# Patent

## Title
Pharmaceutically active disubstituted triazine derivatives

## Abstract

The present invention relates to disubstituted triazine derivatives and/or pharmaceutically acceptable salts thereof, the use of these derivatives as pharmaceutically active agents, especially for the prophylaxis and/or treatment of infectious diseases, including opportunistic diseases, immunological diseases, autoimmune diseases, cardiovascular diseases, cell proliferative diseases, inflammation, erectile dysfunction and stroke, and pharmaceutical compositions containing at least one of said disubstituted triazine derivatives and/or pharmaceutically acceptable salts thereof. Furthermore, the present invention relates to the use of said disubstituted triazine derivatives as inhibitors for a protein kinase.

## Background
The present invention relates to disubstituted triazine derivatives and/or pharmaceutically acceptable salts thereof, the use of these derivatives as pharmaceutically active agents, especially for the prophylaxis and/or treatment of cell proliferative diseases, inflammatory and immunological diseases, cardiovascular diseases and infectious diseases. Furthermore, the present invention is directed towards pharmaceutical composition containing at least one of the disubstituted triazine derivatives and/or pharmaceutically acceptable salts thereof.
Cyclin-dependent kinase (CDK) family members that trigger passage through the cell cycle are being considered as attractive therapeutic targets, especially for cancer. CDK family members that control other processes such as transcription and RNA processing have caught less attention so far, although experimental evidence for their involvement in different pathological processes is emerging. As a general regulator of transcription, CDK9 is a therapeutic target for treatment of diseases like inflammation, virus replication such as HIV, EBV, and HCV, cancer and cardiac hypertrophy.
CDK9 regulates transcription by phosphorylation of RNA polymerase II as well as additional regulatory factors, thereby enabling productive elongation of transcription. Certain subgroups of genes, especially genes encoding RNAs or proteins with fast turnover like immediate early genes of the inflammatory response, NF-kappaB activated genes (Brasier 2008, Cell Cycle 7:17, 2661-2666, Hargreaves et al. (2009) Cell 138, 129-145); and antiapoptotic genes such as MCL-1 and Bcl-2 family members appear to be especially sensitive to CDK9 inhibition.
In addition, it has been reported that hypertrophic growth of cardiomyocytes is related to CDK9 activation. Furthermore, viruses like the human immune deficiency virus recruit CDK9 actively to nascent RNA transcripts, facilitating their replicating. The dependency of the expression of antiapoptotic genes on CDK9 activity makes it an attractive therapeutic target for various forms of leukaemia such as chronic lymphocytic leukaemia (CLL), acute myelogenous leukaemia (AML) and acute lymphoblastic leukaemia, and solid tumours like prostate, lung, colon, breast and pancreas cancer. In addition, CDK9 inhibitors have been active in models of stroke (Osuga 2000, PNAS 97 (18): 10254-10259). For reviews, see Wang, 2009 (Trends in Pharmacological Sciences 29:6, 302-313), and Kohoutek, 2009 (Cell Division 2009, 4:19).
DESCRIPTION OF THE STATE OF THE ART
In the prior art there are two documents wherein disubstituted triazine derivatives are used for the inhibition of different enzymes.
In WO 01/25220A1 the scientific work relates to inhibitors of enzymes that catalyze phosphoryl transfer and/or that bind ATP/GTP nucleotides, compositions comprising the inhibitors, and methods of using the inhibitors and inhibitor compositions. The inhibitors and compositions comprising them are useful for treating or modulating disease in which phosphoryl transferases, including kinases, may be involved, symptoms of such disease, or the effect of other physiological events mediated by phosphoryl transferases, including kinases. The invention also provides for methods of making the inhibitor compounds and methods for treating diseases in which one or more phosphoryl transferase, including kinase, activities are involved.
However phosphoryl transferases are a large family of enzymes that transfer phosphorous containing groups from one substrate to another and no emphasis has been given to cyclin-dependent kinase (CDK) family members nor have been made any experiments according to the inhibition of cyclin-dependent kinases.
In WO2010/022055 A2 the scientific work provides compounds that are inhibitors of voltage-gated sodium channel (Nav), in particular Nav 1.7 and are therefore useful for the treatment of diseases treatable by inhibition of these channels, in particular, chronic pain disorders. Also provided are pharmaceutical compositions containing such compounds and processes for preparing such compounds.
However the use of these compounds for the inhibition of cyclin-dependent kinase (CDK) family members has not been mentioned and the citation does not disclose any experiments according to an inhibition of CDK family members.


DETAILED DESCRIPTION OF THE INVENTION
It is object of the present invention to provide compounds and/or pharmaceutically acceptable salts thereof which can be used as pharmaceutically active agents, especially for prophylaxis and/or treatment of cell proliferative diseases, inflammatory diseases, immunological diseases, cardiovascular diseases and infectious diseases, as well as compositions comprising at least one of those compounds and/or pharmaceutically acceptable salts thereof as pharmaceutically active ingredients.
This object is solved by the compounds and/or their pharmaceutically acceptable salts according to independent claim 1, the compounds of the present invention for use as pharmaceutically active agents, the use of the compounds of the present invention for the preparation of a pharmaceutical composition for the prophylaxis and/or treatment of infectious diseases, including opportunistic diseases, immunological diseases, autoimmune diseases, cardiovascular diseases, cell proliferative diseases, inflammation, erectile dysfunction and stroke according to independent claim 6, the use of compounds according to the present invention as inhibitors for the protein kinase CDK9.
Further advantageous features, aspects and details of the invention are evident from the dependent claims, the description, the examples and the drawings.
The novel disubstituted triazine compounds according to the present invention are defined by the general formula (I)




wherein

    R1 is






    L is a bond or —CR5R6—, —CR5R6—CR7R8—, —CR5R6—CR7R8—CR9R10—, —CR5R6—CR7R8—CR9R10—CR11R12—;
    R5-R12 represent independently of each other —H, —CH3, —C2H5, —C3H7, —F, —Cl, —Br, —I;
    R3 is selected from —H, —NO2, —CN, —Br, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5, —CR13R14R21, —CR13R14—CR15R16R21, —CR13R14—CR15R16—CR17R18R21, —CR13R14—CR15R16—CR17R18—CR19R20R21, —SO2R22, —CONR23R24, —NR25SO2NR23R24, —NR25SO2R22, —NR25CONR23R24, —SO2NR23R24, —SO(NR26)R22;
    R13-R21 and R29-R32 represent independently of each other —H, —CH3, —C2H5, —C3H7, —C4H9, —F, —Cl, —Br, —I;
    R22 and R28 are independently selected from R27′, —CR13R14R21, —CR13R14—CR15R16R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30R21, —CR13R14—CR15R16—CR17R18R21, —CR13R14—CR15R16—CR17R18 CR19R20CR21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—CR31R32R21, —CH2Ph; —CH2Ph the phenyl group of which may further be substituted by one, two, three, four or five substituents selected from the group consisting of —CH3, —C2H5, —C3H7, —F, —Br and —I;
    R23 and R24 are independently selected from —H, —CR13R14R21, —CR13R14—CR15R16R21, CR13R14—CR15R16—CR17R18—CR19R20—CR29R30R21, —CR13R14—CR15R16CR17R18R21—CR13R14—CR15R16—CR17R18—CR19R20R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—CR31R32R21, —CR13R14—CR15R16—O—R33, —CR13R14—CR15R16—CR17R18—O—R33, —CR13R14—CR15R16—NR33R34, —CR13R14—CR15R16—CR17R18—NR33R34, —CR13R14CR15R16—CR17R18—CR19R20—NR33R34, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—NR33R34, -Ph, —CH2PH, phenyl group which may further be substituted by one, two, three, four or five substituents selected from the group consisting of —CH3, —C2H5, —C3H7, —F, —Cl, —Br and —I; —CH2Ph the phenyl group of which may further be substituted by one, two, three, four or five substituents selected from the group consisting of —CH3, —C2H5, —C3H7, —F, —Br and —I; or both residues R23 and R24 together form with the nitrogen atom to which they are attached an azetidine, pyrrolidine, piperidine, piperazine, azepane, or morpholine ring;
    R25 is selected from —H, —CH3, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5 or —C(CH3)3;
    R26 is —H, —CH3, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —C(CH3)3, —CH(CH3)—C2H5, —CH(CH3)—C3H7, —CH2—CH(CH3)—C2H5, —CH(CH3)—CH(CH3)2, —C5H11, —C(CH3)2—C2H5, —CH2—C(CH3)3, —CH(C2H5)2, —C2H4—CH(CH3)2, —C6H13, —C3H6—CH(CH3)2, —C2H4—CH(CH3)—C2H5, —CH(CH3)—C4H9, —CH2—CH(CH3)—C3H7, —CH(CH3)—CH2—CH(CH3)2, —CH(CH3)—CH(CH3)—C2H5, —CH2—CH(CH3)—CH(CH3)2, —CH2—C(CH3)2—C2H5, —C(CH3)2—C3H7, —C(CH3)2—CH(CH3)2, —C2H4—C(CH3)3, —CH(CH3)—C(CH3)3, —CR13R14R21—COR28, —CR13R14—CR15R16R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30R21, —CR13R14—CR15R16—CR17R18R21, —CR13R14—CR15R16—CR17R18—CR19R20R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—CR31R32R21, COOR28, —R27;
    R27, R27′′ and R27 independently selected from






    these C3-C10-cycloalkyl groups may further be substituted by one, two, three, four, five or more substituents selected from the group consisting of —F, —Cl, —Br and —I;
    R33 and R34 represent independently of each other —H, —CH3, —C2H5, —C3H7, —C4H9, —CH2Ph, —COOC(CH3)3, —COOCH3, —COOCH2CH3, —COOCH2CH2CH3, —COOCH(CH3)2, —COOCH2Ph, —COCH3;
    R4 is selected from —H, —NO2, —CN, —F, —Cl, —Br, —I, —CR35R36R37, —CR35R36—CR38R39—CR40R41—CR42R43R37, —O—CR35R36CR38R39R37, —O—CR35R36—CR38R39—CR40R41R37, —CR35R36CR38R39—CR40R41R37, —O—CR35R36—CR38R39—CR40R41—CR42R43R37, —CR35R36—CR38R39R37, —O—CR35R36—CR38R39—CR40—R41—CR42R43CR44R45R37, —O—CR35R36R37, —O—CR35R36CR38R39CR40R41—CR42R43—CR44R45—CR46R47R37, —CR35R36—CR38R39—CR40R41—CR42R43—CR44R45R37, OCH2Ph, R27′′, OR27′′, —CR35R36—CR38R39—CR40R41—CR42R43—CR44R45—CR46R47R37,
    R35-R47 represent independently of each other —H, CR48R49R50, —CR48R49—CR51R52R50, —CR48R49—CR51R52—CR53R54R50, —CR48R49—CR51R52—CR53R54—CR55R56R50, —F, —Cl, —Br, —I;
    R48-R56 represent independently of each other —H, —F, —Cl, —Br, —I;
    R4 together with R22 or R23 or R24 or R25 may form a group —CH2CF12 or —CH2CH2CH2— if R4 is attached ortho to -L—R3;
    R2 is selected from












    B is a bond, —CH2—, —CH2CH2—, —CH2CH2CH2—, —CF2—, —CH2O—, —CH2OCH2—, —CH2CH2O—, —CH2NH—, —CH2N(CH3)—, —CH2NHCH2—, —CH2N(CH3)CH2—, —CH2CH2NH—, —CH2CH2N(CH3)—;
    R57 is —H, —CH3, —CF3, —CHF2, —CH2F, —COOCH3, —COOCH2CH3, —COOH, —COOCH2Ph, —COOCH2CH2CH3, —COOCH(CH3)2, —COOCH2CH2CH2CH3, —COOCH(CH3)(CH2CH3), —COOCH2CH(CH3)2, —COOC(CH3)3, —CONH2, -Ph, —CONH(CH3), —CON(CH3)2, —CN, —NH2, —NH(CH3), —N(CH3)2, —NH(CH2CH3), —N(CH3)(CH2CH3), —N(CH2CH3)2, pyrrolidin-1-yl, piperidin-1-yl, azetidin-1-yl, morpholin-4-yl, optionally substituted phenyl or optionally substituted pyridine;
    R58 is —H, —CH3, —F, —Cl, —CF3, —CH2OCH3;
    R59 is —H, —CH3, —F, —Cl, —CF3, -Ph, —CH2OCH3; and
    R60 is —H, —CH3, —CF3, —COCH3, —COOCH3, —COOCH2CH3, —COOC(CH3)3, —COOCH2Ph;
    Y is —O—, —S—, —NH—, or —N(CH3)—;
    R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph in which the phenyl groups may optionally be substituted;
    R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3;
    R64 and R65 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3;
    R66 and R67 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br;
    R68 is selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —OH, —OCH3, —NH2;
    R69 is selected from —H, —CH3, —CH2CH3;
    R70 and R71 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —NH2; and
    R72 is selected from —H, —CH3, —CF3, —CH2CH3;

and enantiomers, stereoisomeric forms, mixtures of enantiomers, diastereomers, mixtures of diastereomers, prodrugs, hydrates, solvates, acid salt forms, tautomers, and racemates of the above mentioned compounds and pharmaceutically acceptable salts thereof.



The expression prodrug is defined as a substance, which is applied in an inactive or significantly less active form. Once applied and incorporated, the prodrug is metabolized in the body in vivo into the active compound.
The expression tautomer is defined as an organic compound that is interconvertible by a chemical reaction called tautomerization. Tautomerization can be catalyzed preferably by bases or acids or other suitable compounds.
Preferred are compounds having the general formula (I):




    wherein
    R1 represents






in which

    L is a bond, —CH2—, —CH2CH2—, or —CF2—, particularly preferred —CH2—;
    R3 is selected from —H, —NO2, —CN, —Br, —I, —SO2R22, —CONR23R24, —NR25SO2NR23R24, —NR25SO2R22, —NR25CONR23R24, —SO2NR23R24, and —SO(NR26)R22;
    R22, R23, R24, R25, R26 are defined as described herein;
    R4 is —H, —CH3, —F, —Cl, or —CF3, particularly preferred —H;
    R2 is defined as described in formula (I) and
    B is a bond, —CH2—, —CH2CH2—, —CH2CH2CH2—, —CF2—, —CH2O—, —CH2OCH2—, —CH2CH2O—, —CH2NH—, —CH2N(CH3)—, —CH2NHCH2—, —CH2N(CH3)CH2—, —CH2CH2NH—, —CH2CH2N(CH3)—;
    R57 is —H, —CH3, —CF3, —CHF2, —CH2F, —COOCH3, —COOCH2CH3, —COOH, —COOCH2Ph, —COOCH2CH2CH3, —COOCH(CH3)2, —COOCH2CH2CH2CH3, —COOCH(CH3)(CH2CH3), —COOCH2CH(CH3)2, —COOC(CH3)3, —CONH2, -Ph, —CONH(CH3), —CON(CH3)2, —CN, —NH2, —NH(CH3), —N(CH3)2, —NH(CH2CH3), —N(CH3)(CH2CH3), —N(CH2CH3)2, pyrrolidin-1-yl, piperidin-1-yl, azetidin-1-yl, morpholin-4-yl, optionally substituted phenyl or optionally substituted pyridine;
    R58 and R59 are independently selected from —H, —F, or —CH3, and
    R60 is —H, —CH3, —CF3, —COCH3, —COOCH3, —COOCH2CH3, —COOC(CH3)3, —COOCH2Ph;
    Y is —O—, —S—, —NH—, or —N(CH3)—;
    R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph in which the phenyl groups may optionally be substituted;
    R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3,
    R64 and R65 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3;
    R66 and R67 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br;
    R68 is —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl;
    R69 is —H or —CH3;
    R70 and R71 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —NH2; and
    R72 is selected from —H, —CH3, —CF3, —CH2CH3.


Preferred are also compounds of the general formula (I), wherein

    R1 represents






    in which
    L is a bond, —CH2—, —CH2CH2—, or —CF2—;
    R3 is SO2NH2, SO2NH(CH3), SO2N(CH3)2, SO2NH(CH2CH2OCH3), NHSO2CH3, NHSO2CH2CH3, NHSO2CH2CH2CH3, NHSO2CF3, SO2CH3, NHSO2NH2, SO(NH)CH3;
    R4 is H, CH3, F, Cl, or CF3;
    R2 represents one of the residues listed under (i) to (xv):






    
    
        the group BR57 is H, CH2OCH3, COOCH3, COOCH2CH3, CH2NHCOOC(CH3)3, phenyl, or CH2CH2N(CH3)2; and
        R58-R60 are independently selected from —H, —F, —CH3, —CH2OH; or
    
    






    
    
        in which the group —B—R57 is —H, —CH3, —CF3, —CH2OCH3, —COOCH3, —COOCH2CH3, —CONH2, benzyl, —CH2-benzyl, —CH2—O—CH2-phenyl,
        CH2NHphenyl, CH2—O-phenyl, phenyl, or
    
    






and

    
    
        R58 and R59 are independently selected from —H, —F, —CH2OCH3, —CH2OH, or —CH3, more preferred —H; or
    
    






    
    
        in which R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3, more preferred both are —H; or
    
    






    
    
        in which R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3, more preferred both are —H; or
    
    






    
    
        in which R64 and R65 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3; and
        R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3; and
        R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, -phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; or
    
    






    
    
        in which R68 is —H, —CH3, —CF3, or —CH2CH3, more preferred —H; or
    
    






    
    
        in which R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —F, —Cl; more preferred both, R62 and R63 are —H; or
    
    






    
    
        in which R68 is —H, —CH3, —CF3, —CH2—CH3, —CN, —F, —Cl, more preferred —H; and
        R69 is —H or —CH3, more preferred —H; or
    
    






    
    
        in which R68 is —H, —CH3, —CF3, or —CH2—CH3, more preferred —H; or
    
    






    
    
        in which —B—R57 is —H, —CH2OCH3, —COOCH3, or phenyl; or
    
    






    
    
        R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
        R70 and R71 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —NH2; or
    
    






    
    
        R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
        R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3; or
    
    






    
    
        R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
        R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3; or
    
    






    
    
        R70 is independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —NH2; or
    
    






    
    
        R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
        R66 and R67 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br;

and enantiomers, stereoisomeric forms, mixtures of enantiomers, diastereomers, mixtures of diastereomers, prodrugs, hydrates, solvates, acid salt forms, tautomers, and racemates of the above mentioned compounds and pharmaceutically acceptable salts thereof.

    
    


As used herein the bonds ending in the middle of a ring system like the bond to the substituents R58 or R59 shall mean that this substituent can be attached to any carbon atom of the ring and that also both substituents can be attached to the same carbon atom.
In more preferred compounds of Formula (I)

    the substituent -L—R3 is —SO2NH2, —CH2SO2NH2, —CH2CH2SO2NH2, —CF2SO2NH2, —NHSO2NH2, —CH2NHSO2NH2, —SO2CH3, —SO(NH)CH3, —CH2SO(NH)CH3; and R4 is —H;
    R2 represents in all general formula disclosed herein preferably






It is preferred if R3 is selected from —H, —NO2, —CN, —SO2R22, —CONR23R24, —NR25SO2NR23R24, —NR25SO2R22, NR25SO2R22—NR25CONR23R24, —SO2NR23R24, and —SO(NR26)R22, and even more preferred if R3 is selected from —H, —SO2R22, —CONR23R24, —NR25SO2NR23R24, —NR25SO2R22, NR25SO2R22, —NR25CONR23R24, —SO2NR23R24, and —SO(NR26)R22, and still more preferred if R3 is selected from —SO2R22, —NR25SO2NR23R24, —NR25SO2R22, —SO2NR23R24, and —SO(NR26)R22, and most preferred if R3 is selected from —SO2R22, —SO2NR23R24, and —SO(NR26)R22.
It is in addition preferred if L represents —CH2—, —CH2CH2—, or —CF2—, and more preferred if L represents —CH2— or —CF2—, and particularly preferred if L represents —CH2—.
The residue -L—R3 represents preferably —CH2—SO2R22, —CH2—CONR23R24, —CH2—NR25SO2NR23R24, —CH2—NR25SO2R22, —CH2—NR25CONR23R24, —CH2—SO2NR23R24, or —CH2—SO(NR26)R22, and still more preferably —CH2—SO2R22, —CH2—NR25SO2NR23R24, —CH2—NR25SO2R22, —CH2—SO2NR23R24, or —CH2—SO(NR26)R22, and most preferably -L—R3 represents —CH2—SO2R22, —CH2—SO2NR23R24, or —CH2—SO(NR26)R22.
In regard to R2 is preferred that the residue —B—R57 is present and more preferably is present in ortho position to the ring nitrogen atom, respectively in position 2 of the nitrogen heterocyclic ring. Moreover it is very important for the biological activity that R2 represents a nitrogen herterocycle which is attached through the nitrogen atom to the rest of the molecule. Thus, it is very important that the residue R2 is linked through a nitrogen atom and that R2 represent a nitrogen heterocyclic ring system. In case R2 represents a bicyclic ring system it is preferred that the second condensed ring is also a nitrogen heterocyclic ring and even more preferred is an aromatic nitrogen heterocyclic ring as second ring of the bicyclic system.
In all general formula disclosed herein it is most preferred if the residue —R—57 represents —H, —CF3, —CH3, -Ph, —CH2OCH3, —COOCH3, —CONH2, -phenyl, —CH2—CH2—N(CH3)2, —CH2-Ph, —CH2—CH2-Ph, —CH2—O-Ph, —CH2—O—CH2-Ph, or —CH2—NH-Ph.
It has been found that compounds of the general formulas disclosed herein are biologically more potent in regard to state of the art compounds due to the nitrogen heterocyclic ring R2 which is linked through the nitrogen atom and due to the residue —B—R57 especially when attached in ortho position or position 2 of the nitrogen heterocyclic ring R2 which might also be a bicyclic ring system.
It is especially preferred if R3 is selected from —H, —NO2, —CN, —SO2—NH2, —SO2NH(CH3), —SO2N(CH3)2, —SO2NH(CH2CH2OCH3), —SO2CH3, or —SO(NH)CH3, and even more preferred if R3 is selected from —H, —SO2NH2, SO2NH(CH3), or —SO2N(CH3)2, and still more preferred if R3 is selected from —SO2NH2, or —SO2N(CH3)2, and most preferred if R3 represents —SO2NH2.
It is in addition to the especially preferred selection for R3 preferred if L represents —CH2—, —CH2CH2—, or —CF2, and more preferred if L represents —CH2— or —CF2—, and particularly preferred if L represents —CH2—.
The residue -L—R3 represents more preferably —H, —CH2—NO2, —CH2—CN, —CH2—SO2NH2, —CH2—SO2NH(CH3), —CH2—SO2N(CH3)2, —CH2—SO2NH(CH2CH2OCH3), —CH2—SO2CH3, or —CH2SO(NH)CH3, and still more preferably —H, —CH2—SO2NH2, —CH2—SO2NH(CH3), or —CH2—SO2N(CH3)2, and still more preferably —H, —CH2—SO2NH2, or —CH2—SO2N(CH3)2, and most preferably -L—R3 represents —CH2—SO2NH2. Moreover it is preferred if the residue -L—R3 is in meta position.
Even more preferred are compounds of the general formula (I), wherein

    R1 represents






    L is —CH2—;
    R3 is —SO2NH2;
    R2 represents:






    —B—R57 is —H, —CF3, —CH3, -Ph, —CH2OH, —CH2OCH3, —COOCH3, —CONH2, —CH2NH—COO—C(CH3)3, -phenyl, —CH2—CH2—N(CH3)2, —CH2-Ph, —CH2—CH2-Ph,
    —CH2—O-Ph,






—CH2—O—CH2-Ph, or —CH2—NH-Ph,

    R58-R60 and R62 represent independently of each other:
    —H, —CH3, —F, —NH2, —CH2OCH3, —COOCH3, —CH2OH, —CF3, —CONH2, —CH2-Ph, —CH2—CH2-Ph, —CH2—O-Ph, —CH2—NH-Ph, —CH2—O—CH2-Ph, -Ph,


In all general formula disclosed herein it is especially preferred if the residue —B—R57 represents —H, —CF3, —CH3, -Ph, —CH2OH, —CH2OCH3, —COOCH3, —CONH2, —CH2NH—COO—C(CH3)3, -phenyl, —CH2—CH2—N(CH3)2, —CH2-Ph, —CH2—CH2-Ph,
—CH2—O-Ph,




—CH2—O—CH2-Ph, or —CH2—NH-Ph.

