WO2011079087A1

WO2011079087A1 - Enac blockers

Info

Publication number: WO2011079087A1
Application number: PCT/US2010/061398
Authority: WO
Inventors: Dramane Ibrahim Laine; Tindy Li; Hongxing Yan
Original assignee: Glaxo Group Limited
Priority date: 2009-12-23
Filing date: 2010-12-21
Publication date: 2011-06-30

Abstract

The present invention relates to benzimidazole analogs, pharmaceutical compositions containing them and their use as ENaC blockers.

Description

ENaC BLOCKERS

FIELD OF THE INVENTION The present invention relates to benzimidazole analogs, pharmaceutical compositions containing them and their use as ENaC blockers.

BACKGROUND OF THE INVENTION

The epithelial sodium channel (ENaC) is a membrane-bound ion-channel that is permeable for Li⁺, protons and especially Na⁺. It is a 'constitutively active' channel, i.e. does not require a gating stimulus and is open at rest. ENaC is a heteromeric protein comprised of three different subunits a SCNN1A), β (SCNN1 B), and γ SCNN1 G).

(Pitkanen, OM, Smith, D, O'Brodovich, H, Otulakowski, G. (2001 ) Expression of α, β, and γ-hENaC mRNA in the human nasal, bronchial and distal lung epithelium. Am J Respir Crit Care Med 163: 273-276.) ENaC is located in the apical membrane of polarized epithelial cells particularly in the kidney, the lung and the colon. It is involved in the transepithelial Na⁺-ion transport which it accomplishes together with the Na+/K+-ATPase. It plays a major role in the Na⁺- and K⁺-ion homeostasis of blood, epithelia and extra epithelial fluids by resorption of Na⁺-ions. The activity of ENaC in colon and kidney is modulated by the mineralcorticoid aldosterone. ENaC can furthermore be found in taste receptor cells, where it plays an important role in salt taste perception.

The airways are lined with a film of liquid about 10 microm deep that is in two layers. Around the cilia is the watery periciliary sol. Over this is a mucous blanket that traps inhaled particles. The mucus layer itself traps inhaled pathogens/particles, allowing their removal via ongoing mucociliary clearance, without the need to trigger a potentially injurious inflammatory response. The low viscosity of the periciliary sol allows the cilia to beat and propel the mucous blanket along airways to the mouth. In large airways, mucus comes predominantly from the mucous glands but also from goblet cells in the surface epithelium. Water is added to the airway surface by gland secretion that is driven by active CI secretion by serous cells. Water is removed by Na transport via ENaC across the surface epithelium. In airway diseases, the balance is shifted from water secretion to mucus secretion. (Widdicombe, JH and Widdicombe JG (2002) Regulation of airway surface liquid. Respir. Physiol. 93: 3-12; Randell, SH and Boucher, RC (2006) Effective Mucus Clearance is Essential for Respiratory Health. Am. J Resp. Cell Mol. Biol. 35: 20- 28, Kunzelmann, K, Schreiber, R, Boucherot, A. (2001 ) Mechanisms of the inhibition of epithelial Na+ channels by CFTR and purinergic stimulation. Kidney Internat. 60:455-461 )

Both periciliary sol volume depletion and mucus layer dehydration contribute to reductions in mucociliary and cough clearance, as cilial movement is restricted as the height of the sol layer becomes more shallow than the height of the cilia themselves, and because the mucus layer is less transportable when dehydrated it ultimately adheres to the airway surface. Mucus stasis results, causing obstruction. Adherent mucus is the nidus for the onset of first intermittent, and then chronic bacterial airway infection.

Blockade of ENaC would reverse this imbalance, thus restoring mucociliary clearance. Blockers of ENaC would be useful in the treatment of cystic fibrosis, COPD, and (non- cystic fibrosis) bronchiectasis. (Sood, N. et al (2003) Increasing Concentration of Inhaled Saline with or without Amiloride. Am J Resp Crit. Care Med. 167:158-163, Brown MA, Leman RJ. Bronchiectasis. In: Chernick V, Boat T, eds. Kendig's disorders of the respiratory tract in children. 6th Edn. Philadelphia, W.B. Saunders, 1998; pp. 538-560, Livraghi, A and Randell, SH (2007) Cystic fibrosis and other respiratory diseases of impaired mucus clearance. Toxicol. Pathol. 35: 1 16-129, Boucher, RC (2007) Evidence for airway surface dehydration as the initiating event in CF airway disease J Int. Med. 261 :5-16, Knowles, MR et al. (1990) A pilot study of aerosolized amiloride for the treatment of lung disease in cystic fibrosis. New Eng. J Med. 322:1 189-1 194, Donaldson, S. (2008) Hydrator therapies for cystic fibrosis lung disease. Ped. Pulmonol. 43:S18-S23, Mall, MA et al. (2008) Development of chronic bronchitis and emphysema in β-epithelial Na+ channel-everexpressing mice. Am J Resp Crit Care Med. 177:730-742, Li, W, XU, YJ, Zhang, ZX. (2004) Detection of the mRNA level of the subunits of amiloride-sensitive Na_ channel in human bronchial epithelium cells from patients with chronic obstructive pulmonary disease. Chinese J Tuberculosis Resp Dis. 27:533-536)

SUMMARY OF THE INVENTION

In one aspect this invention provides for benzimidazole analogs, pharmaceutically acceptable salts thereof, and pharmaceutical compositions containing them.

In a second aspect, this invention provides for the use of the compounds of

Formula (I) as ENaC blockers.

In another aspect, this invention provides for the use of the compounds of Formula (I) for treating and preventing conditions associated with ENaC imbalance. In yet another aspect, this invention provides for the use of the compounds of Formula (I) for the treatment or prevention of cystic fibrosis, COPD, and non-cystic fibrosis bronchiectasis.

The ENaC blocker may be administered alone or in conjunction with one or more other therapeutic agents, eg. agents being selected from the group consisting of: advair, tiotropium, tobi, pulmozyme, azithromycin, hypertonic saline, albuterol, pancreatic enzymes, oral antibiotics, and oral steroids.

DETAILED DESCRIPTION OF THE INVENTION

The present in :

(I)

wherein:

R-i is C_1-6alkyl; wherein the C_1-6alkyl, may be further substituted by phenyl; C_1-6alkyl-OH, or (CH₂)m-C₃-₆cycloalkyl;

R₂ is Ci_-6 alkyl; wherein the C_1-6 alkyl, may be further substituted by phenyl;

(CH₂)_m-C0₂-(CH₂)_m-phenyl, (CH₂)_m-0-R₅, (CH₂)_m-C0₂-R₅, or (CH₂)_m-0-R₄;

R₃ is hydrogen, C_1-6alkyl, halo, CN, OC_1-6alkyl, C_1-6alkyl-OH, or phenyl; wherein the phenyl may be further substituted by -OCi_-6alkyl, -Ci_-6alkyl or -OH;

or R₃ is thiazole, S0₂R₄, (CH₂)_nC0₂R₅, (CH₂)_nC(0)N(R₅)₂, -P-(CH₂)_mC0₂R₅,

-0(CH₂)_nphenyl, -C(0)-Q-oxotetrahydrothienyl, -C(0)-Q-oxotetrahydrofuryl, SCF₃,

-C(0)-Q-(CHR₅)-C0₂R₅, (CH₂)_mC(0)-morpholino, or (CH₂)_n-0-(CH₂)_n-C(0)-Ci_-3alkyl;

R₄ is independently Ci_-6alkyl;

R₅ is independently hydrogen, Ci_-6alkyl, or Ci_-6alkyl-OH;

X is oxygen or NH;

Y is CI or Br;

P is O or S;

Q is NH or S;

m is independently 1 , 2, or 3;

n is independently 0, 1 , 2, or 3; and Z^" is a physiologically acceptable anion selected from the group consisting of: chloride, bromide, iodide, hydroxide, sulfate, nitrate, phosphate, acetate, trifluoroacetate, fumarate, citrate, tartrate, oxalate, succinate, mandelate, methanesulfonate and p-toluenesulfonate. The compounds of Formula I may exist in two different (and equivalent) tautomeric forms as shown below.

"Alkyl" refers to a monovalent saturated hydrocarbon chain having the specified number of member atoms. For example, Ci_-4 alkyl refers to an alkyl group having from 1 to 4 member atoms. Alkyl groups may be straight or branched. Representative branched alkyl groups have one, two, or three branches. Alkyl includes methyl, ethyl, propyl (n- propyl and isopropyl), and butyl (n-butyl, isobutyl, and t-butyl).

"Cycloalkyl" refers to a monovalent saturated or unsaturated hydrocarbon ring having the specified number of member atoms. For example, C₃-₆ cycloalkyl refers to a cycloalkyl group having from 3 to 6 member atoms. Unsaturated cycloalkyl groups have one or more carbon-carbon double bonds within the ring. Cycloalkyl groups are not aromatic. Cycloalkyl includes cyclopropyl, cyclopropenyl, cyclobutyl, cyclobutenyl, cyclopentyl, cyclopentenyl, cyclohexyl, and cyclohexenyl.

When used herein, the terms 'halogen' and 'halo' include fluorine, chlorine, bromine and iodine, and fluoro, chloro, bromo, and iodo, respectively.

"Substituted" in reference to a group indicates that one or more hydrogen atom attached to a member atom within the group is replaced with a substituent selected from the group of defined substituents. It should be understood that the term "substituted" includes the implicit provision that such substitution be in accordance with the permitted valence of the substituted atom and the substituent and that the substitution results in a stable compound (i.e. one that does not spontaneously undergo transformation such as by rearrangement, cyclization, or elimination and that is sufficiently robust to survive isolation from a reaction mixture). When it is stated that a group may contain one or more substituents, one or more (as appropriate) member atoms within the group may be substituted. In addition, a single member atom within the group may be substituted with more than one substituent as long as such substitution is in accordance with the permitted valence of the atom. Suitable substituents are defined herein for each substituted or optionally substituted group.

