ enzymes.

[0125] The term "DGKα and DGKζ inhibitors" refers to agents capable of inhibiting the activity of diacylglycerol kinase α and / or diacylglycerol kinase ζ (DGKα and DGKζ) in T cells, resulting in stimulation of T cells. DGKα and DGKζ inhibitors can be reversible or irreversible DGKα and DGKζ inhibitors. "Reversible DGKα and DGKζ inhibitors" are compounds that reversibly inhibit DGKα and DGKζ enzyme activity at either catalytic or non-catalytic sites, and "irreversible DGKα and DGKζ inhibitors" are compounds that irreversibly impair DGKα and DGKζ enzyme activity by forming a covalent bond with the enzyme.

[0126] The types of cancer that may be treated with the compounds of formula (I) include, but are not limited to, brain cancer, skin cancer, bladder cancer, ovarian cancer, breast cancer, stomach cancer, pancreatic cancer, prostate cancer, colon cancer, blood cancer, lung cancer, and bone cancer. Examples of such cancer types include neuroblastoma, intestinal cancer (e.g., rectal cancer, colon cancer, familial adenomatous polyposis, and hereditary nonpolyposis colorectal cancer), esophageal cancer, lip cancer, laryngeal cancer, hypopharyngeal cancer, tongue cancer, salivary gland cancer, stomach cancer, adenocarcinoma, medullary thyroid cancer, papillary thyroid cancer, kidney cancer, renal parenchymal cancer, ovarian cancer, cervical cancer, uterine cancer, endometrial cancer, choriocarcinoma, pancreatic cancer, prostate cancer, testicular cancer, breast cancer, urinary tract cancer, melanoma, brain Tumors include glioblastoma, astrocytoma, meningioma, medulloblastoma and peripheral neuroectodermal tumors, Hodgkin's lymphoma, non-Hodgkin's lymphoma, Burkitt's lymphoma, acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), adult T-cell leukemia-lymphoma, diffuse large B-cell lymphoma (DLBCL), hepatocellular carcinoma, gallbladder carcinoma, bronchial carcinoma, small cell lung cancer, non-small cell lung cancer, multiple myeloma, basal cell carcinoma, teratoma, retinoblastoma, choroidal melanoma, seminoma, rhabdomyosarcoma, craniopharyngioma, osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma, Ewing's sarcoma and plasmacytoma.

[0127] For the treatment of diseases, disorders or conditions associated with DGKα and DGKζ, one or more additional pharmaceutical agents or therapeutic methods (e.g., antiviral agents, chemotherapeutic agents or other anticancer agents, immune enhancers, immunosuppressants, radiation, antitumor and antiviral vaccines, cytokine therapy (e.g., IL2 and GM-CSF), and / or tyrosine kinase inhibitors) may be used in combination with the compounds of formula (I), as appropriate. The agents may be combined with the compounds in a single dosage form, or the agents may be administered simultaneously or sequentially in different dosage forms.

[0128] Suitable chemotherapeutic or other anti-cancer agents include, for example, alkylating agents (including but not limited to, nitrogen mustards, ethylenimine derivatives, alkylsulfonic acids, nitrosoureas, and triazenes), such as uracil mustard, chlormethine, cyclophosphamide (Cytoxan®), ifosfamide, melphalan, chlorambucil, pipobroman, triethylene-melamine, triethylenethiophosphoramine, busulfan, carmustine, lomustine, streptozocin, dacarbazine, and temozolomide.

[0129] Suitable agents for use in combination with the compounds of formula (I) in the treatment of melanoma include dacarbazine (DTIC), optionally with other chemotherapeutic agents (e.g., carmustine (BCNU) and cisplatin; the "Dartmouth regimen" consisting of DTIC, BCNU, cisplatin and tamoxifen; the combination of cisplatin, vinblastine, and DTIC, temozolomide or Yervoy®). The compounds of formula (I) may also be combined with immunotherapeutic agents (e.g., cytokines such as interferon alpha, interleukin 2, and tumor necrosis factor (TNF)) in the treatment of melanoma.

[0130] The compounds of formula (I) can also be used in combination with vaccine therapy in the treatment of melanoma.Anti-melanoma vaccines are similar in some respects to anti-viral vaccines used to prevent diseases caused by viruses (e.g., polio, measles, and mumps).Attenuated melanoma cells or parts of melanoma cells called antigens can be injected into patients to stimulate the body's immune system to destroy melanoma cells.

[0131] Melanomas of the arms or legs may also be treated using hyperthermic perfusion therapy with a combination of drugs containing one or more compounds of formula (I). This treatment protocol involves temporarily isolating the circulatory system of the affected limb from the rest of the body and injecting high concentrations of chemotherapy drugs into the arteries of the affected limb, delivering high doses to the tumor site that would cause serious side effects if exposed to internal organs. Typically, this treatment involves warming the body fluids to 38.9°C to 40°C. Melphalan is the drug most frequently used in this chemotherapy. Another drug called tumor necrosis factor (TNF) may also be used.

[0132] Suitable chemotherapeutic or other anti-cancer agents include, for example, antimetabolites (including but not limited to folate antagonists, pyrimidine analogs, purine analogs, and adenosine deaminase inhibitors), such as methotrexate, 5-fluorouracil, floxuridine, cytarabine, 6-mercaptopurine, 6-thioguanine, fludarabine phosphate, pentostatin, and gemcitabine.

[0133] Suitable chemotherapeutic or other anti-cancer agents further include, for example, certain natural products and their derivatives (e.g., vinca alkaloids, antitumor antibiotics, enzymes, lymphokines and epipodophyllotoxins), such as vinblastine, vincristine, vindesine, bleomycin, dactinomycin, daunorubicin, doxorubicin, epirubicin, idarubicin, cytarabine, paclitaxel (taxol), mithramycin, deoxycoformycin, mitomycin C, L-asparaginase, interferons (especially IFNα), etoposide, and teniposide.

[0134] Other cytotoxic agents include navelbine, CPT-11, anastrozole, letrozole, capecitabine, raloxifene, and droloxifene.

[0135] Suitable cytotoxic agents also include, for example, epipodophyllotoxins; anti-neoplastic enzymes; topoisomerase inhibitors; procarbazine; mitoxantrone; platinum coordination complexes (e.g., cisplatin and carboplatin); biological response modifiers; growth inhibitory agents; anti-hormonal therapeutics; leucovorin; tegafur; and hematopoietic growth factors.

[0136] Other anti-cancer agents include antibody drugs such as trastuzumab (Herceptin®), antibodies against costimulatory molecules (such as CTLA-4, 4-1BB and PD-1), or antibodies against cytokines (IL-1O or TGF-β).

[0137] Other anti-cancer agents also include those that block immune cell migration, such as antagonists to chemokine receptors (eg, CCR2 and CCR4).

[0138] Other anti-cancer drugs also include anti-cancer drugs that enhance the immune system, such as adjuvants or adoptive T-cell transfer.

[0139] Anti-cancer vaccines include dendritic cells, synthetic peptides, DNA vaccines and recombinant viruses.

[0140] The pharmaceutical composition of the present invention may optionally contain at least one signal transduction inhibitor (STI). A "signal transduction inhibitor" is an agent that selectively inhibits one or more critical steps in a signal transduction pathway in the normal function of cancer cells, thereby inducing apoptosis. Suitable STIs include, but are not limited to, (i) bcr / abl kinase inhibitors (e.g., STI 571 (Gleevec®)); (ii) epidermal growth factor (EGF) receptor inhibitors (e.g., kinase inhibitors (Iressa®, SSI-774) and antibodies (Imclone: ​​C225 [Goldstein et al., Clin. Cancer Res., 1: 1311-1318 (1995)], and Abgenix: ABX-EGF); (iii) her-2 / neu receptor inhibitors (e.g., farnesyltransferase inhibitors (FTIs) (e.g., L-744,832 (Kohl et al., Nat. Med., 1(8):792-797 (1995)); (iv) inhibitors of the Akt family kinases or Akt pathway (e.g., rapamycin (e.g., Sekulic et al., Cancer Res., 60: 3504-3513 (1995)); (2000)); (v) cell cycle kinase inhibitors (e.g., flavopiridol and UCN-O1 (see, e.g., Sausville, Curr. Med. Chem. Anti-Canc. Agents, 3:47-56(2003)); and (vi) phosphatidylinositol kinase inhibitors (e.g., LY294002 (see, e.g., Vlahos et al., J. Biol. Chem., 269: 5241-5248 (1994)). Alternatively, the at least one STI and the at least one compound of formula (I) may be formulated in separate pharmaceutical compositions. In certain embodiments of the present invention, the at least one compound of formula (I) and the at least one STI may be administered to a patient simultaneously or sequentially. In other words, the at least one compound of formula (I) may be administered first, the at least one STI may be administered first, or the at least one compound of formula (I) and the at least one STI may be administered simultaneously.Furthermore, when more than one compound of formula (I) and / or an STI is used, the compounds may be administered in any order.

[0141] The present invention further provides pharmaceutical compositions comprising at least one compound of formula (I), optionally at least one chemotherapeutic agent, and optionally at least one antiviral agent, in a pharma- ceutically acceptable carrier for treating a chronic viral infection in a patient.

[0142] Also provided is a method for treating a chronic viral infection in a patient by administering an effective amount of the above pharmaceutical composition.

[0143] In certain embodiments of the present invention, at least one compound of formula (I) and at least one chemotherapeutic agent are administered to patient simultaneously or sequentially.In other words, at least one compound of formula (I) can be administered first, at least one chemotherapeutic agent can be administered first, or at least one compound of formula (I) and at least one chemotherapeutic agent can be administered simultaneously.In addition, when one or more compounds of formula (I) and / or chemotherapeutic agents are used, the compounds can be administered in any order.Similarly, any antiviral drug or STI can be administered at any time compared to the administration of the compound of formula (I).

[0144] Chronic viral infections that may be treated using the present combination therapy include, but are not limited to, diseases caused by Hepatitis C virus (HCV), human papillomavirus (HPV), cytomegalovirus (CMV), herpes simplex virus (HSV), Epstein-Barr virus (EBV), varicella zoster virus, coxsackievirus, and human immunodeficiency virus (HIV). In particular, parasitic infections (e.g., malaria) may be treated by the above methods, optionally with the addition of compounds known for treating parasitic diseases in place of antiviral drugs.

[0145] Suitable antiviral agents contemplated for use in combination with the compounds of formula (I) may include nucleoside and nucleotide reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), protease inhibitors and other antiviral agents.

[0146] Examples of suitable NRTIs include zidovudine (AZT); didanosine (ddI); zalcitabine (ddC); stavudine (d4T); lamivudine (3TC); abacavir (1592U89); adefovir pivoxil [bis(POM)-PMEA]; lobucavir; BCH-I0652; emtricitabine [(-)-FTC]; β-L-FD4 (also called β-L-D4C, which has the name β-L-2',3'-dideoxy-5-fluoro-cytidine); DAPD, ((-)-β-D-2,6-diamino-purine dioxolane); and rhodenosine (FddA). Representative suitable NNRTIs include nevirapine (BI-RG-587); delavirdine (BHAP, U-90152); efavirenz (DMP-266); PNU-142721; AG-1549; MKC-442 (1-(ethoxy-methyl)-5-(1-methylethyl)-6-(phenylmethyl)-(2,4(1H,3H)-pyrimidinedione); and (+)-calanolide A (NSC-675451) and B. Representative suitable protease inhibitors include saquinavir (Ro 31-8959); ritonavir (ABT-538); indinavir (MK-639); nelfinavir (AG-1343); amprenavir (141W94); lasinavir; DMP-450; BMS-2322623; ABT-378; and AG-1549. Other antiviral drugs include hydroxyurea, ribavirin, IL-2, IL-12, pentafuside, and Yissum Project No. 11607.

[0147] The present invention also includes pharmaceutical kits useful, for example, in the treatment or prevention of DGKα and DGKζ-related diseases or disorders, and other diseases described herein, which include one or more containers containing a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (I). Such kits may further include one or more of a variety of conventional pharmaceutical kit components (e.g., a container containing one or more pharma- ceutically acceptable carriers, another container), which will be readily apparent to those skilled in the art. Instructions, either in the form of a package insert or label, indicating the amounts of components to be administered, administration guidelines, and / or mixing guidelines for the components, may also be included in the kit.

[0148] Combination therapy is intended to include administration of these therapeutic agents in a sequential manner, i.e., each therapeutic agent is administered at various different times, as well as the therapeutic agents or at least two therapeutic agents are administered in a substantially simultaneous manner. Substantially simultaneous administration can be achieved, for example, by administering to the patient a single dosage form with a fixed ratio of each therapeutic agent, or multiple single dosage forms for each therapeutic agent. Sequential or substantially simultaneous administration of each therapeutic agent can be performed by any suitable route, including, but not limited to, oral, intravenous, intramuscular, and direct absorption through mucosal membranes. The therapeutic agents can be administered by the same route or by different routes. For example, a first therapeutic agent in a selected combination can be administered by intravenous injection, while another therapeutic agent in the combination can be administered orally. Alternatively, for example, all therapeutic agents can be administered orally, or all therapeutic agents can be administered by intravenous injection. Combination therapy can also include administration of the above-mentioned therapeutic agents in combination with other biologically active ingredients and non-pharmaceutical therapies, such as surgery or radiation therapy. When the combination therapy further includes a non-drug treatment, the non-drug treatment can be administered at any suitable time, so long as the beneficial effect resulting from the synergistic action of the combination of the therapeutic agent and the non-drug treatment is achieved, e.g., in suitable cases, the beneficial effect is achieved even when the non-drug treatment is temporarily suspended, perhaps for days or weeks, from administration of the therapeutic agent.

[0149] The present invention also provides pharma- ceutically acceptable compositions comprising a therapeutically effective amount of one or more compounds of formula (I), formulated together with one or more pharma- ceutically acceptable carriers (excipients) and / or diluents, and optionally one or more additional therapeutic agents as described above.

[0150] The compounds of the present invention may be administered for any of the uses described herein by any suitable means, for example, oral administration (e.g., tablets, capsules (each including sustained or timed release formulations), pills, powders, granules, elixirs, tinctures, suspensions (including nanosuspensions, microsuspensions, spray dried dispersions), syrups, and emulsions); sublingual administration; buccal administration; parenteral administration (e.g., subcutaneous, intravenous, intramuscular, or intrasternal injection, or infusion techniques (e.g., sterile injectable aqueous or non-aqueous solutions or suspensions); nasal administration, including administration to the nasal membranes (e.g., inhalation sprays); topical administration (e.g., in the form of creams or ointments); or rectal administration (e.g., in the form of suppositories). They may be administered alone, but will generally be administered with a pharmaceutical carrier selected on the basis of the chosen route of administration and standard pharmaceutical practice.

[0151] The term "pharmaceutical acceptable carrier" as used herein means a pharmaceutical acceptable substance, composition, or vehicle, such as a liquid or solid filler, diluent, excipient, processing aid (e.g., lubricant, magnesium talc, calcium or zinc stearate, or stearic acid), or solvent encapsulant, which is involved in the transport or delivery of a particular compound from one organ or part of the body to another organ or part of the body. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients in the formulation (i.e., including adjuvants, excipients, or vehicles (e.g., diluents, preservatives, fillers, flow regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants, and dispensing agents), which vary depending on the method of administration and the nature of the dosage form) and not harmful to the patient.

[0152] The term "pharmaceutical composition" refers to a composition comprising a compound of the present invention in combination with at least one additional pharma- ceutically acceptable carrier.