It has been demonstrated that compounds with the nitrogen containing residue R2 in combination with a sulphur containing residue R3 exhibit better inhibitory effects that compounds wherein R2 does not contain a nitrogen atom or wherein R2 is not linked through a nitrogen atom or wherein R3 does not contain a sulphur atom. Thus it seems to be very advantageous to combine a nitrogen linked residue R2 with a sulphur containing residue R3 in the compounds of general formula (I). The sulphur atom in the residue R3 might preferably be present in form of a sulfoxide or sulphone or sulfoximine or sulfonamide.
The following general formula are especially preferred wherein L, R2 and R3 have the meanings as disclosed herein:




In a further aspect of the present invention, the novel compounds according to the general formula (I) represent chiral compounds. The novel compounds according to the general formula (I) represent a racemate, or a S or a R enantiomer or a mixture of isomers.
In yet another preferred embodiment of the present invention, the compound according to the general formula (I) is selected from the group of compounds depicted in the following Table 1.









TABLE 1





Compound




No.
Structure
Nomenclature







B1




3-[(4-(Piperidin-1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B2




3-[(4-(4,4-Difluoropiperidin-1-yl)-1,3,5- triazin-2-yl)amino] benzenemethanesulfonamide


 


B3




3-[(4-(3,3-Difluoropiperidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B4




rac-3-[(4-(2-Methoxymethylpiperidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B5




(R)-3-[(4-(2-Methoxymethylpiperidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B6




(R)-Methyl 1-[4-((3- (Sulfamoylmethyl)phenyl)amino)-1,3,5- triazin-2-yl]piperidine-2-carboxylate


 


B7




rac-tert-Butyl [(1-(4-((3- (Sulfamoylmethyl)phenyl)amino)-1,3,5- triazin-2-yl)piperidine-2- yl)methyl]carbamate


 


B8




rac-3-[(4-(2-(2- (Dimethylamino)ethyl)piperidin-1-yl)- 1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B9




rac-3-[(4-(2-Phenylpiperidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B10




3-[(4-(Morpholin-4-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B11




3-[(4-(Piperazin-1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B12




3-[(4-(4-Methylpiperazin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B13




rac-3-[(4-(2-(Hydroxymethyl)piperazin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B14




3-[(4-(Pyrrolidin-1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B15




(R)-3-[(4-(2-Methylpyrrolidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B16




(S)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B17




(R)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B18




(S)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B19




3-((4-((2R,5R)- Bis(methoxymethyl)pyrrolidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B20




3-[(4-((2S,5S)- Bis(methoxymethyl)pyrrolidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B21




(R)-Methyl 1-[4-((3- (Sulfamoylmethyl)phenyl)amino)-1,3,5- triazin-2-yl]pyrrolidine-2-carboxylate


 


B22




(S)-Methyl 1-[4-((3- (Sulfamoylmethyl)phenyl)amino)-1,3,5- triazin-2-yl]pyrrolidine-2-carboxylate


 


B23




(R)-1-[4-((3- (Sulfamoylmethyl)phenyl)amino)-1,3,5- triazin-2-yl]pyrrolidine-2-carboxamide


 


B24




(R)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B25




(S)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B26




rac-3-[(4-(2-Benzylpyrrolidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B27




rac-3-[(4-(2-(2-Phenylethyl)pyrrolidin-1- yl)-1,3,5-triazin-2- yl)amino)benzenemethanesulfonamide


 


B28




(R)-3-[(4-(2-(Phenoxymethyl)pyrrolidin- 1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B29




(R)-3-[(4-(2-(Phenylamino- methyl)pyrrolidin-1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B30




(R)-3-[(4-(2-(Benzyloxymethyl)pyrrolidin- 1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B31




(R)-3-[(4-(2-((Pyrrolidin-1- yl)methyl)pyrrolidin-1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B32




(S)-3-[(4-(2-((Pyrrolidin-1- yl)methyl)pyrrolidin-1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B33




rac-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B34




(R)-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B35




(R)-3-[(4-(2-(Methoxymethyl)azetidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B36




(R)-Methyl 1-[4-((3- (Sulfamoylmethyl)phenyl)amino)-1,3,5- triazin-2-yl)azetidine-2-carboxylate


 


B37




rac-3-[(4-(2-Phenylazetidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B38




3-[(4-(7,8-Dihydro-1,6-naphthyridin- yl)amino]benzenemethanesulfonamide


 


B39




3-[(4-(7,8-Dihydropyrido[4,3-d]pyrimidin- 6(5H)-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B40




3-[(4-(3,4-Dihydroisoquinolin-2(1H)-yl)- 1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B41




3-[(4-(5-Amino-3,4-dihydroisoquinolin- 2(1H)-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B42




3-[(4-(3,4-Dihydroquinolin-1(2H)-yl)- 1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B43




3-[(4-(6,7-Dihydrothieno[3,2-c]pyridin- 5(4H)-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B44




3-[(4-(6,7-Dihydro-3H-imidazo[4,5- c]pyridin-5(4H)-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B45




3-[(4-(6,7-Dihydrothiazolo[5,4-c]pyridin- 5(4H)-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B46




3-[(4-(5,6-Dihydroimidazo[1,2-a]pyrazin- 7(8H)-yl)-1,3,5-triazin-2-  yl)amino]benzenemethanesulfonamide


 


B47




3-[(4-(6,7-Dihydropyrazolo[1,5- a]pyrimidin-4(5H)-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B48




3-[(4-(5H-Pyrrolo[3,4-b]pyridin-6(7H)-yl)- 1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B49




3-[(4-(1H-Pyrrolo[3,4-c]pyridin-2(3H)-yl)- 1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B50




3-[(4-(5H-Pyrrolo[3,4-d]pyrimidin-6(7H)- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B51




3-[(4-(Pyrrolo[3,4-c]pyrazol- 5(1H,4H,6H)-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B52




3-[(4-(Indolin-1-yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide


 


B53




(S)-3-[(4-(2-Methylpyrrolidin-1-yl)-1,3,5- triazin-2- yl)amino]benzenemethanesulfonamide


 


B54




(R)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1- yl)-1,3,5-triazin-2- yl)amino]benzenemethanesulfonamide









The compounds of the present invention may form salts with organic or inorganic acids or bases. Examples of suitable acids for such acid addition salt formation are hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, acetic acid, citric acid, oxalic acid, malonic acid, salicylic acid, p-aminosalicylic acid, malic acid, fumaric acid, succinic acid, ascorbic acid, maleic acid, sulfonic acid, phosphonic acid, perchloric acid, nitric acid, formic acid, propionic acid, gluconic acid, lactic acid, tartaric acid, hydroxymaleic acid, pyruvic acid, phenylacetic acid, benzoic acid, p-aminobenzoic acid, p-hydroxybenzoic acid, methanesulfonic acid, ethanesulfonic acid, nitrous acid, hydroxyethanesulfonic acid, ethylenesulfonic acid, p-toluenesulfonic acid, naphthylsulfonic acid, sulfanilic acid, camphorsulfonic acid, china acid, mandelic acid, o-methylmandelic acid, hydrogen-benzenesulfonic acid, picric acid, adipic acid, d-o-tolyltartaric acid, tartronic acid, (o, m, p)-toluic acid, naphthylamine sulfonic acid, trifluoroacetic acid, and other mineral or carboxylic acids well known to those skilled in the art. The salts are prepared by contacting the free base form with a sufficient amount of the desired acid to produce a salt in the conventional manner.
In the case the inventive compounds bear acidic groups, salts could also be formed with inorganic or organic bases. Examples for suitable inorganic or organic bases are, for example, NaOH, KOH, NH4OH, tetraalkylammonium hydroxide, lysine or arginine and the like. Salts may be prepared in a conventional manner using methods well known in the art, for example by treatment of a solution of the compound of the general formula (I) with a solution of an acid, selected out of the group mentioned above.
Syntheses of Compounds
The synthesis of the inventive disubstituted triazines according to the present invention is preferably carried out according to the general synthetic sequences, shown in Schemes 1 and 2.




In a first step 2,4-dichloro-1,3,5-triazine is reacted with anilines R1NH2 to give 2-arylamino-4-chloro-1,3,5-triazines. The reaction is carried out with one equivalent of the aniline in an inert solvent like DMF, THF, DME (1,2-dimethoxyethane), dioxane or an alcohol like isopropanol, or mixtures of such solvents. Preferably the reaction is carried out at a temperature below room temperature in such a way that the reaction mixture is kept homogenous. Preferred conditions use an additional base like triethylamine or N,N-diisopropylethylam ine.
In a second step the intermediate 2-arylamino-4-chloro-1,3,5-triazine is reacted with a cyclic secondary amine H—R2 to give compounds of Formula (I). The reaction is preferably carried out in an inert solvent like THF, dioxane, DME, DMF, DMSO, acetone, isopropanol, ethanol, or methanol or mixtures of such solvents, and at a temperature between 0° C. and reflux temperature of the mixture, preferably at temperatures between 50° C. and 150° C. In preferred procedures at least one equivalent of a base like triethylamine, N,N-diisopropylethylamine, pyridine, potassium carbonate, sodium carbonate, or sodium hydride is added. Preferably the intermediate 2-arylamino-4-chloro-1,3,5-triazine and the amine H—R2 are heated at 60° C. in a mixture of THF and isopropanol and in the presence of an excess of N,N-diisopropylethylamine. In another preferred process the reaction is carried out in DMSO at 80° C. in the presence of potassium carbonate. In a further preferred process the reactants are heated in DMF at 150° C. using a microwave oven.




The synthesis of 1,3,5-triazines of Formula (I) starting from 2,4-dichloro-1,3,5-triazine may be carried out in the inverse order of the reaction steps compared to Scheme 1, in such a manner that in a first step the reaction of a triazine with the amine H—R2 is followed in a second step by the reaction of the intermediate triazine with an aniline H2N—R1. The conditions for both reaction steps are the same as described for the corresponding steps in Scheme 1.
Several compounds of Formula (I) may be prepared by converting substituents which are attached to the aromatic rings R1 and/or of R2 to other substituents using standard reactions which are known to the person skilled in the art. For example, a nitro group can be reduced to an amino group, such an amino group can be converted to a sulfonamide by reaction with a sulfonyl chloride, to a carboxamide by reaction with a carbonyl chloride or another activated derivative of a carboxylic acid, to an urea by reaction with an isocyanate. Carbamate substituents may be cleaved to amino groups, in particular tert-butyl carbamates by reaction with acids like trifluoroacetic acid or hydrochloric acid. Formyl groups may be converted to aminomethyl groups by reaction with primary amines under conditions of a reductive amination.
In a further aspect of the present invention, the novel compounds according to the general formula (I) are used as pharmaceutically active agent.
Pharmaceutical Compositions and Drug Combinations
Another aspect of the present invention relates to drug combinations and pharmaceutical compositions comprising at least one compound of general formula (I) as active ingredient together with at least one pharmaceutically acceptable carrier, excipient and/or diluent and optionally together with one or more other anti-tumor agents or with one or more anti-retroviral drugs. As used herein the term “drug combination” refers to a combination of at least to pharmaceutically active agents or therapeutic agents with or without further ingredients, carrier, diluents and/or solvents. As used herein the term “pharmaceutical composition” refers to a galenic formulation of at least one pharmaceutically active agent together with at least one further ingredient, carrier, diluent and/or solvent.
Compounds of formula (I) may be administered as the sole pharmaceutical agent or in combination with one or more additional therapeutic agents, wherein the drug combination causes no unacceptable adverse effects. This combination therapy includes administration of a single pharmaceutical dosage formulation, which contains a compound of formula (I) and one or more additional therapeutic agents in form of a single pharmaceutical composition, as well as administration of the compound of formula (I) and each additional therapeutic agent in its own separate pharmaceutical dosage formulation, i.e. in its own separate pharmaceutical composition. For example, a compound of formula (I) and a therapeutic agent may be administered to the patient together in a single oral dosage composition such as a tablet or capsule, or each agent may be administered in separate pharmaceutical compositions.
Where separate pharmaceutical compositions are used, the compound of formula (I) and one or more additional therapeutic agents may be administered at essentially the same time (e.g., concurrently) or at separately staggered times (e.g., sequentially).
In particular, the compounds of the present invention may be used in fixed or separate pharmaceutical compositions with other anti-tumor agents such as alkylating agents, anti-metabolites, plant-derived anti-tumor agents, hormonal therapy agents, topoisomerase inhibitors, camptothecin derivatives, kinase inhibitors, targeted drugs, antibodies, interferons and/or biological response modifiers, anti-angiogenic compounds, and other anti-tumor drugs. In this regard, the following is a non-limiting list of examples of secondary agents that may be used in combination with the compounds of the present invention:

    
    
        Alkylating agents include, but are not limited to, nitrogen mustard N-oxide, cyclophosphamide, ifosfamide, thiotepa, ranimustine, nimustine, temozolomide, altretamine, apaziquone, brostallicin, bendamustine, carmustine, estramustine, fotemustine, glufosfamide, mafosfamide, and mitolactol; platinum-coordinated alkylating compounds include, but are not limited to, cisplatin, carboplatin, eptaplatin, lobaplatin, nedaplatin, oxaliplatin, and satraplatin;
        Anti-metabolites include, but are not limited to, methotrexate, 6-mercaptopurine riboside, mercaptopurine, 5-fluorouracil alone or in combination with leucovorin, tegafur, doxifluridine, carmofur, cytarabine, cytarabine ocfosfate, enocitabine, gemcitabine, fludarabin, 5-azacitidine, capecitabine, cladribine, clofarabine, decitabine, eflornithine, ethynylcytidine, cytosine arabinoside, hydroxyurea, melphalan, nelarabine, nolatrexed, ocfosfite, disodium premetrexed, pentostatin, pelitrexol, raltitrexed, triapine, trimetrexate, vidarabine, vincristine, and vinorelbine;
        Hormonal therapy agents include, but are not limited to, exemestane, Lupron, anastrozole, doxercalciferol, fadrozole, formestane, 11-beta hydroxysteroid dehydrogenase 1 inhibitors, 17-alpha hydroxylase/17,20 lyase inhibitors such as abiraterone acetate, 5-alpha reductase inhibitors such as finasteride and epristeride, anti-estrogens such as tamoxifen citrate and fulvestrant, Trelstar, toremifene, raloxifene, lasofoxifene, letrozole, anti-androgens such as bicalutamide, flutamide, mifepristone, nilutamide, Casodex, and anti-progesterones and combinations thereof;
        Plant-derived anti-tumor substances include, e.g., those selected from mitotic inhibitors, for example epothilones such as sagopilone, ixabepilone and epothilone B, vinblastine, vinflunine, docetaxel, and paclitaxel;
        Cytotoxic topoisomerase inhibiting agents include, but are not limited to, aclarubicin, doxorubicin, amonafide, belotecan, camptothecin, 10-hydroxycamptothecin, 9-aminocamptothecin, diflomotecan, irinotecan, topotecan, edotecarin, epimbicin, etoposide, exatecan, gimatecan, lurtotecan, mitoxantrone, pirambicin, pixantrone, rubitecan, sobuzoxane, tafluposide, and combinations thereof;
        Immunologicals include interferons such as interferon alpha, interferon alpha-2a, interferon alpha-2b, interferon beta, interferon gamma-1a and interferon gamma-n1, and other immune enhancing agents such as L19-IL2 and other IL2 derivatives, filgrastim, lentinan, sizofilan, TheraCys, ubenimex, aldesleukin, alemtuzumab, BAM-002, dacarbazine, daclizumab, denileukin, gemtuzumab, ozogamicin, ibritumomab, imiquimod, lenograstim, lentinan, melanoma vaccine (Corixa), molgramostim, sargramostim, tasonermin, tecleukin, thymalasin, tositumomab, Vimlizin, epratuzumab, mitumomab, oregovomab, pemtumomab, and Provenge; Merial melanoma vaccine;
        Biological response modifiers are agents that modify defense mechanisms of living organisms or biological responses such as survival, growth or differentiation of tissue cells to direct them to have anti-tumor activity; such agents include, e.g., krestin, lentinan, sizofiran, picibanil, ProMune, and ubenimex;
        Anti-angiogenic compounds include, but are not limited to, acitretin, aflibercept, angiostatin, aplidine, asentar, axitinib, recentin, bevacizumab, brivanib alaninat, cilengtide, combretastatin, DAST, endostatin, fenretinide, halofuginone, pazopanib, ranibizumab, rebimastat, removab, revlimid, sorafenib, vatalanib, squalamine, sunitinib, telatinib, thalidomide, ukrain, and vitaxin;
        Antibodies include, but are not limited to, trastuzumab, cetuximab, bevacizumab, rituximab, ticilimumab, ipilimumab, lumiliximab, catumaxomab, atacicept, oregovomab, and alemtuzumab;
        VEGF inhibitors such as, e.g., sorafenib, DAST, bevacizumab, sunitinib, recentin, axitinib, aflibercept, telatinib, brivanib alaninate, vatalanib, pazopanib, and ranibizumab; Palladia
        EGFR (HER1) inhibitors such as, e.g., cetuximab, panitumumab, vectibix, gefitinib, erlotinib, and Zactima;
        HER2 inhibitors such as, e.g., lapatinib, tratuzumab, and pertuzumab; mTOR inhibitors such as, e.g., temsirolimus, sirolimus/Rapamycin, and everolimus;
        c-Met inhibitors;
        PI3K and AKT inhibitors;
        CDK inhibitors such as roscovitine and flavopiridol;
        Spindle assembly checkpoints inhibitors and targeted anti-mitotic agents such as PLK inhibitors, Aurora inhibitors (e.g. Hesperadin), checkpoint kinase inhibitors, and KSP inhibitors;
        HDAC inhibitors such as, e.g., panobinostat, vorinostat, MS275, belinostat, and LBH589;
        HSP90 and HSP70 inhibitors;
        Proteasome inhibitors such as bortezomib and carfilzomib;
        Serine/threonine kinase inhibitors including MEK inhibitors (such as e.g. RDEA 119) and Raf inhibitors such as sorafenib;
        Farnesyl transferase inhibitors such as, e.g., tipifarnib;
        Tyrosine kinase inhibitors including, e.g., dasatinib, nilotibib, DAST, bosutinib, sorafenib, bevacizumab, sunitinib, AZD2171, axitinib, aflibercept, telatinib, imatinib mesylate, brivanib alaninate, pazopanib, ranibizumab, vatalanib, cetuximab, panitumumab, vectibix, gefitinib, erlotinib, lapatinib, tratuzumab, pertuzumab, and c-Kit inhibitors; Palladia, masitinib
        Vitamin D receptor agonists;
        Bcl-2 protein inhibitors such as obatoclax, oblimersen sodium, and gossypol;
        Cluster of differentiation 20 receptor antagonists such as, e.g., rituximab;
        Ribonucleotide reductase inhibitors such as, e.g., gemcitabine;
        Tumor necrosis apoptosis inducing ligand receptor 1 agonists such as, e.g., mapatumumab;
        5-Hydroxytryptamine receptor antagonists such as, e.g., rEV598, xaliprode, palonosetron hydrochloride, granisetron, Zindol, and AB-1001;
        Integrin inhibitors including alpha5-beta1 integrin inhibitors such as, e.g., E7820, JSM 6425, volociximab, and endostatin;
        Androgen receptor antagonists including, e.g., nandrolone decanoate, fluoxymesterone, Android, Prost-aid, andromustine, bicalutamide, flutamide, apo-cyproterone, apo-flutamide, chlormadinone acetate, Androcur, Tabi, cyproterone acetate, and nilutamide;
        Aromatase inhibitors such as, e.g., anastrozole, letrozole, testolactone, exemestane, aminoglutethimide, and formestane;
        Matrix metalloproteinase inhibitors;
        Other anti-cancer agents including, e.g., alitretinoin, ampligen, atrasentan bexarotene, bortezomib, bosentan, calcitriol, exisulind, fotemustine, ibandronic acid, miltefosine, mitoxantrone, 1-asparaginase, procarbazine, dacarbazine, hydroxycarbamide, pegaspargase, pentostatin, tazaroten, velcade, gallium nitrate, canfosfamide, darinaparsin, and tretinoin.
    