With regard to stereoisomers, the compounds of Formula (I) may have one or more asymmetric carbon atom and may occur as racemates, racemic mixtures and as individual enantiomers or diastereomers. All such isomeric forms are included within the present invention, including mixtures thereof.

As used herein, "pharmaceutically acceptable" refers to those compounds, materials, compositions, and dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

As used herein, the term "a compound of Formula (I)" or "the compound of Formula (I)" refers to one or more compounds according to Formula (I). The compound of Formula (I) may exist in solid or liquid form. In the solid state, it may exist in crystalline or noncrystalline form, or as a mixture thereof. The skilled artisan will appreciate that pharmaceutically acceptable solvates may be formed for crystalline compounds wherein solvent molecules are incorporated into the crystalline lattice during crystallization.

Solvates may involve non-aqueous solvents such as, but not limited to, ethanol, isopropanol, DMSO, acetic acid, ethanolamine, or ethyl acetate, or they may involve water as the solvent that is incorporated into the crystalline lattice. Solvates wherein water is the solvent incorporated into the crystalline lattice are typically referred to as "hydrates." Hydrates include stoichiometric hydrates as well as compositions containing variable amounts of water. The invention includes all such solvates.

The skilled artisan will further appreciate that certain compounds of the invention that exist in crystalline form, including the various solvates thereof, may exhibit polymorphism (i.e. the capacity to occur in different crystalline structures). These different crystalline forms are typically known as "polymorphs." The invention includes all such polymorphs. Polymorphs have the same chemical composition but differ in packing, geometrical arrangement, and other descriptive properties of the crystalline solid state. Polymorphs, therefore, may have different physical properties such as shape, density, hardness, deformability, stability, and dissolution properties. Polymorphs typically exhibit different melting points, IR spectra, and X-ray powder diffraction patterns, which may be used for identification. The skilled artisan will appreciate that different polymorphs may be produced, for example, by changing or adjusting the reaction conditions or reagents, used in making the compound. For example, changes in temperature, pressure, or solvent may result in polymorphs. In addition, one polymorph may spontaneously convert to another polymorph under certain conditions.

The subject invention also includes isotopically-labelled compounds, which are identical to those recited in Formula (I) and following, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into compounds of the invention and pharmaceutically acceptable salts thereof include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, sulphur, fluorine, iodine, and chlorine, such as 2H, 3H, 1 1 C, 13C, 14C, 15N, 170, 180, 31 P, 32P, 35S, 18F, 36CI, 1231 and 1251.

Compounds of the present invention and pharmaceutically acceptable salts of said compounds that contain the aforementioned isotopes and/or other isotopes of other atoms are within the scope of the present invention. Isotopically-labelled compounds of the present invention, for example those into which radioactive isotopes such as 3H, 14C are incorporated, are useful in drug and/or substrate tissue distribution assays. Tritiated, i.e., 3H, and carbon-14, i.e., 14C, isotopes are particularly preferred for their ease of preparation and detectability. 1 1 C and 18F isotopes are particularly useful in PET

(positron emission tomography), and 1251 isotopes are particularly useful in SPECT (single photon emission computerized tomography), all useful in brain imaging. Further, substitution with heavier isotopes such as deuterium, i.e., 2H, can afford certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life or reduced dosage requirements and, hence, may be preferred in some circumstances. Isotopically labeled compounds of formula I and following of this invention can generally be prepared by carrying out the procedures disclosed in the Schemes and/or in the Examples below, by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent.

Representative Embodiments

In one embodiment:

Ri is Ci_-6alkyl; wherein the Ci_-6alkyl, may be further substituted by phenyl; Ci_-6alkyl-OH, or (CH₂)m-C₃-₆cycloalkyl;

R₂ is C-i-6 alkyl; wherein the Ci_-6 alkyl, may be further substituted by phenyl;

R₃ is hydrogen, Ci_-6alkyl, halo, CN, OCi_-6alkyl, Ci_-6alkyl-OH, or phenyl; wherein the phenyl may be further substituted by -OC_1-6alkyl, -C_1-6alkyl or -OH; or R₃ is thiazole, S0₂R₄, (CH₂)_nC0₂R₅, (CH₂)_nC(0)N(R₅)₂, -P-(CH₂)_mC0₂R₅, -0(CH₂)_nphenyl, -C(0)-Q-oxotetrahydrothienyl, -C(0)-Q-oxotetrahydrofuryl, SCF₃, -C(0)-Q-(CHR₅)-C0₂R₅, (CH₂)_mC(0)-morpholino, or (CH₂)_n-0-(CH₂)_n-C(0)-C_1-3alkyl; R₄ is independently Ci_-6alkyl;

R₅ is independently hydrogen, Ci_-6alkyl, or Ci_-6alkyl-OH;

X is oxygen or NH;

Y is CI or Br;

P is O or S;

Q is NH or S;

m is independently 1 , 2, or 3;

n is independently 0, 1 , 2, or 3; and

Z^" is a physiologically acceptable anion selected from the group consisting of: chloride, bromide, iodide, hydroxide, sulfate, nitrate, phosphate, acetate, trifluoroacetate, fumarate, citrate, tartrate, oxalate, succinate, mandelate, methanesulfonate and p-toluenesulfonate.

In another embodiment:

Ri is C-i-6 alkyl; wherein the Ci_-6 alkyl, may be further substituted by phenyl;

R₂ is C-i-6 alkyl; wherein the C_1-6 alkyl, may be further substituted by phenyl;

R₃ is hydrogen, Ci_-6 alkyl, halo, CN, OCi_-6 alkyl, phenyl; wherein the phenyl may be further substituted by OCi_-6 alkyl;

X is oxygen or NH;

Y is CI or Br; and

It is to be understood that the present invention covers all combinations of particular groups described hereinabove.

Specific examples of compounds of the present invention include the following: 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H- benzimidazol-3-ium iodide;

6-bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 /-/- benzimidazol-3-ium iodide; 6-chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H^ benzimidazol-3-ium iodide;

6-cyano-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 /-/- benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-6-(methyloxy)- 1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3,6-trimethyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H-benzimidazol- 3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-6-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H-benzimidazol-

3- ium trifluoroacetate;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1-ethyl-3-(phenylmethyl)-1 H- benzimidazol-3-ium bromide;

6-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide;

6-Cyano-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1-ethyl-3,6-dimethyl-1 H- benzimidazol-3-ium iodide;

3,5-diamino-N-[(6-bromo-1-ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- pyrazinecarboxamide iodide ;

3,5-diamino-N-[(5-bromo-1-ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- pyrazinecarboxamide iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-(methyloxy)-1 /-/- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-methyl-1 /-/- benzimidazol-3-ium iodide; 2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-propyl-1 /-/- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-(3-thienyl)-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

6-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-1 /-/- benzimidazol-3-ium iodide;

5-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-1 /-/- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-5-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-5- (methylsulfonyl)-l H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-5-[2- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-5-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-(4- methylphenyl)-1 H-benzimidazol-3-ium iodide;

5- carboxy-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3- methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-5- [(ethyloxy)carbonyl]-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-5- [(methyloxy)carbonyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6- [(methyloxy)carbonyl]-1 H-benzimidazol-3-ium iodide;

6- carboxy-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3- methyl-1 H-benzimidazol-3-ium;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-6- [(ethylamino)carbonyl]-3-methyl-1 H-benzimidazol-3-ium iodide; 6-[(butyloxy)carbonyl]-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-^ ethyl-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-{[(2- oxotetrahydro-3-thienyl)amino]carbonyl}-1 H-3,1 -benzimidazol-3-ium iodide;

Methyl N-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- methyl-1 H-3, 1-benzimidazol-3-ium-6-yl]carbonyl}glycinate iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-6- [(ethyloxy)carbonyl]-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-{[(2- oxotetrahydro-3-furanyl)amino]carbonyl}-1 H-3, 1-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-5-[4- (methyloxy)phenyl]-1 H-3, 1 -benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-6-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

3- Butyl-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-1 H- benzimidazol-3-ium iodide;

2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1-ethyl-3-(2-hydroxyethyl)- 1 H-benzimidazol-3-ium bromide;

3- (Cyclopropylmethyl)-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 - ethyl-1 H-benzimidazol-3-ium bromide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3,6-dimethyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- (methyloxy)-l H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-{2-oxo-2- [(phenylmethyl)oxy]ethyl}-1 H-benzimidazol-3-ium;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-dimethyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1-propyl-1 H- benzimidazol-3-ium iodide; 2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-5-(4- hydroxyphenyl)-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1-[2- (methyloxy)ethyl]-1 H-benzimidazol-3-ium iodide;

1-(Cyclobutylmethyl)-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3- methyl-1 H-benzimidazol-3-ium;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-diethyl-6- [(trifluoromethyl)thio]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-dimethyl-6- [(trifluoromethyl)thio]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1 -(phenylmethyl)- 1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1-(2- methylpropyl)-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6- [(phenylmethyl)oxy]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-{[2- (methyloxy)-2-oxoethyl]oxy}-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-[2-(methyloxy)-2- oxoethyl]-1 H-benzimidazol-3-ium bromide;

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-[2-(ethyloxy)-2 oxoethyl]-1 H-benzimidazol-3-ium iodide;

[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-1 H-benzimidazol-

3- ium-3-yl]acetate;

6-bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-[2- (ethyloxy)-2-oxoethyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-[3 (methyloxy)-3-oxopropyl]-1 H-3, 1 -benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6-{[3- (methyloxy)-3-oxopropyl]thio}-1 H-benzimidazol-3-ium iodide;

6- Chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-meth 1 H-benzimidazol-3-ium iodide;

7- Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-meth 1 H-benzimidazol-3-ium iodide; 5- Chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide;

6- Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-propyl- 1 /-/-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6- [(trifluoromethyl)thio]-1 /-/-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-{3-[(3- hydroxypropyl)amino]-3-oxopropyl}-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-[3-(4- morpholinyl)-3-oxopropyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-[3-(ethylamino)-3- oxopropyl]-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-{3-[(2- hydroxyethyl)amino]-3-oxopropyl}-3-methyl-1 H-benzimidazol-3-ium;

6-[(carboxymethyl)oxy]-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 - ethyl-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-(ethyloxy)-3- methyl-1 H-benzimidazol-3-ium;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6- (propyloxy)-1 H-benzimidazol-3-ium iodide;

6-(acetyloxy)-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3- methyl-1 H-benzimidazol-3-ium iodide;

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-(hydroxymethyl)-

3- methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-({[2- (methyloxy)-2-oxoethyl]oxy}methyl)-1 H-benzimidazol-3-ium iodide;

N-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-1 H- 3, 1 -benzimidazol-3-ium-6-yl]carbonyl}homoserine iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1-[3-(methyloxy)- 3-oxopropyl]-1 /-/-benzimidazol-3-ium iodide;

6-((carboxymethyl)carbamoyl)-2-((3,5-diamino-6-chloropyrazine-2-carboxamido)methyl)-

1- ethyl-3-methyl-1 H-benzo[d]imidazol-3-ium iodide;

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-({[2-(ethyloxy)-2- oxoethyl]thio}carbonyl)-3-methyl-1 H-3,1 -benzimidazol-3-ium iodide; 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-m 2-oxotetrahydro-3-furanyl]amino}carbonyl)-1 H-3, 1 -benzimidazol-3-ium iodide; and 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-({[(3R)- 2-oxotetrahydro-3-furanyl]amino}carbonyl)-1 H-3, 1 -benzimidazol-3-ium iodide.

Compound Preparation

The skilled artisan will appreciate that if a substituent described herein is not compatible with the synthetic methods described herein, the substituent may be protected with a suitable protecting group that is stable to the reaction conditions. The protecting group may be removed at a suitable point in the reaction sequence to provide a desired intermediate or target compound. Suitable protecting groups and the methods for protecting and de-protecting different substituents using such suitable protecting groups are well known to those skilled in the art; examples of which may be found in T. Greene and P. Wuts, Protecting Groups in Chemical Synthesis (3rd ed.), John Wiley & Sons, NY (1999). In some instances, a substituent may be specifically selected to be reactive under the reaction conditions used. Under these circumstances, the reaction conditions convert the selected substituent into another substituent that is either useful as an intermediate compound or is a desired substituent in a target compound.

The synthesis of the compounds of the general Formula (I) and pharmaceutically acceptable derivatives and salts thereof may be accomplished as outlined below in Schemes 1- 13. In the following description, the groups are as defined above for compounds of Formula (I) unless otherwise indicated. Starting materials are

commercially available or are made from commercially available starting materials using methods known to those skilled in the art.

Methods for making compounds are illustrated in the following schemes.

As shown in Scheme 1 , the ester derivatives of Formula (I) can be prepared in a multi-step sequence from the commercially available methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate 1.. Hydrolysis of 1. with sodium hydroxide gives the acid intermediate 2, which can be treated with carbonyldiimidazole to give the imidazolide derivative 3. Subsequent treatment of 3 with a suitably substituted benzimidazol-2-yl-methanol derivative 4 results in the formation of the ester 5. Further treatment with an appropriate alkylhalide R₂Z produces the symmetrically substituted quaternary derivatives 6 of Formula (I). Scheme 1

Reagents and conditions: a) 6 N NaOH, MeOH; b) CDI, DCM; c) 4, TEA, DMSO: d) R₂Z

When not commercially available, the substituted benzimidazol-2-yl-methanol derivatives 4 may be prepared by cyclisation of a substituted 1 ,2-diaminobenzene 7 with

2-hydroxyacetic acid 8 as shown in Scheme 2. heme 2

1 4

Alternatively, the ester derivatives of Formula (I) may also be prepared according to Scheme 3.

Treatment of 1-fluoro-2-nitrobenzene derivatives 9 with a primary amine R₂NH₂ affords the corresponding 2-nitroaniline 10, which in turn can be reduced to the corresponding 1 ,2-diaminobenzene Λ Λ_ by hydrogenation. Cyclisation of Λ Λ_ with 2-hydroxyacetic acid 8 gives the corresponding benzimidazol-2-yl-methanol derivative 12. Compound 12 may also be prepared by direct alkylation of the previously described derivative 4.

Subsequent treatment of 12 with the previously described imidazolide derivative 3 results in the formation of the ester 13. Further treatment of 13 with an appropriate alkylhalide R"iZ produces the differentially substituted quaternary derivatives 14 of Formula (I). Scheme 3

Reagents and conditions: a) R₂NH₂, EtOH; b) H₂, Pd/C, EtOH; c) 8,microwave reactor; d) R₂Z, base; e) 3, microwave reactor; f) R^Z

When not commercially available, the aryl or heteroaryl substituted benzimidazol-2-yl- methanol derivatives 4 (R₃ is aryl or heteroaryl) may be prepared by a coupling reaction of 4_(R₃ is Br) with a suitable aryl or heteroaryl boronic acid derivative 38 as shown in Scheme 4.

Scheme 4

4 (R₃=Br) 4 (R₃=Ar/Heteroaryl)

Another method of preparation of the ester derivatives of Formula (I) is shown in Scheme 5. The previously describe benzimidazol-2-yl-methanol derivative 12 is treated with an appropriate alkylhalide R-|Z to gives a quaternary salt intermediate 15, which upon treatment with the imidazolide derivative 3 produces the differentially substituted quaternary derivatives 14 of Formula (I).

Scheme 5

Reagents and conditions: a) F^Z; b) 3, microwave reactor.

As shown in Scheme 6, the amide derivatives of Formula (I) can be prepared in a two-steps sequence from the previously described imidazolide derivative 3. Treatment of 3 with a suitably substituted benzimidazol-2-yl-methylamine derivative 16 results in the formation of the amide 17. Further treatment with an appropriate alkylhalide RiZ

produces the quaternary derivatives 18 of Formula (I). In an analogous reaction sequence, coupling of 3 with an un-substituted benzimidazol-2-yl-methylamine derivative 19 gives the corresponding amide derivative 20, which upon treatment with R-|Z forms the quaternary salts 21 of Formula (I).

Scheme 6

21

Reagents and conditions: a) 16, rt, DMSO; b) RiZ; c) 19, rt, DMSO; d) R₂Z

When not commercially available, the benzimidazol-2-yl-methylamine derivatives 16 and 19 may be prepared as shown in Scheme 7. The previously described 1 ,2- diaminobenzene 1J. is treated with 1 ,1 -dimethylethyl (2-oxoethyl)carbamate 22 to give the BOC protected amine 23. Deprotection of 23 with TFA leads to the substituted benzimidazol-2-yl-methylamine derivatives 16. In an analogous reaction sequence, cyclization of 7 with 22 gives the corresponding derivative 24, which upon treatment with acid affords 19. Compound 24 may also be converted to 23 by monalkylation with an appropriate alkyl halide R₂X. cheme 7

24 19

Reagents and conditions: a) sodium bisulfite, 60°C; b) R₂X, base; c) TFA.

Various R₃ groups can be introduced from the BOC-protected bromide derivative 23a (R₃=Br) as shown in Scheme 8. Compound 23a can be treated with a dioxaborolane 24 in the presence of a Pd catalyst to give a boronate ester 23b, which in turn can be oxidized to the corresponding phenol with hydrogen peroxide to afford 23c.

Hydrogenolysis of 23a leads to the dehydrogenated derivative 23d. Alternatively, 23a may also undergo a Heck reaction with methyl 2-propenoate to give the alkene 23e. Subsequent hydrogenation of 23e leads to the alkane 23f. Compounds 23a, 23c-f may be processed to the corresponding derivatives 16 as previously shown in Scheme 7. heme 8

Reagents and conditions: a) 24, PdCI₂; b) H₂0₂; c) H₂, Pd(OH)₂/C; d) methyl-2- propenoate, PdCI₂; e) H₂, Pd(OH)₂/C

In an analogous manner to that presented in Scheme 8, a variety of R₃ groups can be introduced from the deprotected bromide derivative 16a (R₃=Br). As depicted in Scheme 9, compound 16a can be treated with a sulfide derivative HSR₄ in the presence of a Pd catalyst to give the corresponding aryl sulfide 16b, Scheme 9

Reagents and conditions: a) HSR , Pd₂(dba)₃

Derivatives substituted with a R₃ group incorporating an amide group, an ester group or a carboxylic acid group can be prepared following the reaction sequence depicted in

Scheme 10.

The carboxylic acid intermediate 25, which can be prepared following procedures analogous to that described in Scheme 6, can be esterified to the corresponding ester 26 by treatment with the appropriate alcohol in the presence of an acid. The reverse reaction can be carried out by treating 26 with an appropriate base in an aqueous solvent.

