FIELD OF THE INVENTION [0002] The invention relates to benzimidazole derivatives having antibacterial activity, to compositions of matter comprising the same, and to antibacterial methods of using the same. The invention also contemplates assays and diagnostic methods employing benzimidazole derivatives according to the invention.

BACKGROUND [0003] The emergence of bacterial resistance to various classes of antibiotics, including (3-lactam antibiotics, macrolides, quinolones and vancomycin, is becoming a major worldwide health problem.

The spread of antibiotic resistance among pathogenic bacteria imposes another serious problem for the clinical management of infectious diseases. In particular, antibiotic resistance among Gram-positive bacteria (Staphylococci, Enterococcci, and Streptococci) is becoming increasingly serious. Enterococci, which are generally resistant to most antibiotics, including penicillin, cephalosporin and aminoglycosides, has been previously treated with vancomycin or with a combination of more than one antibiotic. However, with the recent increased use of vancomycin in methicillin-resistant Staphylococcus aureus (MRSA), infections and colitis due to Colstridium difficle, multiple-drug-resistant Enterococcus faecium has been spreading. As such, the last resort for anti-infective diseases, the Vancomycin family of antibiotics, has now been gravely challenged in recent years due to the emergence of MRSA strains in clinical practice. There is an urgent need to discover novel antibacterial agents other than alalogs of existing antibiotics.

[0004] A considerable amount of attention has focused recently on new RNA-binding molecules for drug discovery. The interactions between RNA and biological macromolecules are clearly essential for many vital processes in molecular biology. In addition, the excitement over RNA-based viruses has fueled an interest in the development of potential RNA inhibitors. RNA offers several selective advantages over DNA as a therapeutic agent. First, chromosomal DNA is packaged extensively, significantly limiting its accessibility to small molecule regents. Second, DNA repair systems are available in the cell, whereas analogous enzymes for RNA repair are virtually unknown. Finally, RNA exhibits a high level of diversity in terms of tertiary folding, and therefore will likely have a greater potential for selective targeting based on structure rather than sequence.

[0005] Historically, however, RNA-based drug discovery has proved to be extremely difficult, and only a few classes of compounds are known to bind RNA with S A R information, for example aminoglycosides and cationic peptides. Discovery of RNA binders using traditional high throughput assays such as fluorescence, filter binding, SPA, SPR, etc. has proved to be equally unsuccessful.

[0006] Recently, a MS-based high throughput-screening assay has been developed. See, Hofstadler, A.; Griffey, R. H. Cum. Opin. Drug Discovery Dev. 2000,3, 423-431 ; Hofstadler, S. A.; Griffey, R. H.

Chem. Rev. (Washington. D. C. ) 2001,101, 377-390; Griffey, R. H.; Greig, M. J.; An, H.; Sasmor, H.; Manalili, S. J. Am. Chem. SOC. 1999,122, 474-475 ; Sannes-Lowery, K. A.; Griffey, R. H.; Hofstadler, S. A. Anal. Biochem. 2000,280, 264-271 ; Griffey, R. H.; Sannes-Lowery, K. A.; Drader, J. J.; Mohan, V.; Swayze, E. E.; Hofstadler, S. A. J. Am. Chem. SOC. 2000,122, 9933-9938, and Griffey, R. H.; Hofstadler, S. A.; Sannes-Lowery, K. A.; Ecker, D. J.; Crooke, S. T. Proc. Natl. Acad. Sci. U. S. A. 1999, 96,10129-10133, each of which is incorporated herein by reference in its entirety.

[0007] This assay is extremely sensitive and could detect RNA binders with Kd values ranging from nanomolar to millimolar. Coupled with mass assays to carry out competition experiments and determine the binding locations, such assays can be used to discover of novel compounds that bind to bacterial ribosomal RNA.

[0008] In view of the great importance of antibacterial compounds in animal, and particularly human health, it can be seen that there is a need for novel antibacterial agents. The present invention is therefore directed to, inter alia, such compounds and their uses, as well as other important ends.

SUMMARY OF THE INVENTION [0009] The present invention provides compounds having antibacterial activity. The present invention also provides compositions containing compounds of formula I and methods for using the subject compounds. Methods for making the compounds of the invention are also disclosed. Other useful methods will be apparent to those skilled in the art, upon consideration of the present disclosure, including the appended drawings and claims.

DETAILED DESCRIPTION OF THE INVENTION [0010] In some embodiments, the invention provides compounds of formula I : wherein: R3 and R4 are independently each H, halogen, C,-C, alkyl, C, -C, alkoxy, trihaloakyl, alkoxycarbonyl, alkoxy, NR15RI6, or N02 ; R30, is C1-6 alkyl, heteroarylalkyl, arylalkyl, or heteroaryl, wherein each of said heteroarylalkyl, arylalkyl, or heteroaryl groups each can be optionally substituted with up to three substituents selected from halogen, N02 and haloalkyl, dihaloalkyl, or trihaloalkyl ; or Wo has the structure XX: wherein R3'is alkylamino, aminoalkylamino, poly (alkylamino) amino, heterocycloalkylamino, heterocycloalkyl, -NH-(CHOH), -CH,OH, -NH-(CH2)1-12-heteroaryl, or -NH-(CH2)1-12-heterocycloalkyl.

[0011] In some embodiments, the present invention provides dimeric benzimidazole compounds having structure of Formula II: wherein: R2 is NHZ or piperidin-4-yl; R50 and R51 are each independently selected from H, halogen, C,-C6 alkyl, trihaloalkyl, alkoxycarbonyl, alkoxy, NR15R16, and NO2 ; wherein said C, 6 alkyl, alkoxycarbonyl and alkoxy groups can each be optionally substituted with NR'5Rl6 ; R 15 is H, halogen, C1-12 alkyl, methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, branched-or straight-chain polyaminoalkyl, or a group of formula CH2 (CHOH) 4CH20H, wherein said methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, and branched-or straight-chain polyaminoalkyl groups can be substituted by up to 3 OH groups; Rl6 is H, halogen, or C,-C, alkyl ; or R15 and R16 together with the nitrogen atom to which they are attached can form a succinimido or phthalimido group or a fused ring derivative thereof, wherein said succinimido or phthalimido group, or fused ring derivative thereof, can be optionally substituted by up to three substituents independently selected from NO2 and halogen; Ro is alkylene having from 1 to 18 carbons, or R !-X-R'O-) H; R9 and R° are each independently alkylene having from 1 to about 20 carbons; X is-N (R12)-, -C(R13)(R14)- or O ; and R12, R13 and R14 are each independently H or C1-6 alkyl.

[0012] In some embodiments, the present invention provides compounds of formula: wherein R52 and R53 are each independently selected from H, halogen, C,-C, alkyl, trihaloalkyl, alkoxycarbonyl, alkoxy, NRl5Rl6 and NO2, wherein said Cl 6 alkyl, trihaloalkyl, alkoxycarbonyl, and alkoxy groups can each be optionally substituted with NR'5R'6 ; R'S is H, halogen, C1-12 alkyl, methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, branched-or straight-chain polyaminoalkyl, or a group of formula: CH2 (CHOH) 4CH20H ; wherein said methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, and branched-or straight-chain polyaminoalkyl groups can be substituted by up to 3 OH groups; R'6 is H, halogen, or Cl 6 alkyl ; or R15 and R16 together with the nitrogen atom to which they are attached can form a succinimido or phthalimido group or a fused ring derivative thereof, wherein said succinimido or phthalimido group or fused ring derivative thereof can be optionally substituted by up to three substituents independently selected from NO2 and halogen; and z is 1 to 6.

[0013] Also provided by the present invention are compounds having Formula IV: wherein: R2a is amino, phenyl, monocyclic or bicyclic heterocycloalkyl having 1 or 2 ring nitrogen atoms, monocyclic heteroaryl or bicyclic heteroaryl having 1 or 2 ring nitrogen atoms, cycloalkyl, halogen, heterocycloalkylalkyl (i. e., alkyl substituted with heterocycloalkyl) having 1 or 2 ring nitrogen atoms, monocyclic or bicyclic heterocycloalkylamino having 1 or 2 ring nitrogen atoms or a group of formula- S-alkylene-L'where L'is monocyclic or bicyclic heteroaryl having 1 or 2 ring nitrogen atoms; wherein each of said amino, phenyl, heterocycloalkyl, heteroaryl, cycloalkyl, heterocycloalkylalkyl, or heterocycloalkylamino groups can be optionally substituted with a group selected from amino, OH, C1-12 alkyl, a structure of formula: C (=O) CH (NH2)-L2, where L2 is the side chain of a naturally occurring alpha amino acid,-C (NH2) NH, Cl l2 alkylcarbonyl, monocyclic or bicyclic heteroaryl having 1 or 2 ring nitrogen atoms, monocyclic or bicyclic heteroarylalkyl having 1 or 2 ring nitrogen atoms, or S-alkyl-heteroaryl where said heteroaryl is monocyclic or bicyclic having 1 or 2 ring nitrogen atoms; and R3 and R4 are each independently halogen, amino, NO2, CN, Cl 6 alkoxy or Cl 6 alkyl, optionally substituted with up to 3 halogen atoms; and R30 is H, alkyl, aryl, arylalkyl, heteroaryl; heteroarylalkyl, heterocycloalkyl, arylsulfonyl, aryloxycarbonyl, alkoxyalkoxyalkyl, alkyl-S-R, alkyl-NH-C (=O)-R8 or R9-X-R'°-R")-H ; wherein each of the alkyl, aryl, arylalkyl heteroaryl, heteroarylalkyl, heterocycloalkyl, arylsulfonyl, aryloxycarbonyl and alkoxyalkoxyalkyl moieties in each of the foregoing R'groups can be optionally substituted with up to 3 groups independently selected from the group consisting of C1-6 alkyl, OH, hydroxyalkyl,-C (=O)-R5, CN, aryl, alkoxycarbonyl, alkylaryl, arylalkyl, heteroaryl, S-heteroaryl optionally substituted with halogen, heteroarylalkyl optionally substituted with halogen, heterocycloalkyl optionally substituted with amino, NOz, halogen, monohaloalkyl, dihaloalkyl, trihaloalkyl, perhaloaryl, perhaloalkylaryl, alkyl-NRI5Rl6 or NRI5Rl6 ; or one of said alkyl, aryl, arylalkyl heteroaryl, heteroarylalkyl, heterocycloalkyl, arylsulfonyl, aryloxycarbonyl or alkoxyalkoxyalkyl moieties of one of said R'groups can be attached to a structure of Formula IV at position R'thereof ; Rs is H, NHNHR6, NHN=CH-R6, heteroaryl, heterocycloalkyl, wherein said hereteroaryl group can be optionally substituted with an aryl or heteroaryl group, R6 is aryl, heteroaryl, arylsulfonyl, heteroarylsulfonyl, -C (=S)-NH-aryl, -C (=S) -NH-arylcarbonyl, -C (=S)-NH-heteroarylcarbonyl,-C (=S)-NH-alkylene-R 21,-C (=O)-NH-aryl,- C (=O)-NH-arylcarbonyl,-C (=O)-NH-heteroarylcarbonyl, or-C (=O)-NH-alkylene-R2l where Ruz'ils carboxy, alkoxycarbonyl, aryl, heteroaryl, heterocycloalkyl, arylaminocarbonyl, cycloalkylaminocarbonyl, or a saturated hydrocarbon fused ring system optionally having an aryl ring fused thereto, said ring system being optionally substituted with up to three alkyl groups on the alkyl or aryl rings thereof; wherein any of said R6 groups can be optionally substituted with up to 3 groups selected from NRlsRl6, alkyl, hydroxy, halogen, aryl, alkoxy, trihaloalkoxy, arylalkyloxy, NO2,-SH,-S-alkyl, heteroarylcarbonyl, heteroaryl, alkylheteroaryl, or a moiety of formula-OC2CHz-O-attached to adjacent atoms of said R6 group; R7 is heteroaryl or heterocycloalkyl ; R8 is aryl; R9 and R'° are each independently alkylene having from 1 to about 20 carbons; <BR> <BR> <BR> X is -N(R12)-, -C(R13)(R14)- or O ;<BR> R11 is H, heterocycloaryl or alkoxy, wherein said heterocycloaryl or alkoxy group can be optionally substituted with up to four groups independently selected from halogen, amino, trihaloalkyl, alkoxycarbonyl, and CN; Rl2 is H or Cl 6 alkyl ; and *13 and R14 are each independently H or Cl 6 alkyl ; Rls is H, halogen, C1-12 alkyl, methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, branched-or straight-chain polyaminoalkyl, or a group of formula CH2 (CHOH) 4CH20H, wherein said methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, and branched-or straight-chain polyaminoalkyl groups can be substituted by up to 3 OH groups; Rl6 is H, halogen, or C,-6 alkyl ; or R15 and Rl6 together with the nitrogen atom to which they are attached can form a succinimido or phthalimido group or a fused ring derivative thereof, wherein said succinimido or phthalimido group or fused ring derivative thereof can be optionally substituted by up to three substituents independently selected from NO2 and halogen, or a group of Formula I at position R, threreof; or R 15 and Rl6 together with the nitrogen atom to which they are attached can form a group of Formula I wherein said nitrogen atom is Q4 thereof; [0014] The invention also provides compounds of Formula V: wherein: Q5 is CH or N ; Q6 is C-R61 or N ; Q7 iS C R60 or N ; e and R61 are each independently H, halogen, Cl 6 alkyl, trihaloalkyl, or Cl 6 alkoxy ; provided that when Q6 iS C-R61, Q7 iS C-R60, and Ql is CH, then R6° and R6', are not both H.

[0015] The present invention provides methods of treating a subject with a compound belonging to one of formulae I-V, the method comprising contacting the subject with a detectable amount of an inventive compound of formulae I-V, set forth above. In some embodiments, the subject is an animal, e. g. a mammal. Exemplary mammalian subjects include mouse, rat, monkey, chimpanzee, dog, cat and human.

In some embodiments, the subject has a microbial infection, e. g. a bacterial infection. In certain embodiments, the infective microbe is a Gram-positive bacterium, such as [insert Gram-positive bacteria here. ] In particular embodiments, the method further comprises analyzing the subject or some tissue or fluid extracted therefrom for one or more indicators of pharmacokinetics, pharmacodynamics or toxicology. Exemplary tissues that may be extracted include hepatic, pancreatic, renal, pulmonary, spleen, lymphatic, cardiac, gastrointestinal, esophageal, dermal, epidermal, etc. Exemplary fluids that may be extracted include blood, lymphatic, synovial, and mucus fluids.