[0153] Pharmaceutically acceptable carriers are formulated according to many factors well within the expertise of those skilled in the art. Pharmaceutically acceptable carriers are formulated according to many factors well within the expertise of those skilled in the art. These factors include, but are not limited to, the type and nature of the active agent being formulated, the patient to whom the composition containing the active agent will be administered, the intended route of administration of the composition, and the targeted therapeutic index. Pharmaceutically acceptable carriers include both aqueous and non-aqueous liquid media, as well as a variety of solid and semi-solid dosage forms. Such carriers can include many different components and additives in addition to the active agent, and such additional components are included in the formulation for a variety of reasons, such as stabilization of the active agent, binders, etc., as is well known to those skilled in the art. Descriptions of suitable pharmacopoeia acceptable carriers and factors in selecting them can be found in a variety of readily available literature, such as Allen, LV Jr. et al. Remington: The Science and Practice of Pharmacy (Vol. 2), 22nd Edition (2012), Pharmaceutical Press.

[0154] Dosing regimens for the compounds of the invention will, of course, vary depending on known factors, such as the pharmacodynamic properties of the particular agent and its method and route of administration; the species, age, sex, health, medical condition, and weight of the recipient; the nature and extent of the condition; type of concomitant treatment; frequency of treatment; route of administration, the patient's renal and hepatic function, and the desired effect.

[0155] As a general guidance, the daily oral dose of each active ingredient, when used to obtain the intended effect, will range from about 0.001 to about 5000 mg / day, preferably about 0.01 to about 1000 mg / day, and most preferably between about 0.1 and about 250 mg / day. The most preferred doses for constant rate infusion intravenously range from about 0.01 to about 10 mg / kg / min. The compounds of the present invention may be administered in a single daily dose or in divided doses of two, three, or four total daily doses.

[0156] The compounds will generally be administered in admixture with suitable pharmaceutical diluents, excipients, or carriers (collectively referred to herein as pharmaceutical carriers) appropriately selected for the intended form of administration (e.g., oral tablets, capsules, elixirs, and syrups) and consistent with conventional pharmaceutical standards.

[0157] Dosage forms (pharmaceutical compositions) suitable for administration may contain about 1 mg to about 2000 mg of active ingredient per dosage unit. In these pharmaceutical compositions, the active ingredient is usually present in an amount of about 0.1 to 95% by weight of the total weight of the composition.

[0158] A typical capsule for oral administration contains at least one compound of the invention (250 mg), lactose (75 mg), and magnesium stearate (15 mg), which mixture is passed through a 60 mesh sieve and filled into a No. 1 gelatin capsule.

[0159] A typical injectable formulation is prepared by aseptically adding at least one compound of the present invention (250 mg) to a vial, aseptically lyophilizing and sealing it, and at the time of use, mixing the contents of the vial with saline (2 mL) to prepare the injectable formulation.

[0160] The present invention includes within its scope pharmaceutical compositions comprising a therapeutically effective amount of at least one compound of the invention as an active ingredient, alone or in combination with a pharmaceutical carrier. The compounds of the invention may be used alone, in combination with other compounds of the invention, or in combination with one or more other therapeutic agents (e.g., anti-cancer agents or other pharmacologic active substances), as appropriate.

[0161] Regardless of the route of administration selected, the compounds of the present invention, which may be used in a suitable hydrated form, and / or the pharmaceutical compositions of the present invention are formulated into pharma- ceutical acceptable dosage forms by conventional methods known to those skilled in the art.

[0162] Actual dosage levels of the active ingredients in the pharmaceutical compositions of the present invention may be varied to contain an amount of the active ingredient that is non-toxic to the patient and effective to obtain a therapeutic effect for a particular patient, composition and mode of administration.

[0163] The selected dosage level will depend upon a variety of factors, including the activity of the particular compound of the invention or its esters, salts or amides employed, the route of administration of the particular compound employed, the time of administration, the rate of excretion or metabolism, the rate and extent of absorption, the duration of treatment, other drugs, compounds and / or substances used in combination with the particular compound employed, the age, sex, weight, symptoms, health and medical history of the patient being treated, and factors well known in the medical arts.

[0164] A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, the physician or veterinarian can start the dosage of the compound of the present invention used in the pharmaceutical composition at a level lower than required to achieve a therapeutic effect, and gradually increase the dosage until an effect is achieved.

[0165] In general, the appropriate daily dose of the compound of the present invention is the lowest effective dose of the compound to obtain a therapeutic effect. Such an effective dose is generally determined by the above factors. In general, the dose of the compound of the present invention to a patient is about 0.01 to about 50 mg / kg body weight / day for oral, intravenous, intracerebroventricular and subcutaneous administration.

[0166] If desired, the effective daily dose of the active compound may be administered in two, three, four, five, six or more divided doses at appropriate intervals throughout the day, conveniently in unit dosage forms. In some embodiments of the invention, dosing is once a day.

[0167] While it is possible for a compound of the present invention to be administered alone, it is preferable to administer the compound as a pharmaceutical formulation (composition).

[0168] The other therapeutic agents described above, when used in combination with the compounds of the invention, may be used, for example, in amounts as set forth in the Pharmaceutical and Medical Devices Act (PDR) or as otherwise specifically determined by one of ordinary skill in the art. In the methods of the invention, the other therapeutic agents may be administered prior to, simultaneously with, or after administration of the compounds of the invention.

[0169] Manufacturing method The compounds of the present invention can be synthesized by many methods available to those skilled in the art of organic chemistry. A general synthetic scheme for preparing the compounds of the present invention is described below. This scheme is illustrative and is not intended to limit the possible techniques that a person skilled in the art may use to prepare the compounds described herein. Various methods for preparing the compounds of the present invention will be apparent to those skilled in the art. Examples of compounds of the present invention prepared by the methods described in the general scheme are shown in the Examples section below. Preparation of homochiral examples can be carried out by techniques known to those skilled in the art. For example, homochiral compounds can be prepared by separating racemic products or diastereomers by chiral phase preparative HPLC. Alternatively, the compounds can be prepared by known methods that give enantiomerically enriched or diastereomerically enriched products.

[0170] The reactions and techniques described in this section are carried out in solvents suitable for the reagents and materials used and are suitable for the transformations effected. In addition, in the description of the synthetic methods described below, it is understood that all reaction conditions presented (including the choice of solvent, reaction atmosphere, reaction temperature, experimental time and work-up method) are selected to be conditions that are standard for the reaction, which should be easily recognized by those skilled in the art. It is understood by those skilled in the art of organic synthesis that the functional groups present on various parts of the molecule must be compatible with the reagents and reactions presented. Such restrictions on the substituents that are compatible with the reaction conditions are readily apparent to those skilled in the art, and alternatives are required when the substituents present are not suitable. The reactions may require the decision to change the order of synthetic steps or to select a particular reaction course differently in order to obtain the compounds of the present invention. It is also recognized that another important consideration in planning any synthetic route in this field is the judicious selection of protective groups used to protect reactive functional groups present in the desired compounds described in the present invention. For the experienced experimenter, an authoritative reference describing many protecting group alternatives is Greene's Protective Groups in Organic Synthesis by Wuts and Greene (Fourth Edition, Wiley & Sons, 2007).

[0171] Methods that may be utilized to synthesize intermediates useful in the preparation of embodiments of the present invention are illustrated in the following schemes.

[0172] Benzoxazine-2,4(1H)-diones of the type shown can be treated with a strong base and a methylating agent such as methyl iodide to give 1-methyl-2H-benzo[d][1,3]oxazine-2,4(1H)-diones. These can be treated with activated acetates such as nitroacetate, malonate, and cyanoacetate to give 1-methyl-3-nitroquinoline-2,4(1H,3H)-dione, 1-methyl-3-carboxyethylquinoline-2,4(1H,3H)-dione, and 1-methyl-3-cyanoquinoline-2,4(1H,3H)-dione, respectively. The 3-ester derivatives can be further transformed by basic hydrolysis and acidic decarboxylation to give 6-fluoro-4-hydroxy-1-methylquinolin-2(1H)-ones.

[0173] Scheme 1 [ka]

[0174] Alternatively, quinolinediones can be prepared from substituted 2-aminobenzoic acids. For example, methyl 2-amino-5-bromobenzoate can be converted to 4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-6-carbonitrile in four steps.

[0175] Scheme 2 [ka]

[0176] Such quinolinediones can in turn be converted to the relevant 4-chloro or 4-triflate derivatives and reacted with variously functionalized piperazines to give protected compounds such as tert-butyl 4-(6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate, or compounds of the examples of the present invention.

[0177] Scheme 3 [ka]

[0178] A protected piperazine such as tert-butyl 4-(6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate can be further transformed by deprotecting the piperazine with, for example, TFA in dichloromethane and reacting with an electrophile such as 3-fluorobenzyl bromide to give 6-fluoro-4-{4-[(3-fluorophenyl)methyl]piperazin-1-yl}-1-methyl-3-nitro-1,2-dihydroquinolin-2-one.

[0179] Scheme 4 [ka]

[0180] The compounds of the present invention can also be prepared by reacting quinolinediones or aminoquinolones with electrophiles such as phosphorus oxychloride / DMF, nitric acid / acetic acid, or N-bromosuccinimide / DMF to obtain, for example, 4-(4-benzhydrylpiperazin-1-yl)-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbaldehyde, 4-hydroxy-1-methyl-3-nitroquinolin-2(1H)-one, and tert-butyl 4-(3-bromo-6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate, respectively. These compounds can be further transformed, if necessary, to obtain the compounds of the present invention. Examples of the present invention can be prepared from 4-hydroxy-1-methyl-3-nitroquinolin-2(1H)-one according to the steps described in Scheme 3.

[0181] Scheme 5 [ka]

[0182] tert-Butyl 4-(3-bromo-6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate can be reacted with aryl and heteroaryl boronic acids or stanines in the presence of a suitable catalyst to give, for example, tert-butyl 4-(6-fluoro-1-methyl-2-oxo-3-(pyridin-4-yl)-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate. These examples can be further transformed by deprotecting and alkylating the piperazine as shown in Scheme 3.

[0183] Scheme 6 [ka]

[0184] Working Example The following examples are illustrative of specific and preferred embodiments of the present invention and are not intended to limit the scope of the invention. Chemical and scientific abbreviations and symbols have their common and customary meanings unless otherwise specified. Additional abbreviations used in the examples and elsewhere herein are as defined above. Common intermediates are generally useful in the preparation of one or more examples and are identified sequentially (e.g., intermediate 1, intermediate 2, etc.) and abbreviated as Int.1 or I1, Int.2 or I2, etc. Compounds of the examples are identified by the example and the step by which they are prepared (e.g., "1-A" represents step A of Example 1), or by example only if the compound is the title compound of the example (e.g., "1" represents the title compound of Example 1). In some cases, alternative methods of preparing intermediates or examples are described. Those skilled in the art of synthesis may frequently devise alternative preparation methods that are desirable based on one or more considerations (e.g., shorter reaction times, cheaper starting materials, ease of manipulation and isolation, improved yields, handling of catalysts, avoidance of toxic reagents, availability of specialized equipment, and reduction in the number of linear steps). The intent of describing alternative preparation methods is to further enable the preparation of the embodiments of the present invention. In some cases, some functional groups in the outlined examples and claims may be replaced by well-known bioisostere substitutions known to those skilled in the art (e.g., replacement of carboxylic acid groups with tetrazole or phosphate moieties). Collected in deuterated dimethylsulfoxide. 1 H NMR data used water suppression in the data processing. Spectra are reported without correction for the effects of water suppression. Protons adjacent to the water suppression frequency at 3.35 ppm show a decrease in signal intensity.

[0185] Abbreviation [Table 1] [Table 2]

[0186] Intermediate 1 6-Fluoro-1-methyl-2H-benzo[d][1,3]oxazine-2,4(1H)-dione [ka]

[0187] To a solution of 6-fluoro-1H-benzo[d][1,3]oxazine-2,4-dione (10.2 g, 56.3 mmol) in DMF (60 mL) was added 60% sodium hydride dispersion in mineral oil (2.70 g, 67.6 mmol) portionwise at 0° C. The reaction mixture was allowed to warm to room temperature and stirred for 30 minutes. Methyl iodide (4.23 mL, 67.6 mmol) was then added dropwise. The reaction mixture was stirred at room temperature overnight. The reaction was quenched with water. Diethyl ether was added to the reaction mixture. A light yellow solid was collected as 6-fluoro-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione (9.55 g, 48.9 mmol, 87% yield). LC / MS LC / MS column: Phenomenex LUNA C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 1.982 min, 196.0 (M+H + ); 1 H NMR (400MHz, DMSO-d6)δ 7.82-7.73(m,2H), 7.56-7.46(m,1H), 3.47(s,3H)

[0188] Intermediate 2 6-Fluoro-1-methyl-3-nitroquinoline-2,4(1H,3H)-dione [ka]

[0189] To a solution of ethyl 2-nitroacetate (3.38 g, 25.4 mmol) in NMP (15 mL) at 0° C. in a round-bottom flask was added 60% sodium hydride dispersion in mineral oil (1.107 g, 27.7 mmol) in portions. The reaction mixture was stirred at 0° C. for 5 min and then at room temperature for 15 min. 6-Fluoro-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione (4.5 g, 23.06 mmol) was then added and the reaction mixture was heated at 120° C. for 2 h. LC / MS showed the reaction was complete. The reaction was quenched with ice water. The reaction mixture was acidified with 1N HCl solution and then with 1N HCl solution. Ethyl ether was added and the yellow solid was collected as 6-fluoro-1-methyl-3-nitroquinoline-2,4(1H,3H)-dione (2.69 g, 11.29 mmol, 49.0% yield). LC / MS LC / MS: Column: Phenomenex Luna C18, 2.0x50 mm, 3 mm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Temperature: 40 °C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8 mL / min; Detection: UV (220 nm); Retention time: 1.488 min, 237.1 (MH - ); 1 H NMR (400MHz, DMSO-d6)δ 7.85(dd,J=9.2, 2.6Hz,1H), 7.72-7.47(m,2H), 6.73(brs,1H), 3.58(s,3H)

[0190] Intermediate 3 4-Chloro-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one [ka]

[0191] In a sealed tube, 6-fluoro-1-methyl-3-nitroquinoline-2,4(1H,3H)-dione (1.4 g, 5.88 mmol) and phosphorus oxychloride (10 mL, 107 mmol) were added. The reaction mixture was heated at 95° C. for 4 h. The reaction mixture was concentrated, the residue was poured onto ice water and neutralized with saturated NaHCO3 solution. The mixture was extracted with dichloromethane (2×40 mL), the organic layers were combined, dried (MgSO4) and concentrated to give 4-chloro-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one as an orange solid (1.28 g, 4.99 mmol, 85% yield). LC / MS: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 2.648 min, 257.0 (M+H + ); 1 H NMR (500MHz, DMSO-d6)δ 7.93(d,J=9.5Hz,1H), 7.86-7.79(m,2H), 3.72(s,3H)

[0192] Intermediate 4 tert-Butyl 4-(6-fluoro-1-methyl-3-nitro-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate [ka]

[0193] To a solution of 4-chloro-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one (600 mg, 2.338 mmol) in DMF (8 mL) was added tert-butyl piperazine-1-carboxylate (435 mg, 2.338 mmol) and triethylamine (0.978 mL, 7.01 mmol). The reaction mixture was stirred at room temperature overnight. LC / MS showed the reaction was complete. The reaction was quenched with water. The yellow solid was collected as the product tert-butyl 4-(6-fluoro-1-methyl-3-nitro-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate (765 mg, 1.882 mmol, 81% yield). LC / MS LC / MS: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 3.0 min, 407.1 (M+H + ); 1 H NMR (500MHz, DMSO-d6)δ 7.79-7.63(m,3H), 3.65(s,3H), 3.58(brs,4H), 3.07(brs,4H), 1.43(s,9H)