Coupling of compound 25 with an appropriate amine following procedures well known in the art, such as treatment with T₃P or HATU, gives the amide 27. In an analogous manner to that presented in Scheme 6, compounds 25, 26 and 27 can be treated with an appropriate alkylhalide RiZ to give the corresponding quaternary salts 28, 29 and 30 of general Formula (I). Additionally, hydrolysis of the ester derivatives 29 with an appropriate base in an aqueous solvent may also lead to the corresponding acid 28. Scheme 10

30 Reagents and conditions: a) H⁺, R₄OH; b) OH^", H₂0; c) T₃P or HATU, R₄NH₂; d) R,Z

Derivatives substituted with a R₃ group comprising of a hydroxyl or an ether group can be prepared following the reaction sequence depicted in Scheme 11. The phenol intermediate 31., which can be prepared following procedures analogous to that described in Scheme 6, can be alkylated to the corresponding ether 32 by treatment with an appropriate alkyl halide R₅X in the presence of a base. In an analogous manner to that presented in Scheme 6, compounds 31. and 32 can be treated with an appropriate alkylhalide RiZ to give the corresponding quaternary salts 33 and 34 of general Formula

(I)- Scheme11

Reagents and conditions: a) K₂C0₃, R5 ; b) RiZ

Another method of preparation of compounds of Formula (I) is depicted in Scheme 12. Successive alkylation of a 2-methyl-1 H-benzimidazole derivative 35 with a suitable alkyl halide RiX then with a suitable alkyl halide R₂X, followed by treatment with iodine in the presence of a base give the iodide derivative 36. Treatment of 36 by sodium azide, followed by hydrogenation gives the primary amine 37. Subsequent coupling of 37 with the imidazolide 3 produces compound 18 of Formula (I).

Scheme 12

Reagents and conditions: a) R,X; b) R₂X; c) l₂, TEA; d) NaN₃; e) H₂, Pd(OH)₂; f)3,

DMSO.

Compounds of Formula (I) substituted with a R1 group incorporating an acid can be prepared as depicted in Scheme 13. The ester 38, which can be prepared following procedures analogous to that described in Scheme 6, can be hydrolyzed with an appropriate base in an aqueous solvent to the corresponding acid 39.

Scheme 13

Biological Activity

As stated above, the compounds according to Formula (I) are ENaC blockers, and are useful in the treatment or prevention of cystic fibrosis, COPD, (non-cystic fibrosis) bronchiectasis, and obliterative bronchiolitis.

The biological activity of the compounds according to Formula (I) can be determined using any suitable assay for determining the activity of a candidate compound as an ENaC blockers, as well as tissue and in vivo models.

The biological activity of the compounds of Formula (I) are demonstrated by the following tests.

ENaC lonWorks Assay

Cellular Reagent

The potency of ENaC inhibitors were assessed in recombinant cells expressing the three ENaC subunits required to give functional ENaC sodium currents, the a-subunit (SCNN1A, NCBI Reference Sequence Accession NM_001 159576), the β-subunit (SCNN1 B, NCBI Reference Sequence Accession NM_000336) and the γ-subunit (SCNN1 G, NCBI Reference Sequence Accession NM_001039). A HEK293 cell line was generated which stably co-expressed variants of the SCNN1 B and SCNN1 G subunits, SCNN1 B-(P618A,Y620L) and SCNN1 G-(P624stop). HEK293 cells were grown in DMEM-F12 with 10%FBS and 2mM glutamine. The stable HEK293- SCNN1 B-

(P618A,Y620L)/ SCNN1 G-(P624stop) cell line was generated following transfection (by Amaxa Nucleofection according to manufacturer's protocols) of CMV expression vectors pBacMIRE-SCNN1 B-(P618A,Y620L) and pCIHGW- SCNN1 G-(P624stop) and co- selection in 500micrograms/ml Geneticin-G418 and 200micrograms/ml hygromycin-B. The clonal cell line was isolated by single cell dilution cloning. In order to reconstitute the functional ENaC channel in this cell line the wild-type SCNN1 A subunit was introduced by BacMam virus transduction. The SCNNIA-BacMam virus was prepared according to the protocols described in Condreay et al (1999, Proc. Natl. Acad. Sci. USA 96:127-132). Transduction of the HEK293- SCNN1 B-(P618A,Y620L)/ SCNN1 G-(P624stop) cell line with the SCNNIA-BacMam virus was carried out 24 hours prior to assay by adding 10% virus (at approximately 1 .37 x10⁸ pfu/ml) to 1 x10⁷ cells, and continuing to incubate at 37°C, in the presence of the selection agents (G418 & hygromycin).

I on Works Assay

Recombinant cell cultures were prepared according to the platform manufacturer's protocol (MDC Corp, Sunnyvale, CA / Finkel et al, J. Biomol. Screen. 2006; 1 1 : 488-496) using TrypLE™ Express (Gibco / Invitrogen, Paisley, Renfrewshire, UK, Cat # 12604) in place of the specified Versene solution. Sodium-ion flux across the cell membrane via ENaC generates a measurable current (l_ENac) that is assessed by the lonWorks platform (lonWorks Quattro™ model, MDC Corp, Sunnyvale, CA). Inhibition of the IEN_SC with ENaC blockers was used to assess their potency. l_ENac was measured (lonWorks PPC- mode) under voltage clamp conditions from a mean of up to 64 cells in each of up to 384 wells both before and after addition of compounds. The effect of the compound is calculated by normalizing the post compound addition data to the naive state and comparing this against high (100% block by 10μΜ amiloride gold standard) and low ("zero"-effect; 1 % v/v DMSO) control data.

Stock test compounds were prepared as 10mM solutions in DMSO. Concentration se curves were constructed with appropriate serial dilutions of the stock compound in DMSO. These serial dilutions were further diluted 1 :30 times in to External buffer (composition, milli-molar: sodium gluconate, 120; sodium chloride, 20; potassium chloride, 5; 4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic acid, N-(2- Hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid) (HEPES), 10; calcium chloride, 2; magnesium chloride, 1 , and pluronic F-127 0.03% w/v, pH 7.35 - 7.40), and added to a 384 well disposable plate (compound plate), which was loaded in to the lonWorks platform. The lonWorks was programmed to transfer an appropriate volume from the compound plate to the assay well to give a further 1 :3 dilution, at approximately 3 mins prior to the post-compound reading. The data corresponding to the series of dilutions of test compounds were then used to estimate the concentration of compound required to produce a 50 % decrease of the ENaC-mediated current response (IC₅₀). Potency was reported as plC₅₀ (-log IC₅₀).

In each experiment curves for amiloride and phenamil standards were included, and used to monitor data quality. Internal buffer composition was (milli-molar): potassium gluconate (120); potassium chloride (20); HEPES (10); sodium chloride (5); magnesium chloride (2) & calcium chloride (1 ), pH 7.35 -7.40. All chemical reagents, including standards were purchased from Sigma Chemical Co.

Compounds of the formula I have plC50's > 4.5.

Ussing Chamber Assay

The potency of ENaC inhibitors was assessed in commercially available (MatTek

Corporation) human bronchial epithelial cells grown in Air-Liquid Interface (Cat # AIR- 1 12-SNP). Epithelial cell cultures were mounted in Ussing Chambers filled with Krebs bicarbonate buffer solution (KBS) continuously bubbled with 95% oxygen/5% carbon dioxide. Sodium transport across the epithelium via ENaC generates a measurable current that is assessed my measuring short-circuit current (Isc). Inhibition of the Isc with ENaC blockers was used to assess their potency. Isc was measured (Physiologic Instruments, Model VCC MC6) under voltage clamp conditions so that active ion transport was the dominant ion transport process generating Isc.

Test compounds were prepared as 10mM solutions in DMSO and working solutions were prepared by serial 10 fold dilutions in KBS. Cumulatively increasing concentrations (0.5 logs) of test compound are added to the chamber bathing the apical surface of the epithelium. Concentration response curves were used to calculate the concentration of compound that causes a 50 % decrease of the IsC (EC 50). Potency was reported as plC50 (-log EC50).

In each experiment 1 culture was used to assess the potency of the gold-standard

ENaC blocker Amiloride as a positive control.

Methods of Use

The compounds of the invention are ENac blockers, and are useful in the treatment or prevention of cystic fibrosis, COPD, non-cystic fibrosis bronchiectasis, and obliterative bronchiolitis. Accordingly, in another aspect the invention is directed to methods of treating such conditions.

The methods of treatment of the invention comprise administering a safe and effective amount of a compound according to Formula I or a pharmaceutically-acceptable salt thereof to a patient in need thereof. As used herein, "treat" in reference to a condition means: (1 ) to ameliorate or prevent the condition or one or more of the biological manifestations of the condition, (2) to interfere with (a) one or more points in the biological cascade that leads to or is responsible for the condition or (b) one or more of the biological manifestations of the condition, (3) to alleviate one or more of the symptoms or effects associated with the condition, or (4) to slow the progression of the condition or one or more of the biological manifestations of the condition.

As indicated above, "treatment" of a condition includes prevention of the condition. The skilled artisan will appreciate that "prevention" is not an absolute term. In medicine, "prevention" is understood to refer to the prophylactic administration of a drug to substantially diminish the likelihood or severity of a condition or biological manifestation thereof, or to delay the onset of such condition or biological manifestation thereof.

As used herein, "safe and effective amount" in reference to a compound of the invention or other pharmaceutically-active agent means an amount of the compound sufficient to treat the patient's condition but low enough to avoid serious side effects (at a reasonable benefit/risk ratio) within the scope of sound medical judgment. A safe and effective amount of a compound will vary with the particular compound chosen (e.g.

consider the potency, efficacy, and half-life of the compound); the route of administration chosen; the condition being treated; the severity of the condition being treated; the age, size, weight, and physical condition of the patient being treated; the medical history of the patient to be treated; the duration of the treatment; the nature of concurrent therapy; the desired therapeutic effect; and like factors, but can nevertheless be routinely determined by the skilled artisan.

As used herein, "patient" refers to a human or other animal.

The compounds of the invention may be administered by any suitable route of administration, including both systemic administration and topical administration.

Systemic administration includes oral administration, parenteral administration, transdermal administration, rectal administration, and administration by inhalation.