    


The compounds of the present invention may also be employed in cancer treatment in conjunction with radiation therapy and/or surgical intervention.
Further aspects of the present invention relate to the use of the compounds of general formula (I) for the preparation of a pharmaceutical composition useful for prophylaxis and/or treatment of infectious diseases including opportunistic diseases, immunological diseases, autoimmune diseases, cardiovascular diseases, cell proliferative diseases, inflammation, erectile dysfunction and stroke.
Another aspects of the present invention relate to compounds of general formula (I) for the use for the preparation of a pharmaceutical composition useful for prophylaxis and/or treatment of infectious diseases including opportunistic diseases, immunological diseases, autoimmune diseases, cardiovascular diseases, cell proliferative diseases, inflammation, erectile dysfunction and stroke.
Infectious Diseases Including Opportunistic Infections
In yet another aspect of the present invention, the compounds according to the general formula (I) are for the preparation of a pharmaceutical composition for the prophylaxis and/or treatment of infectious diseases, including opportunistic diseases and opportunistic infections. The term infectious diseases comprises infections caused by viruses, bacteria, prions, fungi, and/or parasites.
Especially, virally induced infectious diseases, including opportunistic diseases are addressed. In a preferred embodiment of this aspect, the virally induced infectious diseases, including opportunistic diseases, are caused by retroviruses, human endogenous retroviruses (HERVs), hepadnaviruses, herpesviruses, flaviviridae, and/or adenoviruses. Preferably, the retroviruses are selected from lentiviruses or oncoretroviruses, wherein the lentivirus is preferably selected from the group comprising: HIV-1, HIV-2, feline immunodeficiency virus (FIV), bovine immunodeficiency virus (BIV), sivian immunodeficiency viruses (SIVs), chimeras of HIV and SIV (SHIV), caprine arthritis encephalitis virus (CAEV), visna/maedi virus (VMV) or equine infectious anemia virus (EIAV), preferably HIV-1 and HIV-2, and the oncoretrovirus is preferably selected from HTLV-I, HTLV-II or bovine leukemia virus (BLV), preferably HTLV-I and HTLV-II.
The hepadnavirus is preferably selected from HBV, ground squirrel hepatitis virus (GSHV) or woodchuck hepatitis virus (WHV), preferably HBV, the herpesvirus is selected from the group comprising: Herpes simplex virus I (HSV I), herpes simplex virus II (HSV II), Epstein-Barr virus (EBV), varicella zoster virus (VZV), human cytomegalovirus (HCMV) or human herpesvirus 8 (HHV-8), preferably HCMV, and the flaviviridae is selected from HCV, West nile or Yellow Fever.
It is to be understood, that all the viruses mentioned above, also comprise drug resistant virus strains.
Examples of infective diseases are AIDS, Alveolar Hydatid Disease (AHD, Echinococcosis), Amebiasis (Entamoeba histolytica Infection), Angiostrongylus Infection, Anisakiasis, Anthrax, Babesiosis (Babesia Infection), Balantidium Infection (Balantidiasis), Baylisascaris Infection (Raccoon Roundworm), Bilharzia (Schistosomiasis), Blastocystis hominis Infection (Blastomycosis), Boreliosis, Botulism, Brainerd Diarrhea, Brucellosis, BSE (Bovine Spongiform Encephalopathy), Candidiasis, Capillariasis (Capillaria Infection), CFS (Chronic Fatigue Syndrome), Chagas Disease (American Trypanosomiasis), Chickenpox (Varicella-Zoster virus), Chlamydia pneumoniae Infection, Cholera, Chronic Fatigue Syndrome, CJD (Creutzfeldt-Jakob Disease), Clonorchiasis (Clonorchis Infection), CLM (Cutaneous Larva Migrans, Hookworm Infection), Coccidioidomycosis, Conjunctivitis, Coxsackievirus A16 (Hand, Foot and Mouth Disease), Cryptococcosis, Cryptosporidium Infection (Cryptosporidiosis), Culex mosquito (Vector of West Nile Virus), Cutaneous Larva Migrans (CLM), Cyclosporiasis (Cyclospora Infection), Cysticercosis (Neurocysticercosis), Cytomegalovirus Infection, Dengue/Dengue Fever, Dipylidium Infection (Dog and Cat Flea Tapeworm), Ebola Virus Hemorrhagic Fever, Echinococcosis (Alveolar Hydatid Disease), Encephalitis, Entomoeba coli Infection, Entomoeba dispar Infection, Entomoeba hartmanni Infection, Entomoeba histolytica Infection (Amebiasis), Entomoeba polecki Infection, Enterobiasis (Pinworm Infection), Enterovirus Infection (Non-Polio), Epstein-Barr Virus Infection, Escherichia coli Infection, Foodborne Infection, Foot and mouth Disease, Fungal Dermatitis, Gastroenteritis, Group A streptococcal Disease, Group B streptococcal Disease, Hansen's Disease (Leprosy), Hantavirus Pulmonary Syndrome, Head Lice Infestation (Pediculosis), Helicobacter pylori Infection, Hematologic Disease, Hendra Virus Infection, Hepatitis (HCV, HBV), Herpes Zoster (Shingles), HIV Infection, Human Ehrlichiosis, Human Parainfluenza Virus Infection, Influenza, Isosporiasis (Isospora Infection), Lassa Fever, Leishmaniasis, Kala-azar (Kala-azar, Leishmania Infection), Leprosy, Lice (Body lice, Head lice, Pubic lice), Lyme Disease, Malaria, Marburg Hemorrhagic Fever, Measles, Meningitis, Mosquito-borne Diseases, Mycobacterium avium Complex (MAC) Infection, Naegleria Infection, Nosocomial Infections, Nonpathogenic Intestinal Amebae Infection, Onchocerciasis (River Blindness), Opisthorciasis (Opisthorcis Infection), Parvovirus Infection, Plague, PCP (Pneumocystis carinii Pneumonia), Polio, Q Fever, Rabies, Respiratory Syncytial Virus (RSV) Infection, Rheumatic Fever, Rift Valley Fever, River Blindness (Onchocerciasis), Rotavirus Infection, Roundworms Infection, Salmonellosis, Salmonella Enteritidis, Scabies, Shigellosis, Shingles, Sleeping Sickness, Smallpox, Streptococcal Infection, Tapeworm Infection (Taenia Infection), Tetanus, Toxic Shock Syndrome, Tuberculosis, Ulcers (Peptic Ulcer Disease), Valley Fever, Vibrio parahaemolyticus Infection, Vibrio vulnificus Infection, Viral Hemorrhagic Fever, Warts, Waterborne infectious Diseases, West Nile Virus Infection (West Nile Encephalitis), Whooping Cough, Yellow Fever.
Immunological Diseases
Another aspect of the present invention is directed to the use of at least one compound of the general formula (I) and/or pharmaceutically acceptable salts thereof for prophylaxis and/or treatment of immunological diseases, neuroimmunological diseases, and autoimmune diseases.
Immunological diseases are, for instance, asthma and diabetes, rheumatic and autoimmune diseases, AIDS, rejection of transplanted organs and tissues (cf. below), rhinitis, chronic obstructive pulmonary diseases, osteoporisis, ulcerative colitis, sinusitis, lupus erythematosus, recurrent infections, atopic dermatitis/eczema and occupational allergies, food allergies, drug allergies, severe anaphylactic reactions, anaphylaxis, and other manifestations of allergic disease, as well as uncommon problems such as primary immunodeficiencies, including antibody deficiency states, cell mediated immunodeficiencies (e.g., severe combined immunodeficiency, DiGeorge syndrome, Hyper-IgE syndrome, Wiskott-Aldrich syndrome, ataxia-telangiectasia), immune mediated cancers, and white cell defects.
In autoimmune diseases, such as systemic lupus erythematosus, rheumatoid arthritis (RA), multiple sclerosis (MS), immune-mediated or type 1 diabetes mellitus, immune mediated glomerulonephritis, scleroderma, pernicious anemia, alopecia, pemphigus, pemphigus vulgaris, myasthenia gravis, inflammatory bowel diseases, Crohn's disease, psoriasis, autoimmune thyroid diseases, and Hashimoto's disease, dermatomyositis, goodpastture syndrome, myasthenia gravis pseudoparalytica, ophtalmia sympatica, phakogene uveitis, chronical agressivce hepatitis, primary billiary cirrhosis, autoimunehemolytic anemy, Werlof disease, specific cells uncontrollably attack the body's own tissues and organs (autoimmunity), producing inflammatory reactions and other serious symptoms and diseases.
Hashimoto's thyroiditis is one of the most common autoimmune diseases. “Autoimmune disease” refers to a category of more than 80 chronic illnesses, each very different in nature, that can affect everything from the endocrine glands (like the thyroid) to organs like the kidneys, as well as to the digestive system.
There are many different autoimmune diseases, and they can each affect the body in different ways. For example, the autoimmune reaction is directed against the brain in multiple sclerosis and the gut in Crohn's disease. In other autoimmune diseases such as systemic lupus erythematosus (lupus), affected tissues and organs may vary among individuals with the same disease. One person with lupus may have affected skin and joints whereas another may have affected skin, kidney, and lungs. Ultimately, damage to certain tissues by the immune system may be permanent, as with destruction of insulin-producing cells of the pancreas in Type 1 diabetes mellitus.
Cardiovascular Diseases
The inventive compounds are also useful for prophylaxis and/or treatment of cardiovascular diseases such as cardiac hypertrophy, adult congenital heart disease, aneurysm, stable angina, unstable angina, angina pectoris, angioneurotic edema, aortic valve stenosis, aortic aneurysm, arrhythmia, arrhythmogenic right ventricular dysplasia, arteriosclerosis, arteriovenous malformations, atrial fibrillation, Behcet syndrome, bradycardia, cardiac tamponade, cardiomegaly, congestive cardiomyopathy, hypertrophic cardiomyopathy, restrictive cardiomyopathy, cardiovascular disease prevention, carotid stenosis, cerebral hemorrhage, Churg-Strauss syndrome, diabetes, Ebstein's Anomaly, Eisenmenger complex, cholesterol embolism, bacterial endocarditis, fibromuscular dysplasia, congenital heart defects, heart diseases, congestive heart failure, heart valve diseases, heart attack, epidural hematoma, hematoma, subdural, Hippel-Lindau disease, hyperemia, hypertension, pulmonary hypertension, hypertrophic growth, left ventricular hypertrophy, right ventricular hypertrophy, hypoplastic left heart syndrome, hypotension, intermittent claudication, ischemic heart disease, Klippel-Trenaunay-Weber syndrome, lateral medullary syndrome, long QT syndrome mitral valve prolapse, moyamoya disease, mucocutaneous lymph node syndrome, myocardial infarction, myocardial ischemia, myocarditis, pericarditis, peripheral vascular diseases, phlebitis, polyarteritis nodosa, pulmonary atresia, Raynaud disease, restenosis, Sneddon syndrome, stenosis, superior vena cava syndrome, syndrome X, tachycardia, Takayasu's arteritis, hereditary hemorrhagic telangiectasia, telangiectasis, temporal arteritis, tetralogy of fallot, thromboangiitis obliterans, thrombosis, thromboembolism, tricuspid atresia, varicose veins, vascular diseases, vasculitis, vasospasm, ventricular fibrillation, Williams syndrome, peripheral vascular disease, varicose veins and leg ulcers, deep vein thrombosis, Wolff-Parkinson-White syndrome.
Preferred are cardiac hypertrophy, adult congenital heart disease, aneurysms, angina, angina pectoris, arrhythmias, cardiovascular disease prevention, cardiomyopathies, congestive heart failure, myocardial infarction, pulmonary hypertension, hypertrophic growth, restenosis, stenosis, thrombosis and arteriosclerosis.
Proliferative Disease
In yet another preferred embodiment, the cell proliferative disease is cancer, which is preferably selected from the group comprising:
The proliferation disorders and cancers are preferably selected from the group comprising or consisting of adenocarcinoma, choroidal melanoma, acute leukemia, acoustic neurinoma, ampullary carcinoma, anal carcinoma, astrocytoma, basal cell carcinoma, pancreatic cancer, desmoid tumor, bladder cancer, bronchial carcinoma, breast cancer, Burkitt's lymphoma, corpus cancer, CUP-syndrome (carcinoma of unknown primary), colorectal cancer, small intestine cancer, small intestinal tumors, ovarian cancer, endometrial carcinoma, ependymoma, epithelial cancer types, Ewing's tumors, gastrointestinal tumors, gastric cancer, gallbladder cancer, gall bladder carcinomas, uterine cancer, cervical cancer, cervix, glioblastomas, gynecologic tumors, ear, nose and throat tumors, hematologic neoplasias, hairy cell leukemia, urethral cancer, skin cancer, skin testis cancer, brain tumors (gliomas), brain metastases, testicle cancer, hypophysis tumor, carcinoids, Kaposi's sarcoma, laryngeal cancer, germ cell tumor, bone cancer, colorectal carcinoma, head and neck tumors (tumors of the ear, nose and throat area), colon carcinoma, craniopharyngiomas, oral cancer (cancer in the mouth area and on lips), cancer of the central nervous system, liver cancer, liver metastases, leukemia, eyelid tumor, lung cancer, lymph node cancer (Hodgkin's/Non-Hodgkin's lymphomas), lymphomas, stomach cancer, malignant melanoma, malignant neoplasia, malignant tumors gastrointestinal tract, breast carcinoma, rectal cancer, medulloblastomas, melanoma, meningiomas, Hodgkin's disease, mycosis fungoides, nasal cancer, neurinoma, neuroblastoma, kidney cancer, renal cell carcinomas, non-Hodgkin's lymphomas, oligodendroglioma, esophageal carcinoma, osteolytic carcinomas and osteoplastic carcinomas, osteosarcomas, ovarial carcinoma, pancreatic carcinoma, penile cancer, plasmocytoma, prostate cancer, pharyngeal cancer, rectal carcinoma, retinoblastoma, vaginal cancer, thyroid carcinoma, Schneeberger disease, esophageal cancer, spinalioms, T-cell lymphoma (mycosis fungoides), thymoma, tube carcinoma, eye tumors, urethral cancer, urologic tumors, urothelial carcinoma, vulva cancer, wart appearance, soft tissue tumors, soft tissue sarcoma, Wilm's tumor, cervical carcinoma, tongue cancer, invasive ductal carcinoma, invasive lobular carcinoma, ductal carcinoma in situ, lobular carcinoma in situ, small-cell lung carcinoma, non-small-cell lung carcinoma, bronchial adenoma, pleuropulmonary blastoma, mesothelioma, brain stem glioma, hypophtalmic glioma, cerebellar astrocytoma, cerebral astrocytoma, neuroectodermal tumours, pineal tumors, sarcoma of the uterus, salivary gland cancers, anal gland adenocarcinomas, mast cell tumors, pelvis tumours, ureter tumours, hereditary papillary renal cancers, sporadic papillary renal cancers, intraocular melanoma, hepatocellular carcinoma (liver cell carcinomas with or without fibrolamellar variant), cholangiocarcinoma (intrahepatic bile duct carcinoma), mixed hepatocellular cholangiocarcinoma, squamous cell carcinoma, malignant melanoma, Merkel cell skin cancer, non-melanoma skin cancer, hypopharyngeal cancer, nasopharyngeal cancer, oropharyngeal cancer, oral cavity cancer, squamous cell cancer, oral melanoma, AIDS-related lymphoma, cutaneous T-cell lymphoma, lymphoma of the central nervous system, malignant fibrous histiocytoma, lymphosarcoma, rhabdomyosarcoma, malignant histiocytosis, fibrosarcoma, hemangiosarcoma, hemangiopericytoma, leiomyosarcoma, canine mammary carcinoma, and feline mammary carcinoma.
Preferred are the following cancer types: Leukemias including but not limited to chronic lymphocytic leukemia, chronic myelogenous leukemia, acute lymphoblastic leukemia, acute myeloid leukemia, mixed lineage leukemia, bladder cancer, breast cancer, breast carcinoma, cancer of the central nervous system, colon carcinoma, gastric cancer, lung cancer, kidney cancer, melanoma, head and neck tumors (tumors of the ear, nose and throat area), ovarian cancer, ovarial carcinoma, cervical cancer, cervix, cervical carcinoma, glioblastomas, pancreatic cancer, pancreatic carcinoma, prostate cancer, stomach cancer, skin cancer, skin testis cancer, Hodgkin's lymphoma, liver cancer, liver metastases and renal cell carcinomas.
Inflammation
In yet another preferred embodiment, said inflammation is mediated preferably by the cytokines TNF-α, IL-1β, GM-CSF, IL-6 and/or IL-8.
As described above, the compounds according to general formula (I) are pharmaceutically active agents for prophylaxis and/or treatment of inflammatory diseases. Thus, these compounds are used for the manufacture of a pharmaceutical formulation for prophylaxis and/or treatment of inflammations and inflammatory diseases in mammals, including humans.
Inflammatory diseases can emanate from infectious and non-infectious inflammatory conditions which may result from infection by an invading organism or from irritative, traumatic, metabolic, allergic, autoimmune, or idiopathic causes as shown in the following list.