[0194] Intermediate 5 6-Fluoro-1-methyl-3-nitro-4-(piperazin-1-yl)quinolin-2(1H)-one TFA [ka]

[0195] To a solution of tert-butyl 4-(6-fluoro-1-methyl-3-nitro-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate (2.1 g, 5.17 mmol) in CHCl (10 mL) was added TFA (5 mL, 64.9 mmol). The reaction mixture was stirred at room temperature overnight. LC / MS showed the reaction was complete. The reaction mixture was concentrated and ethyl acetate was added. A yellow solid was obtained as the product 6-fluoro-1-methyl-3-nitro-4-(piperazin-1-yl)quinolin-2(1H)-one·TFA (1.72 g, 4.09 mmol, 79% yield). LC / MS column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 10:90 methanol:water + 0.1% TFA; Mobile phase B: 90:10 methanol:water + 0.1% TFA; Gradient: 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 1.62 min, 307.2 (M+H + ); 1 H NMR (400MHz, DMSO-d6)δ 7.73-7.67(m,2H), 7.64(d,J=10.0Hz,1H), 3.65(s,3H), 3.11-3.03(m,4H), 2.93(brs,4H)

[0196] Example 1 6-Fluoro-4-{4-[(3-fluorophenyl)methyl]piperazin-1-yl}-1-methyl-3-nitro-1,2-dihydroquinolin-2-one [ka]

[0197] A solution of 6-fluoro-1-methyl-3-nitro-4-(piperazin-1-yl)quinolin-2(1H)-one·TFA (340 mg, 816 μmol) and DIPEA (646 mL, 3.6 mmol) in DMF (17.0 mL) was prepared. To a weighed amount of 3-fluorobenzyl bromide in a 16x100 mm screw-threaded vial was added 500 μL of the 6-fluoro-1-methyl-3-nitro-4-(piperazin-1-yl)quinolin-2(1H)-one·TFA / DIPEA solution. The vial was capped and shaken at room temperature until the reaction was complete (approximately 24 hours) as determined by LCMS analysis. A 500 μL aliquot of DMF was added to the vial and the crude material was purified via preparative LC / MS using the following conditions: Column: XBridge C18, 19x200mm, 5 μm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Gradient: 45-85% B over 15 min, then hold at 100% B for 5 min; Flow: 20 mL / min. Fractions containing product were combined and dried via centrifugal evaporation. LC / MS conditions: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 10:90 methanol:water + 0.1% TFA; Mobile phase B: 90:10 methanol:water + 0.1% TFA; Gradient: 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Measured MS ion: 415.1, retention time: 1.23 min.

[0198] Intermediate 6 Ethyl 6-fluoro-1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinoline-3-carboxylate [ka]

[0199] To a solution of diethyl malonate (0.903 g, 5.64 mmol) in NMP (10 mL) in a round bottom flask at 0° C., 60% sodium hydride dispersion in mineral oil (0.246 g, 6.15 mmol) was added portionwise. The reaction mixture was stirred at 0° C. for 5 min and then at room temperature for 15 min. 6-Fluoro-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione (1 g, 5.12 mmol) was then added and the reaction mixture was heated at 120° C. for 2 h. LC / MS showed the reaction was complete. The reaction was quenched with ice water and acidified with 1N HCl solution. Ethyl ether was added and a beige solid was collected as the product, ethyl 6-fluoro-1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinoline-3-carboxylate (245 mg, 0.924 mmol, 18.03% yield). LC / MS: Column: Phenomenex Luna C18, 2.0x50 mm, 3 mm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Temperature: 40 °C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8 mL / min; Detection: UV (220 nm); Retention time: 1.660 min, 264.1 (MH - ); 1 H NMR(400MHz, DMSO-d6)δ 7.77(dd,J=9.3, 2.9Hz,1H), 7.68-7.54(m,2H), 4.32(q,J=7.1Hz,2H), 3.56(s,3H), 1.30(t,J=7.1Hz,3H)

[0200] Intermediate 7 6-Fluoro-4-hydroxy-1-methylquinolin-2(1H)-one [ka]

[0201] A suspension of ethyl 6-fluoro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (6.5 g, 24.51 mmol) in 2M sodium hydroxide (150 mL, 300 mmol) was heated at 100° C. for 12 hours. The reaction mixture was cooled to room temperature and 50 mL of 6N HCl was added dropwise with stirring. After approximately 30 minutes, the solid was collected by filtration. The filtrate was washed with water and then dried under vacuum overnight. The solid was further dried by suspending in toluene and evaporating to dryness (2×) to give the product as an off-white solid (4.93 g, 24.51 mmol, 100% yield). m / z; 194 (M+H). +

[0202] Intermediate 8 4-Chloro-6-fluoro-1-methylquinolin-2(1H)-one [ka]

[0203] In a dry vial containing a magnetic stir bar, 6-fluoro-1-methylquinoline-2,4(1H,3H)-dione (1000 mg, 5.18 mmol) was added followed by phosphorus oxychloride (5790 μL, 62.1 mmol). The mixture was kept under nitrogen atmosphere and placed in a 100° C. oil bath for 1 h. The reaction mixture was removed from the oil bath and allowed to cool to room temperature before being evaporated under vacuum. The residue was dissolved / suspended in acetonitrile and placed in an ice bath. Ice crystals were added and the reaction mixture was allowed to warm to room temperature overnight. The reaction mixture was concentrated under vacuum to remove acetonitrile. The resulting suspension was filtered and the product was collected by filtration. The tan solid was dried under vacuum at 50° C. to give the title compound as a reddish tan solid (974 mg). 1 H NMR (400MHz, DMSO-d6)δ 7.82-7.58(m,3H), 7.05(s,1H), 3.64(s,3H)

[0204] Intermediate 9 tert-Butyl 4-(6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate [ka]

[0205] 4-Chloro-6-fluoro-1-methylquinolin-2(1H)-one (940 mg, 4.44 mmol) and tert-butyl piperazine-1-carboxylate (910 mg, 4.89 mmol) were dissolved in a mixture of DMA (20 mL) and t-BuOH (5.00 mL). Cesium carbonate (4342 mg, 13.33 mmol) was added along with (2-dicyclohexylphosphino-2',6'-diisopropoxy-1,1'-biphenyl)[2-(2'-amino-1,1'-biphenyl)]palladium(II) methanesulfonate (186 mg, 0.222 mmol). The reaction mixture was degassed and flushed with nitrogen, then heated at 85° C. overnight. The reaction mixture was diluted with 0.1 M EDTA solution and EtOAc was added. The mixture was filtered through Celite to remove colloidal precipitated palladium. The organic layer was then separated and the aqueous mixture was re-extracted with EtOAc (2x). The extracts were combined and washed once with brine, then dried over MgSO4, filtered and concentrated in vacuo to give the crude product as a dark purple viscous oil. The residue was dissolved in DCM and adsorbed onto silica gel. The material was then fractionated using flash chromatography on silica gel, eluting with 90-100% EtOAc in hexanes. Homogeneous fractions were combined to give the product as a colorless oil (960 mg). 1 H NMR (400MHz, DMSO-d6)δ 7.63-7.44(m,3H), 6.13(s,1H), 3.57(s,7H), 2.99(brt,J=4.6Hz,4H), 1.44(s,9H)

[0206] Intermediate 10 tert-Butyl 4-(3-bromo-6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate [ka]

[0207] tert-Butyl 4-(6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate (98 mg, 0.271 mmol) was dissolved in DMF (2 mL) in a round bottom flask maintained under nitrogen atmosphere. The solution was cooled to 0° C. and N-bromosuccinimide (43.9 mg, 0.247 mmol) was added in one portion. The mixture was then removed from the ice bath and allowed to warm gradually over 15 min. The solvent was then removed under high vacuum and the resulting residue was dissolved in DCM, adsorbed onto SiO2, and then chromatographed using 40% EtOAc in hexanes. Homogeneous fractions were combined and evaporated under vacuum to give the product as a colorless film (98 mg, 88% yield). 1 H NMR (400 MHz, chloroform-d) δ 7.80-7.73 (m, 1H), 7.39-7.30 (m, 2H), 3.78 (s, 3H), 1.51 (s, 9H); DCM signal was observed at 5.3 ppm.

[0208] Intermediate 11 tert-Butyl 4-(6-fluoro-1-methyl-2-oxo-3-(pyridin-4-yl)-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate [ka]

[0209] tert-Butyl 4-(3-bromo-6-fluoro-1-methyl-2-oxo-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate (562 mg, 1.276 mmol) and 4-(tributylstannyl)pyridine (0.392 mL, 1.276 mmol) were dissolved in anhydrous DMF (10 mL). The reaction flask was evacuated and flushed with nitrogen (3x). Tetrakis (147 mg, 0.128 mmol) was then added and the evacuation / flush procedure was repeated twice. The reaction mixture was heated at 100° C. under nitrogen in a sealed tube for 48 hours. The reaction mixture was then fractionated via flash chromatography using 5% MeOH in DCM. Homogeneous fractions were combined and evaporated under vacuum to give the product as a yellow oil (928 mg).

[0210] Intermediate 12 6-Fluoro-1-methyl-4-(piperazin-1-yl)-3-(pyridin-4-yl)quinolin-2(1H)-one [ka]

[0211] Trifluoroacetic acid (8.15 mL, 106 mmol) was added to a solution of tert-butyl 4-(6-fluoro-1-methyl-2-oxo-3-(pyridin-4-yl)-1,2-dihydroquinolin-4-yl)piperazine-1-carboxylate (928 mg, 2.116 mmol) in DCM (8 mL). The reaction mixture was stirred at room temperature for 1 h. The mixture was then diluted with water and the strongly acidic aqueous layer was separated from the dichloromethane solution. The organic layer was re-extracted with more water (the aqueous extract remained strongly acidic). The aqueous extracts were combined and then made basic (pH approx. 14) by the addition of solid KOH. The resulting mixture was extracted with DCM (3x) and the extracts were combined, dried over MgSO4, filtered and evaporated under vacuum to give an oil that solidified on standing overnight (243 mg).

[0212] Example 2 6-Fluoro-4-{4-[(3-fluoro-5-methylphenyl)methyl]piperazin-1-yl}-1-methyl-3-(pyridin-4-yl)-1,2-dihydroquinolin-2-one [ka]

[0213] To a solution of 6-fluoro-1-methyl-4-(piperazin-1-yl)-3-(pyridin-4-yl)quinolin-2(1H)-one (22 mg, 0.065 mmol) in DMF (1.5 mL) was added 4-formyl-3-hydroxybenzonitrile (14.35 mg, 0.098 mmol). The reaction mixture was stirred at room temperature for 1 hour, after which sodium cyanoborohydride (8.18 mg, 0.130 mmol) was added. The reaction mixture was stirred at room temperature overnight. The reaction product was purified via preparative LC / MS using the following conditions: Column: X-Bridge C18, 19x200mm, 5μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Gradient: 35-75% B over 20 min, then hold at 100% B for 5 min; Flow: 20mL / min. Fractions containing the product were combined and dried via centrifugal evaporation. The product yield was 13.5mg and its purity estimated by LCMS analysis was 100%. Analytical LC / MS was used to determine the final purity. Conditions for injection 1: Column: Waters X-Bridge C18, 2.1 mm x 50 mm, 1.7 mm particles; Mobile phase A: 5:95 acetonitrile:water + 0.1% trifluoroacetic acid; Mobile phase B: 95:5 acetonitrile:water + 0.1% trifluoroacetic acid; Temperature: 50 °C; Gradient: 0% B to 100% B over 3 min, then hold at 100% B for 0.75 min; Flow: 1 mL / min; Detection: MS and UV (220 nm). Results for injection 1: Purity: 100.0%; Observed mass: 461.22; Retention time: 1.48 min. Conditions for injection 2: Column: Waters X-Bridge C18, 2.1 mm x 50 mm, 1.7 μm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Temperature: 50 °C; Gradient: 0% B to 100% B over 3 min, then hold at 100% B for 0.75 min; Flow: 1 mL / min; Detection: MS and UV (220 nm). Results for injection 2: Purity: 100.0%; Observed mass: 461.2; Retention time: 2.26 min. 1H NMR(500MHz, DMSO-d6)δ 8.74(d,J=6.6Hz,2H), 7.79(d,J=6.6Hz,2H), 7.70-7.63(m,2H), 7.62-7.53(m,1H), 7.00-6.92(m,1H ), 6.89(brd,J=8.8Hz,2H), 3.63(s,3H), 3.49(s,2H), 2.82(brs,4H), 2.50-2.44(m,4H), 2.31(s,3H)

[0214] Intermediate 13 4-(ピペラジン-1-イルメチル)-1H-インドール·2TFA

change

[0215] To a solution of tert-butyl piperazine-1-carboxylate (481 mg, 2.58 mmol) in DMF (1.5 mL) was added 1H-indole-4-carbaldehyde (250 mg, 1.722 mmol). The reaction mixture was stirred at room temperature for 1 h. Sodium cyanoborohydride (325 mg, 5.17 mmol) was then added and the reaction mixture was stirred at room temperature for 6 days. The reaction mixture was then added to methanol, filtered and the filtrate was subjected to preparative HPLC and purified using a CH3CN-H2O-TFA buffer system. The fractions were collected and concentrated under SpeedVac overnight. LC / MS showed the majority of the material was free of boc groups. The material was dissolved in dichloromethane (5 mL) and 1 mL of TFA was added. The reaction mixture was stirred at room temperature for 3 h and concentrated to give 4-(piperazin-1-ylmethyl)-1H-indole·2TFA (420 mg, 0.947 mmol, 55.0% yield) as a white solid. LC / MS: Column: Phenomenex Luna C18, 2.0x50 mm, 3 mm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Temperature: 40 °C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8 mL / min; Detection: UV (220 nm); Retention time: 1.577 min, 216.1 (MH + )

[0216] Example 3 4-(4-((1H-indol-4-yl)methyl)piperazin-1-yl)-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one [ka]

[0217] To a solution of 4-chloro-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one (150 mg, 0.585 mmol) in DMF (5 mL) was added 4-(piperazin-1-ylmethyl)-1H-indole·2TFA (285 mg, 0.643 mmol) and triethylamine (0.326 mL, 2.338 mmol). The reaction mixture was stirred at room temperature over the weekend. LC / MS showed the reaction was complete. The crude product was purified by preparative HPLC using a CH3CN-H2O-TFA buffer system. The fractions were collected and concentrated under SpeedVac overnight. The product was purified again by preparative HPLC using a CH3CN-H2O-ammonium acetate buffer system. The fractions were collected and concentrated under SpeedVac overnight. A yellow solid was obtained as the final product 4-(4-((1H-indol-4-yl)methyl)piperazin-1-yl)-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one (180 mg, 0.413 mmol, 70.7% yield). LC / MS: Column: Phenomenex Luna C18, 2.0x50 mm, 3 mm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Temperature: 40 °C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8 mL / min; Detection: UV (220 nm); Retention time: 3.1 min, 436.1 (MH + )

[0218] Example 4 4-{4-[(1-ethyl-1H-indol-4-yl)methyl]piperazin-1-yl}-6-fluoro-1-methyl-3-nitro-1,2-dihydroquinolin-2-one [ka]