Parenteral administration refers to routes of administration other than enteral,

transdermal, or by inhalation, and is typically by injection or infusion. Parenteral administration includes intravenous, intramuscular, and subcutaneous injection or infusion. Inhalation refers to administration into the patient's lungs whether inhaled through the mouth or through the nasal passages. Topical administration includes application to the skin as well as intraocular, otic, intravaginal, and intranasal

administration. The compounds of the invention may be administered once or according to a dosing regimen wherein a number of doses are administered at varying intervals of time for a given period of time. For example, doses may be administered one, two, three, or four times per day. Doses may be administered until the desired therapeutic effect is achieved or indefinitely to maintain the desired therapeutic effect. Suitable dosing regimens for a compound of the invention depend on the pharmacokinetic properties of that compound, such as absorption, distribution, and half-life, which can be determined by the skilled artisan. In addition, suitable dosing regimens, including the duration such regimens are administered, for a compound of the invention depend on the condition being treated, the severity of the condition being treated, the age and physical condition of the patient being treated, the medical history of the patient to be treated, the nature of concurrent therapy, the desired therapeutic effect, and like factors within the knowledge and expertise of the skilled artisan. It will be further understood by such skilled artisans that suitable dosing regimens may require adjustment given an individual patient's response to the dosing regimen or over time as individual patient needs change.

Typical daily dosages may vary depending upon the particular route of administration chosen. Typical dosages for oral administration range from 1 mg to 1000 mg per person per dose.

Additionally, the compounds of the invention may be administered as prodrugs. As used herein, a "prodrug" of a compound of the invention is a functional derivative of the compound which, upon administration to a patient, eventually liberates the compound of the invention in vivo. Administration of a compound of the invention as a prodrug may enable the skilled artisan to do one or more of the following: (a) modify the onset of the compound in vivo; (b) modify the duration of action of the compound in vivo; (C) modify the transportation or distribution of the compound in vivo; (d) modify the solubility of the compound in vivo; and (e) overcome or overcome a side effect or other difficulty encountered with the compound. Typical functional derivatives used to prepare prodrugs include modifications of the compound that are chemically or enzymatically cleaved in vivo. Such modifications, which include the preparation of phosphates, amides, esters, thioesters, carbonates, and carbamates, are well known to those skilled in the art.

Compositions

The compounds of the invention will normally, but not necessarily, be formulated into pharmaceutical compositions prior to administration to a patient. Accordingly, in another aspect the invention is directed to pharmaceutical compositions comprising a compound of the invention and one or more pharmaceutically-acceptable excipient. The pharmaceutical compositions of the invention may be prepared and packaged in bulk form wherein a safe and effective amount of a compound of the invention can be extracted and then given to the patient such as with powders or syrups. Alternatively, the pharmaceutical compositions of the invention may be prepared and packaged in unit dosage form wherein each physically discrete unit contains a safe and effective amount of a compound of the invention. When prepared in unit dosage form, the pharmaceutical compositions of the invention typically contain from 1 mg to 1000 mg.

The pharmaceutical compositions of the invention typically contain one compound of the invention. However, in certain embodiments, the pharmaceutical compositions of the invention contain more than one compound of the invention. For example, in certain embodiments the pharmaceutical compositions of the invention contain two compounds of the invention. In addition, the pharmaceutical compositions of the invention may optionally further comprise one or more additional pharmaceutically active compounds.

As used herein, "pharmaceutically-acceptable excipient" means a

pharmaceutically acceptable material, composition or vehicle involved in giving form or consistency to the pharmaceutical composition. Each excipient must be compatible with the other ingredients of the pharmaceutical composition when commingled such that interactions which would substantially reduce the efficacy of the compound of the invention when administered to a patient and interactions which would result in pharmaceutical compositions that are not pharmaceutically acceptable are avoided. In addition, each excipient must of course be of sufficiently high purity to render it pharmaceutically-acceptable.

The compound of the invention and the pharmaceutically-acceptable excipient or excipients will typically be formulated into a dosage form adapted for administration to the patient by the desired route of administration. For example, dosage forms include those adapted for (1 ) oral administration such as tablets, capsules, caplets, pills, troches, powders, syrups, elixers, suspensions, solutions, emulsions, sachets, and cachets; (2) parenteral administration such as sterile solutions, suspensions, and powders for reconstitution; (3) transdermal administration such as transdermal patches; (4) rectal administration such as suppositories; (5) inhalation such as dry powders, aerosols, suspensions, and solutions; and (6) topical administration such as creams, ointments, lotions, solutions, pastes, sprays, foams, and gels.

Suitable pharmaceutically-acceptable excipients will vary depending upon the particular dosage form chosen. In addition, suitable pharmaceutically-acceptable excipients may be chosen for a particular function that they may serve in the composition. For example, certain pharmaceutically-acceptable excipients may be chosen for their ability to facilitate the production of uniform dosage forms. Certain pharmaceutically- acceptable excipients may be chosen for their ability to facilitate the production of stable dosage forms. Certain pharmaceutically-acceptable excipients may be chosen for their ability to facilitate the carrying or transporting of the compound or compounds of the invention once administered to the patient from one organ, or portion of the body, to another organ, or portion of the body. Certain pharmaceutically-acceptable excipients may be chosen for their ability to enhance patient compliance.

Suitable pharmaceutically-acceptable excipients include the following types of excipients: Diluents, fillers, binders, disintegrants, lubricants, glidants, granulating agents, coating agents, wetting agents, solvents, co-solvents, suspending agents, emulsifiers, sweeteners, flavoring agents, flavor masking agents, coloring agents, anticaking agents, hemectants, chelating agents, plasticizers, viscosity increasing agents, antioxidants, preservatives, stabilizers, surfactants, and buffering agents. The skilled artisan will appreciate that certain pharmaceutically-acceptable excipients may serve more than one function and may serve alternative functions depending on how much of the excipient is present in the formulation and what other ingredients are present in the formulation.

Skilled artisans possess the knowledge and skill in the art to enable them to select suitable pharmaceutically-acceptable excipients in appropriate amounts for use in the invention. In addition, there are a number of resources that are available to the skilled artisan which describe pharmaceutically-acceptable excipients and may be useful in selecting suitable pharmaceutically-acceptable excipients. Examples include

Remington's Pharmaceutical Sciences (Mack Publishing Company), The Handbook of Pharmaceutical Additives (Gower Publishing Limited), and The Handbook of

Pharmaceutical Excipients (the American Pharmaceutical Association and the

Pharmaceutical Press).

The pharmaceutical compositions of the invention are prepared using techniques and methods known to those skilled in the art. Some of the methods commonly used in the art are described in Remington's Pharmaceutical Sciences (Mack Publishing

Company).

In one aspect, the invention is directed to a solid oral dosage form such as a tablet or capsule comprising a safe and effective amount of a compound of the invention and a diluent or filler. Suitable diluents and fillers include lactose, sucrose, dextrose, mannitol, sorbitol, starch (e.g. corn starch, potato starch, and pre-gelatinized starch), cellulose and its derivatives (e.g. microcrystalline cellulose), calcium sulfate, and dibasic calcium phosphate. The oral solid dosage form may further comprise a binder. Suitable binders include starch (e.g. corn starch, potato starch, and pre-gelatinized starch), gelatin, acacia, sodium alginate, alginic acid, tragacanth, guar gum, povidone, and cellulose and its derivatives (e.g. microcrystalline cellulose). The oral solid dosage form may further comprise a disintegrant. Suitable disintegrants include crospovidone, sodium starch glycolate, croscarmelose, alginic acid, and sodium carboxymethyl cellulose. The oral solid dosage form may further comprise a lubricant. Suitable lubricants include stearic acid, magnesium stearate, calcium stearate, and talc.

Compounds of Formula (I) may be administered parenterally, that is by

intravenous, intramuscular, subcutaneous intranasal, intrarectal, intravaginal or intraperitoneal administration. The subcutaneous and intramuscular forms of parenteral administration are generally preferred. Appropriate dosage forms for such administration may be prepared by conventional techniques. Compounds of Formula (I) may also be administered by inhalation, that is by intranasal and oral inhalation administration.

Appropriate dosage forms for such administration, such as an aerosol formulation or a metered dose inhaler, may be prepared by conventional techniques.

In one embodiment of the present invention, the agents of the present invention are delivered via oral inhalation or intranasal administration. Appropriate dosage forms for such administration, such as an aerosol formulation or a metered dose inhaler, may be prepared by conventional techniques.

For administration by inhalation the compounds may be delivered in the form of an aerosol spray presentation from pressurized packs or a nebulizer, with the use of a suitable propellant, e.g. dichlorodifluoromethane, trichlorofluoromethane,

dichlorotetrafluoroethane, a hydrofluoroalkane such as tetrafluoroethane or

heptafluoropropane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of e.g. gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of a compound of the invention and a suitable powder base such as lactose or starch.

Dry powder compositions for topical delivery to the lung by inhalation may, for example, be presented in capsules and cartridges of for example gelatin or blisters of for example laminated aluminum foil, for use in an inhaler or insufflator. Powder blend formulations generally contain a powder mix for inhalation of the compound of the invention and a suitable powder base (carrier/diluent/excipient substance) such as mono-, di or poly-saccharides (e.g. lactose or starch). Use of lactose is preferred. The compounds of the invention may be milled using known milling procedures such as wet milling to obtain a particle size appropriate for tablet formation and for other formulation types. Finely divided (nanoparticulate) preparations of the compounds of the invention may be prepared by processes known in the art, for example see International Patent Application No. WO 02/00196 (SmithKline Beecham).

The compounds may be administered alone or in conjunction with one or more other therapeutic agents, said agents being selected from the group consisting of advair, tiotropium, tobi, pulmozyme, azithromycin, hypertonic saline, albuterol, pancreatic enzymes, oral antibiotics, and oral steroids.