    I. Acute infections




















A.
Viral
B.
Bacterial










    II. Noninfectious Causes
    III. Chronic (Granulomatous) Diseases




















A.
Bacterial
B.
Spirochetal



C.
Mycotic (Fungal)
D.
Idiopathic










    IV. Allergic, Immune, and Idiopathic Disorders
    
        A. Hypersensitivity reactions
        B. Immune and idiopathic disorders
    
    
    V. Miscellaneous inflammatory conditions



















A.
Parasitic infections




B.
Inhalation causes:
Acute (thermal) injury





Pollution and inhalant allergy





Carcinogens



C.
Radiation injury:
Radionecrosis










Thus, the compounds disclosed herein can be used for prophylaxis and/or treatment of inflammations caused by invading organisms such as viruses, bacteria, prions, and parasites as well as for prophylaxis and/or treatment of inflammations caused by irritative, traumatic, metabolic, allergic, autoimmune, or idiopathic reasons.
Consequently, the disclosed compounds are useful for prophylaxis and/or treatment of inflammatory diseases which are initiated or caused by viruses, parasites, and bacteria which are connected to or involved in inflammations.
The following bacteria are known to cause inflammatory diseases: mycoplasma pulmonis (causes e.g. chronic lung diseases (CLD), murine chronic respiratory disease), ureaplasma urealyticum (causes pneumonia in newborns), mycoplasma pneumoniae and chlamydia pneumoniae (cause chronic asthma), C. pneumoniae (causes atherosclerosis, pharyngitis to pneumonia with empyema, human coronary heart disease), Helicobacter pylori (human coronary heart disease, stomach ulcers). The following viruses are known to cause inflammatory diseases: herpesviruses especially cytomegalovirus (causes human coronary heart disease).
The compounds disclosed herein are useful for prophylaxis and/or treatment of inflammatory diseases caused and/or induced and/or initiated and/or enhanced by the afore-mentioned bacteria or viruses.
Furthermore, the compounds of formula (I) are useful for prophylaxis and/or treatment of inflammatory diseases of the central nervous system (CNS), inflammatory rheumatic diseases, inflammatory diseases of blood vessels, inflammatory diseases of the middle ear, inflammatory bowel diseases, inflammatory diseases of the skin, inflammatory disease uveitis, inflammatory diseases of the larynx.
Examples for inflammatory diseases of the central nervous system (CNS) are algal disorders, protothecosis, bacterial disorders, abscessation, bacterial meningitis, idiopathic inflammatory disorders, eosinophilic meningoencephalitis, feline polioencephalomyelitis, granulomatous meningoencephalomyelitis, meningitis, steroid responsive meningitis-arteritis, miscellaneous meningitis/meningoencephalitis, meningoencephalitis in greyhounds, necrotizing encephalitis, pyogranulomatous meningoencephalomyelitis, shaker dog disease, mycotic diseases of the CNS, parasitic encephalomyelitis, prion protein induced diseases, feline spongiform encephalopathy, protozoal encephalitis-encephalomyelitis, toxoplasmosis, neosporosis, sarcocystosis, encephalitozoonosis, trypanosomiasis, acanthamebiasis, babesiosis, leishmaniasis, rickettsial disorders, rocky mountain spotted fever, canine ehrlichiosis, salmon poisoning, viral disorders, aujeszky's disease, borna disease, canine herpes virus encephalomyelitis, canine distemper encephalomyelitis, canine distemper encephalomyelitis in immature animals, chronic relapsing encephalomyelitis, post-vaccinal canine distemper encephalitis, feline immunodeficiency virus, feline infectious peritonitis, feline leukemia virus, infectious canine hepatitis, La Crosse virus encephalitis, parvovirus encephalitis, rabies, post-vaccinal rabies.
Examples for inflammatory rheumatic diseases are rheumatoid arthritis, scleroderma, lupus, polymyositis, dermatomyositis, psoriatic arthritis, ankylosing spondylitis, Reiters's syndrome, juvenile rheumatoid arthritis, bursitis, tendinitis (tendonitis), and fibromyositis.
Examples for inflammatory diseases of blood vessels are vasculitis, autoantibodies in vasculitis, microscopic polyangiitis, giant cell arteritis, Takayasu's arteritis, vasculitis of the central nervous system, thromboangiitis obliterans (Buerger's Disease), vasculitis secondary to bacterial, fungal, and parasitic infection, vasculitis and rheumatoid arthritis, vasculitis in systemic lupus erythematosus, vasculitis in the idiopathic inflammatory myopathies, relapsing polychondritis, systemic vasculitis in sarcoidosis, vasculitis and malignancy, and drug-induced vasculitis.
Examples for inflammatory diseases of the middle ear are acute suppurative otitis media, bullous myringitis, granular myringitis, and chronic suppurative otitis media, which can manifest as mucosal disease, cholesteatoma, or both.
Examples for inflammatory bowel diseases are ulcerative colitis, Crohn's disease.
Examples for inflammatory diseases of the skin are acute inflammatory dermatoses, urticaria (hives), spongiotic dermatitis, allergic contact dermatitis, irritant contact dermatitis, atopic dermatitis, erythemal multiforme (EM minor), Stevens-Johnson syndrome (SJS, EM major), toxic epidermal necrolysis (TEN), chronic inflammatory dermatoses, psoriasis, lichen planus, discoid lupus erythematosus, and acne vulgaris.
Uveitis are inflammations located in and/or on the eye and may be associated with inflammation elsewhere in the body. In most circumstances, patients who have uveitis as part of a disease elsewhere in the body are aware of that illness. The majority of patients with uveitis do not have an apparent associated systemic illness. Causes of uveitis can be infectious causes, masquerade syndromes, suspected immune-mediated diseases, and/or syndromes confined primarily to the eye.
The following viruses are associated with inflammations: human immunodeficiency virus-1, herpes simplex virus, herpes zoster virus, and cytomegalovirus.
Bacterial or spirochetal caused, induced, initiated and/or enhanced inflammations are tuberculosis, leprosy, proprionobacterium, syphilis, Whipple's disease, leptospirosis, brucellosis, and lyme disease.
Parasitic (protozoan or helminthic) caused, induced, initiated and/or enhanced inflammations are toxoplasmosis, acanthameba, toxocariasis, cysticercosis, onchocerciasis.
Examples of inflammatory diseases caused, induced, initiated and/or enhanced by fungi are histoplasmosis, coccidioidomycosis, candidiasis, aspergillosis, sporotrichosis, blastomycosis, and cryptococcosis.
Masquerade syndromes are, for instance, leukemia, lymphoma, retinitis pigmentosa, and retinoblastoma.
Suspected immune-mediated diseases can be selected from the group comprising ankylosing spondylitis, Behcet's disease, Crohn's disease, drug or hypersensitivity reaction, interstitial nephritis, juvenile rheumatoid arthritis, Kawasaki's disease, multiple sclerosis, psoriatic arthritis, Reiter's syndrome, relapsing polychondritis, sarcoidosis, Sjogren's syndrome, systemic lupus erythematosus, ulcerative colitis, vasculitis, vitiligo, Vogt Koyanagi Harada syndrome.
Syndromes confined primarily to the eye are, for instance, acute multifocal placoid pigmentary epitheliopathy, acute retinal necrosis, birdshot choroidopathy, Fuch's heterochromic cyclitis, glaucomatocyclitic crisis, lens-induced uveitis, multifocal choroiditis, pars planitis, serpiginous choroiditis, sympathetic ophthalmia, and trauma.
Examples for inflammatory diseases of the larynx are gastroesophageal (laryngopharyngeal) reflux disease, pediatric laryngitis, acute laryngeal infections of adults, chronic (granulomatous) diseases, allergic, immune, and idiopathic disorders and miscellaneous inflammatory conditions.
Pediatric laryngitis is known as acute (viral or bacterial) infection such as laryngotracheitis (croup), supraglottitis (epiglottitis), diphtheria, and noninfectious causes are for example spasmodic croup and traumatic laryngitis.
Acute laryngeal infections of adults are, for instance, viral laryngitis, common upper respiratory infection, laryngotracheitis, herpes simplex, bacterial laryngitis, supraglottitis, laryngeal abscess, and gonorrhea.
Chronic (granulomatous) diseases can be selected from the group comprising bacterial diseases, tuberculosis, leprosy, scleroma, actinomycosis, tularemia, glanders, spirochetal (syphilis) diseases, mycotic (fungal) diseases, candidiasis, blastomycosis, histoplasmosis, coccidiomycosis, aspergillosis, idiopathic diseases, sarcoidosis, and Wegener's granulomatosis.
Allergic, immune, and idiopathic disorders are, for example, hypersensitivity reactions, angioedema, Stevens-Johnson syndrome, immune and idiopathic disorders, infections of the immunocompromised host, rheuatoid arthritis, systeic lupus erythematosus, cicatricial pemphigoid, relapsing polychondritis, Sjogren's syndrome, and amyloidosis.
Miscellaneous inflammatory conditions are, for instance, parasitic infections, trichinosis, leishmaniasis, schistosomiasis, syngamus laryngeus, inhalation laryngitis, acute (thermal) injury, pollution and inhalant allergy, carcinogens, radiation injury, radiation laryngitis, radionecrosis, vocal abuse, vocal-cord hemorrhage, muscle tension dysphonias, and contact ulcer and granuloma.
Stroke
The inventive compounds according to the general formula (I) as well as pharmaceutically acceptable salts thereof are also useful for treatment of stroke.
In another aspect of the present invention, the compounds according to the general formula (I) as well as pharmaceutically acceptable salts thereof are used as an inhibitor for a protein kinase, preferably as an inhibitor for a cellular protein kinase.
In a preferred embodiment of this aspect said cellular protein kinase consists of Cyclin-dependent protein kinases (CDKs).
The cyclin-dependent protein kinase can be selected from the group comprising:
CDK1, CDK2, CDK3, CDK4, CDK5, CDK6, CDK7, CDK8, CDK9, CDK10, CDK11, CrkRS (Crk7, CDC2-related protein kinase 7), CDKL1 (cyclin-dependent kinase-like 1); KKIALRE, CDKL2 (cyclin-dependent kinase-like 2), KKIAMRE, CDKL3 (cyclin-dependent kinase-like 3), NKIAMRE, CDKL4, similar to cyclin-dependent kinase-like 1, CDC2L1 (cell division cycle 2-like 1), PITSLRE B, CDC2L1 (cell division cycle 2-like 1), PITSLRE A, CDC2L5 (cell division cycle 2-like 5), PCTK1 (PCTAIRE protein kinase 1), PCTK2 (PCTAIRE protein kinase 2), PCTK3 (PCTAIRE protein kinase 3) or PFTK1 (PFTAIRE protein kinase 1).
In a further preferred embodiment said cyclin-dependent protein kinase is CDK9. Thus, the compounds according to the general formula (I) as well as pharmaceutically acceptable salts thereof are used as an inhibitor for CDK9.
Surprisingly it turned out that the compounds according to the general formula (I) as well as pharmaceutically acceptable salts thereof selectively inhibit CDK9 in comparison to other protein kinases and in comparison to other cyclin-dependent protein kinases. Thus, the compounds according to the general formula (I) as well as pharmaceutically acceptable salts thereof are used as selective inhibitors for CDK9.
As used herein, a kinase “inhibitor” refers to any compound capable of downregulating, decreasing, suppressing or otherwise regulating the amount and/or activity of a kinase. Inhibition of these kinases can be achieved by any of a variety of mechanisms known in the art, including, but not limited to binding directly to the kinase polypeptide, denaturing or otherwise inactivating the kinase, or inhibiting the expression of the gene (e.g., transcription to mRNA, translation to a nascent polypeptide, and/or final polypeptide modifications to a mature protein), which encodes the kinase. Generally, kinase inhibitors may be proteins, polypeptides, nucleic acids, small molecules, or other chemical moieties.
As used herein the term “inhibiting” or “inhibition” refers to the ability of a compound to downregulate, decrease, reduce, suppress, inactivate, or inhibit at least partially the activity of an enzyme, or the expression of an enzyme or protein and/or the virus replication.
In a further aspect of the present invention, a method for preventing and/or treating infectious diseases, including opportunistic diseases, in a mammal, especially in a human, is provided, which method comprises administering to the mammal an amount of at least one compound according to the general formula (I), effective to prevent and/or treat said infectious diseases, including opportunistic diseases. In a preferred embodiment of this method, the infectious diseases, including opportunistic diseases, are virally induced infectious diseases. The virally induced infectious diseases, including opportunistic diseases, are caused by retroviruses, hepadnaviruses, herpesviruses, flaviviridae, and/or adenoviruses. In a further preferred embodiment of this method, the retroviruses are selected from lentiviruses or oncoretroviruses, wherein the lentivirus is selected from the group comprising: HIV-1, HIV-2, FIV, BIV, SIVs, SH IV, CAEV, VMV or EIAV, preferably HIV-1 or HIV-2 and wherein the oncoretrovirus is selected from the group consisting of: HTLV-I, HTLV-II or BLV. In a further preferred embodiment of this method, the hepadnavirus is selected from HBV, GSHV or WHV, preferably HBV, the herpesivirus is selected from the group comprising: HSV I, HSV II, EBV, VZV, HCMV or HHV 8, preferably HCMV and the flaviviridae is selected from HCV, West nile or Yellow Fever.
In a further aspect of the present invention, methods for preventing and/or treating infectious diseases including opportunistic diseases, prion diseases, immunological diseases, autoimmune diseases, cardiovascular diseases, cell proliferative diseases, inflammation, erectile dysfunction and stroke in a mammal, especially in a human, are provided, which methods comprise administering to the mammal an amount of at least one compound according to the general formula (I) and/or pharmaceutically acceptable salts thereof, effective to prevent and/or treat said infectious diseases including opportunistic diseases, prion diseases, immunological diseases, autoimmune diseases, cardiovascular diseases, cell proliferative diseases, inflammation, erectile dysfunction and stroke.
In further preferred embodiments, the specific diseases addressed as infectious diseases including opportunistic diseases, prion diseases, immunological diseases, autoimmune diseases, cardiovascular diseases, cell proliferative diseases, inflammation, erectile dysfunction and stroke are selected from the groups disclosed above.
The compounds shown explicitly in Table 1 are preferred to be used within the methods or indications disclosed herein. Another aspect of the present invention is that at least one compound according to the general formula (I) used as a pharmaceutically active agent may be administered in combination with further therapeutic compounds.
For the indication HIV compounds according to the general formula (I), preferably those falling under the activity range “a” for CDK9 as shown in Table 5, may be administered in combination with anti-retroviral drugs, selected from the following five classes:

    
    
        1) Nucleoside reverse transcriptase inhibitors (NRTIs),
        2) Non-nucleoside reverse transcriptase inhibitors (NNRTIs),
        3) Protease inhibitors (PIs),
        4) Fusion inhibitors or
        5) Immune stimuli.
    
    