[0219] To a solution of 4-(4-((1H-indol-4-yl)methyl)piperazin-1-yl)-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one (15 mg, 0.034 mmol) in DMF (1.5 mL) was added 60% sodium hydride dispersion in mineral oil (2.76 mg, 0.069 mmol). The reaction mixture was stirred at room temperature for 15 minutes. Iodoethane (10.75 mg, 0.069 mmol) was then added. The reaction mixture was stirred at room temperature overnight. LC / MS showed the reaction was complete. The reaction was quenched with methanol. The reaction mixture was filtered and the filtrate was purified via preparative LC / MS using the following conditions: Column: X-Bridge C18, 19x200mm, 5μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Gradient: 20-100% B over 15 min, then hold at 100% B for 5 min; Flow: 20mL / min. Fractions containing the product were combined and dried via centrifugal evaporation. LC / MS conditions: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm): Measured MS ion: 464.1, retention time: 2.28 min; 1 H NMR (500MHz, DMSO-d6)δ 7.80-7.65(m,2H), 7.63-7.49(m,1H), 7.45-7.32(m,2H), 7.17-7.06(m,1H), 7.03-6.94(m,1H), 6.73-6.51(m,1) H), 4.20(q,J=9.3Hz,2H), 3.83(s,2H), 3.64(s,3H), 3.16-3.09(m,4H), 2.71-2.59(m,4H), 1.37(t,J=9.3Hz,3H)

[0220] Example 5 4-(4-benzyl-3-oxopiperazin-1-yl)-6-fluoro-1-methyl-3-nitro-1,2-dihydroquinolin-2-one [ka]

[0221] 4-Chloro-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one (25 mg, 0.097 mmol), 1-benzylpiperazin-2-one (20.39 mg, 0.107 mmol) and Hunig's Base (0.037 mL, 0.214 mmol) were dissolved in DMF (2 mL). The reaction mixture was stirred at room temperature overnight. The solution containing the product was analyzed by preparative LC / MS under the following conditions: Column: X-Bridge C18, 19x200 mm, 5 μm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Gradient: 21-61% B over 20 min, then hold at 100% B for 4 min; Flow: 20 mL / min. Fractions containing the product were combined and dried via centrifugal evaporation. The yield of the product was 24.1 mg and its purity estimated by LCMS analysis was 100%. Final purity was determined using analytical LC / MS. Conditions for injection 1: Column: Waters X-Bridge C18, 2.1 mm x 50 mm, 1.7 μm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Temperature: 50 °C; Gradient: 0% B to 100% B over 3 min, then hold at 100% B for 0.75 min; Flow: 1 mL / min; Detection: MS and UV (220 nm). Results for injection 1: Purity: 100.0%; Observed mass: 411.2; Retention time: 1.6 min. Conditions for injection 2: Column: Waters X-Bridge C18, 2.1mmx50mm, 1.7μm particles; Mobile phase A: 5:95 acetonitrile:water + 0.1% trifluoroacetic acid; Mobile phase B: 95:5 acetonitrile:water + 0.1% trifluoroacetic acid; Temperature: 50°C; Gradient: 0%B to 100%B over 3 min, then hold at 100%B for 0.75 min; Flow: 1mL / min; Detection: MS and UV (220nm). Results for injection 2: Purity: 100.0%; Observed mass: 411.02; Retention time: 1.6 min. 1H NMR (500MHz, DMSO-d6)δ 7.77-7.61(m,3H), 7.43-7.36(m,2H), 7.35-7.27(m,3H), 4.62(s,2H), 3.88(s,2H), 3.66(s,3H), 2.56(s,4H)

[0222] Intermediate 14 4-Hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbonitrile [ka]

[0223] In a round bottom flask, methyl 2-cyanoacetate (4.95 g, 50 mmol) was dissolved in NMP (40 mL) and the mixture was cooled to 0° C. Sodium hydride (1.818 g, 50.0 mmol) was added and the reaction mixture was stirred at 0° C. for 5 min and then at room temperature for 15 min. At room temperature, 1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione (8.86 g, 50.0 mmol) was added and the reaction mixture was heated at 120° C. for 2 h. LCMS analysis showed the reaction was complete. The reaction mixture was diluted with 60 mL of diethyl ether and the mixture was poured onto 50 mL of ice and 100 mL of 1N HCl. The resulting solid was collected (6 g). Evaporation of the ether formed additional solids in the filtrate and these solids were collected separately (2.2 g). NMR and LCMS of both solid samples were identical and consistent with the product. LC / MS 2.15 min, M+1: 201.2, exact mass: 200.1; starting %B = 0, final %B = 100, gradient time = 4 min, flow rate = 0.8 mL / min, wavelength = 220 nm, solvent pair = water-methanol-0.1% TFA, solvent A = 90% water-10% methanol-0.1% TFA, solvent B = 10% water-90% methanol-0.1% TFA, column = Phenomenex Luna 2.0x50mm 3μm; 1H NMR (400MHz, DMSO-d6)δ 8.11(dd,J=8.1, 1.2Hz,1H), 7.77(ddd,J=8.6, 7.2, 1.6Hz,1H), 7.56(d,J=8.6Hz,1H), 7.34(t,J=7.6Hz,1H), 3.57(s,3H)

[0224] Intermediate 15 4-Chloro-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbonitrile [ka]

[0225] In a round bottom flask, 1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinoline-3-carbonitrile (6 g, 30.0 mmol) and phosphorus oxychloride (13.97 ml, 150 mmol) were combined at room temperature. The reaction mixture was heated at 110° C. for 40 minutes. After cooling to room temperature, the volatiles were removed on a Rotovap. To the crude material, 50 mL of diethyl ether was added and the volatiles were removed again on a Rotovap. Water was added slowly with vigorous stirring and the solid was collected by filtration. The residue was placed under vacuum overnight to give the title compound (4.6 g) as a pale yellow solid. 1 H NMR (400 MHz, chloroform-d) δ 8.14 (dd, J = 8.2, 1.3 Hz, 1H), 7.82 (ddd, J = 8.6, 7.1, 1.5 Hz, 1H), 7.51-7.42 (m, 2H), 3.79 (s, 3H)

[0226] Example 6 4-[(2S,5R)-4-[bis(4-fluorophenyl)methyl]-2,5-dimethylpiperazin-1-yl]-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbonitrile (homochiral) [ka]

[0227] A microwave vial was charged with 4-chloro-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbonitrile (25 mg, 0.114 mmol), (2R,5S)-1-(bis(4-fluorophenyl)methyl)-2,5-dimethylpiperazine (36.2 mg, 0.114 mmol) and Hunig's base (0.040 mL, 0.229 mmol). The mixture was dissolved in NMP (2 mL). The reaction mixture was heated at 150° C. for 14 hours and then cooled to room temperature. The mixture was filtered through a 4 μm frit, concentrated and the residue was purified by preparative HPLC (Column: X-Bridge C18, 200 mm x 19 mm, 5 μm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Gradient: 41% B hold for 0 min, 41-81% B over 22 min, then 100% B hold for 6 min; Flow rate: 20 mL / min; Column temperature: 25 °C). Fraction collection was triggered by MS and UV signals. Fractions containing the product were combined and dried via centrifugal evaporation. LC / MS conditions QC-ACN-AA-XB: LCMS: m / z = 499.2 (M+H); Retention time: 2.58 min

[0228] Example 7 4-[(2S,5R)-4-[bis(4-fluorophenyl)methyl]-2,5-dimethylpiperazin-1-yl]-1-methyl-2-oxo-1,2-dihydroquinoline-6-carbonitrile (homochiral) [ka]

[0229] To a stirred solution of 4-((2S,5R)-2,5-dimethylpiperazin-1-yl)-1-methyl-2-oxo-1,2-dihydroquinoline-6-carbonitrile (0.08 g, 0.270 mmol) in acetonitrile (5 mL) at room temperature, DIPEA (0.141 mL, 0.810 mmol) and 4,4'-(bromomethylene)bis(fluorobenzene) (0.115 g, 0.405 mmol) were added. The reaction mixture was heated at 85°C for 16 hours. The reaction mixture was cooled to room temperature and the solvent was removed under reduced pressure. The residue was dissolved in ethyl acetate (100 mL). The organic layer was washed with brine, dried over Na2SO4 and concentrated under reduced pressure to give the crude product, which was purified by reverse-phase preparative HPLC (Method: Column: Waters X-Bridge C18, 150mmx19mm, 5μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Gradient: 15% B hold for 0 min, 15-65% B over 25 min, then hold at 100% B for 5 min; Flow rate: 15mL / min). The fractions were concentrated under reduced pressure and purified to give 4-((2S,5R)-4-(bis(4-fluorophenyl)methyl)-2,5-dimethylpiperazin-1-yl)-1-methyl-2-oxo-1,2-dihydroquinoline-6-carbonitrile (50 mg, 0.100 mmol, 37.2% yield). LCMS: m / z=499.2 (M+H); retention time 2.426 min; LCMS method column: X-Bridge BEH XP C18 (50x2.1) mm, 2.5 μm Mobile phase A: 95% water: 5% acetonitrile; 10 mM NH4OAC, Mobile phase B: 5% water: 95% acetonitrile; 10 mM NH4OAC Flow: 1.1 mL / min, Temperature: 50 °C; 1H NMR(400MHz, DMSO-d6)δppm 8.12(d,J=1.71Hz,1H), 7.97(dd,J=8.80,1.71Hz,1H), 7.66(d,J=8.80Hz,1H), 7.51 -7.62(m,4H), 7.15(dt,J=17.61,8.80Hz,4H), 6.06(s,1H), 4.82(s,1H), 3.75-3.83( m,1H), 3.55-3.65(m,1H), 3.33(s,3H), 3.01-3.11(m,2H), 2.65(dd,J=11.86, 2.57Hz ,1H), 2.27(dd,J=11.49,1.96Hz,1H), 1.09(d,J=6.60Hz,3H), 1.04(d,J=6.36Hz,3H)

[0230] Intermediate 16 6-メトキシ-1-メチル-2H-ベンゾ[d][1,3]オキサジン-2,4(1H)-ジオン

change

[0231] To a solution of 6-methoxy-1H-benzo[d][1,3]oxazine-2,4-dione (3 g, 15.53 mmol) in DMF (20 mL) was added 60% sodium hydride dispersion in mineral oil (0.745 g, 18.64 mmol) portionwise at 0° C. The reaction mixture was allowed to warm to room temperature and stirred for 30 minutes. Methyl iodide (1.165 mL, 18.64 mmol) was then added dropwise. The reaction mixture was then stirred at room temperature overnight. The reaction was quenched with water. Diethyl ether was added to the mixture. The product, 6-methoxy-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione (3.11 g, 15.01 mmol, 97% yield), was collected as a beige solid. LC / MS: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 2.032 min, 208.0 (M+H + ); 1 H NMR (400MHz, DMSO-d6)δ 7.68-7.23(m,3H), 3.85(s,3H), 3.46(s,3H)

[0232] Intermediate 17 6-Methoxy-1-methyl-3-nitroquinoline-2,4(1H,3H)-dione [ka]

[0233] To a solution of ethyl 2-nitroacetate (2.120 g, 15.93 mmol) in NMP (10 mL) in a round bottom flask at 0° C., 60% sodium hydride dispersion in mineral oil (0.695 g, 17.38 mmol) was added in portions. The reaction mixture was stirred at 0° C. for 5 min and then at room temperature for 15 min. 6-Methoxy-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione (3 g, 14.48 mmol) was then added and the reaction mixture was heated at 120° C. for 3 h. LC / MS showed the reaction was complete. The reaction was quenched with ice water. The mixture was acidified with 1N HCl solution. Ethyl ether was added to give 6-methoxy-1-methyl-3-nitroquinoline-2,4(1H,3H)-dione (1.66 g, 6.63 mmol, 45.8% yield) as a yellow solid. LC / MS: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 1.613 min, 251.1 (M+H + ); 1 H NMR (400MHz, DMSO-d6)δ 7.61(d,J=2.7Hz,1H), 7.54(d,J=9.3Hz,1H), 7.39(dd,J=9.2, 2.8Hz,1H), 3.85(s,3H), 3.59(s,3H)

[0234] Intermediate 18 4-Chloro-6-methoxy-1-methyl-3-nitroquinolin-2(1H)-one [ka]

[0235] In a sealed tube, 6-methoxy-1-methyl-3-nitroquinoline-2,4(1H,3H)-dione (1.65 g, 6.59 mmol) and phosphorus oxychloride (10 mL, 107 mmol) were added. The reaction mixture was heated at 95° C. for 4 h. The reaction mixture was then concentrated, and the residue was poured into ice water and neutralized with solid NaHCO3. The mixture was extracted with dichloromethane (2×40 mL), and the organic layers were combined, dried (MgSO4), and concentrated to give 4-chloro-6-methoxy-1-methyl-3-nitroquinolin-2(1H)-one (1.58 g, 5.88 mmol, 89% yield) as an orange solid. LC / MS: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 2.703 min, 269.1 (M+H + ); 1 H NMR (500MHz, DMSO-d6) δ 7.76(d,J=9.2Hz,1H), 7.57(d,J=9.5Hz,1H), 7.47(s,1H), 3.91(s,3H), 3.72(s,3H)

[0236] Example 8 6-Methoxy-1-methyl-4-{4-[(naphthalen-1-yl)methyl]piperazin-1-yl}-3-nitro-1,2-dihydroquinolin-2-one [ka]

[0237] To a solution of 4-chloro-6-methoxy-1-methyl-3-nitroquinolin-2(1H)-one (15 mg, 0.056 mmol) in DMF (1 mL) was added 1-(naphthalen-1-ylmethyl)piperazine (12.64 mg, 0.056 mmol) and triethylamine (0.023 mL, 0.168 mmol). The reaction mixture was stirred at room temperature overnight. LC / MS showed the reaction was complete. CH3CN was then added to the reaction mixture. The reaction mixture was purified via preparative LC / MS using the following conditions (Column: X-Bridge C18, 19x200mm, 5μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Gradient: 50-90% B over 15 min, then hold at 100% B for 5 min; Flow: 20mL / min). Fractions containing the product were combined and dried via centrifugal evaporation. LC / MS: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 40°C; Gradient: 0% B, 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Measured MS ion: 459.2, retention time: 2.34 min; 1 H NMR (500MHz, DMSO-d6)δ 8.31(d,J=8.4Hz,1H), 7.95-7.89(m,1H), 7.88-7.83(m,1H), 7.61-7.37(m,6H), 7.3 6-7.31(m,1H), 4.00(s,2H), 3.88(s,3H), 3.60(s,3H), 3.13(brs,4H), 2.69(brs,4H)

[0238] Intermediate 19 4-Chloro-1-methylquinolin-2(1H)-one [ka]