EXAMPLES

The following examples illustrate the invention. These examples are not intended to limit the scope of the present invention, but rather to provide guidance to the skilled artisan to prepare and use the compounds, compositions, and methods of the present invention. While particular embodiments of the present invention are described, the skilled artisan will appreciate that various changes and modifications can be made without departing from the spirit and scope of the invention.

In the Examples:

Chemical shifts are expressed in parts per million (ppm) units. Coupling constants (J) are in units of hertz (Hz). Splitting patterns describe apparent multiplicities and are designated as s (singlet), d (doublet), t (triplet), q (quartet), dd (double doublet), dt (double triplet), m (multiplet), br (broad).

Flash column chromatography was performed on silica gel.

The naming program used is ACD Name Pro 6.02.

Purifications and analyses of materials were carried out using the following instruments: LC-MS analysis

The LC/MS of Intermediates and Examples were performed using the following equipment and conditions:

Liquid Chromatoqraph:

System: Shimadzu LC system with SCL-10A Controller and dual UV detector Autosampler: Leap CTC with a Valco six port injector

Column: Aquasil/Aquasil (C18 40x1 mm)

Inj. Volume: 2.0 μί

Solvent A: H₂0, 0.02% TFA

Solvent B: MeCN, 0.018% TFA

Gradient: linear Channel A: UV 214 nm

Channel B: ELS

Step Time (min) Dura.(min) Flow (μΙ_/Γηίη) Sol.A Sol.B

0 0.00 0.00 300.00 95.00 5.00

1 0.00 0.01 300.00 95.00 5.00

2 0.01 3.20 300.00 10.00 90.00

3 3.21 1.00 300.00 10.00 90.00

4 4.21 0.10 300.00 95.00 5.00

5 4.31 0.40 300.00 95.00 5.00

Mass Spectrometer:

Instrument: PE Sciex Single Quadrupole LC/MS API-150

Polarity: Positive

Acquisition mode: Profile

Preparatory HPLC

Automated preparatory HPLC purifications were conducted using a Gilson® semi- preparative HPLC system under the following conditions:

Column: 75 x 33mm I. D. , S-5um, 12nm

Flow rate: 30mL/min

Injection Volume: 0.800 mL

Room temperature

The eluent was a mixture composed of solvents A and B. Either one of three different solvent combinations were used:

TFA conditions

Solvent A: 0.1 % trifluoroacetic acid in water

Solvent B: 0.1 % trifluoroacetic acid in acetonitrile

NH₄OH conditions

Solvent A: 0.1 % NH₄OH in water

Solvent B: 0.1 % NH₄OH in acetonitrile

Neutral conditions

Solvent A: 0.1 % NH₄OH in water

Solvent B: 0.1 % NH₄OH in acetonitrile

Automated Flash Chromatography

The automated flash chromatography purifications were conducted with a CombiFlash® Companion® personal flash chromatography system under the following conditions: Silica cartridge: Size, 4, 12, 40, 80 or 120 g depending on the amount of material to be purified Flow rate: Between 4 and 85 mL/min

Room temperature

The eluent was a mixture composed of solvents A and B:

Solvent A: Hexane

Solvent B: Ethyl acetate

Mass-Directed Auto Prep HPLC

The Mass-Directed Auto Prep HPLC (MDAP) purifications were conducted with an Agilent preparatory HPLC-MS system under the following conditions:

Column: ZORBAX Eclipse XDB-C18 (21 .2 ^χ 50 mm)

Flow rate: 20 mL/min

Injection volume: 900 uL

Temperature: 30°C

absorption wavelength: 230 nm

The eluent was a mixture composed of solvents A and B:

Solvent A: 0.1 % trifluoroacetic acid in water

Solvent B: 0.1 % trifluoroacetic acid in acetonitrile

Microwave reactions

The microwave reactions were conducted with a Biotage Initiator™ microwave

synthesizer under the following conditions:

Temperature range: 40-250°C

Temperature increase 2-5°C/sec depending on solvent and power applied

Pressure range: 0-20 bar (2 MPa; 290 PSI)

Power range: 0-400 W from magnetron at 2.45 GHz

Reaction volumes: 0.5-5 mL or 0.2-20 mL depending on amount of material

The following abbreviations and terms had the indicated meanings throughout: aq. Aqueous

bn Benzyl

BOC tert-butyloxycarbonyl

DCM Dichloromethane

DEAD Diethylazodicarboxylate

DMAP Dimethylaminopyridine

DIPEA/DIEA Di-isopropylethylamine

DMF Dimethylformamide

DMSO Dimethylsulfoxide EA Ethyl acetate

ESI Electrospray ionization

eq. Equivalent

HPLC High pressure liquid chromatography

LC/MS Liquid chromatography-Mass spectrometry

Me Methyl

Ph Phenyl

PS Polymer-supported

Rt Retention time

sat. Saturated

SPE Solid phase extraction

TBAF tetra-Butylammonium fluroride

TBS t-Butyldimethyl silyl

TBS-CI t-Butyldimethyl silyl chloride

TEA Triethylamine

TFA Trifluoroacetic acid

THF Tetrahydrofuran

UV Ultraviolet

INTERMEDIATE COMPOUNDS

Intermediate 1

3,5-Diamino-6-chloro-2-pyrazinecarboxylic acid

A mixture of methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate (10 g, 49.4 mmol), 6 N NaOH (9.87 mL, 59.2 mmol) and methanol (50 mL) was stirred at room temperature over the weekend. A solid precipitated out of the solution. 1 N HCI was added to the mixture until pH~2. A solid was filtered off, washed with ethyl ether then collected and dried under high vacuum to give the title compound (9.35 g, 49.6 mmol, 100 %) as a yellow solid. LC/MS: m/z 189.1 (M+H)⁺, Rt 0.32 min. Intermediate 2

-Chloro-5-(1H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine

A mixture of 3,5-diamino-6-chloro-2-pyrazinecarboxylic acid (6 g, 31.8 mmol) and 1 ,1 '- (oxomethanediyl)bis-l H-imidazole (6.7 g, 41 .4 mmol) in DMF (30 mL) was stirred at room temperature for 2 h. The crude was filtered off and washed with DCM. The solid was collected and dried under high vacuum to give the title compound (7.5 g, 31 .5 mmol, 99 %) as a solid. LC/MS: m/z 239.1 (M+H)⁺, Rt 0.35 min. Intermediate 3

1 inecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (100 mg, 0.419 mmol) in DMSO (2 mL) were added triethylamine (0.088 mL, 0.629 mmol) and 1 H- benzimidazol-2-ylmethanol (74.5 mg, 0.503 mmol). The reaction mixture was stirred at 100°C. After 1 h, the reaction mixture was purified by preparatory HPLC (NH₄OH). The desired fractions were concentrated to give the title compound (59.7 mg, 0.187 mmol, 44.7 %) as a white solid. LC/MS: m/z 318.8 (M+H)⁺, Rt 0.61 min.

Intermediate 4

-Chloro-1 H-benzimidazol-2-yl)methanol

To a mixture of 4-chloro-1 ,2-benzenediamine (177 mg, 1 .241 mmol), hydroxyacetic acid (133 mg, 1.489 mmol) and 6 N HCI (0.34 mL, 2.040 mmol), water (1.5 mL) were added. The resulting reaction mixture was placed in a microwave reactor for 30 min at 150°C. A 6 N NaOH aqueous solution was added until the formation of precipitate was observed (0.4-0.9 mL). The formed precipitate was filtered off and then washed with 5 mL of water followed by 5 mL of DCM and 5 mL of ether. The resulting solid was dried in an oven under vacuum for 16 h to give the title compound (129.6 mg, 57.2 %). LC/MS: m/z 183.0 (M+H)⁺, Rt 0.40 min.

Intermediate 5

-(Hydroxymethyl)-1 H-benzimidazole-5-carbonitrile

To a mixture of 3,4-diaminobenzonitrile (183 mg, 1.375 mmol), hydroxyacetic acid (148 mg, 1 .649 mmol), 6 N HCI (0.367 mL, 2.199 mmol) was added water (1.5 mL). The resulting reaction mixture was placed in a microwave reactor for 30 min at 150°C. Then a 6 N NaOH aqueous solution was added to the mixture until the formation of a precipitate was observed (0.4-0.9 mL). The formed precipitate was filtered off and then washed with 5 mL of water followed by 5 mL of DCM and 5 mL of ether. The resulting solid was dried in an oven under vacuum for 16 h to give the title compound (163.2 mg, 68.6 %). LC/MS: m/z 173.8 (M+H)⁺, Rt 0.42 min.

Intermediate 6

-Bromo-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (100 mg, 0.419 mmol) and (5-bromo-1 H-benzimidazol-2-yl)methanol (1 14 mg, 0.503 mmol) in DMSO (2 mL) was added triethylamine (0.1 mL, 0.717 mmol). The reaction mixture was stirred at 100°C for 6 h then cooled to room temperature. The crude product in DMSO was purified by preparatory HPLC (NH₄OH). The desired fractions were concentrated under a stream of nitrogen at 50°C to give the title compound (68.1 mg, 0.171 mmol, 40.9 %). LC/MS: m/z 396.8 (M+H)⁺, Rt 0.72 min.

Intermediate 7

-Methyl-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (100 mg, 0.419 mmol) and (5-methyl-1 H-benzimidazol-2-yl)methanol (82 mg, 0.503 mmol) in DMSO (2 mL) was added triethylamine (0.1 mL, 0.717 mmol). The reaction mixture was stirred at 100°C for 23 h then it was cooled to room temperature. The crude product in DMSO was purified by preparatory HPLC (NH₄OH). The desired fractions were concentrated under a stream of nitrogen at 50°C to give the title compound (43 mg, 0.129 mmol, 30.8 %). LC/MS: m/z 332.9 (M+H)⁺, Rt 0.67 min.