Thus, another aspect of the present invention relates to drug combinations comprising at least one inventive compound according to general formula (I) and/or pharmaceutically acceptable salts thereof together with at least one anti-retroviral drug, especially at least one of the drugs mentioned above.
The pharmaceutical compositions according to the present invention comprise at least one compound according to the present invention as an active ingredient together with at least one pharmaceutically acceptable (i.e. non-toxic) carrier, excipient and/or diluent. The pharmaceutical compositions of the present invention can be prepared in a conventional solid or liquid carrier or diluent and a conventional pharmaceutically-made adjuvant at suitable dosage level in a known way. The preferred preparations are adapted for oral application. These administration forms include, for example, pills, tablets, film tablets, coated tablets, capsules, powders and deposits.
Furthermore, the present invention also includes pharmaceutical preparations for parenteral application, including dermal, intradermal, intragastral, intracutan, intravasal, intravenous, intramuscular, intraperitoneal, intranasal, intravaginal, intrabuccal, percutan, rectal, subcutaneous, sublingual, topical, or transdermal application, which preparations in addition to typical vehicles and/or diluents contain at least one compound according to the present invention and/or a pharmaceutical acceptable salt thereof as active ingredient.
The pharmaceutical compositions according to the present invention containing at least one compound according to the present invention and/or a pharmaceutical acceptable salt thereof as active ingredient will typically be administered together with suitable carrier materials selected with respect to the intended form of administration, i.e. for oral administration in the form of tablets, capsules (either solid filled, semi-solid filled or liquid filled), powders for constitution, gels, elixirs, dispersable granules, syrups, suspensions, and the like, and consistent with conventional pharmaceutical practices. For example, for oral administration in the form of tablets or capsules, the active drug component may be combined with any oral non-toxic pharmaceutically acceptable carrier, preferably with an inert carrier like lactose, starch, sucrose, cellulose, magnesium stearate, dicalcium phosphate, calcium sulfate, talc, mannitol, ethyl alcohol (liquid filled capsules) and the like. Moreover, suitable binders, lubricants, disintegrating agents and coloring agents may also be incorporated into the tablet or capsule. Powders and tablets may contain about 5 to about 95-weight % of the 4,6-disubstituted pyrimdine derivative according to the general formula (I) or analogues compound thereof or the respective pharmaceutically active salt as active ingredient.
Suitable binders include starch, gelatin, natural sugars, corn sweeteners, natural and synthetic gums such as acacia, sodium alginate, carboxymethylcellulose, polyethylene glycol and waxes. Among suitable lubricants there may be mentioned boric acid, sodium benzoate, sodium acetate, sodium chloride, and the like. Suitable disintegrants include starch, methylcellulose, guar gum, and the like. Sweetening and flavoring agents as well as preservatives may also be included, where appropriate. The disintegrants, diluents, lubricants, binders etc. are discussed in more detail below.
Moreover, the pharmaceutical compositions of the present invention may be formulated in sustained release form to provide the rate controlled release of any one or more of the components or active ingredients to optimise the therapeutic effect(s), e.g. antihistaminic activity and the like. Suitable dosage forms for sustained release include tablets having layers of varying disintegration rates or controlled release polymeric matrices impregnated with the active components and shaped in tablet form or capsules containing such impregnated or encapsulated porous polymeric matrices.
Liquid form preparations include solutions, suspensions, and emulsions. As an example, there may be mentioned water or water/propylene glycol solutions for parenteral injections or addition of sweeteners and opacifiers for oral solutions, suspensions, and emulsions. Liquid form preparations may also include solutions for intranasal administration.
Aerosol preparations suitable for inhalation may include solutions and solids in powder form, which may be present in combination with a pharmaceutically acceptable carrier such as an inert, compressed gas, e.g. nitrogen.
For preparing suppositories, a low melting wax, such as a mixture of fatty acid glycerides like cocoa butter is melted first, and the active ingredient is then dispersed homogeneously therein e.g. by stirring. The molten, homogeneous mixture is then poured into conveniently sized moulds, allowed to cool, and thereby solidified.
Also included are solid form preparations, which are intended to be converted, shortly before use, to liquid form preparations for either oral or parenteral administration. Such liquid forms include solutions, suspensions, and emulsions.
The compounds according to the present invention may also be delivered transdermally. The transdermal compositions may have the form of a cream, a lotion, an aerosol and/or an emulsion and may be included in a transdermal patch of the matrix or reservoir type as is known in the art for this purpose.
The term capsule as recited herein refers to a specific container or enclosure made e.g. of methylcellulose, polyvinyl alcohols, or denatured gelatins or starch for holding or containing compositions comprising the active ingredient(s). Capsules with hard shells are typically made of blended of relatively high gel strength gelatins from bones or pork skin. The capsule itself may contain small amounts of dyes, opaquing agents, plasticisers and/or preservatives.
Under tablet a compressed or moulded solid dosage form is understood which comprises the active ingredients with suitable diluents. The tablet may be prepared by compression of mixtures or granulations obtained by wet granulation, dry granulation, or by compaction well known to a person of ordinary skill in the art.
Oral gels refer to the active ingredients dispersed or solubilised in a hydrophilic semi-solid matrix.
Powders for constitution refers to powder blends containing the active ingredients and suitable diluents which can be suspended e.g. in water or in juice.
Suitable diluents are substances that usually make up the major portion of the composition or dosage form. Suitable diluents include sugars such as lactose, sucrose, mannitol, and sorbitol, starches derived from wheat, corn rice, and potato, and celluloses such as microcrystalline cellulose. The amount of diluent in the composition can range from about 5 to about 95% by weight of the total composition, preferably from about 25 to about 75 weight %, and more preferably from about 30 to about 60 weight %.
The term disintegrants refers to materials added to the composition to support break apart (disintegrate) and release the pharmaceutically active ingredients of a medicament. Suitable disintegrants include starches, “cold water soluble” modified starches such as sodium carboxymethyl starch, natural and synthetic gums such as locust bean, karaya, guar, tragacanth and agar, cellulose derivatives such as methylcellulose and sodium carboxymethylcellulose, microcrystalline celluloses, and cross-linked microcrystalline celluloses such as sodium croscaramellose, alginates such as alginic acid and sodium alginate, clays such as bentonites, and effervescent mixtures. The amount of disintegrant in the composition may range from about 2 to about 20 weight % of the composition, more preferably from about 5 to ca. 10 weight %.
Binders are substances which bind or “glue” together powder particles and make them cohesive by forming granules, thus serving as the “adhesive” in the formulation. Binders add cohesive strength already available in the diluent or bulking agent. Suitable binders include sugars such as sucrose, starches derived from wheat corn rice and potato, natural gums such as acacia, gelatin and tragacanth, derivatives of seaweed such as alginic acid, sodium alginate and ammonium calcium alginate, cellulose materials such as methylcellulose, sodium carboxymethylcellulose and hydroxypropylmethylcellulose, polyvinylpyrrolidone, and inorganic compounds such as magnesium aluminum silicate. The amount of binder in the composition may range from about 2 to about 20 weight % of the composition, preferably from about 3 to about 10 weight %, and more preferably from about 3 to about 6 weight %.
Lubricants refer to a class of substances which are added to the dosage form to enable the tablet granules etc. after being compressed to release from the mould or die by reducing friction or wear. Suitable lubricants include metallic stearates such as magnesium stearate, calcium stearate, or potassium stearate, stearic acid, high melting point waxes, and other water soluble lubricants such as sodium chloride, sodium benzoate, sodium acetate, sodium oleate, polyethylene glycols and D,L-leucine. Lubricants are usually added at the very last step before compression, since they must be present at the surface of the granules. The amount of lubricant in the composition may range from about 0.2 to about 5 weight % of the composition, preferably from about 0.5 to about 2 weight %, and more preferably from about 0.3 to about 1.5 weight % of the composition.
Glidents are materials that prevent caking of the components of the pharmaceutical composition and improve the flow characteristics of granulate so that flow is smooth and uniform. Suitable glidents include silicon dioxide and talc. The amount of glident in the composition may range from about 0.1 to about 5 weight % of the final composition, preferably from about 0.5 to about 2 weight %.
Coloring agents are excipients that provide coloration to the composition or the dosage form. Such excipients can include food grade dyes adsorbed onto a suitable adsorbent such as clay or aluminum oxide. The amount of the coloring agent may vary from about 0.1 to about 5 weight % of the composition, preferably from about 0.1 to about 1 weight %.
EXAMPLES
Preparation of Compounds
Abbreviations Used in the Description of the Chemistry and in the Examples that Follow are:
CDCl3 (deuterated chloroform); cHex (cyclohexane); DCM (dichloromethane); DIPEA (di-iso-propylethylamine); DMF (dimethylformamide); DMSO (dimethyl sulfoxide); eq (equivalent); ES (electrospray); EtOAc (ethyl acetate); EtOH (ethanol); MeOH (methanol); MS (mass spectrometry); NMR (nuclear magnetic resonance); Pd(dppf)Cl2 ([1,1′-bis(diphenylphosphino)ferrocene]dichloro palladium(II) complex with dichloromethane); iPrOH (iso-propanol); RT (room temperature); sat. aq. (saturated aqueous); SiO2 (silica gel); TFA (trifluoroacetic acid); THF (tetrahydrofuran).
Preparative Examples
Intermediates
Intermediate 1: 
3-[(4-Chloro-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (A1)
To a solution of 2,4-dichloro-1,3,5-triazine (1.0 eq) in dry DMF (0.7 M) at 0° C. under N2 atmosphere was added a solution of (3-aminophenyl)-methanesulfonamide (1.0 eq) in dry DMF (0.7 M). The reaction mixture was stirred for 2.5 h at 0° C. Then water was added and the aqueous solution was neutralized with sat. aq. NaHCO3 solution. The aqueous layer was extracted with EtOAc and the combined organic layers were dried over Na2SO4. Evaporation of solvent gave A1 as a white solid which was used in the next step without further purification. 1H NMR (400 MHz, d6-DMSO, 300K) δ 4.26 (s, 2H), 6.89 (s, 2H), 7.15 (d, J=7.4 Hz, 1H), 7.36 (d, J=7.4 Hz, 1H), 7.66 (d, J=7.4 Hz, 1H), 8.64 (s, 1H), 9.75 (br. s, 1H), 10.83 (s, 1H). MS (ES) C10H10ClN5O2S requires: 299. found: 300 (M+H)+.
Intermediate 2: 
(R)-2-Methoxymethylpiperidine Hydrochloride (A2)
Step 1: To an ice-cold mixture of D-pipecolic acid (2.0 g, 15.5 mmol) in THF (30 mL) was added dropwise a 1 M solution of borane in THF (46 mL). It was stirred over night at room temperature and the clear solution was carefully quenched with aq. 3 M NaOH (63 mL). The mixture was heated with reflux over night. It was cooled with ice and bis-tert-butyl dicarbonate (3.49 g, 16.0 mmol) was added. It was stirred again over night while another amount of the dicarbonate was added in order to drive the reaction to completion. The organic solvent was removed under reduced pressure and the remaining aqueous mixture was extracted with ether. The organic layer was washed with brine, dried over MgSO4, and concentrated in vacuo.
Step 2: The residue from Step 1, a mixture of the N-Boc and N,O-bis-Boc protected aminoalcohol, was treated with a solution of KOH in MeOH (0.36 M). After stirring over night it was diluted with chloroform and neutralized with aq. 1 N HCl. The aqueous layer was extracted several times with chloroform, the combined organic layers dried over MgSO4, and concentrated under reduced pressure. The residue was chromatographed (silica gel, cyclohexane/EtOAc 90:10 to 80:20) to leave the intermediate (R)—N-Boc-2-hydroxypiperidine (0.97 g, 29%).
Step 3: A suspension of sodium hydride (0.13 g, 5.4 mmol) in DMF (4 mL) was cooled with ice and a solution of the intermediate alcohol of Step 2 (0.97 g, 4.5 mmol) in DMF (1 mL) was added dropwise. After stirring for 30 min at room temperature iodomethane (0.96 g, 6.8 mmol) was added and the mixture was stirred over night. The mixture was quenched with saturated NH4Cl-solution and extracted with EtOAc. The combined organic extracts were washed with brine, dried over MgSO4 and concentrated under reduced pressure to leave the crude (R)—N-Boc-2-methoxymethylpiperidine (0.88 g, 85%).
Step 4: The crude intermediate of Step 3 (0.88 g, 3.8 mmol) was dissolved in chloroform (5 mL) and treated with a 4 M solution of HCl in dioxane (5 mL). After stirring over night the solvent was removed in vacuo to leave the crude title hydrochloride salt which was used for the next step without further purification.
Intermediate 3: 
(R)-2-(Phenoxymethyl)pyrrolidine Trifluoroacetate (A3)
Step 1: To a solution of (R)-2-(hydroxymethyl)pyrrolidine (2.0 g, 19.8 mmol) in DCM (20 mL) was added a 1M aqueous solution of NaOH (20 mL). Bis-tert-butyl dicarbonate (4.32 g, 19.8 mmol) in DCM (10 mL) was added dropwise and the mixture was stirred for 5 hours at room temperature. Both layers were separated, the organic layer washed two times with water and the aqueous layer extracted with DCM. The combined organic layers were dried over MgSO4, and the solvent was removed under reduced pressure.
The intermediate tert-butyl (R)-2-(hydroxymethyl)pyrrolidine-1-carboxylate was isolated by crystallization from hexane; yield: 2.60 g (65%).
Step 2: To an ice-cold solution of the compound from Step 1 (2.60 g, 12.9 mmol), phenol (3.64 g, 38.7 mmol), and triphenylphosphine (6.77 g, 25.8 mmol) in DCM (50 mL) was added a solution of diisopropyl azodicarboxylate (5.22 g, 25.8 mmol) in toluene (30 mL). After stirring at room temperature for 60 hours it was diluted with ether and the solution was extracted with aqueous 3N NaOH. The organic layer was washed with brine, dried over MgSO4, and concentrated in vacuo. The intermediate tert-butyl (R)-2-(phenoxymethyl)pyrrolidine-1-carboxylate was obtained from the residue by column chromatography (silica gel, cHex/EtOAc 29:1); yield: 1.82 g (51%).
Step 3: To solution of the intermediate of Step 2 (1.65 g, 5.9 mmol) in DCM (8 mL) was added trifluoroacetic acid (8 mL). The mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure to afford the crude title trifluoroacetate (1.85 g) which was used for the next step without further purification.
Intermediate 4: 
(R)-2-(Benzyloxymethyl)pyrrolidine (A4)
Step 1: A solution of tert-butyl (R)-2-(hydroxymethyl)pyrrolidine-1-carboxylate
(Intermediate 3, Step 1; 2.60 g, 12.9 mmol) in dry THF (10 mL) was cooled with ice and sodium hydride (0.36 g, 14.9 mmol) was added in small portions. After stirring for 30 min a solution of benzyl bromide (2.54 g, 14.9 mmol) in THF (2 mL) was added dropwise. It was stirred over night, the mixture poured into water (20 ml), and extracted with EtOAc. The combined organic layers were washed with brine and dried over MgSO4. The solvent was removed under reduced pressure to leave crude tert-butyl (R)-2-(benzyloxymethyl)pyrrolidine-1-carboxylate (3.90 g, quant.) which was used for the next step without further purification.
Step 2: The carboxylate from the previous step (3.70 g, 12.7 mmol) was cooled with ice and treated with 90% formic acid. After stirring for 3 hours it was concentrated to dryness under reduced pressure. Saturated K2CO3 solution was added until the mixture reached pH 8-10. It was extracted with EtOAc and the organic layer dried over MgSO4. The solvent was removed in vacuo and the title compound obtained after column chromatography (silica gel, DCM/MeOH/conc.NH3 80:1:1); yield: 1.45 g (60%).
Intermediate 5: 
(R)-2-Phenylpyrrolidine (A5)
The amine A5 was prepared by resolution of racemic 2-phenylpyrrolidine through formation of its (S)-alpha-methoxyphenylacetamide according to the procedure described in WO1995/00508.
Intermediate 6: 
(R)-2-Methoxymethylazetidine Hydrochloride (A6)
The title compound A6 was prepared from (R)-azetidine-2-carboxylic acid by essentially the same procedure as described for A2.
Intermediate 7: 
(R)-Methyl Azetidine-2-carboxylate Hydrochloride (A7)
A suspension of (0.25 g, 2.5 mmol) in MeOH (10 mL) was cooled with ice and thionyl chloride (0.45 g, 3.8 mml) was added. After stirring over night at room temperature the solvent was removed under reduced pressure to leave the crude title amine hydrochloride which was used for the next step without further purification.
Intermediate 2: 
3-[(4-Chloro-1,3,5-triazin-2-yl)amino]benzenesulfonamide (A2)
A2 was prepared following the general procedure reported for A1 using 2,4-dichloro-1,3,5-triazine and 3-aminobenzenesulfonamide as reacting agents. The crude product was purified by flash chromatography on silica gel (cHex/EtOAc=20:1 to 1:20) to yield the desired product A2 as a white solid (35%). 1H NMR (400 MHz, d6-DMSO, 300K) δ 7.41 (s, 2H), 7.59 (m, 2H), 7.85 (d, J=6.9 Hz, 1H), 8.21 (br.s, 1H), 8.69 (s, 1H), 11.02 (s, 1H). MS (ES) C9H8ClN5O2S requires: 285. found: 286 (M+H)+.
Intermediate 3: 
2-(3-Aminophenyl)ethanesulfonamide (A3)
The compound A3 has been obtained by reduction of 2-(3-nitrophenyl)-ethanesulfonamide according to the procedure described in WO2009/076140, and the nitro compound from 2-(3-nitrophenyl)ethanol according to J.Med.Chem. 45 (2002), 567-583.
Intermediate 4: 
2-[3-((4-Chloro-1,3,5-triazin-2-yl)amino)phenyl]ethanesulfonamide (A4)
A4 was prepared following the general procedure reported for A1 using 2,4-dichloro-1,3,5-triazine and 2-(3-aminophenyl)ethanesulfonamide A3 as reacting agents. The crude product was purified by flash chromatography on silica gel (DCM/MeOH=100:0 to 4:1) to yield the desired product A5 (54%, 65% purity) as a brown solid. MS (ES) C11H12ClN5O2S requires: 313. found: 314 (M+H)+.
Example Compounds
Example 1
3-[(4-(Piperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethane sulfonamide (B1)
A mixture of triazine A1 (99 mg, 0.33 mmol) and piperidine (42.6 mg, 0.50 mmol) in dry
DMF (3 mL) was heated for 90 min at 150° C. in a microwave oven. The mixture was concentrated under reduced pressure and the residue was purified by reverse phase RP-HPLC (column: C18), using H2O (0.1% TFA) and MeOH (0.1% TFA) as eluents. The desired fractions were lyophilized to afford the titled compound (B1) (91 mg, 79%) as a white powder. 1H NMR (400 MHz, d6-DMSO, 300K) δ 1.50 (m, 4H), 1.61 (m, 2H), 3.73 (m, 4H), 4.18 (s, 2H), 6.82 (s, 2H), 6.98 (d, J=7.8 Hz, 1H), 7.26 (t, J=7.8 Hz, 1H), 7.60 (d, J=7.8 Hz, 1H), 7.76 (s, 1H), 8.19 (s, 1H), 9.59 (s, 1H). MS (ES) C15H20N6O2S requires: 348. found: 349 (M+H)+.
Example 2
3-[(4-(4,4-Difluoropiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B2)
To a solution of triazine A1 (270 mg, 0.9 mmol) in DMSO (0.9 mL) were added K2CO3 (370 mg, 2.7 mmol) and 4,4-difluoropiperidine hydrochloride (140 mg, 0.9 mmol). The mixture was stirred for 2 hours at 80° C. After cooling to room temperature it was diluted with EtOAc and extracted with water. The organic layer was washed with brine, dried over MgSO4 and concentrated under reduced pressure. The residue was stirred with DCM to give the solid title compound B2 which was isolated by filtration and dried in vacuo (white powder, 110 mg, 32%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.90-2.15 (m, 4H), 3.81-3.98 (m, 4H), 4.21 (s, 2H), 6.88 (s, 2H), 6.98-7.04 (m, 1H), 7.21-7.34 (m, 1H), 7.44-7.60 (m, 1H), 7.78-7.90 (m, 1H), 8.25 (s, 1H), 9.77 (s, 1H). MS (ES) C15H18F2N6O2S requires: 384. found: 385 (M+H)+.
Example 3
3-[(4-(3,3-Difluoropiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B3)
B3 was obtained as a white crystalline powder by following the procedure reported for B2 using A1 and 3,3-difluoropiperidine hydrochloride; yield: 125 mg (36%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.62-1.79 (m, 2H), 2.00-2.21 (m, 2H), 3.82 (bs, 2H), 4.05-4.25 (m, 2H), 4.22 (s, 2H), 6.87 (bs, 2H), 6.95-7.08 (m, 1H), 7.23-7.35 (m, 1H), 7.48-7.88 (m, 2H), 8.27 (s, 1H), 9.78 (s, 1H). MS (ES) C15H18F2N6O2S requires: 384. found: 385 (M+H)+.
Example 4
rac-3-[(4-(2-Methoxymethylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B4)
B4 was prepared following the procedure reported for B2 using A1 and rac-2-methoxymethylpiperidine hydrochloride. The title compound was purified by thick-layer chromatography (silica gel, chloroform/MeOH 19:1) and obtained as a white crystalline solid; yield: 235 mg (67%). MS (ES) C17H24N6O3S requires: 392. found: 393 (M+H)+.
Example 5
(R)-3-[(4-(2-Methoxymethylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B5)
B5 was prepared following the procedure reported for B4 using the intermediates A1 and A2 and obtained as a white crystalline solid; yield: 90 mg (25%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.27-1.90 (m, 4H), 2.48-2.60 (m, 2H), 2.90-3.03 (m, 1H), 3.38 (s, 3H), 3.61 (bs, 2H), 4.26 (s, 2H), 4.61-4.75 (m, 1H), 5.03-5.15 (m, 1H), 6.90 (bs, 2H), 7.03-7.10 (m, 1H), 7.28-7.40 (m, 1H), 7.62-7.73 (m, 1H), 7.82 (bs, 1H), 8.28 (s, 1H), 9.68 (bs, 1H). MS (ES) C17H24N6O3S requires: 392. found: 393 (M+H)+.
Example 6
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]piperidine-2-carboxylate (B6)
B6 was prepared following the procedure reported for B2 using A1 and (R)-methyl piperidine-carboxylate hydrochloride. The title compound was purified by thick-layer chromatography (silica gel, chloroform/MeOH 9:1) and obtained as a white crystalline solid; yield: 160 mg (44%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.23-1.57 (m, 2H), 1.65-1.85 (m, 2H), 2.17-2.32 (m, 1H), 2.92-3.08 (m, 1H), 3.71 (s, 3H), 4.18-4.31 (m, 2H), 4.60-4.77 (m, 1H), 5.45-5.60 (m, 1H), 5.82 (s, 1H), 6.88 (bs, 2H), 7.00-7.10 (m, 1H), 7.26-7.40 (m, 1H), 7.57-7.88 (m, 2H), 8.30 and 8.38 (2s, 1H), 9.83 (bs, 1H). MS (ES) C17H22N6O4S requires: 406. found: 407 (M+H)+.
Example 7
rac-tert-Butyl[(1-(4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl)piperidine-2-yl)methyl]carbamate (B7)
B7 was prepared following the procedure reported for B4 using A1 and rac-tert-butyl (piperidin-2-yl)methylcarbamate and obtained as a white crystalline solid; yield: 280 mg (65%). MS (ES) C21H31N7O4S requires: 477. found: 478 (M+H)+.
Example 8
rac-3-[(4-(2-(2-(Dimethylamino)ethyl)piperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B8)
B8 was prepared following the procedure reported for B4 using A1 and rac-2-(2-(dimethylamino)ethyl)piperidine and obtained as a white crystalline solid; yield: 230 mg (61%). MS (ES) C19H29N7O2S requires: 419. found: 420 (M+H)+.
Example 9
rac-3-[(4-(2-Phenylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B9)
B9 was obtained as a white crystalline solid by following the procedure reported for B4 using A1 and rac-2-phenylpiperidine; yield: 100 mg (26%). MS (ES) C21H24N6O2S requires: 424. found: 425 (M+H)+.
Example 10
3-[(4-(Morpholin-4-yl)-1,3,5-triazin-2-yl)amino]benzenemethane sulfonamide (B10)
To a solution of triazine A1 (300 mg, 1.0 mmol) in a mixture of THF (4 mL) and iPrOH (4 mL) were added morpholine (87 mg, 1.0 mmol) and DIPEA (200 μL, 1.15 mmol). The mixture was heated for 2 hours at 60° C. and the solvent removed under reduced pressure. The residue was treated with MeOH and the solid title compound (B10) collected by filtration, washed with MeOH, and dried in vacuo. Yield: 93.3 mg (27%), colorless amorphous solid. MS (ES) C14H18N6O3S requires: 350. found: 351 (M+H)+.
Example 11
3-[(4-(piperazin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethane sulfonamide (B11)
B11 was prepared following the procedure reported for B1 using A1 and piperazine. MS (ES) C14H19N7O2S requires: 349. found: 350 (M+H)+.
Example 12
3-[(4-(4-Methylpiperazin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B12)
B12 was prepared following the procedure reported for B1 using A1 and N-methylpiperazine. MS (ES) C15H21N7O3S requires: 363. found: 364 (M+H)+.
Example 13
rac-3-[(4-(2-(Hydroxymethyl)piperazin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B13)
B13 was prepared following the procedure reported for B1 using A1 and rac-2-(hydroxymethyl)piperazine. MS (ES) C15H21N7O3S requires: 379. found: 380 (M+H)+.
Example 14
3-[(4-(Pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethane sulfonamide (B14)
B14 was prepared following the general procedure reported for B10 using A1 and pyrrolidine. Yield: 96.6 mg (29%), colorless amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 1.84-1.96 (m, 4H), 3.43-3.55 (m, 4H), 4.18 (s, 2H), 6.81 (s, 2H), 6.96 (d, J=7.6 Hz, 1H), 7.25 (t, J=7.9 Hz, 1H), 7.69 (d, J=9.5 Hz, 1H), 7.86 (s, 1H), 8.18 (s, 1H), 9.59 (s, 1H). MS (ES) C14H18N6O2S requires: 334. found: 335 (M+H)+.
Example 15
(R)-3-[(4-(2-Methylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B15)
B15 was prepared following the general procedure reported for B10 using A1 and (R)-2-methylpyrrolidine. Yield: 31.5 mg (9%), colorless amorphous solid. MS (ES) C15H20N6O2S requires: 348. found: 349 (M+H)+.
Example 16
(S)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B16)
B16 was prepared following the general procedure reported for B10 using A1 and (S)-2-(trifluoromethyl)pyrrolidine. The title compound (B16) was purified by reverse phase RP-HPLC (column: C18), using H2O (0.1% TFA) and MeOH (0.1% TFA) as eluents. Yield: 58.7 mg (15%), colorless amorphous solid. MS (ES) C15H17F3N6O2S requires: 402. found: 403 (M+H)+.
Example 17
(R)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B17)
B17 was prepared following the procedure reported for B1 using A1 and (R)-2-(methoxymethyl)pyrrolidine. 1H NMR (400 MHz, DMSO, 300K) δ 1.85-2.01 (m, 4H), 3.25 (s, 3H), 3.31-3.63 (m, 4H), 4.18-4.29 (m, 3H), 6.81 (s, 2H), 6.98 (t, J=8.4 Hz, 1H), 7.25 (dt, J=4.7 Hz, J=7.8 Hz, 1H), 7.65-7.87 (m, 2H), 8.20 (s, 1H), 9.63 (s, 1H). MS (ES) C16H22N6O3S requires: 378. found: 379 (M+H)+.
Example 18
(S)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B18)
B18 was prepared following the procedure reported for B1 using A1 and (S)-2-(methoxymethyl)pyrrolidine. MS (ES) C16H22N6O3S requires: 378. found: 379 (M+H)+.
Example 19
3-[(4-((2R,5R)-Bis(methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B19)
B19 was prepared following the procedure reported for B4 using A1 and (2R,5R)-bis(methoxymethyl)pyrrolidine and obtained as a white crystalline solid; yield: 230 mg (60%). MS (ES) C18H26N6O4S requires: 422. found: 423 (M+H)+ and 445 (M+Na)+.
Example 20
3-[(4-(2S,5S)-Bis(methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B20)
B20 was prepared following the procedure reported for B4 using A1 and (2S,5S)-bis(methoxymethyl)pyrrolidine. It was obtained as a white crystalline solid; yield: 170 mg (45%). MS (ES) C18H26N6O4S requires: 422. found: 423 (M+H)+.
Example 21
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]pyrrolidine-2-carboxylate (B21)
B21 was obtained as a white crystalline solid by following the procedure reported for B4 using A1 and (R)-proline methyl ester hydrochloride; yield: 158 mg (45%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.88-2.08 (m, 3H), 2.22-2.42 (m, 1H), 3.50-3.77 (m, 2H), 3.55 and 3.60 (2 s, 3H), 4.22 (2 s, 2H), 4.50-4.67 (m, 1H), 6.88 (bs, 2H), 6.97-7.05 (m, 1H), 7.20-7.32 (m, 1H), 7.60-7.72 (m, 1H), 7.68 and 7.89 (2bs, 1H), 8.19 and 8.26 (2 s, 1H), 9.73 and 9.78 (2 s, 1H). MS (ES) C16H20N6O4S requires: 392. found: 393 (M+H)+.
Example 22
(S)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]pyrrolidine-2-carboxylate (B22)
B22 was obtained as a white crystalline solid by following the procedure reported for B4 using A1 and (S)-proline methyl ester hydrochloride; yield: 169 mg (48%). MS (ES) C16H20N6O4S requires: 392. found: 393 (M+H)+.
Example 23
(R)-1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]pyrrolidine-2-carboxamide (B23)
B23 was obtained as a white powder by following the procedure reported for B4 using A1 and (R)-proline amide; yield: 65 mg (19%). MS (ES) C15H19N7O3S requires: 377. found: 378 (M+H)+.
Example 24
(R)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B24)
B24 was prepared following the procedure reported for B1 using Al and (R)-2-(hydroxymethyl)pyrrolidine. MS (ES) C15H20N6O3S requires: 364. found: 365 (M+H)+.
Example 25
(S)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B25)
B25 was prepared following the general procedure reported for B1 using A1 and (S)-2-(hydroxymethyl)pyrrolidine. MS (ES) C15H20N6O3S requires: 364. found: 365 (M+H)+.
Example 26
rac-3-[(4-(2-Benzylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B26)
B26 was prepared following the procedure reported for B4 using A1 and rac-2-benzylpyrrolidine. It was obtained as a white crystalline solid; yield: 160 mg (42%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.67-1.92 (m, 4H), 2.57-2.77 (m, 1H), 3.03-3.21 (m, 1H), 3.42-3.62 (m, 2H), 4.20 (2 s, 2H), 4.28-4.48 (m, 1H), 6.88 (bs, 2H), 6.93-7.08 (m, 1H), 7.12-7.38 (m, 6H), 7.58 and 7.71 (2 m, 1H), 7.82-8.01 (m, 1H), 8.23 and 8.30 (2 s, 1H), 9.66 (bs, 1H). MS (ES) C21H24N6O2S requires: 424. found: 425 (M+H)+.
Example 27
rac-3-[(4-(2-(2-Phenylethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B27)
B27 was prepared following the procedure reported for B4 using A1 and rac-2-(2-phenylethyl)pyrrolidine and obtained as a white crystalline solid; yield: 140 mg (35%). MS (ES) C22H26N6O2S requires: 438. found: 439 (M+H)+.
Example 28
(R)-3-[(4-(2-(Phenoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B28)
B28 was prepared following the procedure reported for B4 using the intermediates A1 and A3. It was obtained as a white crystalline powder; yield: 140 mg (35%). MS (ES) C21H24N6O3S requires: 440. found: 441 (M+H)+.
Example 29
(R)-3-[(4-(2-(Phenylamino-methyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B29) 
B29 was prepared following the procedure reported for B4 using A1 and (R)-2-(phenylamino-methyl)pyrrolidine. It was obtained as a white crystalline solid; yield: 245 mg (62%). MS (ES) C21H25N7O2S requires: 439. found: 440 (M+H)+.
Example 30
(R)-3-[(4-(2-(Benzyloxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B30)
B30 was prepared following the procedure reported for B4 using A1 and A4 and obtained as a white powder; yield: 270 mg (66%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.82-2.08 (m, 4H), 3.42-3.78 (m, 4H), 4.20 (2 s, 2H), 4.24-4.39 (m, 1H), 4.39-4.58 (m, 2H), 6.85 (bs, 2H), 6.93-7.02 (m, 1H), 7.18-7.39 (m, 6H), 7.67-7.77 (m, 1H), 7.73 and 7.88 (2bs, 1H), 8.21 (2 s, 1H), 9.67 (s, 1H). MS (ES) C22H26N6O3S requires: 454. found: 455 (M+H)+.
Example 31
(R)-3-[(4-(2-((Pyrrolidin-1-yl)methyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B31)
B31 was prepared following the procedure reported for B4 using A1 and (R)-2-((pyrrolidin-1-yl)methyl)pyrrolidine. It was obtained as a white crystalline solid; yield: 65 mg (17%). MS (ES) C19H27N7O2S requires: 417. found: 418 (M+H)+.
Example 32
(S)-3-[(4-(2-((Pyrrolidin-1-yl)methyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B32)
B32 was prepared following the procedure reported for B4 using A1 and (S)-2-((pyrrolidin-1-yl)methyl)pyrrolidine and obtained as a white crystalline solid; yield: 87 mg (23%). MS (ES) C19H27N7O2S requires: 417. found: 418 (M+H)+.
Example 33
rac-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B33)
B33 was prepared following the procedure reported for B4 using A1 and rac-2-phenylpyrrolidine, and it was obtained as a white powder; yield: 90 mg (24%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.77-2.05 (m, 3H), 2.26-2.48 (m, 1H), 3.61-3.80 (m, 1H), 3.80-3.98 (m, 1H), 4.07 and 4.22 (2 s, 2H), 5.28 and 5.30 (2 s, 1H), 6.75-6.92 (m, 3H), 6.94-7.07 (m, 1H), 7.09-7.42 (m, 6H), 7.70 and 7.90 (2 s, 1H), 8.06 and 8.27 (2 s, 1H), 9.55 and 9.72 (2bs, 1H). MS (ES) C20H22N6O2S requires: 410. found: 411 (M+H)+.
Example 34
(R)-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B34)
B34 was prepared following the procedure reported for B4 using A1 and A5 and obtained as white crystals; yield: 170 mg (46%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 1.73-2.02 (m, 3H), 2.23-2.45 (m, 1H), 3.60-3.79 (m, 1H), 3.79-3.97 (m, 1H), 4.06 and 4.22 (2bs, 2H), 5.22-5.34 (m, 1H), 6.82 and 6.88 (2bs, 2H), 6.95-7.08 (m, 1H), 7.09-7.42 (m, 7H), 7.65-7.77 and 7.92 (m and bs, 1H), 8.05 and 8.26 (2 s, 1H), 9.57 and 9.72 (2bs, 1H). MS (ES) C20H22N6O2S requires: 410. found: 411 (M+H)+.
Example 35
(R)-3-[(4-(2-(Methoxymethyl)azetidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B35)
B35 was prepared following the general procedure reported for B4 using A1 and A6 and obtained as white crystals; yield: 85 mg (26%). MS (ES) C15H20N6O3S requires: 364. found: 365 (M+H)+.
Example 36
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]azetidine-2-carboxylate (B36)
B36 was prepared following the general procedure reported for B4 using A1 and A7 and obtained as white crystals; yield: 60 mg (18%). MS (ES) C15H18N16O4S requires: 378. found: 379 (M+H)+.
Example 37
rac-3-[(4-(2-Phenylazetidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B37)
B37 was prepared following the general procedure reported for B4 using A1 and rac-2-phenylazetidine and obtained as white crystals; yield: 120 mg (34%). 1H NMR (300 MHz, d6-DMSO, 300K) δ 2.05-2.20 (m, 1H), 2.69-2.90 (m, 1H), 3.88-4.29 (m, 4H), 5.42 (bs, 1H), 6.72-7.08 (m, 3H), 7.20-7.55 (m, 6H), 7.58-7.72 and 7.82-8.00 (2 m, 1H), 8.11 and 8.25 (2bs, 1H), 9.65 and 9.78 (2bs, 1H). MS (ES) C19H20N6O2S requires: 396. found: 397 (M+H)+.
Example 38
3-[(4-(7,8-Dihydro-1,6-naphthyridin-6(5H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B38)
B38 was prepared following the general procedure reported for B10 using A1 and 5,6,7,8-tetrahydro-1,6-naphthyridine hydrochloride. The title compound (B38) was purified by reverse phase RP-HPLC (column: C18), using H2O (0.1% TFA) and MeOH (0.1% TFA) as eluents; yield: 67.6 mg (17%), colorless amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 2.98 (bs, 2H), 4.12 (t, J=6.0 Hz, 2H), 4.26 (bs, 2H), 4.95 (s, 2H), 6.86 (bs, 2H), 7.02 (d, J=7.9 Hz, 1H), 7.25 (dd, J=7.8 Hz, J=4.7 Hz, 1H), 7.31 (t, J=7.9 Hz, 1H), 7.47-7.80 (bm, 2H), 7.83-8.10 (bm, 1H), 8.39 (dd, J=4.8 Hz, J=1.6 Hz, 1H), 8.30 (s, 1H), 9.77 (s, 1H). MS (ES) C18H19N7O2S requires: 397. found: 398 (M+H)+.
Example 39
3-[(4-(7,8-Dihydropyrido[4,3-d]pyrimidin-6(5H)-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B39)
B39 was prepared following the general procedure reported for B10 using A1 and 5,6,7,8-tetrahydropyrido[4,3-d]pyrimidine hydrochloride; yield: 29.7 mg (7%), pale yellow solid. MS (ES) C17H18N8O2S requires: 398. found: 399 (M+H)+.
Example 40
3-[(4-(3,4-Dihydroisoquinolin-2(1H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B40)
B40 was prepared following the general procedure reported for B10 using A1 and 1,2,3,4-tetrahydroisoquinoline; yield: 23.6 mg (6%), colorless amorphous solid. MS (ES) C19H20N6O2S requires: 396. found: 397 (M+H)+.
Example 41
3-[(4-(5-Amino-3,4-dihydroisoquinolin-2(1H)-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B41)
B41 was prepared following the general procedure reported for B10 using A1 and 5-amino-1,2,3,4-tetrahydroisoquinoline; yield: 61.5 mg (15%), pale yellow solid. MS (ES) C19H21N7O2S requires: 411. found: 412 (M+H)+.
Example 42
3-[(4-(3,4-Dihydroquinolin-1(2H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B42)
B42 was prepared following the general procedure reported for B10 using A1 and 1,2,3,4-tetrahydroquinoline; yield: 11.9 mg (3%), colorless amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 1.91 (quint, J=6.3 Hz, 2H), 2.73 (t, J=6.6 Hz, 2H), 3.98 (t, J=6.2 Hz, 2H), 4.16 (s, 2H), 6.82 (s, 2H), 6.99 (d, J=7.6 Hz, 1H), 7.05 (t, J=7.2 Hz, 1H), 7.13-7.19 (m, 2H), 7.23 (t, J=7.8 Hz, 1H), 7.62-7.69 (m, 2H), 7.72 (d, J=7.8 Hz, 1H), 8.32 (s, 1H), 9.81 (s, 1H). MS (ES) C19H20N6O2S requires: 396. found: 397 (M+H)+.
Example 43
3-[(4-(6,7-Dihydrothieno[3,2-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B43)
B43 was prepared following the general procedure reported for B10 using A1 and 4,5,6,7-tetrahydrothieno[3,2-c]pyridine; yield: 88.6 mg (22%), white amorphous solid. MS (ES) C17H18N6O2S2 requires: 402. found: 403 (M+H)+.
Example 44
3-[(4-(6,7-Dihydro-3H-imidazo[4,5-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B44)
B44 was prepared following the general procedure reported for B10 using A1 and 4,5,6,7-tetrahydro-3H-imidazo[4,5-c]pyridine. The title compound (B44) was purified by reverse phase RP-HPLC (column: C18), using H2O and MeOH as eluents; yield: 34.3 mg (9%), white amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 2.59-2.71 (m, 2H), 4.07 (t, J=5.7 Hz, 2H), 4.22 (s, 2H), 4.72 (s, 2H), 6.83 and 6.89 (2 s, 2H), 7.00 (d, J=7.5 Hz, 1H), 7.29 (t, J=7.9 Hz, 1H), 7.51 (s, 1H), 7.58 and 7.65 (2 d, J=7.0 Hz and J=7.5 Hz, 1H), 7.74 and 7.88 (2 s, 1H), 8.26 (s, 1H), 9.70 (s, 1H), 11.86 (bs, 1H). MS (ES) C16H18N8O2S requires: 386. found: 387 (M+H)+.
Example 45
3-[(4-(6,7-Dihydrothiazolo[5,4-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B45)
B45 was prepared following the general procedure reported for B10 using A1 and 4,5,6,7-tetrahydrothiazolo[5,4-c]pyridine hydrochloride. The title compound (B45) was purified by column chromatography (silica gel, DCM/MeOH gradient 100:0 to 85:15); yield: 28.9 mg (7%), colorless amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 2.84-2.94 (m, 2H), 4.13 (t, J=5.8 Hz, 2H), 4.23 (s, 2H), 5.03 (s, 2H), 6.83 (s, 2H), 7.00 (d, J=7.1 Hz, 1H), 7.29 (t, J=7.7 Hz, 1H), 7.42-7.63 (bm, 1H), 7.82-7.93 (bm, 1H), 8.28 (s, 1H), 8.96 (s, 1H), 9.77 (s, 1H). MS (ES) C16H17N7O2S2 requires: 403. found: 404 (M+H)+.
Example 46
3-[(4-(5,6-Dihydroimidazo[1,2-a]pyrazin-7(8H)-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B46)
B46 was prepared following the procedure reported for B1 using Al and 5,6,7,8-tetrahydroimidazo[1,2-a]pyrazine. MS (ES) C16H18N8O2S requires: 386. found: 387 (M+H)+.
Example 47
3-[(4-(6,7-Dihydropyrazolo[1,5-a]pyrimidin-4(5H)-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide (B47)
B47 was prepared following the procedure reported for B1 using Al and 4,5,6,7-tetrahydropyrazolo[1,5-a]pyrimidine. 1H NMR (400 MHz, d6-DMSO, 300K) δ 2.11-2.22 (m, 2H), 4.10-4.20 (m, 4H), 4.22 (s, 2H), 6.85 (bs, 3H), 7.06 (d, J=8.2 Hz, 1H), 7.30 (d, J=8.3 Hz, 1H), 7.34 (s, 1H), 7.54-7.82 (bm, 2H), 8.47 (s, 1H), 10.00 (bs, 1H). MS (ES) C16H18N8O2S requires: 386. found: 387 (M+H)+.
Example 48
3-[(4-(5H-Pyrrolo[3,4-b]pyridin-6(7H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B48)
B48 was prepared following the general procedure reported for B10 using A1 and 6,7-dihydro-5H-pyrrolo[3,4-b]pyridine hydrochloride; yield: 92.2 mg (24%), dark amorphous solid. MS (ES) C17H17N7O2S requires: 383. found: 384 (M+H)+.
Example 49
3-[(4-(1H-Pyrrolo[3,4-c]pyridin-2(3H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B49)
B49 was prepared following the general procedure reported for B10 using A1 and 2,3-dihydro-1H-pyrrolo[3,4-c]pyridine hydrochloride; yield: 139.0 mg (36%), brown amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 4.27 (s, 2H), 4.92 (s, 2H), 4.97 and 4.99 (2 s, 2H), 6.87 and 6.88 (2 s, 2H), 7.01 (d, J=7.5 Hz, 1H), 7.30 (t, J=7.9 Hz, 1H), 7.59-7.72 (m, 2H), 7.98 (bs, 1H), 8.31 (d, J=1.4 Hz, 1H), 8.62 (t, J=6.0 Hz, 1H), 8.76 (d, J=5.7 Hz, 1H), 9.82 (s, 1H). MS (ES) C17H17N7O2S requires: 383. found: 384 (M+H)+.
Example 50
3-[(4-(5H-Pyrrolo[3,4-d]pyrimidin-6(7H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B50)
B50 was prepared following the general procedure reported for B10 using A1 and 6,7-dihydro-5H-pyrrolo[3,4-d]pyrim idine hydrochloride; yield: 205.4 mg (53%), colorless amorphous solid. MS (ES) C16H16N8O2S requires: 384. found: 385 (M+H)+.
Example 51
3-[(4-(Pyrrolo[3,4-c]pyrazol-5(1H,4H,6H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B51)
B51 was prepared following the general procedure reported for B10 using A1 and 1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole dihydrochloride; yield: 194.9 mg (52%), colorless amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 4.23 (s, 2H), 4.58 (s, 2H), 4.64 (s, 2H), 6.83 (s, 2H), 7.00 (d, J=7.7 Hz, 1H), 7.26-7.34 (m, 1H), 7.58 (s, 1H), 7.71-7.78 (m, 1H), 7.86 (s, 1H), 8.27 (s, 1H), 9.73 (s, 1H), 12.73 (s, 1H). MS (ES) C15H16N8O2S requires: 372. found: 373 (M+H)+.
Example 52
3-[(4-(Indolin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethane sulfonamide (B52)
B52 was prepared following the general procedure reported for B10 using A1 and indoline; yield: 76.2 mg (20%), pale yellow solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 3.11-3.18 (m, 2H), 4.14-4.26 (m, 2H), 4.23 (s, 2H), 6.84 (s, 2H), 6.97 (t, J=7.1 Hz, 1H), 7.01-7.09 (bm, 1H), 7.13-7.21 (bm, 1H), 7.25 (d, J=7.0 Hz, 1H), 7.33 (t, J=7.7 Hz, 1H), 7.64-7.90 (bm, 2H), 8.29-8.41 (bm, 1H), 8.43 (s, 1H), 9.90 (s, 1H). MS (ES) C18H18N6O2S requires: 382. found: 383 (M+H)+.
Example 53
(S)-3-[(4-(2-Methylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B53)
B53 was prepared following the general procedure reported for B10 using A1 and (S)-2-methylpyrrolidine; yield: 195.5 mg (56%), colorless amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 1.17 and 1.24 (2 d, J=6.3 Hz, 3H), 1.60-1.70 (m, 1H), 1.82-2.08 (m, 3H), 3.38-3.64 (m, 2H), 4.12-4.29 (m, 1H), 4.18 (s, 2H), 6.82 (2 s, 2H), 6.94-7.00 (m, 1H), 7.25 (t, J=7.9 Hz, 1H), 7.68 (d, J=9.0 Hz, 1H), 7.83-7.90 (m, 1H), 8.18 (s, 1H), 9.58 (s, 1H). MS (ES) C15H20N6O2S requires: 348. found: 349 (M+H)+.
Example 54
(R)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide (B54)
B54 was prepared following the general procedure reported for B10 using A1 and (S)-2-(trifluoromethyl)pyrrolidine; yield: 41.3 mg (10%), colorless amorphous solid. 1H NMR (400 MHz, d6-DMSO, 300K) δ 1.95-2.22 (m, 4H), 3.50-3.78 (m, 2H), 4.20 (2bs, 2H), 4.95-5.07 (m, 1H), 6.81 (s, 2H), 7.00 (d, J=7.4 Hz, 1H), 7.27 (t, J=7.8 Hz, 1H), 7.65 (d, J=8.7 Hz, 1H), 7.76-7.91 (m, 1H), 8.31 (s, 1H), 9.85 (s, 1H). MS (ES) C15H17F3N6O2S requires: 402. found: 403 (M+H)+.
Materials and Methods:
1. Measurement of Binding Affinities to CDKs
This protocol describes how the LanthaScreen Eu Kinase Binding Assay was performed to determine dissociation constants (Kd) of compounds of general formula (I) and CDK/Cyclin complexes. The principle behind this assay is based upon the binding and displacement of an Alexa Fluor 647-labeled tracer, which binds to the active site of kinases. Binding of the tracer to the kinase is detected using a Eu-labeled antibody. Simultaneous binding of both the tracer and antibody to the kinase gives rise to a FRET-signal. Binding of an inhibitor to the kinase competes for binding with the tracer, resulting in a loss of FRET.