[0239] 4-Hydroxy-1-methylquinolin-2(1H)-one (1.4 g, 7.99 mmol) and phosphorus oxychloride (7.45 ml, 80 mmol) were combined in a 100 mL round bottom flask equipped with a stir bar and reflux condenser at room temperature. The round bottom flask was placed in a 95° C. oil bath. After 2.5 hours, the reaction mixture was allowed to cool to room temperature. The reaction mixture was slowly added to 300 mL of ice water. The slurry was extracted with 200 mL of dichloromethane and removed due to the presence of very small lumps after rotary evaporation. The aqueous portion was added to excess solid sodium bicarbonate. Dichloromethane (300 mL) was added and the remaining solids were removed by filtration. The organic portion was dried over magnesium sulfate. The solvent was removed by rotary evaporation to give an orange solid (1.1 g). Starting materials: 1 H NMR (400MHz, DMSO-d6)δ 11.30(s,1H), 7.90(dd,J=7.8, 1.5Hz,1H), 7.68-7.59(m,1H), 7.48(d,J=8.6Hz,1H), 7.25(t,J=7.6Hz,1H), 5.88(s,1H), 3.54(s,3H) Product: 1H NMR (400 MHz, DMSO-d6) δ 7.98 (dd, J = 8.1, 1.2 Hz, 1H), 7.81-7.72 (m, 1H), 7.65 (d, J = 8.3 Hz, 1H), 7.46-7.37 (m, 1H), 6.96 (s, 1H), 3.64 (s, 3H); LC / MS 2.32 min, M+1: 194.0, exact mass: 193.03, starting %B = 0, final %B = 100, gradient time = 4 min, flow rate = 0.8 ml / min, wavelength = 220, solvent pair = ACN:water:ammonium acetate, solvent A = 5% ACN:95% water:10 mM ammonium acetate, solvent B = 95% ACN:5% water:10 mM ammonium acetate, column = Phenomenex Luna C18, 30x2, 3μm, oven temp=40°C, LC / MS 3.02 min, M+1:194.1, exact mass:193.03, starting %B=0, final %B=100, gradient time=4 min, flow rate=0.8mL / min, wavelength=220nm, solvent pair=water-methanol-0.1%TFA, solvent A=90% water-10% methanol-0.1%TFA, solvent B=10% water-90% methanol-0.1%TFA, column=Phenomenex-Luna 2.0x50mm 3μm

[0240] Example 9 4-(4-Benzhydrylpiperazin-1-yl)-1-methylquinolin-2(1H)-one [ka]

[0241] In a microwave reaction vial, 1-benzhydrylpiperazine (0.555 g, 2.200 mmol) and 4-chloro-1-methylquinolin-2(1H)-one (0.387 g, 2 mmol) were dissolved in NMP (8 mL). The reaction vial was sealed under argon and heated at 200° C. for 30 min. The crude reaction mixture was directly chromatographed by reverse phase HPLC (start %B=30 to final %B=80, gradient time=10 min, flow rate=35 mL / min, wavelength=220 nm, solvent A=10% methanol, 90% water+0.1% TFA, solvent B=90% methanol, 10% water+0.1% TFA, column=Phenomenex-Luna 30x50 mm S10). The product was obtained as a white solid (250 mg). 1 H NMR (500MHz, DMSO-d6)δ 7.74(d,J=7.7Hz,1H), 7.62-7.55(m,1H), 7.53-7.45(m,5H), 7.33(t,J=7.5Hz,4H), 7.22(q,J=6.8H z,3H), 6.01(s,1H), 4.42(s,1H), 3.55(s,3H), 3.40(d,J=3.7Hz,2H), 3.09(brs,3H), 2.56(brs,3H)

[0242] Example 10 4-(4-Benzhydrylpiperazin-1-yl)-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbaldehyde [ka]

[0243] DMF (1 mL) was cooled in ice water and phosphorus oxychloride (140 μL, 1.500 mmol) was added. After stirring for 10 min, 4-(4-benzhydrylpiperazin-1-yl)-1-methylquinolin-2(1H)-one (205 mg, 0.5 mmol) in 1 mL of DMF was added and the reaction mixture was stirred at room temperature. After 20 min, LC / MS analysis showed the presence of product and the absence of starting material. Excess ice was added and the reaction mixture was stirred until the ice melted. The pH was adjusted to 10 with 1N aqueous NaOH. The yellow precipitate (108 mg) was collected using a Buchner funnel and dried under vacuum overnight. Proton NMR showed it to be the desired aldehyde. 1 H NMR (400 MHz, chloroform-d) δ 10.44 (s, 1H), 8.16-7.89 (m, 5H), 7.39 (brs, 10H), 5.22-5.03 (m, 1H), 4.64-4.29 (m, 2H), 3.66-3.35 (m, 5H).

[0244] Example 11 4-[4-(diphenylmethyl)piperazin-1-yl]-3-(hydroxymethyl)-1-methyl-1,2-dihydroquinolin-2-one [ka]

[0245] In a reaction vial, sodium borohydride (0.011 g, 0.300 mmol) and 4-(4-benzhydrylpiperazin-1-yl)-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbaldehyde (0.044 g, 0.1 mmol) were combined in methanol (0.500 mL). Gas evolution was observed and the solution went from yellow to colorless. The crude material was purified via preparative LC / MS using the following conditions (Column: X-Bridge C18, 19x200 mm, 5 μm particles; Mobile phase A: 5:95 acetonitrile:water + 10 mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10 mM ammonium acetate; Gradient: 45-85% B over 20 min, then hold at 100% B for 5 min; Flow: 20 mL / min). Fractions containing the product were combined and dried via centrifugal evaporation. The yield of the product was 6.3 mg and its purity estimated by LCMS analysis was 100%. Two analytical LC / MS injections were used to determine the final purity. Conditions for injection 1: Column: Waters Acquity UPLC BEH C18, 2.1x50mm, 1.7μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 50°C; Gradient: 0-100% B over 3 min, then hold at 100% B for 0.75 min; Flow: 1.0mL / min; Detection: UV (220nm); LC / MS: 2.2 min, M+1=440.1. Conditions for injection 2: Column: Waters Acquity UPLC BEH C18, 2.1x50mm, 1.7μm particles; Mobile phase A: 5:95 acetonitrile:water + 0.1% trifluoroacetic acid; Mobile phase B: 95:5 acetonitrile:water + 0.1% trifluoroacetic acid; Temperature: 50°C; Gradient: 0-100% B over 3 min, then hold at 100% B for 0.75 min; Flow: 1.0mL / min; Detection: UV (220nm); LC / MS: 1.4 min, M+1=440.3. Proton NMR was acquired in deuterated DMSO with water suppression. No piperazine hydrogens were observed and the integration of the methyl peak was low, consistent with water suppression. 1H NMR (500MHz, DMSO-d6)δ 7.96(d,J=8.1Hz,1H), 7.60-7.54(m,1H), 7.53-7.47(m,5H), 7.33(t,J=7.5Hz,4H), 7.26(t,J=7 .7Hz,1H), 7.23-7.18(m,2H), 4.78-4.71(m,1H), 4.61(d,J=5.1Hz,2H), 4.43(s,1H), 3.61(s,2H)

[0246] Intermediate 20 Ethyl 4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate [ka]

[0247] In a round bottom flask, diethyl malonate (6.8 g, 42.5 mmol) was dissolved in NMP (40 mL) and cooled to 0° C. Sodium hydride (1.08 g, 29.7 mmol) was added and the reaction mixture was stirred at 0° C. for 5 min and then at room temperature for 15 min. At room temperature, 1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione (5.1 g, 28.8 mmol) was added and the reaction mixture was heated at 120° C. for 3 h. Diethyl ether (60 mL) was added and the solid was collected on a filter. The solid was washed with an additional 30 mL of diethyl ether. The solid was suspended in 25 mL of methanol and 50 mL of water. Hydrochloric acid (6 mL, 5 M aqueous) was added and the mixture was extracted three times with 80 mL portions of ethyl acetate. The organic layers were combined and extracted with 1×20 mL of brine. The organic portion was dried over magnesium sulfate. The solvent was removed by rotary evaporation to give the title compound (5.3 g, 51%). 1 H NMR (400 MHz, chloroform-d) δ 14.23 (s, 1H), 8.21 (dd, J = 8.0, 1.5 Hz, 1H), 7.74-7.65 (m, 1H), 7.33 (d, J = 8.5 Hz, 1H), 7.30-7.25 (m, 1H), 4.52 (q, J = 7.2 Hz, 2H), 3.67 (s, 3H), 1.50 (t, J = 7.2 Hz, 3H).

[0248] Intermediate 21 Ethyl 4-chloro-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate [ka]

[0249] In a round bottom flask, ethyl 1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinoline-3-carboxylate (5 g, 20.22 mmol) and phosphorus oxychloride (5 ml, 53.6 mmol) were added. The reaction mixture was heated at 110° C. for 40 min. Volatiles were removed on a rotovap. To the crude residue, 50 mL of ice was added portionwise over 15 min, followed by 150 mL of water. The mixture was stirred for 15 min and ethyl acetate was added. The organic portion was extracted with 20 mL portions of 1× water, 3×1N aqueous sodium hydroxide, 1× water and 1× brine. The organic portion was dried over magnesium sulfate and the solvent was removed by rotary evaporation. The residue was placed under vacuum overnight to give 3.9 g of a pale yellow solid. 1 H NMR (400 MHz, chloroform-d) δ 8.09 (dd, J = 8.2, 1.1 Hz, 1H), 7.73-7.68 (m, 1H), 7.43 (d, J = 8.5 Hz, 1H), 7.38 (t, J = 7.7 Hz, 1H), 4.49 (q, J = 7.0 Hz, 2H), 3.75 (s, 3H), 1.44 (t, J = 7.2 Hz, 3H)

[0250] Intermediate 22 Ethyl 6-fluoro-1-methyl-2-oxo-4-(((trifluoromethyl)sulfonyl)oxy)-1,2-dihydroquinoline-3-carboxylate [ka]

[0251] 1,1,1-Trifluoro-N-phenyl-N-(trifluoromethyl)sulfonyl methanesulfonamide (404 mg, 1.131 mmol) was added to a solution of ethyl 6-fluoro-1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinoline-3-carboxylate (100 mg, 0.377 mmol) and DIEA (0.263 mL, 1.508 mmol) in THF (3 mL). The resulting solution was stirred at room temperature over the weekend. LC / MS showed the reaction was complete. The reaction mixture was concentrated. The crude product was purified on a Biotage column using hexane to 40% ethyl acetate in hexane as eluent. The product came off at approximately 30% ethyl acetate in hexane. The fractions were collected and concentrated to give ethyl 6-fluoro-1-methyl-2-oxo-4-(((trifluoromethyl)sulfonyl)oxy)-1,2-dihydroquinoline-3-carboxylate (71 mg, 0.179 mmol, 47.4% yield) as a light yellow solid. LC / MS: Column: Phenomenex Luna C18, 2.0x50 mm, 3 mm particles; Mobile phase A: 10:90 methanol:water + 0.1% TFA; Mobile phase B: 90:10 methanol:water + 0.1% TFA; Gradient: 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8 mL / min; Detection: UV (220 nm); Retention time: 3.9 min, 397.9 (M+H + ); 1 H NMR (400 MHz, chloroform-d) δ 7.60 (dd, J = 8.3, 2.7 Hz, 1H), 7.55-7.44 (m, 2H), 4.48 (q, J = 7.1 Hz, 2H), 3.78 (s, 3H), 1.44 (t, J = 7.2 Hz, 3H)

[0252] Intermediate 23 1-((2-Methoxynaphthalen-1-yl)methyl)piperazine [ka]

[0253] A solution of 1-BOC-piperazine (2.8 g, 15.03 mmol) and 2-methoxy-1-naphthaldehyde (2.80 g, 15.03 mmol) in DMF (10 mL) was stirred at room temperature for 1 h. Sodium cyanoborohydride (1.417 g, 22.55 mmol) was added and the reaction mixture was stirred at room temperature overnight. LC / MS analysis showed the reaction was complete. The solvent was removed and the residue was purified on a 40 g silica column using 1:1 hexane / ethyl acetate. The fractions containing tert-butyl 4-((2-methoxynaphthalen-1-yl)methyl)piperazine-1-carboxylate were combined and the solvent was removed using a rotary evaporator. The solid material was dissolved in dichloromethane (20 mL) and TFA (5 mL, 64.9 mmol) was added. The reaction mixture was stirred for 1 h and the solvent was removed to give a white solid (3.85 g). LCMS: Column: Phenomenex Luna C18, 2.0x50mm, 3mm particles; Mobile phase A: 10:90 methanol:water + 0.1% TFA; Mobile phase B: 90:10 methanol:water + 0.1% TFA; Gradient: 0-100% B over 4 min, then hold at 100% B for 1 min; Flow: 0.8mL / min; Detection: UV (220nm); Retention time: 0.8 min, M+1=257.3

[0254] Example 12 Ethyl 4-{4-[(2-methoxynaphthalen-1-yl)methyl]piperazin-1-yl}-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate [ka]

[0255] In a microwave reaction vial, 1-((2-methoxynaphthalen-1-yl)methyl)piperazine (1.55 g, 6.05 mmol), ethyl 4-chloro-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate (1.607 g, 6.05 mmol) and triethylamine (0.843 mL, 6.05 mmol) were combined in DMF (12 mL). The reaction mixture was heated in a microwave reaction vessel at 100° C. for 1 h and then allowed to stand at room temperature over the weekend. Over the weekend, precipitated crystals formed that were consistent with triethylammonium chloride by NMR. The liquid portion was poured into 100 mL of water and the precipitated solid was collected. NMR and LCMS of the solid after drying indicated the product was contaminated with traces of starting material and DMF. 1H NMR (400MHz, chloroform-d)δ 8.27(d,J=8.6Hz,1H), 8.01(dd,J=8.1, 1.5Hz,1H), 7.82(dd,J=11.4, 8.7Hz,2H), 7.64-7.48(m,2H), 7.47-7.23(m,5H, CHCl3 overlap ), 4.39(q,J=7.2Hz,2H), 4.09(s,2H), 4.00(s,3H), 3.67(s,3H), 3.22(brt,J=4.5Hz,4H), 2.77(brs,3H), 1.38(t,J=7.1Hz,3H). The crude material was purified via preparative LC / MS using the following conditions (Column: X-Bridge C18, 19x200mm, 5μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Gradient: 50-90% B over 15 min, then hold at 100% B for 5 min; Flow: 20mL / min). Fractions containing the product were combined and dried via centrifugal evaporation. The yield of the product was 22.3mg and its purity estimated by LCMS analysis was 100%. Two analytical LC / MS injections were used to determine the final purity. Conditions for injection 1: Column: Waters BEH C18, 2.0x50mm, 1.7μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 50°C; Gradient: 0% B, 0-100% B over 3 min, then hold at 100% B for 0.5 min; Flow: 1mL / min; Detection: UV (220nm); LCMS: 2.07 min, M+1=486.23, M-1=484.23. Conditions for injection 2: Column: Waters BEH C18, 2.0x50mm, 1.7μm particles; Mobile phase A: 5:95 methanol:water + 10mM ammonium acetate; Mobile phase B: 95:5 methanol:water + 10mM ammonium acetate; Temperature: 50°C; Gradient: 0% B, 0-100% B over 3 min, then hold at 100% B for 0.5 min, then hold at 100% B for 0.5 min; Flow: 0.5mL / min; Detection: UV (220nm); LCMS: 3.22 min, M+1=486.23, M-1=484.23; 1H NMR (500MHz, DMSO-d6)δ 8.19(d,J=8.8Hz,1H), 7.99-7.89(m,2H), 7.85(d,J=8.1Hz,1H), 7.73-7.61(m,1H), 7.57-7.49(m,2H), 7.46(d,J=9.2Hz,1H), 7.36( q,J=7.8Hz,2H), 4.22(q,J=7.0Hz,2H), 4.01(s,2H), 3.94(s,3H), 3.56(s,3H), 3.03(brs,4H), 2.69(brs,4H), 1.23(t,J=7.0Hz,3H)

[0256] Intermediate 24 2,4-Dichloro-3-nitroquinoline [ka]

[0257] To a 100 mL round bottom flask was added 3-nitroquinoline-2,4-diol (2 g, 9.70 mmol) and phosphoryl trichloride (9.04 ml, 97 mmol). To this yellow slurry was added N,N-dimethylaniline (2.485 ml, 19.40 mmol) and the reaction mixture was stirred at 120° C. for 1.5 h. The reaction mixture was cooled and concentrated. The brown residue was dissolved in toluene (10 mL) and concentrated to remove any unreacted POCl3. The residue was treated with ice (25 g) for 20 min and carefully neutralized with solid Na2CO3. The mixture was extracted with ether (3×20 mL). The organic layers were combined and dried to give a light brown product.