[0016] In some embodiments, the present invention also provides a method for treating a patient having a bacterial infection, said comprising administering to said patient a compound of the invention, e. g. a compound belonging to one of formulae I-V. In some embodiments, said patient is a mammal, such as a mouse, rat, monkey, chimpanzee, dog, cat or human.

[0017] Also provided are methods for inhibiting bacterial growth comprising contacting a bacterium, or the environs of the bacterium, with a compound of the invention, e. g. a compound belonging to one of formulae I-V. In some embodiments, said bacterium is a Gram-positive bacteria, e. g. a member of one of the species of staphylococci, enterococci, or streptococci. In some embodiments the bacterium is S. aureus, E. hirae, S. pyogenes, S. pneumoniae, E. coli, P. vulgaris, K. pneumoniae, P. aeruginosa, C. albicans, E. faecalis, E. faecali, or E. faecium.

[0018] The present invention also provides compositions that include at least one compound of the invention. In some embodiments, the compositions further comprise a second ingredient. In some embodiments, the inventive compositions are in a pharmaceutically acceptable form.

[0019] In some embodiments, the compounds of the invention may be present as a salt, e. g. as a pharmaceutically acceptable salt.

BRIEF DESCRIPTION OF THE DRAWINGS [0020] FIG. 1 is a table showing activity of benzimidazoles of Examples 11 and 12 against four strains of Gram positive and four strains of gram negative bacteria.

[0021] FIG. 2 is a table showing activity of benzimidazoles of Examples 11 and 12 against seven clinically important strains of enterococcus.

[0022] FIG. 3 shows the in vitro inhibitory activity of selected benzimidazoles of Example 16 against for Gram positive bacterial strains, four gram negative bacterial strains and one yeast strain.

DETAILED DESCRIPTION OF THE INVENTION [0023] The present invention provides compounds, including salts thereof, of formulae I-V, above. As used hereinafter, the term"inventive compounds"means novel compounds of formulae I-V and novel salts of compounds belonging to formulae I-V.

[0024] The present invention also provides compositions containing the subject compounds, and methods for using the subject compounds. Methods for making the compounds of the invention are also disclosed. Other useful methods will be apparent to those skilled in the art upon consideration of the present disclosure.

[0025] These and other features of the compounds of the subject invention are set forth in more detail below.

[0026] In some embodiments, compounds are provided having the formula I: wherein: R3 and R4 are independently each H, halogen, C,-C, alkyl, C, -C, alkoxy, trihaloakyl, alkoxycarbonyl, alkoxy, NR15RI6, or N02 ; R30, is C1-6 alkyl, heteroarylalkyl, arylalkyl, or heteroaryl, wherein each of said heteroarylalkyl, arylalkyl, or heteroaryl groups each can be optionally substituted with up to three substituents selected from halogen, N02 and haloalkyl, dihaloalkyl, or trihaloalkyl ; or Wo has the structure XX : wherein R3'is alkylamino, aminoalkylamino, poly (alkylamino) amino, heterocycloalkylamino, heterocycloalkyi,-NH- (CHOH),-CH, OH,-NHCHz-u-heteroaryl, or-NH- (CH2)).) 2-heterocycloalkyl. [0027] In further embodiments, the present invention provides dimeric benzimidazole compounds structure of Formula II : wherein: W is NH2 or piperidin-4-yl; R50 and R51 are each independently selected from H, halogen, Cl-C6 alkyl, trihaloalkyl, alkoxycarbonyl, alkoxy, NR15R16, and NO2 ; wherein said CI_6 alkyl, alkoxycarbonyl and alkoxy groups can each be optionally substituted with NRI5Rl6 ; Rl5 is H, halogen, C1-12 alkyl, methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, branched-or straight-chain polyaminoalkyl, or a group of formula CH2 (CHOH) 4CH20H, wherein said methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, and branched-or straight-chain polyaminoalkyl groups can be substituted by up to 3 OH groups; R'6 is H, halogen, or Cl 6 alkyl ; or R'S and R'6 together with the nitrogen atom to which they are attached can form a succinimido or phthalimido group or a fused ring derivative thereof, wherein said succinimido or phthalimido group, or fused ring derivative thereof, can be optionally substituted by up to three substituents independently selected from NO2 and halogen; R6° is alkylene having from 1 to 18 carbons, or-R9-X-R'°-)-H ; R9 and R'° are each independently alkylene having from 1 to about 20 carbons; X is -N(R12)-, -C(R13)(R14)- or O ; and R12, R'3 and R14 are each independently H or C1-6 alkyl.

[0028] In some embodiments, W is piperidin-4-yl. E other embodiments, Wo and W'are each halogen, e. g. chlorine.

[0029] In some embodiments, R60 is alkylene having from 1 to 6 carbons or from 1 to 4 carbons. In some embodiments, R60 is-CH2-C6H4-CH2-, e. g. in which - CH2-C6H4-CH2- is a para-a, a-xylene radical.

[0030] In some of the foregoing embodiments, R2 is NH2. In further embodiments, R50 and W'are each independently selected from H, halogen, methyl, COOCH3, CN and CF3.

[0031] In some embodiments, R6° is-R9-X-R'°. In further embodiments, X is-N (R12)-. In some embodiments, R 12 is methyl and R9 and R'° are each- (CH2) 2- or- (CH2) 3-, e. g. wherein R50 and Rus are each halogen, or where wherein Wo and Rus are each H, or where Wo is Br and R51 is H, or where Wo is CH3 and R51 is H, or where R50 is COOCH3 and R51 is H, or where Wo is CF3 and R51 is H, or where Wo is CN and R51 is H.

[0032] In some embodiments, X is O. In some such embodiments R9 and R'° are each- (CH2) 2- or- (CH2) 3-, e. g. where R50 and Rus are each halogen, or where R50 and R51 are each H, or where R50 is Br and Wu ils H, or where Wo is CH3 and R"is H, or where Wo is COOCH3, and Wu ils H, or where R50 is CF3 and R"is H, or where R50 is CN and R51 is H.

[0033] In some further embodiments, the present invention provides compounds of formula III: wherein: R52 and R53 are each independently selected from H, halogen, C,-C, alkyl, trihaloalkyl, alkoxycarbonyl, alkoxy, NR15R16 and NO2, wherein said C1-6 alkyl, trihaloalkyl, alkoxycarbonyl, and alkoxy groups can each be optionally substituted with NR15R16 ; Ris is H, halogen, C1-12 alkyl, methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, branched-or straight-chain polyaminoalkyl, or a group of formula: CH2 (CHOH) 4CH20H ; wherein said methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, and branched-or straight-chain polyaminoalkyl groups can be substituted by up to 3 OH groups; R'6 is H, halogen, or CI 6 alkyl ; or R'S and R'6 together with the nitrogen atom to which they are attached can form a succinimido or phthalimido group or a fused ring derivative thereof, wherein said succinimido or phthalimido group or fused ring derivative thereof can be optionally substituted by up to three substituents independently selected from NOz and halogen; and z is 1 to 6.

[0034] In some embodiments, R'S and R'6 are each methyl, e. g. wherein z is 2 or 3, further for example where R52 and Rs3 are each independently, H, Cl 6 alkyl, alkoxy optionally substituted with dialkylamino, or alkylamino. In further embodiments, R52 is H, e. g. where Rs3 methyl, methoxy, alkoxy optionally substituted with dialkylamino, or alkylamino, e. g. wherein R53 is-OCH3 or-O (CH2) 3N (CH3) 2.

[0035] In some embodiments, where RIS and R are each methyl, z is 2 or 3, W is H, C1-6 alkyl, alkoxy optionally substituted with dialkylamino, or alkylamino, W 3 is H. In some such embodiments, R2 is methyl, methoxy, alkoxy optionally substituted with dialkylamino, or alkylamino. In further emboidments, R52 is-OCH3, is-O (CHz) 3N (CH3) z.

[0036] The invention also provides compounds having the Formula IV: wherein: Rua ils amino, phenyl, monocyclic or bicyclic heterocycloalkyl having 1 or 2 ring nitrogen atoms, monocyclic heteroaryl or bicyclic heteroaryl having 1 or 2 ring nitrogen atoms, cycloalkyl, halogen, heterocycloalkylalkyl (i. e., alkyl substituted with heterocycloalkyl) having 1 or 2 ring nitrogen atoms, monocyclic or bicyclic heterocycloalkylamino having 1 or 2 ring nitrogen atoms or a group of formula- S-alkylene-L'where L'is monocyclic or bicyclic heteroaryl having 1 or 2 ring nitrogen atoms; wherein each of said amino, phenyl, heterocycloalkyl, heteroaryl, cycloalkyl, heterocycloalkylalkyl, or heterocycloalkylamino groups can be optionally substituted with a group selected from amino, OH, CI-12 alkyl, a structure of formula:-C (=O) CH (NH2)-L, where L'is the side chain of a naturally occurring alpha amino acid,-C (NH2) NH, C1-12 alkylcarbonyl, monocyclic or bicyclic heteroaryl having 1 or 2 ring nitrogen atoms, monocyclic or bicyclic heteroarylalkyl having 1 or 2 ring nitrogen atoms, or S-alkyl-heteroaryl where said heteroaryl is monocyclic or bicyclic having 1 or 2 ring nitrogen atoms; and R3 and R4 are each independently halogen, amino, NO2, CN, C1-6 alkoxy or C1-6 alkyl, optionally substituted with up to 3 halogen atoms; and R30 is H, alkyl, aryl, arylalkyl, heteroaryl; heteroarylalkyl, heterocycloalkyl, arylsulfonyl, aryloxycarbonyl, alkoxyalkoxyalkyl, alkyl-S-R7,-alkyl-NH-C (=0)-R' or-R"-X-R"'-R")-H ; wherein each of the alkyl, aryl, arylalkyl heteroaryl, heteroarylalkyl, heterocycloalkyl, arylsulfonyl, aryloxycarbonyl and alkoxyalkoxyalkyl moieties in each of the foregoing R'groups can be optionally substituted with up to 3 groups independently selected from the group consisting of CI-6 alkyl, OH, hydroxyalkyl,-C (=O)-R5, CN, aryl, alkoxycarbonyl, alkylaryl, arylalkyl, heteroaryl, S-heteroaryl optionally substituted with halogen, heteroarylalkyl optionally substituted with halogen, heterocycloalkyl optionally substituted with amino, NO2, halogen, monohaloalkyl, dihaloalkyl, trihaloalkyl, perhaloaryl, perhaloalkylaryl, alkyl-NR15R16 or NR15R16; or one of said alkyl, aryl, arylalkyl heteroaryl, heteroarylalkyl, heterocycloalkyl, arylsulfonyl, aryloxycarbonyl or alkoxyalkoxyalkyl moieties of one of said R'groups can be attached to a structure of Formula IV at position R1 thereof; Rs is H, NHNHR6, NHN=CH-R6, heteroaryl, heterocycloalkyl, wherein said hereteroaryl group can be optionally substituted with an aryl or heteroaryl group, R6 is aryl, heteroaryl, arylsulfonyl, heteroarylsulfonyl, -C (=S)-NH-aryl, -C (=S) -NH-arylcarbonyl,-C (=S)-NH-heteroarylcarbonyl,-C (=S)-NH-alkylene-R2l,-C (=O)-NH-aryl,- C (=O)-NH-arylcarbonyl,-C (=O)-NH-heteroarylcarbonyl, or-C (=O)-NH-alkylene-R2l where Wu ils carboxy, alkoxycarbonyl, aryl, heteroaryl, heterocycloalkyl, arylaminocarbonyl, cycloalkylaminocarbonyl, or a saturated hydrocarbon fused ring system optionally having an aryl ring fused thereto, said ring system being optionally substituted with up to three alkyl groups on the alkyl or aryl rings thereof ; wherein any of said R6 groups can be optionally substituted with up to 3 groups selected from NRl5Rl6, alkyl, hydroxy, halogen, aryl, alkoxy, trihaloalkoxy, arylalkyloxy, NO2,-SH,-S-alkyl, heteroarylcarbonyl, heteroaryl, alkylheteroaryl, or a moiety of formula-OC2CH2-O-attached to adjacent atoms of said R6 group; R7 is heteroaryl or heterocycloalkyl ; Rus ils aryl; R9 and R'° are each independently alkylene having from 1 to about 20 carbons; X is-N (R'Z)-,-C (R'3) (R'a)- or O ; R"is H, heterocycloaryl or alkoxy, wherein said heterocycloaryl or alkoxy group can be optionally substituted with up to four groups independently selected from halogen, amino, trihaloalkyl, alkoxycarbonyl, and CN; R'2 is H or Cl 6 alkyl ; and R'3 and R'4 are each independently H or C1-6 alkyl ; Rls is H, halogen, C1-12 alkyl, methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, branched-or straight-chain polyaminoalkyl, or a group of formula CH2 (CHOH) 4CHzOH, wherein said methylcarbonyl, heterocycloalkyl, arylsulfonyl, heteroarylalkyl, aminoalkyl, arylcarbonyl, and branched-or straight-chain polyaminoalkyl groups can be substituted by up to 3 OH groups; R 16 is H, halogen, or Cl 6 alkyl ; or R'S and R'6 together with the nitrogen atom to which they are attached can form a succinimido or phthalimido group or a fused ring derivative thereof, wherein said succinimido or phthalimido group or fused ring derivative thereof can be optionally substituted by up to three substituents independently selected from NO2 and halogen, or a group of Formula I at position RI thereof ; or Ris and R'6 together with the nitrogen atom to which they are attached can form a group of Formula I wherein said nitrogen atom is Q4 thereof.

[0037] In some embodiments R3 and R4 are each halogen, e. g. chlorine. In further embodiments, R a is amino, CI, phenyl, monocyclic heterocycloalkyl having 1 or 2 ring nitrogen atoms, monocyclic heteroaryl having 1 ring nitrogen atom, cyclopentyl, cyclohexyl, heterocycloalkyl-methyl, piperidin-4-yl-amino or a group of the formula-S- (C2-4 alkylene) -N-phthalimido; wherein each of said phenyl, heterocycloalkyl, heteroaryl, cyclopentyl, cyclohexyl, heterocycloalkyl-methyl, and piperidin-4-yl-amino groups can be optionally substituted with a group selected from NH2, OH, CH3, COOCH3, a structure of the formula:- C (=O) CH (NH2)-L2, where L2 is a serine or threonin side chain,-C (NH2) NH, OH, benzimidazolyl, and benzimidazolylmethyl.