TABLE 2







Reagents, stock concentrations and final assay concentrations

















Kinase-


Tracer-


Antibody-


Kinase
Supplier
conc. [nM]
Tracer
Supplier
conc. [nM]
Antibody
Supplier
conc.


















CDK2/Cyclin A
Proqinase
5
236
Invitrogen
20
Eu-Anti-GST
Cisbio
1:750


(135 kDa)


CDK7/CyclinH/MAT1
Carna
5
236
Invitrogen
60
Eu-Anti-His
Invitrogen
2 nM


(126 kDa)
Biosciences


CDK8/Cyclin C
Invitrogen
5
236
Invitrogen
20
Eu-Anti-His
Invitrogen
2 nM


(97 kDa)


CDK9/Cyclin T1
Invitrogen
5
236
Invitrogen
30
Eu-Anti-His
Cisbio
1:250


(132 kDa


CDK9/Cyclin K
Invitrogen
10
236
Invitrogen
35
Eu-Anti-His
Invitrogen
4 nM


(92 kDa)









The compounds of general formula (I) summarized in Table 1 were diluted from a 10 mM DMSO stock solution 1:10 in a total volume of 15 μL DMSO. This compound predilution was then serial diluted 1:3 over 8 steps in DMSO and briefly spun down. Each compound solution was now diluted 1:33.33 in kinase buffer (HEPES: 20 mM, pH: 8.0; MgCl2: 10 mM; DTT: 1 mM; Brij-35: 0,01%), mixed thoroughly and spun down. For every sample, 5 μL of the diluted compound were mixed with 5 μL tracer working solution (e.g. 60 nM tracer 236 in kinase buffer for CDK2/Cyclin A) and 5 μL CDK/Cyclin/Anti-GST-AB-working solution (e.g. 15 nM CDK2/Cyclin A, 1:250 dilution of Anti-GST-AB in kinase buffer) in a well of a small volume 384 well plate (Corning Incorporated, Corning, N.Y., USA; order no. 3673). The tracer concentration was adjusted to its dissociation constant (Kd) for the CDK/Cyclin, which was 30 nM for CDK2/Cyclin A, CDK7/Cyclin H and CDK9/Cyclin T1, 20 nM for CDK8/Cyclin C, and 35 nM for CDK9/Cyclin K. For negative controls, in each well 5 μL of DMSO working solution (3% DMSO diluted in kinase buffer) was mixed with 5 μL Anti-GST-AB working solution (e.g. 1:250 dilution of Anti-GST-AB in kinase buffer for CDK2/Cyclin A) and 5 μL Tracer working solution (e.g. 60 nM Tracer 236 in kinase buffer for CDK2/Cyclin A). For positive controls, in each well 5 μL of DMSO working solution (3% DMSO diluted in kinase buffer) was mixed with 5 μL CDK/Cyclin/Anti-GST-AB-working solution: (e.g. 15 nM CDK2/Cyclin A, 1:250 dilution of Anti-GST-AB in kinase buffer) and 5 μL Tracer working solution (e.g. 60 nM Tracer 236 in kinase buffer for CDK2/Cyclin A). Positive and negative controls were calculated from at least 8 different sample wells. The 384 well plates were mixed in a Teleshaker plate mixer (Beckman Coulter, Brea, Calif., USA) at 2000 rpm for 40 sec, and incubated for 1 h at room temperature before reading. The FRET signal was measured at 340 nm excitation, 665 nm and 615 nm emission (for the kinase tracer and LanthaScreen Eu-AB, respectively) with an Envision spectrophotometer (Perkin Elmer, Waltham, Mass., USA) with 50 ps delay and 300 ps integration time. Kd values were determined from the sigmoidal dose response curves with the software Quattro Workflow (Quattro GmbH, Munich, Germany). Results are presented in Table 4.
2. Measurement of half maximal inhibitory concentration to CDKs This protocol describes how the Lance Ultra KinaSelect Assay was performed to determine half maximal inhibitory concentration (1050) of compounds of general formula (I) and CDK/Cyclin complexes. The principle behind this enzymatic assay is based upon the phosphorylation of the ULight-Peptide Substrat. It is detected by using a specific
EU-labeled anti-phospho peptide antibody. The binding of the Eu-labeled anti-phospho peptide antibody to the phosphorylated ULight labeled peptide gives rise to a FRET-signal. Binding of an inhibitor to the kinase prevents phosphorylation of the ULight-MBP Substrat, resulting in a loss of FRET.