Intermediate 8

[5-(Methyloxy)-1H-benzimidazol-2-yl]methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (100 mg, 0.419 mmol) and [5-(methyloxy)-1 H-benzimidazol-2-yl]methanol (90 mg, 0.503 mmol) in DMSO (2 mL) was added triethylamine (0.1 mL, 0.717 mmol). The reaction mixture was stirred at 100°C for 23 h then it was cooled to room temperature. The crude product in DMOS was purified by preparatory HPLC (NH₄OH). The desired fractions were concentrated under a stream of nitrogen at 50°C to give the title compound (50.4 mg, 0.145 mmol, 34.5 %). LC/MS: m/z 348.9 (M+H)⁺, Rt 0.54 min.

Intermediate 9

-Chloro-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (100 mg, 0.419 mmol) and(5-chloro-1 H-benzimidazol-2-yl)methanol (92 mg, 0.503 mmol) in

DMSO (2 mL) was added triethylamine (0.1 mL, 0.717 mmol). The reaction mixture was stirred at 100°C for 91 h then cooled to room temperature. The reaction mixture was filtered through a 0.45 μηη acrodisc, diluted with 2 mL of DMSO and purified by preparatory HPLC (NH₄OH). The desired fractions were concentrated under a stream of nitrogen at 50°C to give the title compound (43.8 mg, 0.120 mmol, 28.7 %). LC/MS: m/z 352.8 (M+H)⁺, Rt 0.80 min Intermediate 10

(5-Cyano-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (200 mg, 0.838 mmol) and 2-(hydroxymethyl)-1 H-benzimidazole-5-carbonitrile ( 160 mg, 0.924 mmol) in DMSO (4 mL) was added triethylamine (0.1 17 mL, 0.838 mmol). The reaction mixture was stirred at 100°C for 91 h then cooled to room temperature. The crude product was dissolved in DMSO (1 mL), filtered through a 0.45 μηη acrodisc, and purified by preparatory HPLC (neutral conditions). The desired fractions were concentrated under a stream of nitrogen at 50°C to give the title compound (69.7 mg, 0.203 mmol, 24.19 %). LC/MS: m/z 343.7 (M+H)⁺, Rt 0.67 min

Intermediate 11

3,5-diamino-N-[(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- pyrazinecarboxamide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (239 mg, 1 mmol) in DMSO (5 mL) was added 1 -(6-bromo-1-ethyl-1 H-benzimidazol-2- yl)methanamine dihydrochloric salt (327 mg, 1 .000 mmol) then TEA (0.418 mL, 3.00 mmol). The resulting reaction mixture was stirred at room temperature for 17 h and then purified by preparatory HPLC (with 0.1 % NH₄OH) to afford the title compound (261 .6 mg, 0.616 mmol, 61.6 % yield). LC/MS: m/z 424 (M+H)⁺, Rt 0.79 min.

Intermediate 12

3,5-diamino-N-[(5-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- razinecarboxamide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (239 mg, 1 mmol) in DMSO (5 mL) was added 1 -(5-bromo-1-ethyl-1 H-benzimidazol-2- yl)methanamine (254 mg, 1 .000 mmol) then TEA (0.139 mL, 1 .000 mmol). The resulting reaction mixture was stirred at room temperature for 17 h and then purified by preparatory HPLC (with 0.1 % NH₄OH) to afford the title compound (235.4 mg, 0.554 mmol, 55.4 % yield). LC/MS: m/z 424 (M+H)⁺, Rt 0.76 min.

LC/MS: m/z 189.1 (M+H)⁺, Rt 0.32 min.

Intermediate 13

(5-Bromo-1 H-benzimidazol-2-yl)methanol

A mixture of 4 N HCI (8.55 ml_, 51 .3 mmol), (2-amino-4-bromophenyl)amine (8 g, 42.8 mmol) and hydroxyacetic acid (4.88 g, 64.2 mmol) in water (3 ml.) was heated in a microwave reactor at 120°C for 1 h. NaOH (6 N, 2 ml.) was added. Solid precipitated out. Solid was filtered and washed with ether to give the title compound (7.3g, 32.2 mmol, 75 % yield). LC/MS: m/z 226.7 (M+H)⁺, Rt 0.61 min.

Intermediate 14

{5-[4-(Methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol

A mixture of (5-bromo-1 H-benzimidazol-2-yl)methanol (800 mg, 3.52 mmol), [4- (methyloxy)phenyl]boronic acid (803 mg, 5.28 mmol), PdCI₂(dppf) (25.8 mg, 0.035 mmol) and sodium carbonate (1 120 mg, 10.57 mmol) was dissolved in a mixture of 1 ,4-Dioxane (1 .500 ml.) and water (0.5ml_) in a microwave vial. It was degassed for 5 min and then heated in a microwave reactor for 10 min at 100°C. The crude product was filtered through a PL-Thiol MP SPE+ column. Then solvents were concentrated to give the title compound (793 mg, 3.12 mmol, 89 % yield). LC/MS: m/z 255.1 (M+H)⁺, Rt 0.60 min.

Intermediate 15

{5-[4-(Methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate

A mixture of 3-chloro-5-(1 H-imidazol-1-ylcarbonyl)-2,6-pyrazinediamine (450 mg, 1.886 mmol), {5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol (719 mg, 2.83 mmol) and TEA (0.789 mL, 5.66 mmol) in DMSO (4 mL) in a microwave vial was heated in a microwave reactor at 120°C for 1 h. The crude product was purified by preparatory HPLC (with 0.1 % NH₄OH). The desired fractions were concentrated under a stream of nitrogen at 50°C to give the title compound (327 mg, 0.770 mmol, 40.8 % yield). LC/MS: m/z 425.0 (M+H)⁺, Rt 0.88 min.

Intermediate 16

1 ,3-Diethyl-2-(hydroxymethyl)-1 H-benzimidazol-3-ium iodide

A mixture of (1 -ethyl-1 H-benzimidazol-2-yl)methanol (200 mg, 1 .135 mmol) and iodoethane (0.917 mL, 1 1.35 mmol) in THF (2 mL) was heated in a microwave reactor for 40 min at 100°C. Solid was filtered and then washed with ether to give the title compound (269 mg, 0.810 mmol, 71 .3 % yield) as solid. LC/MS: m/z 205.1 (M)⁺, Rt 0.54 min.

Intermediate 17

-nitrophenyl)amine

To a mixture of 1-fluoro-2-nitrobenzene (2.242 mL, 21.26 mmol) and potassium carbonate (5.88 g, 42.5 mmol) in Ethanol (10 mL) was added dropwise 70 % ethylamine in water (5.16 mL, 63.8 mmol). The resulting mixture was heated in a microwave reactor for 10 min at 100°C. Water (20 mL) was added and extracted with DCM (2 x 40 mL). The combined organic layer was washed with saturated NaCI (20 mL), dried over sodium sulfate, filtered and then concentrated under vacuum to give the title compound (3.5 g, 21 .06 mmol, 99 % yield). LC/MS: m/z 166.7 (M+H)⁺, Rt 1 .05 min.

Intermediate 18

( -Aminophenyl)ethylamine

A solution of ethyl(2-nitrophenyl)amine (3.5 g, 21.06 mmol) in Ethanol (50 mL) was passed through an H-Cube hydrogenation apparatus (1 mL/min, 25°C, 2 bar) for 3 h. LC/MS showed only 20 % product conversion. 10 % Pd/C (0.336 g, 0.316 mmol) was added to the solution. Hydrogen in balloon was applied and stirred at room temperature over the weekend. The reaction mixture was filtered through celite, washed with methanol (10 mL), followed by ethyl acetate (20 mL). Solvents were evaporated under reduced pressure to give the title compound (2.82 g, 20.71 mmol, 98 % yield) as oil. LC/MS: m/z 136.9 (M+H)⁺, Rt 0.60 min.

Intermediate 19

( -Ethyl-1 H-benzimidazol-2-yl)methanol

To a solution of (2-aminophenyl)ethylamine (2.82 g, 20.71 mmol) and hydroxyacetic acid (1 .890 g, 24.85 mmol) in water (10 mL) was added 6 N HCI,aq (5.18 mL, 31.1 mmol). The resulting reaction mixture was heated in a microwave reactor for 30 minutes at 150°C. NaOH 6 N was added to the reaction mixture until pH=12-13. The formed precipitate was filtered off and then washed with ether (10 mL) to give the title compound (3.62 g, 20.54 mmol, 99 % yield) as brown solid. LC/MS: m/z 177.1 (M+H)⁺, Rt 0.39 min.

Intermediate 20

(1 -Ethyl-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (1000 mg, 4.19 mmol) and (1 -ethyl-1 H-benzimidazol-2-yl)methanol (738 mg, 4.19 mmol) in DMF (9 mL) was added triethylamine (1 .168 mL, 8.38 mmol). The reaction mixture was heated in a microwave reactor for 1 h at 120°C. LC/MS showed the major peak was the desired product and minor amount of starting material remaining. It was re-submitted back to a microwave reactor and heated at normal absorption for 1 h at 120°C. Ether (5 mL) was added. Solid was filtered and washed with ether to give the title compound (746 mg, 2.151 mmol, 51.3 % yield) as yellow solid. The mother liquor was filtered and washed with ether again to give another batch of the title compound (485 mg, 1.399 mmol, 33.4 % yield) as yellow solid. LC/MS: m/z 346.7 (M+H)⁺, Rt 0.74 min. Intermediate 21

(1 -Ethyl -6-methyl-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2- razinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (150 mg,

0.629 mmol) and (1-ethyl-6-methyl-1 H-benzimidazol-2-yl)methanol (120 mg, 0.629 mmol) in DMF (3 mL) was added triethylamine (0.175 mL, 1 .257 mmol). The reaction mixture was heated in a microwave reactor for 1 h at 120°C. The crude product was purified purified by automated flash chromatography to give the title compound (55 mg, 0.152 mmol, 24.25 % yield). LC/MS: m/z 361 .1 (M+H)⁺, Rt 0.74 min.