[0038] In still further embodiments Rua ils amino, Cl, piperidinyl, pyridinyl, phenyl, cyclopentyl, cyclohexyl, pyrrolidinyl, piperazinyl,-CH2-piperazinyl, piperidin-4-yl-amino, or S-alkyl-phthalyl, wherein said piperidinyl, pyridinyl, phenyl, cyclopentyl, cyclohexyl, pyrrolidinyl, piperazinyl,-CH2- piperadinyl, piperidin-4-yl-amino, or S-alkyl-phthalyl groups can be optionally substituted with a group <BR> <BR> <BR> <BR> selected from NH2, methylcarbonyl, -C (=O) CH (NH2)-CH20H, methyl, OH,-C (NH2) =NH, OH, benzimidazol-2-yl, and-CH2-benzimidazol-2-yl.

[0039] In still further embodiments R2a is amino, Cl, piperidinyl, pyridinyl, phenyl, cyclopentyl, cyclohexyl, pyrrolidinyl, piperazinyl,-CH2-piperazinyl, piperidin-4-yl-amino, or S-alkyl-phthalyl, wherein said piperidinyl, pyridinyl, phenyl, cyclopentyl, cyclohexyl, pyrrolidinyl, piperazinyl,-CH2- piperadinyl, piperidin-4-yl-amino, or S-alkyl-phthalyl groups can be optionally substituted with a group selected from NHz, methylcarbonyl,-C (=O) CH (NH2)-CH20H, methyl, OH,-C (NH2) =NH, OH, benzimidazol-2-yl, and-CH2-benzimidazol-2-yl.

[0040] In further embodiments R2a is amino, Cl, phenyl substituted with amino, cyclopentyl substituted with amino, cyclohexyl, cyclohexyl substituted with amino, pyrrolidin-2-yl optionally substituted with hydroxy, piperazin-1-yl optionally substituted at the 4-position with benzimidazole-2-yl, piperazin-1-yl- methyl optionally substituted at the 4-yl position by-CH2-benzimidazol-2-yl, piperidin-4-yl-amino, S- alkyl-phthalyl, or said Wu ils piperidin-4-yl optionally substituted at the 1-yl position with-C (=O) CH3, - C (=O) CH (NH2)-CH20H,-C (NH2) =NH or CH3.

[0041] In still further embodiments, Rua ils amino, piperidin-4-yl, piperazin-1-yl optionally substituted with benzimidazol-2-yl, pyridin-4-yl, piperidin-4-yl optionally substituted with benzimidazol-2-yl, pyridin-4-yl, piperidin-4-yl optionally substituted at the 1-yl position with-C (=O) CH3, -C (=O) CH (NH2)- CH2OH, -c (NH2) =NH, or CH3, 4-amino-piperidin-1-yl, 3-aminophen-1-yl, 3-aminocyclopent-1-yl, cyclohexyl optionally substituted at the 3-or 4-position with NH2, 4-hydroxypyrrolidin-2-yl, piperazin-l- yl-methyl, 4-(benzimidazol-2-ylmethyl)piperazin-1-yl-methyl, or S-alkylphthalyl where said alkyl has from 2 to 4 carbons.

[0042] In still further embodiments, R2a is piperidin4-yl option substituted at the 1-yl position with- C (=O) CH3, -C (=O) CH (NH2) CH2OH,-C (NHz) =NH, or CH3.

[0043] In further embodiments where R3 or R4 are each chlorine, Rua ils piperidin-4-yl optionally substituted at the 1-yl position with-C (=O) CH3, -C (=O) CH (NH2) CH20H,-C (NH2) =NH, or CH3.

[0044] In some embodiments, R2a is piperidin-4-yl, e. g. when R3 and R4 are each chlorine. In some embodiments, R2a is NH2, e. g. wherein R3 and R4 are each chlorine.

[0045] In some embodiments, where R3 and R4 are each chlorine and R2a is piperidin-4-yl, Wo is alkyl substituted with-C (=O) R5, e. g. wherein RS is NHNHR6 or NHN=CH-R6.

[0046] In some embodiments, Rs is-NHNHR6 where R6 is (C=O)-NH-aryl,- (C=O)-NH-cycloalkyl,- C9=S) -NH-aryl, arylsulfonyl, heterarylsulfonyl, heterocycloalkyl, arylaminocarbonyl, cycloalkylaminocarbonyl,-C9=S)-NH-alkylene-RZ'where R2'is heteroaryl or heterocycloaryl, or a saturated hydrocarbon fused ring system optionally having an aryl ring fused thereto, said ring system being optionally substituted with up to three alkyl groups on the alkyl or aryl rings thereof; wherein any of said R6 groups can be optionally substituted wth up to 3 groups selected from NR15R16, NO2, a moiety of formula-OCH2CH2O-attached to adjacent atoms of said R6 group, aryl, CI-6 alkoxy, carboxy, or CI-6 trihaloalkoxy.

[0047] In some embodiments Rs is-NHN=CH-R6. In some embodiments, R6 is aryl or heteroaryl optionally substituted with up to 3 groups selected from OH, C1-6 alkoxy, NO2, CI-6 trihaloalkoxy, CI-6 trihaloalkyl, aryl, arylalkyloxy, and a moiety of the formula-OCH2CH2O-attached to adjacent atoms of said R6 group.

[0048] In some embodiments wherein Rua is piperidin-4-yl, R30 has the formula :- (CH2) q-L4 where q is 0 to 6 and L4 is aryl, heteroaryl or heterocycloalkyl, arylsulfonamino, arylcarboxyamino or S-heteroaryl, where each of said L4 is optionally substituted with up to three substituents selected from halogen and NO2. Preferably, said L4 is maleimido, succinimido, phthalimido, naphthalimido, pyromellitic diimido, phenylsulfonamido, phenylcarboxamido, benzopyrrolidine, benzimidazole, triazole or-S-benzimidazole.

[0049] The invention also provides compounds of Formula V: wherein: Q5 is CH or N ; Q6 is C-R6'or N ; Q7 is C-R60, or N ; R6° and R6'are each independently H, halogen, C, 6 alkyl, trihaloalkyl, or C1-6 alkoxy; provided that when Q6 is C-R61, Q7 is C-R60, and Q5 is CH, then R60 and R6', are not both H.

[0050] In some embodiments, Qu vis N. In further embodiments, wherein Q5 is N, Q6 is also N.

[0051] In some embodiments, Q6 is N. In further embodiments, wherein is Q6 N, Q5 is also N.

[0052] In some embodiments, Q7 is N.

[0053] In some embodiments, Q'is N, Q6 is CR61, and Q7 is CR60. In other embodiments, Q7 is N, Q6 is CR6'and Q5is CH. In further embodiments, Q'is N, Q6 is N and Q7 is CWO. In further embodiments, Q is CH, Q6 is CR61 and Q7 is C60.

[0054] In some embodiments where QS is CH, Q6 is R61 and Q7 is CR60, R60 and R61 are each independently H, Br, Cl, methoxy, methyl or trifluoromethyl. In further such embodiments, Wo is OCH3 and R6'is H, or Wo is CH3 and R6'is H, or R60 Br and R6'is H, or R60 is Cl and R6'is H, or R60 is CF3 and Wl is H, or R60 is Cl and R6'is CH3, or R60 and R61 are both Cl.

[0055] In some embodiments, the invention provides methods of contacting a subject with a compound of one of formulae I, II, III, IV or V, as described herein. In some embodiments, the invention provides methods for treating a patient having a bacterial infection, the methods in general comprising administering to said patient a compound of claim 1, or of one of formulae I, II, III, IV or V. In some embodiments, said subject or patient is a human. Also provided are methods for inhibiting bacterial growth comprising contacting a bacterium with a compound of the invention, e. g. one of compounds of formulae I, II, III IV or V. In some embodiments, said bacterium is S. aureus, E. hirae, S. pyogenes, S. pneumoniae, E. coli, P. vulgaris, K. pneumoniae, P. aeruginosa, C. albicans, E. faecalis, E. faecali, or E. faecium.

[0056] The present invention also provides compositions that include at least one compound of the invention, e. g. of one of formulae I, II, III, IV or V.

[0057] As used herein the term alkyl is intended to have its accustomed meaning of a straight or branched chain hydrocarbon, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, t-butyl, n-pentyl, sec-pentyl, t-pentyl, neopentyl, and the like.

[0058] As used herein the term aryl is intended to mean an aromatic hydrocarbon system for example phenyl, naphthyl, phenanthrenyl, anthracenyl, pyrenyl, and the like. In some embodiments, aryl groups have from 6 to 10 carbon atoms. As used herein, the term arylalkyl (or"aralkyl") is intended to mean an alkyl group that has an aryl group appended thereto, for example benzyl and naphthylmethyl groups. In some embodiments, arylalkyl groups have from 7 to 11 carbon atoms.

[0059] As used herein, the term alkylql (or"alkaryl") is intended to mean an aryl group that has one or more alkyl groups appended thereto, for example a 4-methylphen-1-yl group, or a xylyl group attached through the phenyl ring thereof.

[0060] As used herein, the term heteroaryl means an aryl group that contains one or more ring hetero (i. e. , non-carbon) atoms, which are preferably 0, N or S, more preferably N. In some embodiments, heteroaryl groups are monocyclic or bicyclic, and have up to four ring nitrogen atoms. Examples of some preferred heteroaryl groups include radicals derived from pyrrole, pyrazole, imidazole, triazoles, tetrazole, pyridine, pyrazine, pyndazine, pyrimidine, triazines, quinolines, indoles, benzimidazoles, and the like.

[0061] As used herein, the term heteroarylalkyl is intended to mean an alkylene group that has a heteroaryl group appended thereto, for example a group of formula-CH,-benzimidazol-2-yl.

[0062] As used herein, the term cycloalkyl refers to nonaromatic hydrocarbon ring systems, for example cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, including multiple ring systems such as decahydronaphthalene and adamantane. Cycloalkyl groups can also include points of unsaturation, and therefor also include cyclopentenyl, and cyclohexenyl groups.

[0063] As used herein, the term heterocycloalkyl is intended to mean a group that contains a nonaromatic ring which contains one or more ring hetero (i. e. , non-carbon) atoms which are preferably 0, N or S, more preferably N. Also included in the definition of heterocycloalkyl are moieties that contain exocyclic heteroatoms, for example a cycloalkyl ring having a ring carbon attached to an exocyclic 0 or S atom through a double bond. Also included in the definition of heterocycloalkyl are moieties that having one or more aromatic rings fused (ie. , having a bond in common with) to the nonaromatic heterocyclic ring, for example phthalimidyl, naphthalimidyl pyromellitic diimidyl, phthalanyl, and benzo derivatives of saturated heterocycles such as indolene and isoindolene groups.

[0064] As used herein, the term arylsulfonyl is intended to mean a moiety of formula-S (=O),-aryl, for example phenylsulfonyl. The term heteroarylsulfonyl means a moiety of formula-S (=O),-heteroaryl, for example pyridinesulfonyl.

[0065] As used herein the term aryloxy is intended to mean an aryl group attached through an oxygen atom, for example phenoxy. As used herein, the term aryloxycarbonyl is intended to men a moiety of formula-C (=O)-O-aryl, for example phenoxycarbonyl. As used herein, the term alkoxyalkoxyalkyl is intended to mean a moiety of formula-alkylene-O-alkylene-O-alkyl. As used herein, the term hydroxyalkyl is intended to mean an alkyl group that has a hydrogen atom thereof replaced with OH.

[0066] As used herein, the term alkoxycarbonyl is intended to mean a moiety of formula-C (=O)-O- alkyl.

[0067] As used herein, the term amino refers to NH2. The term halogen includes F, C1, Br and I. The prefix"halo"is intended to denote a halogen atom. The term"perhalo"is intended to refer to the substitution of all hydrogen atoms for halogen atoms. Thus, the term"perhaloaryl"indicated a fully halogenated moiety, for example a pentafouorophenyl radical, and the term"perhaloalkylaryl"would be understood to indicate a full halogenated alkylaryl group, for example a 2,3, 5,6, tetrafluoro-4- trifluoromethyl-phenyl radical.

[0068] In some embodiments, the present invention provides dimeric compounds wherein two benzimidazole core structures are joined, preferably at the 1-position, by a tether. Thus, in certain embodiments, various moieties appended to the 1-position of the benzimidazole core can be appended to a benzimidazole core structure at the 1-position thereof.

[0069] As used herein, the term alkoxy means moieties of formula-O-alkyl. The term arylcarbonyl means a moiety of formula-C (=O)-aryl. The term heteroarylcarbonyl means a moiety of formula- C (=O)-heteroaryl.

[0070] The term arylaminocarbonyl means a moiety of formula-C (=O)-NH-aryl.

[0071] The term cycloalkylaminocarbonyl means a moiety of formula-C (=O)-NH-cycloalkyl.

[0072] The phrase"saturated hydrocarbon fused ring system optionally having an aryl ring fused thereto"is intended to denote saturated hydrocarbon ring systems having up to three fused rings, for example decalin, which can optionally have an aryl ring fused thereto, for example benzo derivatives of cycloalkyl groups. The term arylalkyloxy denotes a fioup of formula-O-alkyl-aryl, for example a benzyloxy group. The term alkylheteroaryl denotes a group of formula-heteroaryl-alkyl, for example a 4- methyl-pyrid-2-yl group.

[0073] The phrase"moiety of formula-OCH2CH2-O-attached to adjacent atoms of"is intended to mean that the-OCH2CH2-O-oxygen atoms are attached to adjacent atoms an indicated moiety (which may be a cyclic group) to form a 6 membered fused ring comprising the-OCHzCH2-O-group and the two atoms to which it is attached.

[0074] The term methylcarbonyl is intended to denote an acetoyl (i. e. , CH3C (=O)-) group-. The term aminoalkyl denotes a group of formula-alkyl-NH2.