TABLE 3







Reagents, stock concentrations and final assay concentrations


















Kinase-
ATP-


Substrat-


Antibody-


Kinase
Supplier
conc. [nM]
conc. [μM]
Substrat
Supplier
conc. [nM]
Antibody
Supplier
conc. [nM]



















CDK1/CyclinB1
Carna
2
20
Ulight MBP
Perkin Elmer
50
Eu-anti-P-MBP
Perkin Elmer
0.25


(91 kDa)


CDK2/CyclinA
Proqinase
5
3
Ulight MBP
Perkin Elmer
50
Eu-anti-P-MBP
Perkin Elmer
0.25


[135 kDa)


CDK4/CyclinD1
Invitrogen
10
90
Ulight MBP
Perkin Elmer
50
Eu-anti-P-MBP
Perkin Elmer
0.25


(123 kDa)


CDK6/CyclinD3
Carna
5
55
Ulight MBP
Perkin Elmer
50
Eu-anti-P-MBP
Perkin Elmer
0.025


(123 kDa)


CDK7/CyclinH/
Invitrogen
10
25
Ulight MBP
Perkin Elmer
50
Eu-anti-P-MBP
Perkin Elmer
0.25


MAT1 (126 kDa)


CDK9/CyclinT1
Invitrogen
10
25
Ulight MBP
Perkin Elmer
50
Eu-anti-P-MBP
Perkin Elmer
0.25


(132 kDa)


CDK9/CyclinK
Invitrogen
10
125
Ulight MBP
Perkin Elmer
50
Eu-anti-P-MBP
Perkin Elmer
0.25


(92 kDa)









The compounds of general formula (I) summarized in Table 5 were diluted from a 10 mM DMSO stock solution 1:10 in a total volume of 15 μL DMSO. This compound predilution was then serial diluted 1:3 over 8 steps in DMSO and briefly spun down. Each compound solution was now diluted 1:20 in Enzymatic Buffer (HEPES: 50 mM, pH: 7.5; MgCl2: 10 mM; EGTA: 1 mM; DTT: 2 mM; Tween-20: 0.01%), mixed thoroughly and spun down. For every sample, 2 μL of the diluted compound were mixed with 6 μL CDK/Cyclin/Substrat solution and 2 μL ATP solution in a well of a small volume 384 well plate (Corning Incorporated, Corning, N.Y., USA; order no. 3673). The CDK/Cyclin was diluted to the appropriate concentration (see Table 3) and the ATP concentration was adjusted to its IC50 concentration for the CDK/Cyclin, which was 3 μM for CDK2/Cyclin A, 20 μM for CDK1/Cyclin B1, 25 μM for CDK7/Cyclin H and CDK9/Cyclin T1, 55 μM for CDK6/Cyclin D3, 90 μM for CDK4/Cyclin D1 and 125 μM for CDK9/Cyclin K. For negative controls, in each well 2 μL of DMSO solution (1% final DMSO assay concentration) was mixed with 6 μL substrate solution (50 nM ULight MBP final assay concentration) and 2 μL ATP solution (appropriate final concentration see Table 3). For positive controls, in each well 2 μL of DMSO solution (1% final DMSO assay concentration) was mixed with 6 μL CDK/Cyclin/Substrat (appropriate final concentration see Table 3) and 2 μL Tracer ATP solution (appropriate final concentration see Table 3). Positive and negative controls were calculated from at least 8 different sample wells. The 384 well plates were mixed in a Teleshaker plate mixer (Beckman Coulter, Brea, Calif., USA) at 2000 rpm for 40 sec, and incubated for 1 h at room temperature. Before reading, 10 μL the detection buffer (Lance Detection Buffer 1×; EDTA: 20 nM; Eu-Anti-P-MBP: see Table 3) was added. The FRET signal was measured at 340 nm excitation, 665 nm and 615 nm emission (for the kinase tracer and LanthaScreen Eu-AB, respectively) with an Envision spectrophotometer (Perkin Elmer, Waltham, Mass., USA) with 50 μs delay and 300 μs integration time. IC50 values were determined from the sigmoidal dose response curves with the software Quattro Workflow (Quattro GmbH, Munich, Germany). Results are presented in Table 5.
3. Cellular Assays
3.1 RNA-Polymerase II Ser2 Cellular Phosphorylation Assay:
HCT-116 cells (DSMZ, Braunschweig, Germany) were maintained in Mc Coy's cell culture medium+glutamine (PAN Biotech GmbH, Aidenbach, Germany) supplemented with 10% fetal calf serum (PAA Laboratories GmbH, Pasching, Austria) and grown at 37° C., 5% CO2. For the cellular phosphorylation assay, cells were seeded with 2×105 cells/well/1 ml in 24-well plates (Greiner Bio One, Frickenhausen, Germany; catalog #662160). The compounds of general formula (I) summarized in Table 6 were diluted from a 10 mM DMSO stock solution 1:10 in a total volume of 15 μL DMSO. After overnight incubation at 37° C./5% CO2, 1.5 μL of a compound diluted in DMSO was added to each sample well. Wells with cells and 0.15% DMSO in culture medium were used as positive controls, wells without cells and 0.15% DMSO in culture medium were used as negative controls. The cells were incubated with the compounds for 72 h at 37° C./5% CO2. Before lysis, cells were washed with phosphate buffered saline. Phosphorylation of RNA Polymerase II Ser2 and tubulin levels for normalization were analyzed afterwards with the Multi-Array technology (Meso Scale Discovery, Gaithersburg, Md., USA), a combination of antibody coupled electrochemiluminescence detection and patterned arrays. Manufacturer's instructions were followed and all solutions were purchased from Meso Scale Discovery. In brief, cells were lysed by 30 min incubation in CLB1 lysis buffer (Zeptosens, Witterswil, Switzerland; 60 μL per well), and supernatants were cleared by centrifugation. For analysis of RNA Polymerase II Ser2-phosphorylation, lysates were diluted 1:50 with Meso Scale Lysis Buffer supplemented with phosphatase- and protease-inhibitors, and 25 μL of each sample was pipetted in a well of a MSD Multi-Array 96-Well Plate Sector® Imager High Bind Plate (Meso Scale Discovery; catalog #L15XB-3/L11XB-3), and incubated for 2 h at room temperature. 150 μL Meso Scale Tris Wash Buffer supplemented with 3% w/v Meso Scale Blocker A were added per well, then plates were sealed and incubated 1 h with vigorous shaking. Plates were washed with 1× Tris Wash Buffer (10× Meso Scale Wash Buffer diluted 1:10 in destilled water), 25 μL of antibody solution was added (CTD7 3E10 antibody from Helmholtz Zentrum Munich, Germany, diluted 1:100 in Meso Scale Tris Wash Buffer supplemented with 1% w/v Meso Scale Blocker A), and plates were washed three times in 1× Tris Wash Buffer. 25 μL of MSD® SULFO-TAGTM Goat—Anti—Rat—Antibody (Meso Scale Discovery, atalog # R32AH-1, diluted 1:125 in Tris Wash Buffer with 1% (w/v) blocker A) were added per well, plates were sealed and incubated with vigorous shaking for 1 h at room temperature. Finally, plates were washed three times with Tris Wash Buffer, 150 μl 2× Read Buffer (Meso Scale Discovery) were added per well and plates were analyzed immediately in a Sector Imager from Meso Scale Discovery. For determination of tubulin protein levels, samples were analyzed with the protocol for RNA Polymerase II Ser2-phosphorylation, with an anti-tubulin antibody (rabbit; BIODESIGN International, catalog #T59840R, diluted 1:100) and a MSD® SULFO-TAGTM Goat—Anti—Rat—Antibody (Meso Scale Discovery, catalog #R32AH-1, diluted 1:125). RNA Polymerase II Ser2 phosphorylation was normalized with tubulin protein levels, and IC50 values were calculated with the software XLFit (IDBS, Guildford, UK) from 2-fold dilution series comprising 6 concentrations in duplicates. Results are presented in Table 6.
3.2 NF-kappaB Reporter Assay
Cells were maintained in RPMI cell culture medium+glutamine (PAN Biotech GmbH, Aidenbach, Germany) supplemented with 10% fetal calf serum (PAA Laboratories GmbH, Pasching, Austria) and grown at 37° C., 5% CO2. HEK293 cells grown to 50% confluence were transfected with the Amaxa® Cell Line Nucleofector® Kit V (Lonza, Basel, Switzerland, catalog #VCA-1003). Transfections were performed according to manufacturer's optimized protocol for transfection of HEK293 cells. In brief, 2×105 cells were transfected with 5 μg highly purified plasmid DNA. Cells were transfected with a NF-kappa B reporter plasmid (pNFkBluc), pTALluc for control, or pMAXGFP for transfection control. After transfection, cells were taken up in 500 μL RPMI1640 cell culture medium, incubated for 1 h at 37° C., and 4.5 ml DMEM without phenol red were added per transfection. Transfected cells were seeded in 96 well plates (Greiner Bio-One, Frickenhausen, Germany, catalog #655098) with 100 μL cell suspension per well and incubated for 48 h. To each well, 100 μL DMEM with 2× concentrated compound diluted from 10 mM DMSO stocks, or 100 μL DMEM with 0.4% DMSO for control wells, was added. The compounds of general formula (I) summarized in Table 6 were used in this assay. Cells were stimulated with 20 ng/ml TNF alpha, and plates were incubated for 5 h at 37° C./5% CO2. Cell culture supernatants were removed to leave 100 μl medium per well, followed by addition of 100 μl Bright Glo luciferase assay reagent (Promega, Madison, Wis., USA, catalog #E2620), and shaking for 5 minutes in the dark. Luminescence was measured with a Victor Photospectrometer (Perkin Elmer, Waltham, Mass., USA). IC50 values were calculated with the software Excel Fit (IDBS, Guildford, UK) from 2-fold dilution series comprising at least 10 concentrations in duplicates. Results are presented in Table 6.
3.3 TNF alpha Release Assay
Freshly isolated peripheral blood mononuclear cells (PBMCs) were seeded in 96-well cell culture plates with 200,000 cells in 100 μl cell culture medium (DMEM cell culture medium+glutamine from PAN Biotech GmbH, Aidenbach, Germany) supplemented with 10% fetal calf serum (PAA Laboratories GmbH, Pasching, Austria) per well and incubated overnight at 37° C., 5% CO2. To each well, 100 μL cell culture medium with 2× concentrated test compounds diluted from 10 mM DMSO stocks, or 100 μL DMEM with 0.4% DMSO for control wells, was added. The compounds of general formula (I) summarized in Table 6 were used in this assay. After incubation for 1 h at 37° C., 5% CO2, cells were stimulated with 1 μg/mL LPS (Lipopolysaccharides, Sigma, catalog #L4391-1 MG; 1 mg/ml stock solution), or left untreated for negative controls, and plates were incubated for 6 h at 37° C./5% CO2. The cell culture plates were centrifuged at 2000 rpm for 5 minutes, and supernatants were transferred to fresh 96-well polypropylene plates. 25 μL of supernatants were transferred into 96-well-plates of the human TNF alpha-tissue culture kit (Meso Scale Discovery, Gaithersburg, Md., USA), and manufacturer's instructions were followed for analysis of TNF alpha levels. Chemoluminescence was measured in the Mesoscale Sector Imager, and IC50 values were calculated with the software Excel Fit (IDBS, Guildford, UK) from 2-fold dilution series comprising at least 6 concentrations in duplicates. Results are presented in Table 6.
3.4 Cell Viability Assays
Hela- or MDAMB468-Cells were maintained in RPMI 1640 or McCoy's 5A cell culture medium+glutamine (PAN Biotech GmbH, Aidenbach, Germany; order no. PO4-22100; PO4-05500) supplemented with 10% fetal calf serum “Gold” (PAA Laboratories GmbH, Pasching, Austria; order no. A15-151) and grown at 37° C., 5% CO2. For the cell viability assay, cells were seeded with a density of 400 (Hela cells, DSMZ Braunschweig order no. ACC57) or 800 (MDAMB468 cells, ATCC order no. HTB-132) per well in 25 μL in 384-well plates (Greiner Bio-One, Frickenhausen, Germany; order no. 781080). After overnight incubation at 37° C./5% CO2, 25 nL or 75 nL compound were added to each sample well by using BIOMEK FXP Laboratory Automation Workstation (Beckman Coulter, USA). Wells with cells and 0.1% or 0,3% DMSO in culture medium were used as positive controls, wells with cells and 10 μM staurosporine in culture medium were used as negative controls. The cells were incubated with the compounds for 72 h at 37° C./5% CO2. For measurement of cell viability 25 μL Cell Titer Glo reagent (Promega, Madison, USA; order no. G7573), 1:2 diluted with cell culture medium, was added to each well. The 384we11-plates were placed for 2 min on a orbital microplate shaker and incubated for further 10 min at room temperature to stabilize the luminescence signal.
Luminescence was measured by Envision Plate Reader (Perkin Elmer, USA). IC50 values were calculated with the software Excel Fit (IDBS, Guildford, UK) from 3-fold dilution series comprising at least 8 concentrations in duplicates. Results are presented in Table 6.
Results:
1. Measurement of Binding Affinities to CDKs
The dissociation constants Kd of the compounds according to the present invention for binding to CDK9, CDK7, and CDK2, respectively, are summarized in Table 4. Comparison of binding constants of a special compound of form lua (I) for a number of different CDKs shows that binding of a compound to CKD9 is always stronger than binding to other CDKs. Thus, a compound of form lua (I) binds or interacts specificially with CKD9 and at least selectively with CDK9.







TABLE 4







Affinity for CDK9, CDK7, and CDK2 of compounds according to the present invention












{circle around (1)}
Nomenclature
{circle around (2)}
{circle around (3)}
{circle around (4)}
{circle around (5)}





B1
3-[(4-(Piperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
d



benzenemethanesulfonamide


B2
3-[(4-(4,4-Difluoropiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
n.t.



benzenemethanesulfonamide


B3
3-[(4-(3,3-Difluoropiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
n.t.



benzenemethanesulfonamide


B4
rac-3-[(4-(2-Methoxymethylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
n.t.



benzenemethanesulfonamide


B5
(R)-3-[(4-(2-Methoxymethylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
d



benzenemethanesulfonamide


B6
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]-
a
b
d
n.t.



piperidine-2-carboxylate


B7
rac-tert-Butyl [(1-(4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-
d
d
d
n.t.



2-yl)piperidine-2-yl)methyl]carbamate


B8
rac-3-[(4-(2-(2-(Dimethylamino)ethyl)piperidin-1-yl)-1,3,5-triazin-2-
c
c
n.t.
n.t.



yl)amino]benzenemethanesulfonamide


B9
rac-3-[(4-(2-Phenylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
n.t.



benzenemethanesulfonamide


B10
3-[(4-(Morpholin-4-yl)-1,3,5-triazin-2-yl)amino]-
b
b
c
c



benzenemethanesulfonamide


B11
3-[(4-(Piperazin-1-yl)-1,3,5-triazin-2-yl)amino]-
d
d
d
n.t.



benzenemethanesulfonamide


B12
3-[(4-(4-Methylpiperazin-1-yl)-1,3,5-triazin-2-yl)amino]-
d
d
d
n.t.



benzenemethanesulfonamide


B13
rac-3-[(4-(2-(Hydroxymethyl)piperazin-1-yl)-1,3,5-triazin-2-yl)amino]-
d
d
n.t.
n.t.



benzenemethanesulfonamide


B14
3-[(4-(Pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
c
d
n.t.



benzenemethanesulfonamide


B15
(R)-3-[(4-(2-Methylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
c
d
n.t.



benzenemethanesulfonamide


B16
(S)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
c
d
n.t.



benzenemethanesulfonamide


B17
(R)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
a
b
d
c



benzenemethanesulfonamide


B18
(S)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
c
d
d
d



benzenemethanesulfonamide


B19
3-[(4-((2R,5R)-Bis(methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
d
d
d
n.t.



yl)amino]benzenemethanesulfonamide


B20
3-[(4-((2S,5S)-Bis(methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
d
d
d
n.t.



yl)amino]benzenemethanesulfonamide


B21
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]-
a
b
d
c



pyrrolidine-2-carboxylate


B22
(S)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]-
d
d
d
n.t.



pyrrolidine-2-carboxylate


B23
(R)-1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]-
d
d
d
n.t.



pyrrolidine-2-carboxamide


B24
(R)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
c
c
d
d



benzenemethanesulfonamide


B25
(S)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
c
c
d
d



benzenemethanesulfonamide


B26
rac-3-[(4-(2-Benzylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
n.t.



benzenemethanesulfonamide


B27
rac-3-[(4-(2-(2-Phenylethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
c
c
d
n.t.



benzenemethanesulfonamide


B28
(R)-3-[(4-(2-(Phenoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
c
c
d
n.t.



benzenemethanesulfonamide


B29
(R)-3-[(4-(2-(Phenylamino-methyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
c
c
d
n.t.



yl)amino]benzenemethanesulfonamide


B30
(R)-3-[(4-(2-(Benzyloxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
d



benzenemethanesulfonamide


B31
(R)-3-[(4-(2-((Pyrrolidin-1-yl)methyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
c
d
d
d



yl)amino]benzenemethanesulfonamide


B32
(S)-3-[(4-(2-((Pyrrolidin-1-yl)methyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
d
d
d
n.t.



yl)amino]benzenemethanesulfonamide


B33
rac-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
a
a
d
n.t.



benzenemethanesulfonamide


B34
(R)-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
a
a
d
c



benzenemethanesulfonamide


B35
(R)-3-[(4-(2-(Methoxymethyl)azetidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
c
d
d



benzenemethanesulfonamide


B36
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]-
c
c
d
n.t.



azetidine-2-carboxylate


B37
rac-3-[(4-(2-Phenylazetidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
b
d
n.t.



benzenemethanesulfonamide


B38
3-[(4-(7,8-Dihydro-1,6-naphthyridin-6(5H)-yl)-1,3,5-triazin-2-yl)amino]-
a
a
c
b



benzenemethanesulfonamide


B39
3-[(4-(7,8-Dihydropyrido[4,3-d]pyrimidin-6(5H)-yl)-1,3,5-triazin-2-
b
c
d
n.t.



yl)amino]benzenemethanesulfonamide


B40
3-[(4-(3,4-Dihydroisoquinolin-2(1H)-yl)-1,3,5-triazin-2-yl)amino]-
c
c
d
n.t.



benzenemethanesulfonamide


B41
3-[(4-(5-Amino-3,4-dihydroisoquinolin-2(1H)-yl)-1,3,5-triazin-2-
c
c
d
n.t.



yl)amino]benzenemethanesulfonamide


B42
3-[(4-(3,4-Dihydroquinolin-1(2H)-yl)-1,3,5-triazin-2-yl)amino]-
a
a
d
b



benzenemethanesulfonamide


B43
3-[(4-(6,7-Dihydrothieno[3,2-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-
b
b
c
n.t.



yl)amino]benzenemethanesulfonamide


B44
3-[(4-(6,7-Dihydro-3H-imidazo[4,5-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-
a
a
c
a



yl)amino]benzenemethanesulfonamide


B45
3-[(4-(6,7-Dihydrothiazolo[5,4-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-
a
n.t.
c
a



yl)amino]benzenemethanesulfonamide


B46
3-[(4-(5,6-Dihydroimidazo[1,2-a]pyrazin-7(8H)-yl)-1,3,5-triazin-2-
c
c
d
n.t.



yl)amino]benzenemethanesulfonamide


B47
3-[(4-(6,7-Dihydropyrazolo[1,5-a]pyrimidin-4(5H)-yl)-1,3,5-triazin-2-
a
a
b
a



yl)amino]benzenemethanesulfonamide


B48
3-[(4-(5H-Pyrrolo[3,4-b]pyridin-6(7H)-yl)-1,3,5-triazin-2-yl)amino]-
c
b
d
n.t.



benzenemethanesulfonamide


B49
3-[(4-(1H-Pyrrolo[3,4-c]pyridin-2(3H)-yl)-1,3,5-triazin-2-yl)amino]-
a
a
d
b



benzenemethanesulfonamide


B50
3-[(4-(5H-Pyrrolo[3,4-d]pyrimidin-6(7H)-yl)-1,3,5-triazin-2-yl)amino]-
b
c
d
n.t.



benzenemethanesulfonamide


B51
3-[(4-(Pyrrolo[3,4-c]pyrazol-5(1H,4H,6H)-yl)-1,3,5-triazin-2-yl)amino]-
a
a
c
c



benzenemethanesulfonamide


B52
3-[(4-(lndolin-1-yl)-1,3,5-triazin-2-yl)amino]-
a
a
d
b



benzenemethanesulfonamide


B53
(S)-3-[(4-(2-Methylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
a
n.t.
d
c



benzenemethanesulfonamide


B54
(R)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
n.t.
d
c



benzenemethanesulfonamide





Activity range “a” means, that the compounds do have a Kd (dissociation constant) < 100 nM, activity range “b” means, that the compounds do have a Kd between 100 and 1000 nM, activity range “c” means that the compounds do have an Kd between 1000 and 10000 nM, activity range “d” means that the compounds do have an Kd > 10000 nM; “n.t.” means that the compounds have not been tested in this assay.


{circle around (1)}: Compound Number


{circle around (2)}: CDK9/Cyclin T1 (activity range)


{circle around (3)}: CDK9/Cyclin K (activity range)


{circle around (4)}: CDK7 (activity range)


{circle around (5)}: CDK2 (activity range)






2. Measurement of Half Maximal Inhibitory Concentration to CDKs in Enzymatic Assays

The inhibitory activities of the compounds according to the present invention are shown in Table 5 as half-maximal inhibition constant (IC50) values for inhibition of CDK9, CDK1, CDK2, CDK4, CDK6, and CDK7, respectively.