[0258] Intermediate 25 4-{4-[bis(4-fluorophenyl)methyl]piperazin-1-yl}-2-chloro-3-nitroquinoline [ka]

[0259] To a solution of 2,4-dichloro-3-nitroquinoline (1 g, 4.11 mmol) in DMF (10 mL) was added 1-(bis(4-fluorophenyl)methyl)piperazine (1.186 g, 4.11 mmol) and triethylamine (1.147 mL, 8.23 ​​mmol). The reaction mixture was stirred at room temperature overnight. LC / MS analysis showed the reaction was complete. The mixture was diluted with ethyl acetate, washed with water, brine, and dried. The residue was purified via silica gel chromatography (1:2 hexane:ethyl acetate; 24 g column). Fractions were collected to give the product as a yellow solid (2 g). The product (0.1 mmol) was purified via preparative LC / MS using the following conditions: Column: X-Bridge C18, 19x200mm, 5μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Gradient: 80-100% B over 15 min, then hold at 100% B for 5 min; Flow: 20mL / min. Fractions containing the product were combined and dried via centrifugal evaporation. The product yield was 5.0mg and its purity estimated by LCMS analysis was 100%. Two analytical LC / MS injections were used to determine the final purity. Conditions for injection 1: Column: Waters BEH C18, 2.0x50mm, 1.7μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 50°C; Gradient: 0-100% B over 3 min, then hold at 100% B for 0.5 min; Flow: 1.0mL / min; Detection: UV (220nm). Conditions for injection 2: Column: Waters BEH C18, 2.0x50mm, 1.7μm particles; Mobile phase A: 5:95 methanol:water + 10mM ammonium acetate; Mobile phase B: 95:5 methanol:water + 10mM ammonium acetate; Temperature: 50°C; Gradient: 0-100% B over 3 min, then hold at 100% B for 0.5 min; Flow: 0.5mL / min; Detection: UV (220nm); 1H NMR (500MHz, DMSO-d6)δ 8.11(d,J=8.4Hz,1H), 8.00-7.95(m,1H), 7.94-7.88(m,1H), 7.73(t,J=7.7Hz,1H), 7.51 (dd,J=8.1, 5.9Hz,4H), 7.16(t,J=8.6Hz,4H), 4.53(s,1H), 3.31(brs,4H), 2.57(brs,4H)

[0260] Example 13 4-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-3-nitroquinolin-2(1H)-one [ka]

[0261] To 4-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-2-chloro-3-nitroquinoline (600 mg, 1.212 mmol) was added AcOH (10 mL, 175 mmol) and water (2.5 mL). The reaction mixture was heated at 100° C. for 5 h. The solvent was removed and dichloromethane was added to give 450 mg of a yellow solid (3:1 product:starting material). The remaining material was purified via Biotage (1:4 to 1:1 hexanes / EtOAc; 40 g silica column). Fractions were collected to give the product (190 mg) as a yellow solid.

[0262] Example 14 4-{4-[bis(4-fluorophenyl)methyl]piperazin-1-yl}-1-(2-fluoroethyl)-3-nitro-1,2-dihydroquinolin-2-one [ka]

[0263] To a solution of Example 13 (20 mg, 0.042 mmol) in DMF was added sodium hydride (2.015 mg, 0.084 mmol) at 0°C. After 5 min, 1-fluoro-2-iodoethane (14.60 mg, 0.084 mmol) was added and the reaction vessel was shaken at room temperature under N2 overnight. LC / MS showed conversion to product. The crude material was purified via preparative LC / MS using the following conditions: Column: X-Bridge C18, 19x200mm, 5μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Gradient: 60-100% B over 15 min, then hold at 100% B for 5 min; Flow: 20mL / min. Fractions containing the desired product were combined and dried via centrifugal evaporation. The calculated molecular weight was 522.528. LC / MS conditions: Waters BEH C18, 2.0x50mm, 1.7μm particles; Mobile phase A: 5:95 acetonitrile:water + 10mM ammonium acetate; Mobile phase B: 95:5 acetonitrile:water + 10mM ammonium acetate; Temperature: 50°C; Gradient: 0% B, 0-100% B over 3 min, then hold at 100% B for 0.5 min; Flow: 1mL / min; Detection: UV (220nm). Measured MS ion: 523.1, retention time: 2.41 min.

[0264] The following compounds were prepared in a library fashion from the appropriate alkyl halides using the procedure above. Analytical LCMS conditions used to characterize the products were the same as in the procedure above, except that acetonitrile was used instead of methanol in the mobile phase where indicated.

[0265] [ka] [Table 3]

[0266] Biological assays The pharmacological properties of the compounds of the invention may be confirmed by a number of biological assays. The following exemplary biological assays have been carried out with the compounds of the invention.

[0267] Assay 1: In vitro DGK inhibition assay (Method A) DGKα and DGKζ reactions were performed with extruded liposomes (LIPGLO assay for DGKα and DGKζ). Reactions were performed in 50 mM MOPS pH 7.5, 100 mM NaCl, 10 mM MgCl2, 1 μM CaCl2, and 1 mM DTT (assay buffer). Lipid substrate concentrations were 2 mM PS, 0.25 mM DAG, and 2.75 mM PC for extruded liposome reactions. Reactions were performed in 150 μM ATP. Enzyme concentrations were 5 nM for DGKα and DGKζ.

[0268] Compound inhibition experiments were performed as follows: 50 nL droplets of each test compound dissolved in DMSO (11-point 3-fold serial dilutions for each compound from a top concentration of 10 mM) were transferred to wells of a white 1536-well plate (Corning 3725). 5 mL of enzyme / substrate solution was prepared at 2x final reaction concentration by combining 2.5 mL of 4x enzyme solution (20 nM DGKα or DGKζ in assay buffer (prepared as described below)) with 2.5 mL of 4x liposome solution (composition described below) and incubated at room temperature for 10 minutes. 1 μL of 2x enzyme / substrate solution was then added to the wells containing the test compound and 1 μL of 300 μM ATP was added to initiate the reaction. The reaction was allowed to proceed for 1 hour, after which 2 μL of Glo reagent (Promega V9101) was added and incubated for 40 minutes. 4 μL of kinase detection reagent was then added and incubated for 30 minutes. Luminescence was recorded using an EnVision microplate reader. Percent inhibition was calculated from ATP conversion generated by enzyme-free control reactions set at 100% inhibition and vehicle-only reactions set at 0% inhibition. Compounds were evaluated at 11 concentrations and IC 50 It was decided.

[0269] Preparation of 4x liposomes The lipid composition was 5 mol% DAG (Avanti 800811O), 40 mol% PS (Avanti 840035P), and 55 mol% PC (Avanti 850457) for the 4x liposome solution, with a total lipid concentration of 15.2 mg / mL. PC, DAG, and PS were dissolved in chloroform, combined, and dried under vacuum to a thin film. Lipids were neutralized to 20 mM in 50 mM MOPS pH 7.5, 100 mM NaCl, 5 mM MgCl2, and subjected to five freeze-thaw cycles. The lipid suspension was extruded 11 times through a 100 nm polycarbonate filter. Dynamic light scattering was performed to confirm the size of the liposomes (radius 50-60 nm). The liposome preparation was stored for a long period of time, up to 4 weeks, at 4 °C.

[0270] Baculovirus expression of human DGKα and DGKζ Baculovirus samples of human DGK-alpha-TVMV-His-pFB gate and human DGK-zeta-transcription variant-2-TVMV-His-pFB gate were prepared using Bac-to-Bac® Baculovirus Expression System (Invitrogen) according to the manufacturer's protocol. The DNA used for the expression of DGK-alpha and DGK-zeta are SEQ ID NOs: 1 and 3, respectively. Baculovirus amplification was performed using Sf9 cells infected with virus / cell at a ratio of 1:1500, and grown at 27°C for 65 hours after transfection. Scale-up of the expression of each protein was performed in a Cellbag 50L WAVE bioreactor system 20 / 50 (GE Healthcare Bioscience). 2x10 6Sf9 cells (Expression Systems, Davis, CA) at 1000 cells / mL were infected with virus stock solution at a virus / cell ratio of 1:200 and grown at 27°C for 66-68 hours post-infection. Infected cell cultures were harvested by centrifugation in a SORVALL® RC12BP centrifuge (2000 rpm, 20 min, 4°C). Cell pellets were stored at -70°C until purification.

[0271] Purification of human DGK-alpha and DGK-zeta Full-length human DGKα and DGKζ, expressed containing a TVMV-cleavable, C-terminal Hex-His tag sequence (SEQ ID NOs: 2 and 4, respectively) and produced as described above, were purified from Sf9 baculovirus-infected insect cell paste. Cells were lysed with a nitrogen disrupter (Parr Instruments) using nitrogen cavitation and the lysate was clarified by centrifugation. The clarified lysate was purified to approximately 90% homogeneity using three consecutive column chromatography steps on an AKTA Purifier Plus system. The three column chromatography steps included nickel affinity resin capture (i.e., HisTrap FF crude, GE Healthcare) followed by size exclusion chromatography (i.e., HiLoad 26 / 600 Superdex 200 preparative grade for DGK-α, GE Healthcare; HiPrep 26 / 600 Sephacryl S 300_HR for DGK-ζ, GE Healthcare). The third step was ion exchange chromatography, which differed between the two isoforms. DGKα was purified using Q-Sepharose anion exchange chromatography (GE Healthcare). DGKζ was purified using SP-Sepharose cation exchange chromatography (GE Healthcare). The proteins were > They were delivered at a concentration of 2 mg / mL. The formulation buffer was the same for both proteins: 50 mM Hepes, pH 7.2, 500 mM NaCl, 10% v / v glycerol, 1 mM TCEP, and 0.5 mM EDTA.

[0272] Assay 2: In vitro DGK inhibition assay (Method B) DGKα and DGKζ reactions were performed with extruded liposomes (LIPGLO assays for DGKα and DGKζ). Reactions were performed in 50 mM MOPS pH 7.5, 100 mM NaCl, 10 mM MgCl2, 1 μM CaCl2, and 1 mM DTT (assay buffer). Lipid substrate concentrations were 2 mM PS, 0.25 mM DAG, and 2.75 mM PC for extruded liposome reactions (5 mM lipid total). Reactions were performed in 150 μM ATP. Enzyme concentrations were 5 nM for DGKα and DGKζ.

[0273] Compound inhibition experiments were performed as follows: 25 nL droplets of each test compound dissolved in DMSO (11-point 3-fold serial dilutions for each compound from a top concentration of 10 mM) were transferred to wells of a white 1536-well plate (Corning 3725). 5 mL of enzyme / lipid substrate solution was prepared at 2x final reaction concentration by combining 2.5 mL of 4x enzyme solution (20 nM DGKα or DGKζ in assay buffer (prepared as described below)) with 2.5 mL of 4x detergent / lipid micelle solution (composition described below) and incubated at room temperature for 10 minutes. 1 μL of 2x enzyme / lipid substrate solution was then added to the wells containing the test compound and 1 μL of 300 μM ATP was added to initiate the reaction. The reaction was allowed to proceed for 2 hours, after which 2 μL of Glo reagent (Promega V9101) was added and incubated for 40 minutes. 4 μL of kinase detection reagent was then added and incubated for 30 minutes. Luminescence was recorded using an Envision microplate reader. Percent inhibition was calculated from ATP conversion generated by enzyme-free control reactions set at 100% inhibition and vehicle-only reactions set at 0% inhibition. Compounds were evaluated at 11 concentrations and IC 50 It was decided.

[0274] Preparation of 2x liposomes The lipid composition of the liposome solution was 5 mol% DAG (Avanti 800811O), 40 mol% PS (Avanti 840035P), and 55 mol% PC (Avanti 850457) with an overall lipid concentration of 7-8 mg / mL. PC, DAG, and PS were dissolved in chloroform, combined, and dried under vacuum to a thin film. The lipids were hydrated to 20 mM in 50 mM MOPS, pH 7.5, 100 mM NaCl, 5 mM MgCl2, and freeze-thawed five times. The lipid suspension was extruded 10-12 times through a 100 nm polycarbonate filter. Dynamic light scattering was performed to confirm the size of the liposomes (radius 50-60 nm). The liposome preparations were stored for an extended period of 4 weeks at 4°C.

[0275] Baculovirus expression of near-full-length human DGKα and full-length human DGKζ Baculovirus samples of human MA-hDGKα-(S9-S727)-Ct-TVMV-His-pFB gate and full-length human DGK-ζ-transcript variant-2-TVMV-His-pFB gate were prepared using the Bac-to-Bac baculovirus expression system (Invitrogen) according to the manufacturer's protocol (the "MA-" in the name of the DGKα reagent indicates that two additional amino acids were added before Ser-9). The DNA sequences used for the expression of DGKα(9-727) and DGKζ are SEQ ID NOs: 5 and 3, respectively. Baculovirus amplification was achieved using Sf9 cells infected with a virus / cell ratio of 1:1500 and grown at 27°C for 65 hours after transfection.

[0276] Scale-up of expression for the near-full-length DGK-α(9-727) protein was performed in flasks (2 L) and for full-length DGKζ in a Cellbag 50L WAVE bioreactor system (GE Healthcare Bioscience). Proteins were expressed using similar conditions but at different volumes. For expression of DGKα(9-727), 2x2L flasks were used, each containing a final volume of 0.8L of medium, while DGKζ was cultivated at a 12L scale in a 50L Cellbag. For each, the initial density was 2x10 6 Sf9 cells (Expression System, Davis, CA) at 1000 cells / mL were seeded in ESF921 insect cell medium (Expression System), infected with virus stocks at a ratio of 1:200 virus / cell, and grown at 27°C for 66-68 hours post-infection. Infected cell cultures were harvested by centrifugation at 2000 rpm for 20 minutes at 4°C in a SORVALL® RC12BP centrifuge. Cell pellets were stored at -80°C until purification.

[0277] Purification of human DGK-alpha and DGK-zeta Human DGKα(9-727) and full-length DGKζ were each expressed containing a TVMV-cleavable, C-terminal Hex-His tag sequence (SEQ ID NOs: 2 and 4, respectively) and were produced and purified from Sf9 baculovirus-infected insect cell paste as described above. Cell paste was thawed and suspended in buffer (50 mM HEPES, pH 7.2, 300 mM NaCl, 10% v / v glycerol, 1 mM TCEP, containing benzonase and protease inhibitors) at 1:10 v / v of the original culture volume. Lysis was achieved with a nitrogen disrupter (Parr Instruments) using nitrogen cavitation method, and the lysate was clarified by high-speed centrifugation. The clarified lysate was purified to approximately 90% homogeneity on an AKTA Purifier Plus system using two or three successive column chromatography steps, respectively. Both isoforms were purified by nickel affinity purification with imidazole gradient elution (i.e., HisTrap FF, GE Healthcare) followed by size exclusion chromatography (i.e., for DGKα(9-727), HiLoad 26 / 600 Superdex 200 preparative grade, GE Healthcare; for DGKζ, HiPrep 26 / 600 Sephacryl S 300 HR, GE Healthcare). These two steps yielded DGKα(9-727) in >90% purity. To achieve comparable purity with full-length DGKζ, a third step was required utilizing cation exchange chromatography (SP Sepharose FF, GE Healthcare) and eluting with a NaCl gradient. The final formulation buffers were similar for both proteins: for DGKα(9-727) they were prepared in 50 mM Hepes, pH 7.3, 300 mM NaCl, 10% v / v glycerol, and 1 mM TCEP, and for full-length DGKζ they were prepared in 50 mM Hepes, pH 7.3, 500 mM NaCl, 5% v / v glycerol, and 1 mM TCEP. Proteins were concentrated to 1-2 mg / mL, flash frozen, and stored at -80°C for long-term storage.