[0075] The phrase"branched-or straight-chain polyaminoalkyl"is intended to mean a group of formula - ( (CH2) n-NH) m-H wherein n can be from 1 to 6 and m can be from 2 to about 12, in any one or more of the hydrogens attached to nitrogen can be replaced with a group of formula-((CH2) p-NH) q-H where p is 1 to 6 and q is 1 to 12.

[0076] In some embodiments, compounds of the invention contain simple polyalchol moieties of formula-CH2 (CHOH) 4CH2OH. It is intended that each such group specifically include each individual stereoisomer of such formula, as well as racemic forms of the same.

[0077] In some embodiments, variables Rl5 and R16 together with the nitrogen atom to which they are attached can form a nitrogen heterocycle which can be aromatic or aliphatic, or aliphatic having one or more aromatic rings fused thereto (i. e. , a fused ring derivative). Thus, in some embodiments, Rls and R ; 6 together with the nitrogen atom to which they are attached can form, for example, an N-maleimidyl, N- succinimidyl, N-phthalimidyl, N-naphthalimidyl, N-pyromellitic diimidyl, N-benzopyrrolidinyl, or benzimidazol-1-yl group.

[0078] The term alkylamino is intended to denote a group of formula-NH-alkyl. The term aminoalkylamino is intended to denote a group of formula-NH-alkyl-NH2. The term poly (aminoalkyl) amino is intended to denote a group of formula-NH- (alkyl-NH) x-H where x is from 2 to about 12, and wherein any one or more of the hydrogens attached to nitrogen can be replaced with a group of formula- ( (CH2) p-NH) q-H where p is 1 to 6 and q is 1 to 12.

[0079] The term heterocycloalkylamino is intended to denote a group of formula-NH-heterocycloalkyl.

The term heterocycloalkylalkyl is intended to denote a group of formula alkyl-heterocycloalkyl).

[0080] The term"side chain of a naturally occurring alpha amino acid"is intended to mean the side chain of naturally occurring alpha amino acids, with the exception of glycine, that are known to have the formula H, N-CHR-COOH, where R is the side chain.

[0081] Examples of such naturally occurring amino acids include the 20 so called"essential"amino acids, for example serine and threonine. Further side chains of naturally occurring alpha amino acids can be found in Bikochemistry, 3rd Edition, Matthews, Van Holde, and Ahern, Addison Wesley Longman, San Francisco, CA, incorporated by reference herein in its entirety.

[0082] As used herein, the term alkoxyalkoxyalkyl is intended to denote a group of formula alkyl-O- allcyl-O-alkyl. The term hydroxyalkyl is intended to mean a hydroxyl group that is substituted with up to 3 hydroxy groups. The term heteroarylcarbonyl denotes a moiety of formula-C (=O)-heteroaryl. The term arylaminocarbonyl denotes a moiety of formula-C (=O)-NH-aryl. The term cycloalkylaminocarbonyl denotes a moiety of formula-C (=O)-NH-cycloalkyl.

[0083] The compounds of the present invention and their pharmaceutically acceptable salts are useful in for the treatment of bacterial infections in animal and human subjects. The compounds of the invention can be used alone, or in a pharmaceutical composition containing one or more compounds of the invention, in combination with one or more pharmaceutically acceptable carriers. Thus, in further aspects, the present invention includes pharmaceutical compositions and methods of treating bacterial infections utilizing as an active ingredient the novel compounds described herein.

[0084] In some embodiments, the compounds of the invention can be prepared as amine salts, which can contain any of a variety of pharmaceutically acceptable counterions. Suitable counterions include acetate, adipate, aminosalicylate, anhydromethylenecitrate, ascorbate, aspartate, benzoate, benzenesulfonate, bromide, citrate, camphorate, camphomlfonate, chloride, estolate, ethanesulfonate, fumarate, glucoheptanoate, gluconate, glutamate, lactobionate, malate, maleate, mandelate, methanesulfonate, pantothenate, pectinate, phosphate/diphosphate, polygalacturonate, propionate, salicylate, stearate, succinate, sulfate, tartrate and tosylate. Other suitable anionic species will be apparent to the person skilled in the art.

[0085] Representative examples of compounds of the invention are shown below.

[0086] It is contemplated that the present invention include all possible protonated and unprotonated forms of the compounds disclosed herein.

[0087] The compounds of the invention can be formulated in pharmaceutical compositions which can include one or more compounds of the invention and one or more pharmaceutically acceptable carriers.

The compounds of the invention can be administered in powder or crystalline form, in liquid solution, or in suspension. They may be administered by a variety of means known to be efficacious for the administration of antibiotics, including without limitation topically, orally and parenterally by injection (e. g. , intravenously or intramuscularly).

[0088] When administered by injection, a preferred route of delivery for compounds of the invention is a unit dosage form in ampules, or in multidose containers. The injectable compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain various formulating agents. Alternatively, the active ingredient may be in powder (lyophlllized or non- lyophillized) form for reconstitution at the time of delivery with a suitable vehicle, such as sterile water.

In injectable compositions, the carrier is typically comprised of sterile water, saline or another injectable liquid, e. g. , peanut oil for intramuscular injections. Also, various buffering agents, preservatives and the like can be included.

[0089] Topical applications may be formulated in carriers such as hydrophobic or hydrophilic bases to form ointments, creams, lotions, in aqueous, oleaginous or alcoholic liquids to form paints or in dry diluents to form powders.

[0090] Oral compositions may take such forms as tablets, capsules, oral suspensions and oral solutions.

The oral compositions may utilize carriers such as conventional formulating agents, and may include sustained release properties as well as rapid delivery forms.

[0091] The dosage to be administered depends to a large extent upon the condition and size of the subject being treated, the route and fiequency of administration, the sensitivity of the pathogen to the particular compound selected, the virulence of the infection and other factors. Such matters, however, are left to the routine discretion of the physician according to principles of treatment well known in the antibacterial arts, Another factor influencing the precise dosage regimen, apart from the nature of the infection and peculiar identity of the individual being treated, is the molecular weight of the compound.

[0092] The compositions for human delivery per unit dosage, whether liquid or solid, may contain from about 0.01% to as high as about 99% of active material, the preferred range being from about 10-60%.

The composition will generally contain from about 15 mg to about 2.5 g of the active ingredient; however, in general, it is preferable to employ dosage amounts in the range of from about 250 mg to 1000 mg. In parenteral administration, the unit dosage will typically include the pure compound in sterile water solution or in the form of a soluble powder intended for solution, which can be adjusted to neutral pH and isotonic.

[0093] The invention described herein also includes a method of treating a bacterial infection in a mammal in need of such treatment comprising administering to said mammal a compound of the invention in an amount effective to treat said infection. One preferred method of administration of the antibacterial compounds of the invention include oral and parenteral, e. g. , i. v. infusion, i. v. bolus and i. m. injection.

[0094] Compounds provided herein can be formulated into pharmaceutical compositions by admixture with pharmaceutically acceptable nontoxic excipients and carriers. As noted above, such compositions may be prepared for use in parenteral administration, particularly in the form of liquid solutions or suspensions; or oral administration, particularly in the form of tablets or capsules; or intranasally, particularly in the form of powders, nasal drops, or aerosols; or dermally, via, for example, transdermal patches; or prepared in other suitable fashions for these and other forms of administration as will be apparent to those skilled in the art.

[0095] The composition may conveniently be administered in unit dosage form and may be prepared by any of the methods well known in the pharmaceutical art, for example, as described in Remington's Pharmaceutical Sciences (Mack Pub. Co. , Easton, PA, 1980). Formulations for parenteral administration may contain as common excipients sterile water or saline, polyalkylene glycols such as polyethylene glycol, oils and vegetable origin, hydrogenated naphthalenes and the like. In particular, biocompatible, biodegradable lactide polymer, lactide/glycolide copolymer, or polyoxyethylene-polyox ypropylene copolymers may be useful excipients to control the release of the active compounds. Other potentially useful parenteral delivery systems for these active compounds include ethylene-vinyl acetate copolymer particles, osmotic pumps, implantable infusion systems, and liposomes.

[0096] Formulations for inhalation adrmnistration contain as excipients, for example, lactose, or may be aqueous solutions containing, for example, polyoxyethylene-9-lauryl ether, glycocholate and deoxycholate, or oily solutions for administration in the form of nasal drops, or as a gel to be applied intranasally. Formulations for parenteral administration may also include glycocholate for buccal administration, a salicylate for rectal administration, or citric acid for vaginal administration.

Formulations for transdermal patches are preferably lipophilic emulsions.

[0097] The materials of this invention can be employed as the sole active agent in a pharmaceutical or can be used in combination with other active ingredients, e. g. , other growth factors which could facilitate neuronal survival or axonal regeneration in diseases or disorders.

[0098] The concentrations of the compounds described herein in a therapeutic composition will vary depending upon a number of factors, including the dosage of the drug to be administered, the chemical characteristics (e. g. , hydrophobicity) of the compounds employed, and the route of administration. In general terms, the compounds of this invention may be provided in effective inhlbitory amounts in an aqueous physiological buffer solution containing about 0.1 to 10% w/v compound for parenteral administration.

[0099] Typical dose ranges are from about 1 m a g to about 1 g/kg of body weight per day; a preferred dose range is from about 0.01 mg/kg to 100 mg/kg of body weight per day. Such formulations typically provide inhibitory amounts of the compound of the invention. The preferred dosage of drug to be administered is likely, however, to depend on such variables as the type and extent of progression of the disease or disorder, the overall health status of the particular patient, the relative biological efficacy of the compound selected, and formulation of the compound excipient, and its route of administration.

[0100] As used herein, the term"contacting"means directly or indirectly causing at least two moieties to come into physical association with each other. Contacting thus includes physical acts such as placing the moieties together in a container, or administering moieties to a patient. Thus, for example administering a compound of the invention to a human patient evidencing a disease or disorder associated with abnormal and/or aberrant activity of such proteases falls within the scope of the definition of the term"contacting".

[0101] Compounds of the invention also are useful for in silico studies to determine potential binding to binding pockets present in a variety of bacteria, including those disclosed in the Examples herein. Thus, the present invention further provides methods for determining binding a h t i e s for classes of compounds in silico. In the methods, representations of the compounds of the invention can be used in molecular modeling studies to determine such binding affinities, and therefore aid in the design of therapeutics.

[0102] While the present invention has been described with specificity in accordance with certain of its preferred embodiments, the following examples serve only to illustrate the invention and are not intended to limit the same.

EXAMPLES [0103] Example 1 [0104] General Procedure for Preparation of Benzimidazoles Having Modification to the Phenyl (B) Ring. The general procedure for preparation of benzimidazoles having modification to the phenyl (B) ring is shown in Scheme 1, below: Hop N NH2 1. Coupling N---_c + HOZC NBoc I//+ \\,, oc R/NHZ R/NH a 2. Base OzN /\, NH 3. TFA (-\, N'Boc N/N R H TFA H 5. H2CO C02H 4. Pd/C, HZ I 6. Pd/C, HZ N N N N /NH N N TFA Han H H2N H Scheme 1 [0105] Generally, the steps of the synthesis are: [0106] (1) N-Boc isonipecotic acid (25 g, 109 mmol) was dissolved in DMF (500 mL). HATU (49.5 g, 130 mmol) was added, followed by DMAP (16.0 g, 150 mmol) and DIEA (45 mL, 260 mmol). After the mixture was stirred for 30 minutes, diamine (105 mmol) was added and the resulting reaction mixture stirred overnight. The mixture was concentrated to one fourth of the volume, and poured into brine, extracted with dichloromethane (3x150 mL). The combined organic solution was dried over magnesium sulfate and concentrated too give black oil.

[0107] (2) The black oil was dissolved in ethanol (250 mL) and 2 M sodium hydroxide (250 mL). The mixture was refluxed overnight, cooled to room temperature and poured into saturated citric acid solution. The resulting mixture was extracted with dichloromethane (4x150 mL), and the combined organic solution was dried over magnesium sulfate and concentrated too give a black oil, which was purified on silica gel with ethyl acetate and dichloromethane to give the desired product.

[0108] (3) N-Boc compound (0.02 mmol) was placed in a 2-dram vial with a stir bar, and hydrochloric acid in dioxane (6.0 M, 500 mL) was added. The mixture was stirred at room temperature for 30 minutes to give the corresponding product as precipitate (hydrochloride salt). The mixture was centrifuged, the solution removed using a pipette, and the solid salt was dried under vacuum overnight.

[0109] (4) 4-Nitro Benzimidazole hydrochloride salt (0.02 mmol), prepared by the procedure above was dissolved in methanol (2.0 mL), followed by the addition of palladium on carbon (10%, 5 mg). The resulting mixture was hydrogenated with a hydrogen balloon at room temperature for two hours. The catalyst was filtered off and washed with methanol (3x1. 0 mL). The combined methanol solution was concentrated and dried under vacuum overnight.

[0110] (5) N-Boc benzimidazole (0.04 mmol) was dissolved in formic acid (1.0 mL) and formaldehyde (1.0 mL, 37%), and the mixture was heated at 120°C with an oil bath for 3 hours. Ethyl acetate (5.0 mL) was added, followed by excess solid sodium bicarbonate to neutralize the acid. The mixture was extracted with ethyl acetate (4x5 mL), and the combined organic solution was dried over magnesium sulfate and concentrated too give the crude product, which was purified on silica gel with methanol in chloroform (5%, 10% and 20%, 20%) and 2% NH3H2O and 20% MeOH in CHC13. (1694-5)'H NMR (200 MHz, CDC13) : 8.44 (s, 1H), 8.12 (d, J = 9.0 Hz, 1H), 7.56 (d, J = 9.0 Hz, 1H) 3.60-3. 40 (m, 1H), 3.20-2. 90 (m, 2H), 2.35 (s, 3H), 2.30-2. 00 (m, 5H). LC/MS: M+H+= 261.

[0111] (6) 4-Nitro Benzimidazole from step (4) (0.02 mmol) was dissolved in methanol (2.0 mL), followed by the addition of palladium on carbon (lo%, 5 mg). The resulting mixture was hydrogenated with a hydrogen balloon at room temperature for two hours. The catalyst was filtered off and washed with methanol (3x1. 0 mL). The combined methanol solution was concentrated and dried under vacuum overnight.

[0112] Example 2 General Procedure for Synthesis of Benzimidazoles Having Modification to the Imidazole (A) Ring.