TABLE 5







Inhibition for CDK9, CDK1, CDK2, CDK4, CDK6, and CDK7 of compounds


according to the present invention














{circle around (1)}
Nomenclature
{circle around (2)}
{circle around (3)}
{circle around (4)}
{circle around (5)}
{circle around (6)}
{circle around (7)}





B1
3-[(4-(Piperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
c
d
d
d
d
d



benzenemethanesulfonamide


B8
rac-3-[(4-(2-(2-(Dimethylamino)ethyl)piperidin-1-yl)-1,3,5-
c
d
d
d
d
d



triazin-2-yl)amino]benzenemethanesulfonamide


B17
(R)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
b
c
c
d
d
d



yl)amino]-benzenemethanesulfonamide


B21
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-
b
n.t.
n.t.
n.t.
n.t.
n.t.



triazin-2-yl]pyrrolidine-2-carboxylate


B33
rac-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
c
d
d
d
d



benzenemethanesulfonamide


B38
3-[(4-(7,8-Dihydro-1,6-naphthyridin-6(5H)-yl)-1,3,5-triazin-2-
a
b
b
d
d
d



yl)amino]-benzenemethanesulfonamide


B44
3-[(4-(6,7-Dihydro-3H-imidazo[4,5-c]pyridin-5(4H)-yl)-1,3,5-
a
n.t.
n.t.
n.t.
n.t.
n.t.



triazin-2-yl)amino]benzenemethanesulfonamide


B45
3-[(4-(6,7-dihydrothiazolo[5,4-c]pyridin-5(4H)-yl)-1,3,5-
a
n.t.
a
n.t.
n.t.
c



triazin-2-yl)amino]benzenemethanesulfonamide


B47
3-[(4-(6,7-Dihydropyrazolo[1,5-a]pyrimidin-4(5H)-yl)-1,3,5-
a
b
a
c
c
c



triazin-2-yl)amino]benzenemethanesulfonamide


B51
3-[(4-(Pyrrolo[3,4-c]pyrazol-5(1H,4H,6H)-yl)-1,3,5-triazin-2-
a
c
b
d
d
d



yl)amino]-benzenemethanesulfonamide


B53
(S)-3-[(4-(2-Methylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
b
n.t.
c
n.t.
n.t.
n.t.



benzenemethanesulfonamide


B54
(R)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
c
n.t.
c
n.t.
n.t.
n.t.



yl)amino]-benzenemethanesulfonamide





Activity range “a” means, that the compounds do have a IC50 (inhibitory concentration at 50% of maximal effect) <100 nM,


activity range “b” means, that the compounds do have a IC50 between 100 and 1000 nM,


activity range “c” means that the compounds do have an IC50 between 1000 and 10000 nM,


activity range “d” means that the compounds do have an IC50 >10000 nM;


“n.t.” means that the compounds have not been tested in this assay.


{circle around (1)}: Compound Number


{circle around (2)}: CDK9 LANCE assay (activity range)


{circle around (3)}: CDK1 LANCE assay (activity range)


{circle around (4)}: CDK2 LANCE assay (activity range)


{circle around (5)}: CDK4 LANCE assay (activity range)


{circle around (6)}: CDK6 LANCE assay (activity range)


{circle around (7)}: CDK7 LANCE assay (activity range)






3. Cellular Assays

The cellular activity of the compounds according to the present invention are shown in Table 6 as half-maximal inhibition constant (1050) values on LPS-induced TNF alpha release in PBMCs, NF-kappaB reporter gene activation, cellular CDK9 activity (RNA Polymerase II Ser2 phosphorylation), and cell viability in Hela- or MDAMB468-Cells, respectively.







TABLE 6







Inhibition of LPS-induced TNF alpha release in PBMCs, NF-kappaB reporter


gene activation, cellular CDK9 activity (RNA Polymerase II Ser2 phosphorylation), and


cell viability in Hela- or MDAMB468-Cells by compounds according to the present


invention.













{circle around (1)}
Nomenclature
{circle around (2)}
{circle around (3)}
{circle around (4)}
{circle around (5)}
{circle around (6)}





B1
3-[(4-(Piperidin-1-yl)-1,3,5-triazin-2-yl)amino]-
d
d
n.t.
n.t.
n.t.



benzenemethanesulfonamide


B17
(R)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
c
d
n.t.
n.t.
n.t.



yl)amino]-benzenemethanesulfonamide


B21
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-
n.t.
d
n.t.
n.t.
n.t.



2-yl]-pyrrolidine-2-carboxylate


B30
(R)-3-[(4-(2-(Benzyloxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-
n.t.
d
n.t.
n.t.
n.t.



yl)amino]-benzenemethanesulfonamide


B33
rac-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
n.t.
c
n.t.
n.t.
n.t.



benzenemethanesulfonamide


B34
(R)-3-[(4-(2-Phenylpyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-
n.t.
n.t.
n.t.
d
c



benzenemethanesulfonamide


B38
3-[(4-(7,8-Dihydro-1,6-naphthyridin-6(5H)-yl)-1,3,5-triazin-2-
n.t.
c
d
n.t.
n.t.



yl)amino]-benzenemethanesulfonamide


B44
3-[(4-(6,7-Dihydro-3H-imidazo[4,5-c]pyridin-5(4H)-yl)-1,3,5-triazin-
n.t.
n.t.
n.t.
d
d



2-yl)amino]benzenemethanesulfonamide


B45
3-[(4-(6,7-Dihydrothiazolo[5,4-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-
n.t.
n.t.
n.t.
c
c



yl)amino]benzenemethanesulfonamide


B47
3-[(4-(6,7-Dihydropyrazolo[1,5-a]pyrimidin-4(5H)-yl)-1,3,5-triazin-2-
n.t.
c
n.t.
n.t.
n.t.



yl)amino]benzenemethanesulfonamide


B49
3-[(4-(1H-Pyrrolo[3,4-c]pyridin-2(3H)-yl)-1,3,5-triazin-2-yl)amino]-
n.t.
n.t.
n.t.
d
d



benzenemethanesulfonamide


B51
3-[(4-(Pyrrolo[3,4-c]pyrazol-5(1H,4H,6H)-yl)-1,3,5-triazin-2-
n.t.
n.t.
n.t.
d
d



yl)amino]-benzenemethanesulfonamide





Activity range “a” means, that the compounds do have a IC50 <100 nM,


activity range “b” means, that the compounds do have a IC50 between 100 and 1000 nM,


activity range “c” means that the compounds do have an IC50 between 1000 and 10000 nM,


activity range “d” means that the compounds do have an IC50 >10000 nM;


“n.t.” means that the compounds have not been tested in this assay.


{circle around (1)}: Compound Number


{circle around (2)}: TNF alpha release (activity range)


{circle around (3)}: NF-kappaB activation (activity range)


{circle around (4)}: RNA Polymerase II Ser2 Phosphorylation (activity range)


{circle around (5)}: Cell Viability - Hela cells (activity range)


{circle around (6)}: Cell Viability - MDAMB468 cells (activity range)

## Claims

1. A compound having the general formula (I)




wherein



R1 is
L is a bond or —CR5R6—, —CR5R6—CR7R8—, —CR5R6—CR7R8—CR9R10—, —CR5R6—CR7R8—CR9R10—CR11R12—;
R5 —R12 represent independently of each other —H, —CH3, —C2H5, —C3H7, —F, —CI, —Br, —I;
R3 is selected from —H, —NO2, —CN, —Br, —I, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5, —CR13R14R21, —CR13R14—CR15R16R21, —CR13R14—CR15R16—CR17R18R21, —CR13R14—CR15R16—CR17R18—CR19R20 R21, —SO2R22, —CONR23R24, —NR25SO2NR23R24, —NR25SO2R22, —NR25CONR23R24, —SO2NR23R24, —SO(NR26)R22; 
R13 - R21 and R29 - R32 represent independently of each other —H, —CH3, -C2H5, -C3H7, -C4H9, —F, —CI, —Br, —I;
R22 and R28 are independently selected from R27′, —CR13R14R21, —CR13R14—CR15R16R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30R21, —CR13R14—CR15R16—CR17R18R21, —CR13R14—CR15R16—CR17R18—CR19R20R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—CR31R32R21,—CH2Ph; —CH2Ph the phenyl group of which may further be substituted by one, two, three, four or five substituents selected from the group consisting of —CH3, —C2H5, —C3H7, —F, —CI, —Br and —I;
R23 and R24 are independently selected from —H, —CR13R14R21, —CR13R14—CR15R16R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30R21, —CR13R14—CR15R16—CR17 R18 R21, —CR 13R14—CR15R16—CR17R18—CR19R20R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—CR31R32R21, —CR13R14—CR15R16—O—R33, —CR13R14—CR15R16 —CR17R18—O—R33, —CR13R14—CR15R16—NR33R34, —CR13R14—CR15R16—CR17R18—NR33R34, —CR13R14—CR15R16—CR17R18—CR19R20—NR33R34, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—NR33R34, —Ph, —CH2PH, phenyl group which may further be substituted by one, two, three, four or five substituents selected from the group consisting of —CH3, —C2H5, —C3H7, —F, —CI, —Br and —I; —CH2Ph the phenyl group of which may further be substituted by one, two, three, four or five substituents selected from the group consisting of —CH3, —C2H5, —C3H7, —F, —CI, —Br and —I; or both residues R23 and R24 together form with the nitrogen atom to which they are attached an azetidine, pyrrolidine, piperidine, piperazine, azepane, or morpholine ring;
R25 is selected from —H, —CH3, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5 or —C(CH3)3; 
R26 is —H, —CH3, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5, —C(CH3)3, —C5H11, —CH(CH3)—C3H7, —CH2—CH(CH3)—C2H5, —CH(CH3)—CH(CH3)2, —C(CH3)2—C2H5, —CH2—C(CH3)3, —CH(C2H5)2, —C2H4—CH(CH3)2, —C6H13, —C3H6—CH(CH3)2, —C2H4—CH(CH3)—C2H5, —CH(CH3)—C4H9, —CH2—CH(CH3)—C3H7, —CH(CH3)—CH2—CH(CH3)2, —CH(CH3)—CH(CH3)—C2H5, —CH2—CH(CH3)—CH(CH3)2, —CH2—C(CH3)2C2H5, —C(CH3)2—C3H7, —C(CH3)2—CH(CH3)2, —C2H4—C(CH3)3, —CH(CH3)—C(CH3)3, —CR13R14R21, —COR28, —CR13R14—CR15R16R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30R21, —CR13R14—CR15R16—CR17R18R21, —CR13R14—CR15R16—CR17R18—CR19R20R21, —CR13R14—CR15R16—CR17R18—CR19R20—CR29R30—CR31R32R21, —COOR28, —R27; 
R27, R27′ and R27′′ are independently selected from




these C3—C10-cycloalkyl groups may further be substituted by one, two, three, four, five or more substituents selected from the group consisting of —F, —CI, —Br and —I;
R33 and R34 represent independently of each other —H, —CH3, —C2H5, —C3H7, —C4H9, —CH2Ph, —COOC(CH3)3, —COOCH3, —COOCH2CH3, —COOCH2CH2CH3, —COOCH(CH3)2, —COOCH2Ph, —COCH3; 
R4 is selected from —H, —NO2, —CN, —F, —CI, —Br, —I, —CR35R36R37, —CR35R36—CR38R39—CR40R41—CR42R43R37, —O—CR35R36—CR38R39R37, —O—CR35R36—CR38R39—CR40R41R37, —CR35R36—CR38R39—CR40R41R37, —O—R35R36—CR38R39—CR40R41—CR42R43R37, —CR35R36—CR38R39R37, —O—CR35R36—CR38R39—CR40R41—CR42R43—CR44R45R37, —O—CR35R36R37, —O—CR35R36—CR38R39—CR40R41—CR42R43—CR44R45—CR46R47R37, —CR35R36—CR38R39—CR40R41—CR42R43—CR44R45R37—OCH2Ph, —R27′′, —O—R27′′, —CR35R36—CR38R39—CR40R41—CR42R43—CR44R45—CR46R47R37,
R35 —R47 represent independently of each other —H, —CR48R49R50, —CR48R49—CR51R52R50, —CR48R49—CR51R52—CR53R54R50, —CR48R49—CR51R52—CR53R54—CR55R56 R50, —F, —CI, —Br, —I;
R48 R56 represent independently of each other —H, —F, —Cl, —Br, —I;
R2 is selected from










B is a bond, —CH2, —CH2CH2—, —CH2CH2CH2—, —CF2—, CH2O—, —CH2OCH2—, —CH2CH2O—, CH2NH—, CH2N(CH3)—, CH2NHCH2, —CH2N(CH3)CH2—, CH2CH2NH—, CH2CH2N(CH3)—;
R57 is —H, CH3, —CF3, —CHF2, —CH2F, —COOCH3, —COOCH2CH3, —COOH, —COOCH2Ph, —COOCH2CH2CH3, —COOCH(CH3)2, —COOCH2CH2CH2CH3, —COOCH(CH3)(CH2CH3), —COOCH2CH(CH3)2, —COOC(CH3)3, —CONH2, —Ph, —CONH(CH3), —CON(CH3)2, —CN, —NH2, —NH(CH3), —N(CH3)2, —NH(CH2CH3), —N(CH3)(CH2CH3), —N(CH2CH3)2, pyrrolidin—1—yl, piperidin—1—yl, azetidin—1—yl, morpholin—4—yl;
R58 is —H, CH3, —F, —CI, —CF3, —CH2OCH3; 
R59 is —H, CH3, —F, —CI, —CF3, —Ph—, CH2OCH3; and
R60 is —H, —CH3, —CF3, —COCH3, —COOCH3, —COOCH2CH3, —COOC(CH3)3, —COOCH2Ph;
Y is —O—, —SO—, —NH—, or —N(CH3)—;
R61 is —H, CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, CH2OCH3, —CH2OH, phenyl, —CH2Ph, CH2OPh, —CH2CH2Ph ;
R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —S02CH3; 
R64 and R65 are independently selected from —H—, CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3; 
R66 and R67 are independently selected from —H—, CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br;
R68 is selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —OH, —OCH3, —NH2; 
R69 is selected from —H, —CH3, —CH2CH3; 
R70 and R71 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —NH2; and
R72 is selected from —H, —CH3, —CF3, —CH2CH3; 
or a stereoisomer or mixture thereof, a solvate, a tautomer, or a pharmaceutically acceptable salt thereof.

2. The compound according to claim 1, wherein



R1 represents

in which
L is a bond, —CH2—, —CH2CH2—, or —CF2—;
R3 is —SO2NH2, —SO2NH(CH3), —SO2N(CH3)2, —SO2NH(CH2CH2OCH3), —NHSO2CH3, —NHSO2CH2CH3, —NHSO2CH2CH2CH3, —NHSO2CF3, —SO02CH3, —NHSO2NH2, —SO(NH)CH3; 
R4 is —H, —CH3, —F, —CI, or —CF3; 
R2 represents one of the residues listed under (i) to (xv):




the group —B—R57 is —H, —CH2OCH3, —COOCH3, —COOCH2CH3, —CH2NH—000—C(CH3)3, —phenyl, or —CH2—CH2—N(CH3)2; and
R58 and R59 are independently selected from —H, —F, and —CH3; 
R69 is selected from —H and —CH3; 




in which the group —B—R57 is —H, —CH3, —CF3, —CH2OCH3, —COOCH3, —COOCH2CH3, —CONH2, benzyl, —CH2—benzyl, —CH2—O—CH2—phenyl,
—CH2—NH—phenyl, —CH2—O—phenyl, phenyl, or





and
R58 and R59 are independently selected from —H, —F, —CH2OCH3, or —CH3; 




in which R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —CI, —Br, —OH, —NH2, —OCH3, —SCH3, —SO2CH3; 




in which R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —S02CH3; 




in which R64 and R65 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3; and
R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —S02CH3; and
R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, —phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph;




in which R68 is —H, —CH3, —CF3, or —CH2CH3; 




in which R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —F, —Cl;




in which R68 is —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl; and
R69 is —H or —CH3; 




in which R68 is —H, —CH3, —CF3, or —CH2CH3; 




in which —B—R57 is —H, —CH2OCH3, —COOCH3, or phenyl;




R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
R79 and R71 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —NH2;




R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —SO02CH3;




R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
R62 and R63 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —OH, —NH2, —OCH3, —SCH3, —S02CH3;




R70 is independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br, —COOH, —COOCH3, —COOCH2CH3, —CONH2, —NH2;




R61 is —H, —CH3, —CF3, —CH2CH3, —CH2CH2CH3, —COOCH3, —COCH2CH3, —COOH, —CONH2, —CN, —CH2OCH3, —CH2OH, phenyl, —CH2Ph, —CH2OPh, —CH2CH2Ph; and
R66 and R67 are independently selected from —H, —CH3, —CF3, —CH2CH3, —CN, —F, —Cl, —Br.

3. The compound according to claim 1, wherein the substituent
—L—R3 is —SO2NH2, —CH2SO02NH2, —CH2CH2SO02NH2, —CF2SO02NH2, —NHSO2NH2, —CH2NHSO2NH2, —SO02CH3, —SO(NH)CH3, —CH2SO(NH)CH3;
R4 is —H; and
R2 is

4. The compound according to claim 1, wherein the substituent
R1 represents




L is —CH2—;
R3 is —SO2NH2;
R2 represents:




—B—R57 is —H, —CF3, —CH3, —Ph, —CH2OH, —CH2OCH3, —COOCH3, —CONH2, —CH2NH—000—C(CH3)3, —CH2—CH2—N(CH3)2, —CH2—Ph,





—CH2—CH2—Ph, —CH2—O—Ph, —CH2—O—CH2—Ph or —CH2—NH—Ph,
R58 —R6° and R62 have the meanings as defined in claim 1 .

5. The compound according to claim 1, wherein the compound is selected from the group consisting of :
3-[(4-(Piperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(4,4-Difluoropiperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
3-[(4-(3,3-Difluoropiperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
rac-3-[(4-(2-Methoxymethylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(R)-3-[(4-(2-Methoxymethylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(R)-Methyl 1-[4-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]piperidine-2-carboxylate
rac-tert-Butyl [(1-(44(3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl)-piperidine-2-yl)methyl]carbamate
rac-3-[(4-(2-(2-(Dimethylamino)ethyl)piperidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
rac-3-[(4-(2-Phenylpiperidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
3-[(4-(Morpholin-4-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide
3-[(4-(Piperazin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide
3-[(4-(4-Methylpiperazin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
rac-3-[(4-(2-(Hydroxymethyl)piperazin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(Pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide
(S)-3-[(4-(2-Methylpyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
(R)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(R)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(S)-3-[(4-(2-(Methoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-((2R,5R)-Bis(methoxymethyl)pyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)am ino]-benzenemethanesulfonamide
3-[(4-((2S,5S)-Bis(methoxymethyl)pyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
(R)-Methyl 1 -[44(3-(Sulfamoylmethyl)phenyl)amino)-1 ,3,5-triazin-2-yl]pyrrolidine-2-carboxylate
(S)-Methyl 1 44-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]pyrrolidine-2-carboxylate
(R)-144-((3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]pyrrolidine-2-carboxamide
(R)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(S)-3-[(4-(2-(Hydroxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
rac-3-[(4-(2-Benzylpyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
rac-3-[(4-(2-(2-Phenylethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(R)-3-[(4-(2-(Phenoxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(R)-3-[(4-(2-(Phenylamino-methyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
(R)-3-[(4-(2-(Benzyloxymethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(R)-3-[(4-(2-((Pyrrolidin-1-yl)methyl)pyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
(S)-3-[(4-(2-((Pyrrolidin-1-yl)methyl)pyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
rac-3-[(4-(2-Phenylpyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
(R)-3-[(4-(2-Phenylpyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
(R)-3-[(4-(2-(Methoxymethyl)azetidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
(R)-Methyl 1-[44(3-(Sulfamoylmethyl)phenyl)amino)-1,3,5-triazin-2-yl]azetidine-2-carboxylate
rac-3-[(4-(2-Phenylazetidin-1-yl)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
3-[(4-(7,8-Dihydro-1,6-naphthyridin-6(5H)-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(7,8-Dihydropyrido[4,3-d]pyrimidin-6(5H)-yI)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
3-[(4-(3,4-Dihydroisoquinolin-2(1 H)-yI)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
3-[(4-(5-Amino-3,4-dihydroisoquinolin-2(1 H)-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(3,4-Dihydroisoquinolin-1 (2H)-yI)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
3-[(4-(6,7-Dihydrothieno[3,2-c]pyridin-5(4H)-yI)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(6,7-Dihydro-3H-imidazo[4,5-c]pyridin-5(4H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
3-[(4-(6,7-dihydrothiazolo[5,4-c]pyridin-5(4H)-yI)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(5,6-Dihydroimidazo[1,2-a]pyrazin-7(8H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
3-[(4-(6,7-Dihydropyrazolo[1,5-a]pyrimidin-4(5H)-yl)-1,3,5-triazin-2-yl)amino]-benzenemethanesulfonamide
3-[(4-(5H-Pyrrolo[3,4-b]pyridin-6(7H)-yI)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(1 H-Pyrrolo[3,4-c]pyridin-2(3H)-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(5H-Pyrrolo[3,4-d]pyrimidin-6(7H)-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(Pyrrolo[3,4-c]pyrazol-5(1 H,4H,6H)-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide
3-[(4-(Indolin-1-yl)-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide
(S)-3-[(4-(2-Methylpyrrolidin-1-yl)-1 ,3,5-triazin-2-yl)amino]benzenemethane-sulfonamide
(R)-3-[(4-(2-(Trifluoromethyl)pyrrolidin-1-yl)-1,3,5-triazin-2-yl)amino]benzene-methanesulfonamide.

6. A method for the treatment of a cell proliferative disease selected from the group consisting of:
breast cancer,  colorectal cancer,  cervical cancer, and comprising administering to a patient in need thereof a therapeutically effective amount of a compound according claim 1.

7. A pharmaceutical composition comprising at least one compound according to claim 1 as an active ingredient, together with at least one pharmaceutically acceptable carrier, excipient and/or diluent.

8. The pharmaceutical composition according to claim 7 further comprising one or more further anti-tumor agents.