[0278] Assay 3: Raji CD4 T cell IL2 assay 1536-well IL-2 assays were performed with pre-activated CD4 T cells and Raji cells in a volume of 4 μL. Prior to the assay, CD4 T cells were pre-activated by treatment with α-CD3, α-CD28, and PHA (1.5 μg / mL, 1 μg / mL, and 10 μg / mL, respectively). Raji cells were treated with Staphylococcal Enterotoxin B (SEB, 10,000 ng / mL). Serially diluted compounds were first transferred to a 1536-well assay plate (Corning, #3727), followed by the addition of pre-activated CD4 T cells (2 μL, final density: 6000 cells / well) and SEB-treated Raji cells (2 μL, 2000 cells / well). After 24 h incubation in a 37°C / 5% CO2 incubator, IL-2 detection reagent (4 μL) was added to the assay plate (Cisbio, #64IL2PEC). The assay plate was read on an Envision reader. To assess compound cytotoxicity, either Raji or CD4 T cells were incubated with serially diluted compounds. After 24 h incubation, CellTiter-Glo (4 μL, Promega, #G7572) was added and the plate was read on an Envision reader. The 50% effective concentration (IC 50 ) into a four-variable logistic equation: y=A+((BA) / (1+((C / x)^D))) (where A and B represent the minimum and maximum % activation or inhibition, respectively, and C is the IC 50 , D is the slope of the curve, and x is the compound concentration). The calculation was carried out using

[0279] Assay 4: CellTiter-Glo CD8 T cell proliferation assay Frozen naive human CD8 T cells were thawed in RPMI + 10% FBS, incubated at 37°C for 2 hours, and counted. 384-well tissue culture plates were coated overnight at 4°C with anti-human CD3 (20 μL, 0.1 μg / mL in plain RPMI), which was removed from the plate, after which CD8 T cells (20k / 40 μL) were added to each well along with soluble anti-human CD28 (0.5 μg / mL). Compounds were echoed to the cell plates immediately after the cells were plated. After 72 hours of incubation in a 37°C incubator, CellTiter-Glo reagent (10 μL, Promega catalog number G7570) was added to each well. Plates were shaken vigorously for 5 minutes, incubated at room temperature for an additional 15 minutes, and read on the Envision for CD8 T cell proliferation. The analysis included background CD8 T cell signals stimulated with anti-CD3 (0.1 μg / mL) and anti-CD28 (0.5 μg / mL). The control compound, 8-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-5-methyl-7-nitro-6-oxo-5,6-dihydro-1,5-naphthyridine-2-carbonitrile, had an EC50 concentration of 3 μM to set the 100% range. 50 was used to normalize the data at absolute 50%.

[0280] Assay 5: DGK AP1 reporter assay The Jurkat AP1 luciferase reporter was generated using the Cignal Lenti AP1 Reporter (luc) kit from SABiosciences (CLS-011L). Compounds were transferred from the Echo LDV plate to each well of a 384-well plate (white solid bottom, opaque PE CulturPlate 6007768) using the Echo 550 instrument. Sample size was 30nL / well, one destination plate per source plate. Cell suspension was prepared by transferring 40mL of cells (2x20mL) to a clean conical tube (50mL). Cells were concentrated by centrifugation (1200rpm, 5min, ambient temperature). The supernatant was removed and all cells were suspended in RPMI (Gibco 11875) + 10% FBS and diluted to 1.35x10 6 The concentration was 4.0x104 cells / mL. Cells were manually added to compound-containing 384-well TC plates at 30μL / well of cell suspension using a multichannel pipette. The cell plates were incubated at 37℃ and 5% CO2 for 20 minutes.

[0281] During incubation, αCD3 (3 μL, 1.3 mg / mL) was mixed with medium (10 mL) to prepare an anti-CD3 antibody (αCD3) solution [final concentration = 0.4 μg / mL]. Then, αCD3 (1.5 μL, 1.3 mg / mL) was mixed with medium (0.5 mL) [final concentration = 4 μg / mL]. After 20 min, for reference, medium (10 μL) was added to all wells in the first column (wells A-M) and 10 μL of αCD3 (4 μg / mL) per well was added to rows N-P of the first column. Then, using a multichannel pipette, αCD3 (10 μL, 0.4 μg / mL) was added per well. αCD3 stimulated + / - compound-treated cells were incubated for 6 h at 37 °C, 5% CO2.

[0282] During this incubation, Steady-Glo (Promega E2520) reagent was slowly thawed to ambient temperature. Then, 20 μL of Steady-Glo reagent / well was added using a Multidrop Combi dispenser. Air bubbles were removed by centrifugation (2000 rpm, ambient temperature, 10 s). Cells were incubated at room temperature for 5 min. Samples were characterized by measuring the relative light units (RLU) using an Envision plate reader instrument following the luminescence protocol. Data was analyzed using the reference compound (8-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-5-methyl-7-nitro-6-oxo-5,6-dihydro-1,5-naphthyridine-2-carbonitrile) and normalized to 100% inhibition.

[0283] Assay 6: Murine cytotoxic T lymphocyte assay An antigen-specific cytotoxic T cell (CTL) assay was developed to functionally evaluate the ability of DGKα and DGKζ inhibitors to enhance effector T cell-mediated tumor cell killing. CD8+ T cells isolated from OT-1 transgenic mice recognize MC38, an antigen-presenting cell that displays the ovalbumin-derived peptide SIINFEKL. Recognition of the cognate antigen initiates the cytolytic activity of OT-1 antigen-specific CD8+ T cells.

[0284] Functional CTL cells were prepared as follows. OT-1 splenocytes were isolated from 8-12 week old mice and expanded in the presence of SIINFEKL peptide (1 μg / mL) and mIL2 (10 U / mL). After 3 days, fresh medium containing mIL2 (U / mL) was added. On the 5th day of expansion, CD8+ T cells were isolated and prepared for use. Activated CTL cells can be cryopreserved for 6 months. Separately, 1 million MC38 tumor cells were pulsed with SIINFEKL-OVA peptide (1 μg / mL) for 3 hours at 37°C. The cells were washed (3x) with fresh medium to remove excess peptide. Finally, in a 96-well plate (U-bottom), CTL cells previously treated with DGK inhibitor for 1 hour were combined with antigen-loaded MC38 tumor cells at a ratio of 1:10. The cells were then spun at 700 rpm for 5 minutes and left overnight at 37°C in an incubator. After 24 hours, supernatants were collected and IFN-γ cytokine levels were analyzed by AlphaLisa (purchased from Perkin Elmer).

[0285] Assay 7: PHA proliferation assay Phytohemagglutinin (PHA)-stimulated blast cells derived from frozen stocks were incubated for 1 h in RPMI medium (Gibco, ThermoFisher Scientific, Waltham, MA) supplemented with 10% fetal bovine serum (Sigma Aldrich, St. Louis, MO) and then added to each well of a 384-well plate (10,000 cells / well). Compounds were transferred to each well of the 384-well plate and treated cells were maintained in medium containing human IL2 (20 ng / mL) at 37°C and 5% CO2 for 72 h, after which proliferation was measured using the MTS reagent [3-(4,5-dimethyl-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] according to the manufacturer's instructions (Promega, Madison, WI). Percentage of inhibition was calculated relative to the values ​​between IL2 stimulation (0% inhibition) and non-stimulated control (100% inhibition). Inhibitory concentration (IC 50 ) was calculated based on 50% inhibition of fold induction between IL2 stimulated and unstimulated treatments.

[0286] Assay 8: Human CD8 T cell IFN-γ assay Frozen naive human CD8 T cells were thawed in AIM-V medium, incubated at 37°C for 2 hours, and counted. 384-well tissue culture plates were coated with 20 μL of anti-human CD3 at 0.05 μg / mL in PBS overnight at 4°C, which was removed from the plate, after which 40,000 cells per 40 microliters of CD8 T cells with 0.1 μg / mL of soluble anti-human CD28 were added to each well. Compounds were transferred to the cell plate using an Echo liquid handler immediately after plating the cells. After 20 hours of incubation in a 37°C incubator, 3 microliters / well of supernatant was transferred to a new 384-well white assay plate and cytokines were measured.

[0287] Interferon-γ (IFN-γ) was quantified using an AlphaLISA kit (catalog no. AL217) as described by the manufacturer (Perkin Elmer). Counts from each well were converted to IFN-γ concentration (pg / mL). EC 50 Values ​​were determined by setting anti-CD3 (0.05 μg / mL) + anti-CD28 (0.1 μg / mL) as baseline and co-stimulation with 3 μM of the reference compound 8-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-5-methyl-7-nitro-6-oxo-5,6-dihydro-1,5-naphthyridine-2-carbonitrile and anti-CD3 + anti-CD28 as 100% activity.

[0288] Assay 9: Human CD8 T cell pERK assay Frozen naive human CD8 T cells were thawed in AIM-V medium, incubated at 37°C for 2 hours, and counted. CD8 positive T cells were added to 384-well tissue culture plates at 20,000 cells / well in AIM-V medium. One compound was added to each well, followed by bead-immobilized anti-human CD3 and anti-CD28 mAbs at a final concentration of 0.3 μg / mL. Cells were incubated for 10 minutes at 37°C. The reaction was stopped by adding lysis buffer from the AlphaLISA Surefire kit (Perkin Elmer, Cat. No. ALSU-PERK-A). Lysates (5 μL / well) were transferred to a new 384-well white assay plate to measure pERK activation. EC of the compound 50 set anti-CD3 + anti-CD28 as baseline and determined costimulation with 3 μM 8-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-5-methyl-7-nitro-6-oxo-5,6-dihydro-1,5-naphthyridine-2-carbonitrile and anti-CD3 + anti-CD28 as 100% activation.

[0289] Assay 10: Human whole blood IFN-γ assay Human venous whole blood (22.5 μL / well) obtained from healthy donors was pretreated with compounds for 1 h at 37° C. in a humidified incubator with 95% air / 5% CO2. Blood was stimulated with 2.5 μL of anti-human CD3 and anti-CD28 mAbs at a final concentration of 1 μg / mL each for 24 h each at 37° C. IFN-γ in the supernatant was measured using an AlphaLISA kit (catalog no. AL217). EC of the compound 50 was determined by setting anti-CD3 + anti-CD28 as baseline and costimulation with 3 μM of the reference compound 8-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-5-methyl-7-nitro-6-oxo-5,6-dihydro-1,5-naphthyridine-2-carbonitrile and anti-CD3 + anti-CD28 as 100% activation.

[0290] Assay 11: DGK human whole blood pERK assay Human whole blood ERK phosphorylation assays were performed using human venous whole blood (drawn with heparin as anticoagulant) obtained from healthy donors. Serially diluted compounds (11 points, 3-fold dilutions) in DMSO were added at 20 nL / well to a 384-well plate using an ECHO 550 acoustic dispenser (Labcyte) to achieve a final starting concentration of 20 μM in the assay. Heparinized human whole blood was added to the compound plate at 9 μL per well and incubated for 1 h at 37°C in a 95% humidified, 5% CO2 incubator. After 1 h of compound incubation, 1 μL of human anti-CD3 antibody (made in-house) was added to the well at a final concentration of 1 μg / mL in the presence of the bridging antibody goat anti-mouse IgG (4 μg / mL) to stimulate the pathway and further incubated for 15 min at 37°C. Stimulation was stopped by adding 90 μL of Fix / Lyse buffer (BD 558049). Cells were washed and stained with anti-CD8 PE (BD 555635) antibody for 60 min at room temperature, washed again and permeabilized with Perm III buffer (BD 558050) for 30 min on ice. Cells were then stained with Alexa Fluor™ 647 anti-ERK1 / 2 phospho (Thr202 / Tyr204) antibody (Bioleged 675504) at a dilution of 1:50 for 60 min. Samples were washed and resuspended in dPBS containing 1% BSA (dPBS, Gibco 14190136; BSA, Sigma-Aldrich A9205). Samples were analyzed using Intellicyt™ iQue Screener PLUS. pERK activation was quantified by the percentage of pERK positive population in the CD8 positive population. The efficacy of the compound was calculated based on the value obtained using the internal compound at a concentration of 20 μM, which was taken as a 100% activation baseline, and the value obtained using the anti-CD3 control, which was taken as a 0% activation baseline.

[0291] Table A In vitro DGK inhibition IC 50 Activity value [Table 4]

[0292] Table A shows the in vitro DGK inhibition IC measured in the DGKα and DGKζ liposomal (LIPGLO) assay. 50 Activity values ​​are listed. The compounds of the present invention have activity as inhibitors of either or both of the DGKα and DGKζ enzymes and may therefore be used in the treatment of diseases associated with the inhibition of DGKα and DGKζ activity.

[0293] Nucleotide sequence encoding hDGKα-(M1-S735)-Ct-TVMV-His: [Table 5] [Table 6]

[0294] Amino acid sequence of hDGKα-(M1-S735)-Ct-TVMV-His: [Table 7] (SEQ ID NO:2)

[0295] Nucleotide sequence encoding hDGKζ-(M1-A928)-transcript variant-2Ct-TVMV-His: [Table 8] [Table 9]

[0296] Amino acid sequence of hDGKζ-(M1-A928)-transcript variant-2Ct-TVMV-His: [Table 10]

[0297] Nucleotide sequence encoding MA-hDGKα-(S9-S727)-Ct-TVMV-His:

Table 11

Table 12

Table 13

Claims

1. Formula (I): 【Chemistry 1】 [In the formula: R １ is F, Cl, Br, -CN, C １－３ Alkyl (0 to 4 R １ａ substituted with), C ３－４ Cycloalkyl (0 to 4 R １ａ substituted with), C １－３ Alkoxy (0 to 4 R １ａ substituted with —C(O)NR ａ R ａ , -NR ａ R ａ , -S(O) ｎ R ｅ or -P(O)R ｅ R ｅ and Each R １ａ are independently F, Cl, —CN, —OH, —OCH ３ or -NR ａ R ａ and Each R ａ are independently H or C １－３ is alkyl; Each R ｅ is independently ３－４ Cycloalkyl or C １－３ Alkyl (0 to 4 R １ａ substituted with; R ２ is H, C １－３ Alkyl (0 to 4 R ２ａ substituted with), C ２－３ Alkenyl (0 to 4 R ２ａ substituted with), or C ３－４ Cycloalkyl (0 to 4 R ２ａ substituted with; Each R ２ａ are independently F, Cl, -CN, -OH, -O(C １－２ Alkyl), C ３－４ Cycloalkyl, C ３－４ Alkenyl, or C ３－４ alkynyl; R ３ is H, F, Cl, Br, -CN, C １－３ Alkyl, C １－２ Fluoroalkyl, C ３－４ Cycloalkyl, C ３－４ Fluorocycloalkyl, -NO ２ or pyridinyl (0 to 2 R ３ａ substituted with; Each R ３ａ is halo, -CN, C １－３ Alkyl, or C １－３ is alkoxy; R ４ is -CH ２ R ４ａ , -CH ２ CH ２ R ４ａ , -CH ２ CHR ４ａ R ４ｄ , -CHR ４ａ R ４ｂ , or -CR ４ａ R ４ｂ R ４ｃ and R ４ａ and R ４ｂ is, independently, (i) C １－６ Alkyl (F, Cl, -CN, -OH, -OCH ３ , -SCH ３ , C １－３ Fluoroalkoxy, -NR ａ R ａ , -S(O) ２ R ｅ or -NR ａ S (O) ２ R ｅ substituted with 0 to 4 substituents independently selected from the group consisting of: (ii) C ３－６ cycloalkyl, heterocyclyl, phenyl, or heteroaryl, each of which is F, Cl, Br, —CN, —OH, C １－６ Alkyl, C １－３ Fluoroalkyl, C １－４ Hydroxyalkyl, -(CH ２ ) １－２ O (C １－３ Alkyl), C １－４ Alkoxy, -O(C １－４ Hydroxyalkyl), —O(CH １－３ O (C １－３ Alkyl), C １－３ Fluoroalkoxy, —O(CH) １－３ N.R. ｃ R ｃ , -OCH ２ CH=CH ２ , -OCH ２ C≡CH, -C(O)(C １－４ alkyl), -C(O)OH, -C(O)O(C １－４ alkyl), -NR ｃ R ｃ , -NR ａ S (O) ２ (C １－３ alkyl), -NR ａ C(O)(C １－３ alkyl), -NR ａ C(O)O(C １－４ alkyl), -P(O)(C １－３ Alkyl) ２ , -S(O) ２ (C １－３ alkyl), -O(CH ２ ) １－２ (C ３－６ cycloalkyl), -O(CH ２ ) １－２ (morpholinyl), cyclopropyl, cyanocyclopropyl, methylazetidinyl, acetylazetidinyl, (tert-butoxycarbonyl)azetidinyl, triazolyl, tetrahydropyranyl, morpholinyl, thiophenyl, methylpiperidinyl, and R ｄ substituted with 0 to 4 substituents independently selected from the group consisting of (iii) naphthalenyl substituted with 0 to 2 substituents independently selected from F, -CH 3 , -CH 2 CH 3 , and -OCH 3 , or (iv) C １－４ alkyl, C ３－６ cycloalkyl, heterocyclyl, aryl, and heteroaryl, wherein the ring group is F, Cl, Br, -OH, -CN, C １－６ Alkyl, C １－３ Fluoroalkyl, C １－３ Alkoxy, C １－３ Fluoroalkoxy, -OCH ２ CH=CH ２ , -OCH ２ C≡CH, -NR ｃ R ｃ , -NR ａ S (O) ２ (C １－３ alkyl), -NR ａ C(O)(C １－３ alkyl), -NR ａ C(O)O(C １－４ alkyl), and C ３－６ substituted with 0 to 3 substituents independently selected from cycloalkyl; Or R ４ａ and R ４ｂ together with their attached carbon atoms, C ３－６ cycloalkyl, or 3- to 6-membered heterocyclyl, each of which is 0 to 3 R ｆ is replaced by; Each R ｆ are independently F, Cl, Br, -OH, -CN, C １－６ Alkyl, C １－３ Fluoroalkyl, C １－３ Alkoxy, C １－３ Fluoroalkoxy, -OCH ２ CH=CH ２ , -OCH ２ C≡CH, -NR ｃ R ｃ , or C ３－６ A ring group selected from cycloalkyl, 3- to 6-membered heterocyclyl, phenyl, monocyclic heteroaryl, and bicyclic heteroaryl, each of which is selected from F, Cl, Br, —OH, —CN, C １－６ Alkyl, C １－３ Fluoroalkyl, C １－３ Alkoxy, C １－３ Fluoroalkoxy, and -NR ｃ R ｃ substituted with 0 to 3 substituents, more independently selected; R ４ｃ is C １－６ Alkyl or C ３－６ cycloalkyl, each of which is F, Cl, —OH, C １－２ Alkoxy, C １－２ substituted with 0 to 4 substituents independently selected from: fluoroalkoxy, and —CN; R ４ｄ Ha-OCH ３ and Each R ｃ are independently H or C １－２ is alkyl; R ｄ is phenyl, and F, Cl, —CN, —CH ３ , and -OCH ３ substituted with 0 to 1 substituents selected from: Each R ５ are independently F, Cl, -CN, -OH, C １－６ Alkyl (0 to 4 R ｇ substituted with), C １－３ Alkoxy (0 to 4 R ｇ substituted with), C ２－４ Alkenyl (0 to 4 R ｇ substituted with), C ２－４ Alkynyl (0 to 4 R ｇ substituted with), C ３－４ Cycloalkyl (0 to 4 R ｇ phenyl (substituted with 0 to 4 R ｇ oxadiazolyl (substituted with 0 to 3 R ｇ substituted with 0 to 4 R ｇ substituted with -(CH ２ ) １－２ (0 to 4 R ｇ heterocyclyl substituted with -(CH ２ ) １－２ N.R. ｃ C(O)(C １－４ alkyl), -(CH ２ ) １－２ N.R. ｃ C(O)O(C １－４ alkyl), -(CH ２ ) １－２ N.R. ｃ S (O) ２ (C １－４ alkyl), -C(O)(C １－４ alkyl), -C(O)OH, -C(O)O(C １－４ alkyl), -C(O)O(C ３－４ cycloalkyl), -C(O)NR ａ R ａ or -C(O)NR ａ (C ３－４ cycloalkyl) or two R ５ are attached to the same carbon atom to form =O; Each R ｇ are independently F, Cl, -CN, -OH, C １－３ Alkoxy, C １－３ Fluoroalkoxy, —O(CH ２ ) １－２ O (C １－２ Alkyl), C ３－５ Cycloalkyl, or -NR ｃ R ｃ and Each R ６ is H, F, Cl, -CN, -CH ３ , -CH ２ F, -CHF ２ , -CF ３ , or -OCH ３ and m is 0, 1, 2, or 3; and n is 0, 1, or 2. or a salt thereof.

2. R １ F, Cl, Br, -CN, C １－３ Alkyl (0 to 4 R １ａ cyclopropyl (substituted with 0 to 3 R １ａ substituted with), C １－３ Alkoxy (0 to 3 R １ａ substituted with —C(O)NR ａ R ａ , -NR ａ R ａ , -S(O) ｎ CH ３ or -P(O)(CH ３ ) ２ and Each R １ａ is independently F, Cl, or -CN; Each R ａ are independently H or C １－３ is alkyl; R ２ H, C １－２ Alkyl (0 to 2 R ２ａ substituted with), or C ２－３ Alkenyl (0 to 2 R ２ａ substituted with; Each R ２ａ are independently F, Cl, -CN, -OH, -O(C １－２ Alkyl), cyclopropyl, C ３－４ Alkenyl, or C ３－４ alkynyl; R ３ H, F, Cl, Br, -CN, C １－２ Alkyl, C １－２ Fluoroalkyl, C ３－４ Cycloalkyl, -NO ２ or pyridinyl (0 to 2 R ３ａ substituted with; R ４ａ and R ４ｂ But independently, (i) C １－４ Alkyl (F, Cl, -CN, -OH, -OCH ３ , -SCH ３ , C １－３ Fluoroalkoxy, and -NR ａ R ａ substituted with 0 to 4 substituents independently selected from the group consisting of: (ii) C ３－６ cycloalkyl, heterocyclyl, phenyl, or heteroaryl, each of which is F, Cl, Br, —CN, —OH, C １－６ Alkyl, C １－３ Fluoroalkyl, -CH ２ OH, -(CH ２ ) １－２ O (C １－２ Alkyl), C １－４ Alkoxy, —O(C １－４ Hydroxyalkyl), —O(CH １－２ O (C １－２ Alkyl), C １－３ Fluoroalkoxy, —O(CH) １－２ N.R. ｃ R ｃ , -OCH ２ CH=CH ２ , -OCH ２ C≡CH, -C(O)(C １－４ alkyl), -C(O)OH, -C(O)O(C １－４ alkyl), -NR ｃ R ｃ , -NR ａ S (O) ２ (C １－３ alkyl), -NR ａ C(O)(C １－３ alkyl), -NR ａ C(O)O(C １－４ alkyl), -P(O)(C １－２ Alkyl) ２ , -S(O) ２ (C １－３ alkyl), -O(CH ２ ) １－２ (C ３－４ cycloalkyl), -O(CH ２ ) １－２ (morpholinyl), cyclopropyl, cyanocyclopropyl, methylazetidinyl, acetylazetidinyl, (tert-butoxycarbonyl)azetidinyl, triazolyl, tetrahydropyranyl, morpholinyl, thiophenyl, methylpiperidinyl, and R ｄ or substituted with 0 to 4 substituents selected independently from (iii) C １－３ alkyl, C ３－６ Cycloalkyl, heterocyclyl, phenyl, and heteroaryl are substituted with one ring group selected from F, Cl, Br, —OH, —CN, C １－３ Alkyl, C １－２ Fluoroalkyl, C １－３ Alkoxy, C １－２ Fluoroalkoxy, -OCH ２ CH=CH ２ , -OCH ２ C≡CH, -NR ｃ R ｃ , -NR ａ S (O) ２ (C １－３ alkyl), -NR ａ C(O)(C １－３ alkyl), -NR ａ C(O)O(C １－４ alkyl), and C ３－４ substituted with 0 to 3 substituents independently selected from cycloalkyl; Or R ４ａ and R ４ｂ together with the carbon atom to which they are attached, C ３－６ cycloalkyl or 3- to 6-membered heterocyclyl, each of which is 0 to 3 R ｆ is replaced by; Each R ｆ are independently F, Cl, Br, -OH, -CN, C １－４ Alkyl, C １－２ Fluoroalkyl, C １－３ Alkoxy, C １－２ Fluoroalkoxy, -OCH ２ CH=CH ２ , -OCH ２ C≡CH, -NR ｃ R ｃ , or C ３－６ A ring group selected from cycloalkyl, 3- to 6-membered heterocyclyl, phenyl, monocyclic heteroaryl, and bicyclic heteroaryl, each of which is F, Cl, Br, —OH, —CN, C １－４ Alkyl, C １－２ Fluoroalkyl, C １－３ Alkoxy, C １－２ Fluoroalkoxy, and -NR ｃ R ｃ substituted with 0 to 3 substituents, more independently selected; R ４ｃ But, C １－４ Alkyl or C ３－６ cycloalkyl, each of which is F, Cl, —OH, C １－２ Alkoxy, C １－２ substituted with 0 to 4 substituents independently selected from: fluoroalkoxy, and —CN; Each R ５ are independently F, -CN, -OH, C １－５ Alkyl (0 to 4 R ｇ substituted with), C １－２ Alkoxy (0 to 3 R ｇ substituted with), C ２－３ Alkenyl (0 to 4 R ｇ substituted with), C ２－３ Alkynyl (0 to 4 R ｇ substituted with), C ３－４ Cycloalkyl (0 to 4 R ｇ phenyl (substituted with 0 to 3 R ｇ oxadiazolyl (substituted with 0 to 3 R ｇ substituted with 0 to 3 R ｇ substituted with -(CH ２ ) １－２ (0 to 4 R ｇ heterocyclyl substituted with -(CH ２ ) １－２ N.R. ｃ C(O)(C １－４ alkyl), -(CH ２ ) １－２ N.R. ｃ C(O)O(C １－４ alkyl), -(CH ２ ) １－２ N.R. ｃ S (O) ２ (C １－４ alkyl), -C(O)(C １－４ alkyl), -C(O)OH, -C(O)O(C １－４ alkyl), -C(O)O(C ３－４ cycloalkyl), -C(O)NR ａ R ａ or -C(O)NR ａ (C ３－４ cycloalkyl) or two R ５ are attached to the same carbon atom to form =O; Each R ６ is H, F, or -CH ３ Wherein; m is 0, 1, 2, or 3; 2. The compound according to claim 1 or a salt thereof.

3. R １ is F, -CN, or -OCH ３ and R ２ is H, -CH ３ , -CH ２ CN, -CH ２ CH ２ F, or -CH ２ CH=CH ２ and R ３ H, -CN, -NO ２ or pyridinyl; R ４ Ga-CH ２ R ４ａ or -CHR ４ａ R ４ｂ and R ４ａ is phenyl, naphthalenyl, or indolyl, each of which is F, —CH ３ , -CH ２ CH ３ , and -OCH ３ substituted with 0 to 2 more independently selected substituents; R ４ｂ is phenyl or fluorophenyl; Each R ５ Ga-CH ３ or two R ５ are attached to the same carbon atom to form =O; m is 0, 1, or 2; 2. The compound according to claim 1 or a salt thereof.

4. Structural formula: 【Chemistry 2】 [In the formula: R ５ａ and R ５ｂ is R ５ are more independently selected] The compound according to claim 1, which is represented by the following formula: or a salt thereof.

5. Structural formula: 【Chemistry 3】 The compound according to claim 1, which is represented by the following formula: or a salt thereof.

6. R ４ Ga-CH ２ R ４ａ or -CH ２ CH ２ R ４ａ 2. The compound according to claim 1, wherein:

7. R ４ Ga-CH ２ R ４ａ 2. The compound according to claim 1, wherein:

8. R ４ -CHR ４ａ R ４ｂ Or -CR ４ａ R ４ｂ R ４ｃ 2. The compound according to claim 1, wherein:

9. R ４ -CHR ４ａ R ４ｂ 2. The compound according to claim 1, wherein:

10. R ４ Ga-CH ２ R ４ａ or -CHR ４ａ R ４ｂ 2. The compound according to claim 1, wherein: Claim 11: 6-fluoro-4-{4-[(3-fluorophenyl)methyl]piperazin-1-yl}-1-methyl-3-nitro-1,2-dihydroquinolin-2-one (Example 1); 6-Fluoro-4-{4-[(3-fluoro-5-methylphenyl)methyl]piperazin-1-yl}-1-methyl-3-(pyridin-4-yl)-1,2-dihydroquinolin-2-one (Example 2); 4-(4-((1H-indol-4-yl)methyl)piperazin-1-yl)-6-fluoro-1-methyl-3-nitroquinolin-2(1H)-one (Example 3); 4-{4-[(1-ethyl-1H-indol-4-yl)methyl]piperazin-1-yl}-6-fluoro-1-methyl-3-nitro-1,2-dihydroquinolin-2-one (Example 4); 4-(4-benzyl-3-oxopiperazin-1-yl)-6-fluoro-1-methyl-3-nitro-1,2-dihydroquinolin-2-one (Example 5); 4-[(2S,5R)-4-[bis(4-fluorophenyl)methyl]-2,5-dimethylpiperazin-1-yl]-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbonitrile (Example 6); 4-[(2S,5R)-4-[bis(4-fluorophenyl)methyl]-2,5-dimethylpiperazin-1-yl]-1-methyl-2-oxo-1,2-dihydroquinoline-6-carbonitrile (Example 7); or 6-Methoxy-1-methyl-4-{4-[(naphthalen-1-yl)methyl]piperazin-1-yl}-3-nitro-1,2-dihydroquinolin-2-one (Example 8) or a salt thereof.

12. 10. A pharmaceutical composition comprising a compound of claim 1 or a salt thereof, and a pharma- ceutically acceptable carrier.

13. A pharmaceutical composition comprising a compound or a salt thereof according to any one of claims 1 to 11 for the treatment of cancer or viral infection.

14. 14. The pharmaceutical composition of claim 13, wherein the cancer is selected from colon cancer, pancreatic cancer, breast cancer, prostate cancer, lung cancer, ovarian cancer, cervical cancer, renal cancer, head and neck cancer, lymphoma, leukemia and melanoma.

15. A pharmaceutical composition comprising the compound according to any one of claims 1 to 11 or a salt thereof for inhibiting the activity of at least one diacylglycerol kinase selected from diacylglycerol kinase alpha (DGKα) and diacylglycerol kinase zeta (DGKζ).

Images