[0113] The general procedure for preparation of benzimidazoles having modification to the imidazole (A) ring is shown in Schemes 2 and 3, below: PPA, 180 C, 2 h - (-N-Boc -- (-NH 1ANH (Basic work-up) R H R H R H 50-80% Scheme 2 [0114] Aryl diamines (1.0 mmol) and isonipecotic acid (129 mg, 1.2 mmol) were ground into powder and well mixed. Polyphosphoric acid (PPA, 1.0 g) was then added. The mixture was heated in an oil- bath at 180°C for two hours. The syrup was cooled to room temperature, and saturated sodium hydroxide was added to make the resulting mixture basic. The mixture was extracted with 30% isopropanol ("i-propanol", or"i-PrOH") in chloroform (5 x 30 mL), and the combined organic solution was dried over magnesium sulfate and concentrated. The crude product was then purified by silica gel chromatography using methanol in chloroform (5%, 10% and 15%).

[0115] Rf= 0. 15 (2% NH3 H2O and 20% i-PrOH in CHC13) ; LCMS: M+H+-202 (2G column) ;'H NMR (200 MHz, CDC13) : 7.55-7. 44 (m, 2H), 7.23-7. 13 (m, 2H), 3.34-2. 98 (m, 3H), 2.83-2. 66 (m, 2H), 2.14- 2.00 (m, 2H), 1.96-1. 72 (m, 2H).

Scheme 3 [0116] (1) Sodium hydride (24 mg, 60%, 1 mmol) was washed with hexane (3 X 1 mL). Anhydrous CH, CN (2.0 mL) was added, followed by N-Boc-4,5-dichlorobenzimidazole (37 mg, 0.1 mmol) portion wise under argon. AAer the slurry was stirred at room temperature for 30 minutes, the ablating halide (0.15 mmol) was added, and the reaction mixture was stirred at room temperature for another 30 minutes (The reaction progress was monitored by TLC). The reaction was cooled with an ice bath, and ice water (2.0 mL) was carefully added. The resulting crude mixture was extracted with ethyl acetate (3 X 10 mL), the combined organic solution was washed with brine (2 X 2 mL) and dried over magnesium sulfate and concentrated. The crude product was then purified by silica gel chromatography using ethyl acetate in hexane (lo%, 20% and 30%).

[0117] Yield: 95%; Rf= 0. 15 (2% NH3H20 and 20% i-PrOH in CHC13) ; LCMS: M+H+= 516 (2CN column) ;'H NMR (200 MHz, CD30D): 7.29 (s, 1H), 7.62 (s, 1H), 7.42-7. 40 (m, 2H), 7.05-6. 85 (m, 2H), 5.5 (s, 2H), 4. 20-4. 02 (m, 2H), 3.25-3. 10 (m, 2H), 2.92-2. 72 (m, 2H), 1.45 (s, 9H), 1.28 (s, 3H).

[0118] (2) N-Boc compound (0.02 mmol) was placed in a 2-dram vial with a stir bar, and hydrochloric acid in dioxane (6.0 M, 500 mL) was added. The mixture was stirred at room temperature for 30 minutes to give the corresponding product as precipitate (hydrochloride salt). The mixture was centrifuged, the solution removed using a pipette, and the solid salt was dried under vacuum over night.

[0119] Yield: 90%; LCMS: M+W= 202 (2G column).

[0120] Example 4 [0121] General Procedure for Preparation of Benzimidazole Derivatives via Solid Phase Synthesis: [0122] The general procedure for preparation of benzimidazole derivatives via solid phase synthesis is shown in Scheme 4, below: X N \ HCl/dioxane ) dBoc -- (_NH HCl X/H X//H O TEA ImS 0 3 x 0 1. RX, NaHxN 0 \ \ j E or KZC03 I ?-N 2. TFA/---/N /N -- X H X X = H, Cl ; R as in Scheme 3 [0123] (1) N-Boc-4, 5-dichlorobenzimidazole (3.0 mg, 10. 7-01) was powdered and treated, with hydrogen chloride in dioxane (6 N) for 2 h. Dioxane was then evaporated and the corresponding hydrochloride salt was dried under vacuum overnight, which was directly used to attach to the Wang resin.

[0124] (2) Wang resin (15.0 g, 5.70 mmol) (Sigma-Aldrich 2000-2001 Catalog, item #47,703-6) was swollen in DMF (120 mL), and carbonyl diimidazole (1. 84 g, 11.4 mmol) was added and the resulting mixture stirred at room temperature overnight. The resin was iltered off and washed successively with DMF (3x30 mL), dichloromethane (3x30 mL), diethyl ether (3x30 mL) and dried overnight. The resulting resin was again suspended in DMF (200 mL), all the benzimidazole hydrochloride salt obtained in step (1) was added, followed by triethylamine (3.0 mL, 21.6 mmol). The resulting mixture was stirred at room temperature overnight. The resin was filtered off and washed successively with DMF (3x30 mL), dichloromethane (3x30 mL), methanol (3x30 mL), diethyl ether (3x30 mL) and dried overnight.

[0125] (3) Benzimidazole on Wang resin obtained in step (2) (100 mg,-0. 0324 mmol) was suspended in DMF (2.0 mL), sodium hydride (60%, 50 mg, 1.25 mmol) was added and the mixture stirred for minutes at room temperature. Alkylating halide (0.0972 mmol) was added, and the mixture was stirred for 2 hours at room temperature. The reaction flask was then cooled with ice bath, and water (100 L) was carefully added to react with the excess sodium hydride. The resin was filtered off and washed successively with water (3X1. 0 mL), DMF (3x1. 0 mL), dichloromethane (3X3=1.0 mL), methanol (3X1. 0 mL), diethyl ether (3X1. 0 mL) and dried overnight.

[0126] (4) The resin obtained in step (3) was suspended in dichloromethane (1.4 mL), trifluoroacetic acid (600 L) was added and the mixture was gently stirred for 30 minutes at room temperature. The resin was then filtered off and washed with dichloromethane (5X 1.0 mL). The dichloromethane solution was dried to give the benzimidazoles as trifluoroacetic acid salt.

[0127] Example 5 [0128] General Procedure for Preparation of Xylene-1-yl Benzimidazole Derivatives via Solid-Phase Synthesis: [0129] The general procedure for preparation of xylene-1-yl benzimidazole derivatives via solid phase synthesis is shown in Scheme 5, below: Scheme 5 [0130] (1) Benzimidazole on Wang resin (2.0 g,-0. 65 mmol) was suspended in DMF (20.0 mL), saturated potassium carbonate (2.0 mL) was added, followed by a, a'-dibromo-p-xylene (860 mg, 3.25 mmol), and the resulting mixture was gently stirred at room temperature for 5 hours. The resin was filtered off and washed successively with water (3X10. 0 mL), DMF (3X10. 0 mL), dichloromethane 3X10. 0 mL), methanol (3X10. 0 d), diethyl ether (3X10. 0 mL) and dried overnight.

[0131] (2) Benzyl bromide on resin obtained instep (1) (1 00 mg, 0.0324 mmol) was suspended in DMF (2.0 mL), and amine (0.324 mmol) was added. The reaction mixture was gently stirred at room for six hours. The resin was filtered off and washed successively with DMF (3x1. 0 mL), dichloromethane (3X1. 0 mL), methanol (3X1. 0 mL), diethyl ether (3x1. 0 mL) and dried overnight.

[0132] (3) The resin obtained in step (1) was suspended in dichloromethane (1.4 mL), trifluoroacetic acid (600 L) was added and the mixture was gently stirred for 30 minutes at room temperature. The resin was then filtered off and washed with dichloromethane (5X1. 0 mL). The dichloromethane solution was dried to give the benzimidazoles as trifluoroacetic acid salt.

[0133] Example 6 [0134] General Procedure for Preparation of Benzimidazole Derivatives Having Urea or Thiourea Functionality.

[0135] The general procedure for preparation of benzimidazole derivatives having urea or thiourea functionality is shown below in Scheme 6: O CI 1. BpMe CI ci 1. 6 N (MBoc------- )-- NBcc cri H 2. H2NNHZ CI /N 0====/ HN NH2 ArNCX X=O, S CI \ N CI \ N -- (NH HCI/dioxane -- (NBoc _NH HCI/dioxane I/-- JdBoc a \ cr \ CI CI O O NH NH HN/Ar HN/Ar eNH eNH x x ticneme 6 [0136] (1) To a solution of N-Boc benzimidazole (200 mg, 0.54 mmol) in DMF (2.0 mL) was added sodium hydride (60%, 65 mg, 1.40 mmol) portion-wise, and the resulting mixture was stirred for 20 minutes. Methyl bromoacetate (214 mg, 1.40 mmol) was then added and the reaction mixture was stirred at room temperature for 2 hours. The reaction flask was then cooled with ice bath, and water (100 AL) was carefully added to react with the excess sodium hydride. The resulting mixture was then diluted with ethyl acetate (30 mL), washed with brine (5x2 mL) and dried over magnesium sulfate. The crude product was purified on silica gel with 50% ethyl acetate in hexane to give 210 mg (88%) of the desired methyl ester. (1815-41) ; Rf = 0.40 (AcOEt: Hexane = 1 : 1); LCMS: M+H+ = 442 (2CN column) ; +H NMR (200 MHz, CDC13) : 7.80 (s, 1H), 7.38 (s, 1H), 4.95 (s, 2H), 4.40-4. 15 (m, 3H), 3.78 (s, 3H), 3.00-2. 78 (m, 2H), 2.10-1. 80 (m, 6H).

[0137] (2) To the solution of methyl ester (160 mg, 0.36 mmol) obtained form step 1) in methanol (5.0 mL) was added hydrazine (46.0 mg, 1.44 mmol), and the resulting mixture was stirred at room temperature for three hours. The reaction was then concentrated and the crude product purified on silica gel with 10% methanol in chloroform to give the corresponding acyl hydrazine (152 mg, 92%) Rf= 0.15 (MeOH :, CHC1, = 1 : 1).

[0138] (3) To the solution of acyl hydrazine (30 mg, 0.068 mmol) obtained from step 2) in chloroform (2.0 mL) was added isocynate or isothiocynate (0.068 mmol) at OC, and the reaction was worked up and stirred at room temperature for one hour. TLC and LCMS indicated complete conversion of the starting material and the product has more than 90% purity.

[0139] (4) Urea or thiourea (0.02 mmol) obtained in step 3) was powdered and treated with hydrogen chloride in dioxane (6 N) for 2 h. Dioxane was then evaporated and the corresponding hydrochloride salt was dried under vacuum overnight. LCMS indicated complete conversion of starting material and desired product has over 90% purity.

[0140] Example 7 [0141] General Procedure for Preparation of Benzimidazole Derivatives Having Hydrazone Functionality [0142] The general procedure for preparation of benzimidazole derivatives having hydrazone functionality is shown below in Scheme 7: ci N ci N 04 lq-Boc N CI CI O O HN-NH2 HAN-TZAR | HCI/dioxane cl N NH OHCI N J ci tzar HN-N Ar Scheme 7 [0143] (1) To the solution of acyl hydrazine (20 mg, 0.045 mmol) and aldehyde (0.0475 mmol) in THF (1.0 mL) was added catalytic amount of p-tolunesulfonic acid. The reaction mixture was stirred at room temperature for two hours, and dried. TLC and LC/MS indicated complete conversion of starting material and desired product has over 90% purity. [0144] (2) Half of the N-Boc hydrazone obtained above (0.023 mmol) was powdered and treated with hydrogen chloride in dioxane (6 N) for 30 min. Dioxane was then evaporated and the corresponding hydrochloride salt was dried under vacuum overnight. LC/MS indicated complete conversion of starting material and desired product has over 90% purity.

[0145] Example 8 [0146] General Procedure for Preparation of Benzimidazole Derivatives Having Sulfonamide Functionality [0147] The general procedure for preparation of benzimidazole derivatives having sulfonamide functionality is shown below in Scheme 8: Scheme 8 [0148] (1) To the solution of acyl hydrazine (20 mg, 0.045 mmol) and pyridine (6.0 mg, 0.072 mmol) and DMAP (catalytic) in 30% THF in CH2CI2 was added sulfonyl chloride (0.0475 mmol). The reaction mixture was stirred at room temperature overnight, and dried. TLC and LCMS indicated complete conversion of starting material and desired product has over 90% purity.

[0149] (2) Half of the N-Boc sulfonamide obtained above (0.023 mmol) was powdered and treated with hydrogen chloride in dioxane (6 N) for 2 h. Dioxane was then evaporated and the corresponding hydrochloride salt was dried under vacuum overnight. LC/MS indicated complete conversion of starting material and desired product has over 90% purity.

[0150] Example 9 [0151] General Procedure for Preparation of Benzimidazole Derivatives Having Substituted Alkyl Functionality [0152] The general procedure for preparation of benzimidazole derivatives having substituted alkyl functionality is shown below in Scheme 9. While the procedure is illustrated for phthalimidyl-alkyl functionalization, the procedure is generally applicable to the preparation of benzimidazoles having a wide variety of heterocycles attached through alkyl spacers. cl N ci N NaH, DMF CI cNBoc Boc R R X 0 0 na nu Cl NH ci N O CI N NH CI N c. (D, Boc X O NaH XNH R N O I N O R = e. g. 1, 6-diiodohexane Scheme 9 [0153] To the solution of N-Boc benzimidazole (1 10 mg, 0.30 mmol), and diiodohexane (1.50 mmol) in DMF (3.0 mL) was added sodium hydride (60%, 120 mg, 3.0 mmol) portion-wise. After the reaction mixture was stirred at room temperature for 20 minutes, the reaction flask was then cooled with ice bath, and water (100 mL) was carefully added to react with the excess sodium hydride. The resulting mixture was then extracted with ethyl acetate (3x10 mL) and the combined organic solution was washed with brine and dried over magnesium sulfate. The crude product was purified on silica gel with 50% ethyl acetate in hexane.

[0154] Rf= 0.55 (AcOEt: Hexane = 1: 1); LCMS: M+H+ = 567 (2CN column) ;'H NMR (200 MHz, CDC13) : 7.75 (s, 1H), 7.35 (s, 1H), 4.35-4. 15 (m, 1H), 4.20-3. 95 (m, 2H), 3.20-3. 00 (m, 2H), 2. 00-1. 20 (m, 12 H).

[0155] (2) To the solution of iodo benzimidazole obtained in step 1) (0.044 mmol), phthalimide or amide (0.066 mmol) in DMF (1.5 mL) was added potassium carbonate (11 mg, 0. 066mmol). After being stirred at room temperature overnight, the reaction mixture was diluted with 50 mL of ethyl acetate, washed with brine (5x2 mL), dried over magnesium sulfate and concentrated. The crude product was purified on silica gel with 50% ethyl acetate in hexane.

[0156] Rf= 0.40 (AcOEt: Hexane = 1 : 1); LCMS: M+H+= 631 (2CN column) ;'H NMR. (200 MHz, CDC13) : 8.70-8. 55 (m, 3H), 8.10-8. 00 (m, 1H), 7.78 (s, 1H), 7.41 (s, 1H), 4.40-4 : 00 (m, 3H), 3.82-3. 70 (m, 2H), 3.08-2. 80 (m, 2H), 2.00-1. 20 (m, 12 H).

[0157] Example 10 [0158] Biological Evaluation of Compounds [0159] Compounds were evaluated for in vitro antibacterial activity (referred to MIC, the minimum concentration inhibiting fungal cell growth) against S. aureus and E. coli. Table 2 shows the in vitro inhibitory activity of selected benzimidazoles against additional pathogenic strains of bacteria (four Gram positive strains, four gram negative strains and one yeast strain). The assays are carried out in 150 mL volume in duplicate in 96-well clear flat-bottom plates. The bacterial or yeast suspension from an overnight culture growth in appropriate medium is added to a solution of test compound in 2.5% DMSO in water. Final bacterial or yeast inoculum is approximately 102-103 CFU/well. The percentage growth of the bacteria or yeast in test wells relative to that observed for a control well containing no compound is determined by measuring absorbance at 595 nm (A595) after 20-24 hours at 37°C (bacteria) or 40-48 hours (yeast) at 25°C. The MIC is determined as a range of concentration where complete inhibition of growth is observed at the higher concentration and bacteria/yeast cells are viable at the lower concentration. Ampicillin and tetracycline are used as antibiotic positive controls for bacterial MIC assays. Amphotericin B is used as a positive control for yeast MIC assay.

[0160] Example 11 [0161] Preparation of 5, 6-dichloro-2-piperidin-4-yl Benzimidazole Derivatives [0162] Using the procedures described above, the following 5, 6-dichloro-2-piperidin4-yl- benzimidazole derivatives prepared according to Scheme 11 as described below: Scheme 11 [0163] Treatment of commercially available 4, 5-dichloro-1, 2-phenylenediamine (1) and N-Boc- isonipecotic acid (2) with EDC in the presence of catalytic amount of DMAP led to the formation of the corresponding amide. The crude mixture was then refluxed in aqueous sodium hydroxide solution to give cyclized intermediate 3, which was reacted with various alkyl, benzyl and aryl halides to give 4a-41.

Treatment of compound 4g with various amines or nitrogen-containing heterocylces provided 6a-x.

Deprotection of the Boc group with anhydrous hydrogen chloride (HC1, 4.0 M) in dioxane at room temperature for 30 minutes led to formation of benzimidazoles 7a-x. In a similar manner, 4a-41 were treated with hydrogen chloride to give benzimidazoles 5a-i.

[0164] This procedure was employed to prepare the following compounds: Compound Compound [0165] These compounds were evaluated for their ability to inhibit S. aureus and E. coli growth and bacterial Transcription/Translation according to the procedures described herein. In addition, all benzimidazoles were also screened for their ability to inhibit bacterial translation and transcription using a combined assay. Several compounds (e. g. 7c, 7d, 7j-l, l0a-b) were found to posses low micromolar ICso values. Since most of the ICso values are much higher than the corresponding MICs for S. aureus and E. coli., it's unlikely that the antibacterial activities are the direct results of bacterial transcription/translation inhibition. However they could be a result from the combination of multiple mechanisms of action, including transcription/translation inhibition. The results are presented in Table 1 below: Table 1 Inhibitory Effects of Benzimidazoles on S. aureus and E. coli Growth and Bacterial Transcription/translation. Compounds S. aureus E. Coli T/T ICso MIC MIC MIC (AM) (AM) (µM 5a >100 >100 >100 5b >100 >100 >100 5c 75. 00 94 >100 5d 75.00 86 >100 5e >100 >100 >100 5f >100 >100 >100 5g 52. 00 >100 >100 5h >100 >100 >100 5i >100 >100 >100 7a 6-12 12-25 100 7b 3-6 6-23 >100 7c 6-12 12-25 12 7d 12-25 50-100 20 7e 6-12 25-50 50 7f 12-25 25-50 >100 Compounds S. aureus E. Coli T/T ICso MIC MIC MIC (µM) (µM) (µM) 7g 6-12 12-25 100 7h 12-25 50-100 >100 7i 6-12 6-12 >100 7j 6-12 50-100 10 7k 3-6 25-50 10 71 3-6 12-25 10 7m 6-12 12-25 >100 7n 12-25 12-25 >100 To6-1212-2525 7p 12-25 12-25 >100 7q 7r 6-12 12-25 >100 7s 6-12 12-25 >100 7t 6-12 50-100 60 7u 6-12 25-50 >100 7v >100 >100 12-25 7w 12-25 12-25 >100 7x 6-12 6-12 >100 10a 25-50 25-50 25 10b 12-25 12-25 35 10c6-1212-25 >100 12 3-6 6-12 >100 Example 12 [0166] Preparation of Benzimidazole Dimers [0167] Several benzimidazole dimers were prepared according to the procedures of Schemes 12 and 13, below, and evaluated for their antibacterial activity.

10a : n=1, 99% 9a: n=l, 61% 10b : n=3, 98% 9b: n=3,65% 10c : n=4, 99% 9c: n=4, 62 % Scheme 12 [0168] Synthesis of Benzimidazole Dimers 10a-c [0169] Reagents and conditions: a) NaH, DMF, 0°C, 2 h, 1,3-diiodopropane, 8a, 54%; 1, 5diodopentane, 8b, 66%; 1,6-diodohexane, 8c, 70%; (b) 3, NaH, DMF, 0°C, 2 h, 61 % for 9a, 65% for 9b, 62% for 9c ; (c) 6 MHC1/dioxane, 25°C, 2h.

Scheme 13 [0170] Synthesis of Benzimidazole Dimer 12 [0171] Reagents and conditions: a) 0.5 equivalents ca-dibromo-p-xylene, NaH, DMF, 0°C, 2 h, 56%; (b) 4 M HCl / dioxane, RT, 2h, 98%.

[0172] Mono alkylation of benzimidazole with diiodo alkanes provided intermediates 8a-c, which were then reacted again with 3 to provide the corresponding dimers 9a-c. The Boc protecting groups were cleanly removed using hydrogen chloride in dioxane to give the final dimer analogs 10a, 10b and 10c in almost quantitative yield (Scheme 12). The xylene-spaced dimer 12 was prepared from intermediate 3 by first reacting 0.5 equivalents of a, a-dibromo-p-xylene (11), followed by deprotection using hydrogen chloride (Scheme 13).

[0173] The inhibitory effects of benzimidazole dimers on S. aureus and E. coli growth and bacterial transcription/translation are shown in Table 2 below.

Table 2 Inhibitory Effects of Benzimidazole Dimers on S. aureus and E. coli Growth and Bacterial Transcription/Translation. Compounds S. aureus E. coli T/T ICso MIC MIC MIC (, M) M) (µM 10a 25-50 25-50 25 10b 12-25 12-25 35 10c 6-12 12-25 >100 12 2-6 6-12 >100 [0174] To test effectiveness of benzimidazoles of Examples 11 and 12 against other bacteria, the active compounds from the preliminary screening were screened against additional four strains of Gram positive and four strains of Gram negative bacteria, and the results are shown in Figure 1. These compounds exhibited higher potencies against Gram position bacteria (S. aureus 13709, E. hirae 29121, S. pyogenes 49399, and S. pneumoniae 6303) as compared to Gram-negative bacteria (E. coli 25922, P. vulgaris 8427, K pneumoniae 1338, P. aeruginosa 25416). Several benzimidazoles (7b, 7g-k, 12) showed particularly strong activity against E. hirae. To study the selectivity, these compounds were also screened against yeast cell line C. albicans 10231. These compounds are clearly much less effective as compared to their inhibition of bacterial growth.

[0175] Since entercocccus infection is emerging, and presents a major threat to human health, compounds were screened against seven additional clinically important strains of entercocccus and the results are shown in Figure 2. As mentioned previously, all these selected compounds are very effective against E. Hirae_ATCC_29212. In some embodiments, compounds of the invention are active against other strains, e. g. the six compounds (7a, 7b, 7x, lOb, 10c, 12) exhibited strong inhibitory activities against all eight strains.

[0176] Example 13 [0177] Preparation of Alkyl Spaced Benzimidazole Derivatives [0178] Several alkyl spaced benzimidazole derivatives were prepared according to the procedures of Scheme 14, below: O NOz O O O zN I. N _ N_ _ N_ _'N_ _ //ON / z e O f O g O h O Scheme 14 oo ' . Schème 14 0 O O. I k i i VS N "LL H Nf/> ZC > » 3 ¢$N N N H N C ! 0 m P o q 9 Scheme 14, Continued [0179] Alkyl Spaced Benzimidazole Derivatives [0180] Reagents and conditions: a) EDC, DMAP; b) NaOH, H2O, 65% over 2 steps; c) ICHn (CHn) nCHnI, NaH or K2CO3 ; d) ArH, NaH or K2CO3 ; e) 4.0 M HC1/dioxane, CH2C12, 24°C, 0.5 h, >95%.

[0181] 4, 5-dichloro-1, 2-dianiline (1) reacted smoothly with N-Boc-isonipecotic acid to give the corresponding amide, which cyclized upon treatment with sodium hydroxide to give benzimidazole 5.

Reaction of 5 with different diiodides furnished 6-10 in good yields. A variety of nitrogen-containing heterocycles were then introduced by simple alkylation to give the target molecules 11-15.

[0182] Example 14 [0183] Preparation of Hydrazone Benzimidazole Derivatives. Several hydrazone benzimidazole derivatives were prepared according to the procedures of Scheme 15 below: Ar = Scheme 15 Scheme 15, Continued [0184] Preparation of Hydrazone Benzimidazole Derivatives [0185] Reagents and conditions: a) NaH (3.0 equiv), BrCH2CO2Me (1.2 equivalents), DMF, 25°C, 0.5 h, 92%; b) HzNNHz (5.0 equivalents), DMF, 25°C, 2.0 h, 98%; c) ArCHO (1.02 equivalents), CH2C12, 25°C, 0.5 h, >95%; d) 4.0 M HCVdioxane, CH2C12, 25°C, 0.5 h, >95%.

[0186] Acylhydrazide 17 was synthesized as a key intermediate for the combinatorial generation of benzimidazoles. Since the acylhydrazide could serve as both a hydrogen donor and acceptor to add additional contacts with the target, analogs based on 17 could be potentially more potent than the parent benzimidazoles. Acylhydrazide 17 was easily prepared from 3 in gram quantity in excellent overall yield from 5 by alkylation with methyl a-bromoacetate followed by a nucleophilic displacement of the methoxy group. Many derivatives could be easily synthesized from 17 without the need of vigorous purification. The first series of analogs has the general structure 18 and was prepared by simply reacting 17 with different aldehydes followed by the removal of the Boc protecting group with hydrogen chloride.

All the benzimidazole analogs obtained this way have more than 95% purity based on TLC and LC/MS analysis and were used directly for MS-based screening and antibacterial assays.

[0187] Example 15 [0188] Preparation of Hydrazine Benzimidazole Derivatives [0189] Several hydrazine benzimidazole derivatives were prepared according to the procedures of Scheme 16 below : Scheme 16 Scheme 16, Continued [0190] Hydrazine Benzimidazole Derivatives [0191] Synthesis of Benzimidazoles 19a-19y. Reagents and conditions: a) For 19a-o, RNCO or RNCS (1.05 equivalents), CH2CH2, 25°C, 0.5 h, >95%.; for 19p-y, RSO2CI (1.05 equivalents), Et3N (1.5 equivalents), DMAP (catalyst),-95% ; b) 4.0 M HCI/dioxane, CH2C12, 25°C, 0.5 h, >95%.

[0192] A variety of isocynates, isothiocyanates and sulfonyl chlorides were reacted with acyl hydrazide 17, and the corresponding ureas, thioureas and sulfonates were obtained in excellent yields and purity as shown in Scheme 16. The resulting N-Boc protected intermediates were treated with hydrogen chloride to give the corresponding products of general structure 19 in almost quantitative yields and more than 95% purity. These products were used directly for antibacterial assays.

[0193] Example 16 [0194] Inhibitory Effects of Benzimidazoles on S. aureus and E. coli Growth and Bacterial Transcription/Translation For Compounds of Examples 13-15.

[0195] Table 3 shows the in vitro antibacterial activity (referred to as MIC, the minimum concentration inhibiting fungal cell growth) of the benzimidazoles against S. aureus and E. coli. Figure 3 shows the in vitro inhibitory activity of selected benzimidazoles against additional pathogenic strains of bacteria (four Gram positive strains, four Gram negative strains and one yeast strain). The assays are carried out in 150 AL volume in duplicate in 96-well, clear, flat-bottom plates. The bacterial or yeast suspension from an overnight culture growth in appropriate medium is added to a solution of test compound in 2.5% DMSO in water. Final bacterial or yeast inoculum is approximately 102-103 CFU/well. The percentage growth of the bacteria or yeast in test wells relative to that observed for a control well containing no compound is determined by measuring absorbance at 595 nm (As9s) after 20-24 hours at 37°C (bacteria) or 40-48 hours (yeast) at 25°C. The MIC is determined as a range of concentration where complete inhibition of growth is observed at the higher concentration and bacteria/yeast cells are viable at the lower concentration. Ampicillin and tetracycline are used as antibiotic positive controls for bacterial MIC assays. Amphotericin B is used as a positive control for yeast MIC assay.

Table 3 Inhibitory Effects of Benzimidazoles on S. aureus and E. coli Growth and Bacterial Transcription/Translation. Compound S. aureus E. coli MIC (mM) MIC (mM) 6 >100 >100 11 >100 >100 12 >100 >100 13a >100 >100 130 >100 >100 Compound S. aureus E. coli MIC MIC 13p >100 >100 14a 12-50 25-50 14b 6-12 >100 14c 50-100 25-50 14d 12-25 50-100 14e >100 50-100 14f >100 >100 14 25-50 50-100 14h >100 >100 14i >100 >100 14j >100 >100 14k >100 >100 141 >100 >100 141 50-100 50-100 14q >100 >100 15a >100 >100 18a 12-25 >100 18b 12-25 >100 18c 25-50 >100 18d 25-50 >100 18e 25-50 >100 18f 25-50 >100 18h 25-50 >100 18i 25-50 >100 18'3-6 >100 18k 50-100 >100 181 50-100 >100 18m 6-12 >100 18n >100 >100 18o >100 >100 18p >100 >100 18q 25-50 >100 19b 12-25 25-50 19c 25-50 >100 19c 12-25 25-50 19d 25-50 50-100 19e 50-100 >100 19f 6-12 12-25 19g 6-12 12-25 19h 6-12 25-50 19h 50-100 50-100 19i 25-50 50-100 19j 50-100 >100 19k >100 >100 191 >100 >100 19m >100 >100 190 25-50 25-50 19 12-25 >100 19 50-100 >100 19r >100 >100 19s >100 >100 Compound S. aureus E. coli MIC MIC 19t >100 >100 19u >100 >100 19v >100 >100 19w >100 >100 19x >100 >100 19y >100 >100 [0196] Examplel7 [0197] Synthesis of Piperidine Modified Benzimidazoles and their Binding Affinities For E. coli 16S A- site [0198] It has been established that the 16S A-site is involved in bacterial translation, and that aminoglycosides are known to bind to the region. Thus, the bacterial 16S A-site represents a prime target for discovering antibacterial agents, and much work has focused on the modification of-the natural aminoglycosides. In accordance with the present invention, several small molecules were synthesized that were shown to bind to the 16S A-site of E. coli ribosome RNA. These are shown below in Scheme 17. 0 OzN NH2 EDC OzN NH. 1 + HOZC- (NBoc----I"Y\\ii I NHZ DMAP N (i2 NBoc 2 3a 4a NaOH, H2O 166% 02N TFA, CH2C'2 66% H 02N N 95%/'/N 6a H Pd/C, H2 1. HCI soo/2. Ae20, TEA 87%, 2 steps HZN \ I/'VNH QxN \ O N 1 H 7a Scheme 17 [0199] Synthesis of piperidine-modified benzimidazoles and their binding affinities for E. coli 16S A- site [0200] MS-based competition experiments were used to determine the binding location of 1 to target RNA. Glucosamine is the A-ring of paromomycin that is known to bind to the target RNA and inhibits bacterial translation. Data suggest that 1 and glucosamine compete for the same binding site on the target RNA. Since glucosamine binds to the target RNA at the same location as it is in paromomycin binding, it is believed the 1 binds to the desired RNA decoding region and could potentially inhibit bacterial translations.

[0201] After establishing the binding of 1 to the correct location on the target RNA, systematic chemical modifications were carried out to study the structure activity relationship (SAR) around the benzimidazole. The synthesis of compound 1 and pipendine-modified benzimidazoles are shown above in Scheme 17. Treatment of commercially available 5-nitro-1, 2-dianiline (2) and N-Boc-isonipecotic acid (3) with EDC in the presence of catalytic amount of DMAP led to the formation of the corresponding amide as a mixture of two regioisomers (4a, b). The crude mixture was then refluxed in aqueous sodium hydroxide solution for 2 hours gave the cyclized intermediate 6a. Treatment of compound 5 with 20% TFA in dichloromethane at room temperature for 30 minutes led to the formation of compound 6, which was then hydrogenated over Pd/C to give 1. MS-based assay suggested that an electron withdrawing nitro group at C5 position (6) is preferred over the corresponding amino group (1), and almost doubled the binding affinity for the target 16S RNA A-site. Thus, in order to establish the SAR of the benzimidazoles, a series of pipendine-modified analogs with a nitro substitution at the 5- position (6a-7b) were synthesized by following the same synthetic route and all these compounds were screened against 16S RNA A-site. A basic NH group with the correct orientation in this region is required to maintain the affinity, since acetylation (7a), methylation (7b), removal (6b) of the free NH group and unsaturation of the piperidine ring (6c) all diminished the binding affinity. The NH group is critical, presumably because it forms a hydrogen bond with the negatively charged phosphate in the RNA backbone. The extended piperidine analogs (6g-6t) showed improved affinities, which, are believed to better orient the NH groups to contact the phosphate backbone.

[0202] Example 18 [0203] One-Pot Synthesis of Benzimidazoles and Their Binding Affinities for E. coli 16S A-site [0204] A series of piperidine substituted benzimidazoles were prepared according to the one-pot procedure of Scheme 18: Scheme 18 Scheme 18, Continued [0205] A series of benzimidazole-modified analogs were prepared as shown above. The procedure required the simple heating of a suitable 1,2-dianiline (8) with isonipecotic acid (9) in the presence of polyphosphoric acid. The free benzimidazoles were then isolated in good to excellent yields after basic work-up. From a MS-based assay, it was established that 1) electron donating groups such as, NH, and OMe reduced the affinities (1, 10a) ; 2) Certain hydrophobic substitutions such as a methyl (lOb), bromo (10c) and chloro (lOd) are tolerated; 3) Insertion of nitrogen atoms into the aromatic moiety (l0i-lOk), particularly at the C4 position (lOj) reduced activities; and 4) Electron-withdrawing groups enhanced the affinities (ICe-lOg).

[0206] Example 19 [0207] Synthesis of Additional N-1 Substituted Benzimidazoles and Their Binding Affinities for E. coli 16S A-site [0208] A further series of N-1 benzimidazole analogs were prepared according to Scheme 19 below: >80% overall yield ; >95% purity Scheme 19 Scheme 19, Continued [0209] Solid-phase synthesis of N1 substituted benzimidazoles and their binding affinities for E. coli 16S A-site [0210] This series of compounds was efficiently synthesized by employing the solid-phase chemistry shown in Scheme 19. Wang resin was first converted into an imidazole carbonyl derivative, which was then allowed to react with compound lOg to give common intermediate 32. Compound 11 reacted readily with a variety of alkylating or acylating reagents to give the corresponding alkyl or acyl products, which after removal of Boc group with 50% TFA in dichloromethane led to the desired N-1 substituted analogs in excellent yields and purity.

[0211] Example 20 [0212] Synthesis of Additional Benzimidazole Dimers and MIC and their Transcription/Translation activity [0213] This series of assays is known to those of skill in the art, and other assays may be substituted therefore without deviating from the spirit and scope hereof. The DNA template, pBestLucTM (Promega), is a plasmid containing a reporter gene for firefly luciferase fused to a strong tac promoter and ribosome binding site. Messenger RNA from 1 Ag pBestLuc is transcribed and translated in E. coli S30 bacterial extract in the presence or absence of test compound. Compounds are tested in a black 96 well microtiter plate with an assay volume of 35 p. L. Each test well contains: 5 yL test compound, 13 AL S30 premix (Promega), 4 L 10X complete amino acid mix (1 mM each), 5 AL E. coli S30 extract and 8 AL of 0.125 , ug/yL pBestLuc. The transcription/translation reaction is incubated for 35 minutes at 37°C followed by detection of functional luciferase with the addition of 30 uL LucLitelM (Packard). Light output is quantitated on a Packard Topcount.

[0214] The assays are carried out in 150, uL volume in duplicate in 96-well clear flat-bottom plates.

The bacterial suspension from an overnight culture growth in appropriate medium is added to a solution of test compound in 4% DMSO in water. Final bacterial inoculum is approximately l0s-106 CFU/well.

The percent growth of the bacteria in test wells relative to that observed for a well containing no compound is determined by measuring absorbance at 595 nm (As95) after 24 h. The MIC is determined as a range of single compound where the complete inhibition of growth is observed at the higher concentration and cells are viable at the lower concentrations. Both ampicillin and tetracycline are used as antibiotic-positive controls in each screening assay for S. pyogenes, E. coli, S. auras, E. faecalis, K. pneumoniae and P. vulgaris. Ciprofloxacin is used as an antibiotic positive control in each screening assay for P. aeruginosa.

[0215] Biological activities of selected compounds according to the present invention were assayed according to techniques known in the art.

[0216] A series of 2-aminobenzimidazole dimers were synthesized according to the procedure described in Example 11. A series of 5-and 6-substituted-2-aminobenzimidazoles also were synthesized, and all were evaluated for biological activity. Tables 4-7 report MIC and transcription/translation activity for the dimer compounds by the outlined procedure. Tables 8 and 9 report the MASS screening of 2- aminobenzimidazoles against the AgIIa HCV-IRES target. The reported selectivity was determined by mass spectral analysis of any associations and provides information about the relative binding affinities.

Table 4 Structure MIC, E. coli MIC, S. aureus Transcription/ (yM) (AM) Translation ICso ci 1 12-25 12-25 >100 ,--NYN NEZ C'raj C \ N />--NH2 C1 N 25-50 75-100 25 r--N y N r-j NH2 r_JrN\ OzN Br 25-50 75-100 >100 Ny N NYN rj-N\ N Hz N I/NH2 Br N > >100 >100 6-12 _Ns N Zu r\ \ N N XCO2Me >200 >200 >100 nu NU2 N NEZ (/6NH2 N\ NH2 Structure MIC, E. coli MIC, S. aureus Transcription/ ( (M) Translation ICso cF3 25-50 50-100 >100 N N Hz N Hz N N NHz F3C N N >200 >200 >200 nu N rj_N\ . MHz N rw NEC N >50 >50 >100 N\ N Hz C N Table 5 Structure MIC, E. coli MIC, S. aureus Transcription/ (/tM) (, am) Translation Q icso (/tM) . /NH., EX N X x= - N (CH3) 2- 50-75 25-50 20 - CHz->100 >100 >100 - 0->100 >100 >100 Table 6 Structure MIC, E. coli MIC, S. aureus Transcription/ (gA4) (IAM) Translation < ICso (Fw X NH2 N N cul N Cl N x= - N (CH3)- 25-50 75-100 25 -CH2->100 >100 >100 -O-6-12 >100 >100 Table 7 Structure MIC, E. coli MIC, S. aureus Transcription/ x (IJM) (AM) Translation r ICso (hM) x N X / X x H >100 >100 >100 Cl >100 >100 >100 Table 8 Structure Target Ligand % % Selectivity Concentration Concentration Complex Dimer IM) g _C_CN R= R= H 2. 5 50 125 35 4. 30 H 2. 5 7. 5 33 3 7. 49 CH3 2. 5 50 136 31 6. 55 CH3 2.5 7.5 26 0 8.44 OCH3 2.5 50 286 39 10.67 OCH3 2.5 7.5 52 4 12. 89 O (CH2) 3N (CH3) 2 2.5 50 862 72 96.88 O (CH2) 3N (CH3) 2 2.5 7.5 46 0 34.53 Table 9 Structure Target Ligand % % Selectivity Concentration Concentration Complex Dimer u N R-N R N R-- H 2. 5 50 125 35 4. 30 H 2. 5 7. 5 33 3 7. 49 CHzNHz 2. 5 7. 5 14. 7 0 2. 0 CH2NH2 2. 5 50 109 80 2. 3 [0217] Example 21 [0218] Synthesis of Further Benzimidazole Dimers and MIC and their Transcription/Translation activity [0219] Additional dimers of benzimidazoles according to embodiments of the present invention were prepared as per Scheme 20: H2N RtuR2 Ri F HNN, -a 1 CaC03 HN N02 rl, 14h g la, R, =R2=H r lb, Rl=H, R2=CI (\ le, Rt=H, R2=CF3 ld, Rl=H, R2=Br Rl NH le, Rt=H, RZ=CN If, Rt=H, RZ=COZMe R NO ex lg, Rt=Me, Rz=H lh, Rl--Me, R2=H 3a-i H2balloon li, Rì=R2=CI X Rt R2 R1 R2 HN NH2 CNBr NHz E 20-30% (3 steps) Ri N I nu2 R/'-N'RNH, Rz N Rz NHz 5a-i 4a-i Scheme 20 [0220] Synthesis of 2-aminobenzimidazole dimers 5a-i was accomplished by SNAR displacement of fluorine in la-i with amine 2 to provide nitroanilines 3a-i. The nitroanalines 3a-I were reduced using Rainey Nickel to provide the corresponding phenylene diamines 4a-I, which were cyclized using CNBr to provide the desired 2-aminobenzimidazle dimers 5a-i in moderate yield (Scheme 20,20-30% overall yield over three steps). All dimers prepared were purified (>95% purity) by reverse phas preparative HPLC prior to biological testing.

[0221] Synthesis of 5i : 3, 3'-diamino-N-methyldipropylamine 3 (2mmol, 0. 28g) was added to a mixture of 1, 2-dichloro-3-fluoro-4-nitrogenzene li (0.26 mL, lnumol) and CaC03 (0.3 g, 4 mmol) in CH2C12 (2 mL). After stirring for 12 at rt, the reaction was filtered through celite and the filter bed was washed with additional CH2C12. The filtrate was concentrated under reduced pressure to provide 2i as a yellow solid that was used without any further purification. Crude 2i (1 mmol) obtained above was hydrogenated using Raney Nickel (catalytic) and H2 gas (balloon) in EtOH (10 mL) for 12 h at rt. The reaction was filtered through celite and the filter bed was washed with additional EtOH. The filtrated was concentrated under reduced pressure to provide phenylenediamine 3i as a dark oil that was used without any further purification. Crude phenylenediamine 4i (1 mmol) obtained above was dissolved in EtOH (5 mL) and treated with CNBr (3 mmol, 0.31 g). The reaction was stirred for 12 h at rt after which it was basified using 4M NaOH (ph > 12). The aqueous layer was extracted with EtOAc (2X) and the combined organic layers were washed with brine, dried (MgS04) and concentrated under reduced pressure. Crude dimer 5i was purified by reverse-phase preparative HPLC using a Gilson 215 system with a Waters PrepPak (25 X 100mm) C18 column, eluting with a linear gradient of 20-40% mobile phase B for 30 min. with a flow rate of 5 mL/min (A = 0.1% TFA in water, B = 0.1% TFA in acetonitrile). 5i :'H NMR (300 MHz, DMSO-d6) 6 9.03 (s, 4H), 7.97 (s, 2H), 7.68 (s, 2H), 4.18 (t, 4H, J = 6. 9), 3.19 (m, 4H), 2.78 (s, 3H), 2.09 (m, 4H). LCMS: LC 2.44 min, m/z 515. lli :'H NMR (300 MHz, DMSO-d6) 6 8.99 (s, 4H), 7.81 (s, 2H), 7.58 (s, 2H), 4.32 (m, 4H), 3.19 (m, 4H), 2.78 (s, 3H).

LCMS: LC 2.75 min. , m/z 488.

[0222] Dimers 5a-i were evaluated for their ability to inhibit bacterial growth against both Gram positive and Gram negative organisms (Table 10). Initial tests revealed that dimers 5b and 5h that possess an electron donating substituent at C-6 were also devoid of anti-bacterial activity (entries 7-8). A dramatic improvement in antibacterial activity was seen for dimer 5i, which has a chloro substituent at both the 5-and 6-positions of the benzimidazole's nucleus (entry 9).

[0223] Due to the enhanced biological activity of dichloro-substituted 5i, it was decided to maintain this substitution pattern for further synthetic and biological studies. To explore the importance of the teriary nitrogen atom in the alkyl spacer, dimers 8i and 9i were prepared as shown in scheme 21: ci ci Han NON H2N-, R y H2N R 1 Cl : : aF 6 R = CH2 NH2 j 7R=0 7R=O IR C, N02 I-CaC03, CH2C'2 11 2. Raney Ni, H2 Balloon 3. CNBr Cl N cl N 8i = CH2 9i=0 Scheme 21 [0224] Screening of dimers 8i and 9i indicated that they did not inhibit bacterial growth at 100, uM. To ascertain the significance of the alkyl spacer length, Ili was also prepared (Scheme 22). ci cl Ni2 W N N 6R=CH2 N NH2 I 7R=O b CI<No2 I CaCO3, CH2CI2, rt, 14h cl 1 i 2. Raney Ni, HZ Balloon I NHZ 3. CNBr CI lli Scheme 22 [0225] Biological screening indicated that dimer Ili also possessed moderate to good antibacterial activity against both Gram positive and Gram negative bacteria (entry 12. ) To examine the optimal position of the alkyl spacer, dimers 13 and 14 were prepared from 2,5, 6-trichlorobenzimidazole (Scheme 23). C1 CI ClgCN 10 (0. 5 eq.) CI \/ Cl N Seadd Tube HN N \ 12 rNH jan 2 Sealed Tube Cl r/ 0. 5 eq. (130 C, 14h) /-NH N 13 Ci c NH H ci NN-NN-< Ji J N HHHN, 14 14 Scheme 23 [0226] Biological screening indicated that dimers 13 and 14 had comparable activity to dimers lli and 5i, respectively (entries 13 and 14).

[0227] Synthesis of dimer 13: A mixture of 2, 4,5-trichlorobenzimidazole 12 (0.15 g, 0.68 mmol), 2,2'- diamino-N-methyldiethylamine (0.029 mL, 0.23 mmol), triethylamine (0.031 mL, 0.23 mmol) in EtOH (0.5 mL) was heated in a sealed tube at 140 °C for 6 h. The reaction was then concentrated and purified by reverse-phase preparative HPLC to provide 13 :'H NMR (300 MHz, DMSO-dó) 8 7.27 (s, 4H), 6.88 (m, 2H), 3.4 (m, 4H), 2.63 (t, 4H, J=6.3), 2.30 (s, 3H). LCMS: LC 2.93 min. , m/z 488.14 : 'H NMR (300 MHz, DMSO-d6) E 7.29 (s, 4H), 7.15 (m, 2H), 3.34 (m, 4H), 2.77 (t, 4H, J=7.0), 2.46 (s, 3H), 1.85 (m, 4H). LCMS: LC 2.14 min. , m/z 515.

[0228] Lastly, the importance of the 2-aminobenzimidazole dimers pharmacophore was probed by evaluating 2-aminobenzimidazoles 15 and 16 and benzimidazole dimer 17 for antibacterial activity.

Benzimidazole dimers 17, which was missing the 2-amino group, showed only weak antibacterial activity (entry 13) against both E. coli and S. aureus.

[0229] Compounds 15,16 and 17 are set forth in Scheme 24: Scheme 24 [0230] MIC assays were carried out in a 150 yL volume in duplicate in 96 well, clear, flat-bottom plates.

The bacterial suspension from an overnight culture growth in the appropriate medium was added to a solution of test compound in 2.5% DMSO in water. Final bacterial innoculum was approximately 102- 103 CFU/well. The percentage growth of the bacteria in the test wells relative to that observed for a control well containing no compound was determined by measuring absorbance at 595 nm (As95) after 20-24 h at 37 °C. The MIC was determined as a range of concentrations where complete inhibition was observed at the higher concentration and the bacterial cells were viable at the lower concentration. The bacterial strains used for the assays include E. coli ATCC 25922, S. aureus ATCC 13709, E. Hirae ATCC 29212, S. pyrogenes ATCC 49399. The results of these assays are set forth in Table 10: Table 10 Entry Compounds MIC MIC MIC MIC (yM) (AM) (AM) (AM) E. coli S. aureus S. pyrogenes E. Hirae 1 5a (Ri = H, R2 = H) 50-100 50-100 NT NT 2 5b (Rl = H, R2 = Cl) 25-50 25-50 25-50 12-25 3 5c (R, = H, R2 = CF3) 12-25 25-50 12-25 25-50 4 5d (R, = H, R2 = Br) >100 >100 NT NT 5 5e (R, = H, R2 = CN) >100 >100 NT NT 6 5f (R, = H, R2 = CO2Me >100 >100 NT NT Entry Compounds MIC MIC MIC MIC (IlM) (yM) (ItM) (yM) E. coli S. aureus S. pyrogenes E. Hirae 7 5g (R, = Me, R2 = H >100 >100 NT NT 8 5h (Rl = OMe, R2 = H >100 >100 NT NT 9 5i (R, = Cl, R2 = Cl) 3-6 6-12 3-6 3-6 10 8i (R, = Cl, R2 = Cl) >100 >100 NT NT 11 9i (R, = Cl, R2 = C1) >100 >100 NT NT 12 lli 12-25 6-12 3-6 1. 5-3 13 13 6-12 12-25 6-12 1.5-3 14 14 6-12 3-6 3-6 3-6 15 17 50-100 25-50 NT NT [0231] All dimers were purified by preparative reverse-phase HPLC and tested as their mono- trifluoroacetate or acetate salts. MIC means minimal inhibitory concentration of compound that inhibits visible growth. Ciprofoxacin provided MIC values of 0.04-0. 07 µM (E. coli), 0.3-0. 6, uM (S. aureus), 1.2-2. 5 uM (S. pyrogezles), 1.2-2. 5, uM (E. Hirae) in the assay.

[0232] Analysis of the biological data from Table 10 suggests that a free or secondary 2-amino group and the dimeric structure are crucial for antibacterial activity. Removal of the 2-amino group 17 results in a 16-fold reduction in biological activity while removal of the dimeric structure (15 and 16) abolishes activity completely. Replacement of the tertiary amine in the tether with an oxygen 9i or methylene 8i abolishes biological activity. This result suggests that a positively charged nitrogen (at physiological pH) in the tether is important for antibacterial activity. Activity is also retained when the alkyl spacer chain is moved from the N1 ring nitrogen to the 2-amino group on the benzimidazole nucleus 13 and 14. The length of the spacer between the aromatic rings also appears to have some effect on the biological activity. In general, dimers with a three carbon spacer (5i and 14) between the aromatic ring and the basic amine possess better activity compared to similar dimers (lli and 13) with a two-carbon spacer.

Substitution at the 5-position of the benzimidazole nucleus with non-polar groups such as chloro and trifluoromethyl is tolerated and results in a modest increase in MIC activity (5b-c). The same beneficial effect was not seen when the substituents at C-5 was an electron withdrawing group (EWG) such as cyano or ester or a larger halogen such as bromine. Similarly, substitution at C-6 with a halogen such as chlorine is beneficial (5i) while substitution with an electron donating substituent such as methyl or methoxy (5g-h) abolishes activity. Best results were seen within both C-5 and C-6 were substituted with a chlorine atom (5i). The improved biological activity of the poly-halo-substituted benzimidazole dimers could be attributed to the ability of these dimers to penetrate bacterial cell membranes and reach their cellular targets. It is conceivable that dimers with the dichloro substitution patter are able to penetrate the bacterial cell membranes more efficiently due to their increased lipohilicity. This result may suggest that substituting antibacterial agents with multiple halogen substituents may improve their uptake through bacterial cell membranes and/or reduce their efflux from within the cells.

[0233] In summary, dimers 5i, Ili, 13 and 14 represent a novel class of compounds possessing good antibacterial activity.

[0234] Additional biological analysis were performed on select compounds according to the invention as presented in tables 11-13.

Table 11 Minimal Inhibitory Concentrations (MIC mM) of Benzimidazoles against Bacteria and Yeast Gram+ Gram-Yeast Compound SA1 EH2 SP4 SP6 EC2 PV8 KP1 PA2 CA1 7a 6-12 1-3 3-6 12-25 12-26 25-50 6-12 25-50 50-100 7b 3-6 1-3 3-6 6-12 6-12 12-25 6-12 25-50 25-50 7c 6-12 3-6 6-12 12-25 12-25 25-50 12-25 12-25 >100 7d 12-25 6-12 6-12 25-50 50-100 NT 25-50 25-50 >100 7e 6-12 3-6 6-12 25-50 25-50 NT 25-50 25-50 >100 7g 6-12 1-3 3-6 12-25 12-25 25-50 12-25 25-50 >100 7i 6-12 1-3 3-6 6-12 6-12 12-25 6-12 25-50 >100 7j 6-12 1-3 6-12 12-35 50-100 50-100 12-25 12-25 50-100 7k 3-6 1-3 6-12 6-12 25-50 >100 12-25 25-50 25-50 71 3-6 3-6 3-6 6-12 12-25 25-50 12-25 12-25 >100 7m 6-12 3-6 6-12 12-25 12-25 25-50 12-25 12-25 50-100 7o 6-12 3-6 6-12 12-25 12-25 25-50 12-25 25-50 >100 7r 6-12 3-6 6-12 12-25 12-25 12-25 6-12 12-25 50-100 7s 6-12 3-6 6-12 12-25 12-25 25-50 12-25 25-50 >100 7t 6-12 3-6 6-12 25-50 50-100 25-50 25-50 12-25 50-100 7u 6-12 3-6 6-12 12-25 25-50 25-50 12-25 25-50 >100 7x 6-12 3-6 12-6-12 6-12 25-50 25-50 12-25 50-100 25 lOb 3-7 0. 75- 25-6-12 12-26 25-50 25-50 6-12 6-12 1.5 50 10c 6-12 1-3 3-6 6-12 12-25 NT 6-12 12-25 12-25 12 3-6 1-3 3-6 6-12 6-12 NT 6-12 12-25 50-100 SA1 : S. aureus 13709; EH2: E. hirae 29212; SP4: S. pyogenes 49399; SP6 : S. pneumoniae 6303; EC2: E. coli 25922; PV8 : P. vulgaris 8427; KP1 : K. pheumoniae 13383; PA2: P. aeruginosa 25416; CA1 : C. albicans 10231 ; NT: Not tested. Table 12 Minimal Inhibitory Concentrations (MIC, mM) of Benzimidazoles against Enterococcus Com E. faecalis E. faeca E. faecali E. faecali E. faeca E. faeciu E. faecium E. hirae poun ATCC-11823 li ATCC-ATCC-lis m ATCC-ATCC- d ATCC-4200 49757 ATCC-ATCC 882 29212 23241 828 6569 7a 3-6 6-12 6-12 6-12 6-12 3-6 6-12 1-3 7b 3-6 3-6 3-6 3-6 3-6 3-6 3-6 1-3 7c 6-12 12-25 12-25 12-25 12-25 6-12 12-25 3-6 7d 12-25 25-50 25-50 12-25 12-25 12-25 25-50 6-12 7e 12-25 12-25 12-25 12-25 12-25 12-25 12-25 3-6 7i 6-12 NT 25-50 50-100 50-100 12-25 NT 1-3 7k NT NT NT 25-50 NT 50-100 NT 1-3 7m 12-25 12-25 12-25 12-25 12-25 12-25 12-25 3-6 7p 6-12 12-25 12-25 12-25 6-12 6-12 12-25 3-6 7r 12-25 6-12 12-25 12-25 6-12 6-12 6-12 3-6 7s 12-25 12-25 12-25 6-12 12-25 6-12 12-25 3-6 7t 12-25 12-25 12-25 12-25 12-25 6-12 12-25 3-6 7u 12-25 12-26 12-25 12-26 12-26 6-12 12-25 3-6 7x 3-6 6-12 6-12 6-12 6-12 3-6 6-12 3-6 10b 3-6 6-12 6-12 6-12 6-12 3-6 6-12 0. 7-1.5 10c 3-6 3-6 6-12 3-6 6-12 3-6 3-6 1-3 12 6-12 6-12 6-12 6-12 3-6 3-6 6-12 1-3 NT: Not tested.

Table 13 Minimal Inhibitory Concentrations (MIC) of Selected Benzimidazoles against Bacteria and Yeast Gram+ Gram- Yeast Compound SA1 EH2 SP4 SP6 EC2 PV8 KP1 PA2 14a 6-12 25-50 25-50 12-25 25-50 25-50 25-50 50-100 14b 6-12 25-50 25-50 6-12 >100 >100 50-100 >100 19f 6-12 3-6 6-12 12-25 12-25 12-25 12-25 25-50 17g 6-12 3-6 6-12 12-25 12-25 25-50 6-12 25-50 17h 6-12 1-3 6-12 12-25 25-50 >100 12-25 >100 18j 3-6 3-6 6-12 12-25 >100 50-100 6-12 50-100 18m 6-12 1-3 6-12 12-25 >100 25-50 25-50 50-100 SA1 : S. aureus 13709; EH2: E. hirae 29212; SP4: S. pyogenes 49399; SP6: S. pneumoniae 6303; EC2: E. coli 25922; PV8 : P. vulgaris 8427; KP1 : Kpheumoniae 13383; PA2: P. aeruginosa 25416 ; CA1 : C. albicans 10231 [0235]