Intermediate 22

-Bromo-2-nitrophenyl)ethylamine

To a mixture of 4-bromo-1 -fluoro-2-nitrobenzene (0.4 mL, 3.29 mmol) and potassium carbonate (1 136 mg, 8.22 mmol) in ethanol (3 mL) was added dropwise a 70% solution of EtNH₂ in water (1.2 mL, 14.83 mmol). The resulting mixture was stirred at room temperature for 1 hour. The solvent was evaporated to dryness. To the resulting residue were added 20ml of water (complete dissolution of potassium carbonate) and 50 ml of DCM. The layers were separated, and the aqueous layer was extracted with DCM (2 x 20 mL). The combined organic layers were washed with saturated NaCI (1 x 20 mL), dried over magnesium sulfate, and concentrated under reduced pressure. The residue was dried to give the title compound (720.3 mg, 2.94 mmol, 89 % yield). LC/MS: m/z 244.7 (M+H)⁺, Rt 1.14 min.

Intermediate 23

-Amino-4-bromophenyl)ethylamine

Method 1

700 mg of (4-bromo-2-nitrophenyl)ethylamine were dissolved in 400 mL of a 3/1 mixture of EtOH/THF. The reaction was hydrogenated using an H-Cube hydrogenation apparatus (settings: 60°C, 60 bar, 1 ml_/min flow rate) with a Raney Ni CatCart 30 cartridge. The solvent was evaporated to dryness to give the title compound (512.9 mg, 2.385 mmol, 83 % yield). LC/MS: m/z 215 (M+H)⁺, Rt 0.66 min.

Method 2

To a cold suspension of (4-bromo-2-nitrophenyl)ethylamine (40 g, 163 mmol), and tin(ll) chloride (155 g, 816 mmol) in HCI (4 M) (15 mL). The reaction mixture was stirred at 85°C for 2 h. After the reaction was completed, the mixture was diluted with saturated aqueous NaOH (500 mL) and EOAc(150 mL). The organic layers were separated, and the aqueous layer was extracted with EtOAc(2 x 150 mL). Isolute was added to the combined organic layers, and the mixture was dried over Na₂S0₄ and concentrated under reduced pressure to give the crude product. It was purified by column chromatography, giving the title compound (20 g, 93 mmol, 57.0 % yield). LC/MS: m/z 215 (M+H)⁺, Rt 1.60 min.

Intermediate 24

( -Bromo-2-nitrophenyl)ethylamine

To a mixture of 4-bromo-2-fluoro-1 -nitrobenzene (660 mg, 3 mmol) and potassium carbonate (1037 mg, 7.50 mmol) in ethanol (2.5 mL) was added dropwise a 70 % solution of EtNH₂ in water (1.5 mL, 18.54 mmol). The resulting mixture was stirred at room temperature for 1 hour. The solvent was evaporated to dryness. To the resulting residue were added 20 ml of water (complete dissolution of potassium carbonate) and 50 ml of DCM. The layers were separated, and the aqueous layer was extracted with DCM (2 x 20 mL). The combined organic layers were washed with saturated NaCI (1 x 20 mL), dried over magnesium sulfate, and concentrated under reduced pressure. The residue was dried under high vacuum to give the title compound (655.5 mg, 2.67 mmol, 89 % yield). LC/MS: m/z 244.8 (M+H)⁺, Rt 1.14 min. Intermediate 25

-Amino-5-bromophenyl)ethylamine

655 mg of (4-bromo-2-nitrophenyl)ethylamine were dissolved in 200ml of a 1/1 mixture of EtOH/THF. The reaction was hydrogenated using an H-Cube hydrogenation apparatus (settings: 50°C, 60 bar, 1 ml_/min flow rate) and a Raney Ni CatCart 30 cartridge. The solvent was evaporated to dryness to give the title compound (459.3 mg, 2.135 mmol, 81 % yield). LC/MS: m/z 214.6 (M+H)⁺, Rt 0.80 min.

Intermediate 26

(5-Bromo-1 -ethyl-1 H-benzimidazol-2-yl)methanol

Method 1

A mixture of 6N HCI (0.581 ml_, 3.49 mmol), (2-amino-4-bromophenyl)ethylamine (500 mg, 2.325 mmol) and hydroxyacetic acid (265 mg, 3.49 mmol) in water (3 ml.) in a microwave vial was heated at 150°C for 30 minutes. NaOH (6N, 0.4 ml.) was added then DCM (10 ml). The layers were separated and then the aqueous layer was extracted twice. The combined organic layers were evaporated and the residue was purified by automated flash chromatography to give the title compound (204 mg, 0.800 mmol, 34.4 % yield, purity ca 40%). LC/MS: m/z 255.1 (M+H)⁺, Rt 0.88 min.

Method 2

A mixture of (2-amino-4-bromophenyl)ethylamine (10 g, 46.5 mmol), hydroxyacetic acid (3.54 g, 46.5 mmol) in 4 M HCI (200 ml.) was heated to reflux for 4h. After the reaction was completed, the saturated aqueous NaHC0₃ (1500 ml.) was added to the reaction mixture. The layers were separated. The aqueous layer was extracted with EtOAc (3 x 150 ml_). The combined organic layer was dried over Na₂S0₄ and concentrated under reduced pressure. The crude product was then purified by column chromatography on silica gel (200-300μηι), eluting by 30 % EtOAc / Petrolumn ether, giving the title compound (7.5 g, 29.4 mmol, 63.2 % yield). LC/MS: m/z 255 (M+H)⁺, Rt 1.52 min. Intermediate 27

(6-Bromo-1 -ethyl-1H-benzimidazol-2-yl)methanol

Method 1

A mixture of 6N HCI (0.523 mL, 3.14 mmol), (2-amino-5-bromophenyl)ethylamine (450 mg, 2.092 mmol) and hydroxyacetic acid (239 mg, 3.14 mmol) in water (3 mL) in a microwave vial was heated at 150°C for 30 minutes. NaOH (6N, 0.4 mL) then DCM (10mL) were added. The layers were separated and then the aqueous layer was extracted twice. The combined organic layers were evaporated and the residue was purified by automated flash chromatography to give the title compound (91 mg, 0.357 mmol, 17.05 % yield). LC/MS: m/z 255.0 (M+H)⁺, Rt 0.61 min.

Method 2

To a solution of (5-bromo-2-nitrophenyl)ethylamine (15 g, 61.2 mmol) and tin(ll) chloride (58.0 g, 306 mmol) in 4 N HCI (500 mL) was added hydroxyacetic acid (13.96 g, 184 mmol). The resulting suspension was stirred at 85°C for 3 h. The reaction solution was neutralized with NaOH to pH = 8, then extracted with ethyl acetate (3 x 500 mL). The organic phase was dried with anhydrous Na₂S0₄, and then filtered. The filtrate was concentrated to obtain the crude product. To the crude product was added 150 mL ethyl acetate, sonicated for 10 mins, then filtered to afford the title compound (7.5 g, 28.6 mmol, 46.7 % yield) as white solid. LC/MS: m/z 255.0 (M+H)⁺, Rt 1.408 min.

Intermediate 28

5-Bromo-1 -ethyl-2-(hydroxymethyl)-3-methyl-1 H-benzimidazol-3-ium iodide

A mixture of iodomethane (0.184 mL, 2.94 mmol) and (5-bromo-1-ethyl-1 H-benzimidazol- 2-yl)methanol (150 mg, 0.588 mmol) in THF (2 mL) was heated in a microwave reactor at normal absorption for 30 at 100°C. Solid was filtered and then washed with ether to give the title compound (218 mg, 0.549 mmol, 93 % yield) as solid. LC/MS: m/z 269.0 (M)⁺, Rt 0.58 min. Intermediate 29

6-Bromo-1 -ethyl-2-(hydroxymethyl)-3-methyl-1 H-benzimidazol-3-ium iodide

I

A mixture of iodomethane (0.184 mL, 2.94 mmol) and (6-bromo-1-ethyl-1 H-benzimidazol- 2-yl)methanol (150 mg, 0.588 mmol) in THF (2 mL) was heated in a microwave reactor at normal absorption for 30 at 100°C. Solid was filtered and then washed with ether to give the title compound (219 mg, 0.552 mmol, 94 % yield) as solid. LC/MS: m/z 269.0 (M)⁺, Rt 0.55 min.

Intermediate 30

{1 -Ethyl-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol

A mixture of (6-bromo-1-ethyl-1 H-benzimidazol-2-yl)methanol (200 mg, 0.784 mmol), [4- (methyloxy)phenyl]boronic acid (179 mg, 1.176 mmol), sodium carbonate (249 mg, 2.352 mmol) and PdCI₂(dppf) (57.4 mg, 0.078 mmol) in 1 ,4-Dioxane (2 mL) and water (0.667 mL) in a microwave vial was degassed for 2 min and then heated in a microwave reactor at 100°C for 10 min. Water (1 mL) and ethyl acetate (4 mL) were added to the reaction mixture. Solid precipitated out from the solution. Solid was filtered and then washed with water (2 mL) followed by ethyl acetate (5 mL). Solid was dried to give the title compound (152 mg, 0.538 mmol, 68.7 % yield) as solid. The mother liquor was extracted with ethyl acetate. The organic layer was dried over sodium sulfate, filtered and then concentrated to give another batch of the title compound (58 mg, 0.158 mmol, 20.18 % yield). LC/MS: m/z 283.1 (M+H)⁺, Rt 0.73 min.

Intermediate 31

1 ,3-Diethyl-2-(hydroxymethyl)-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

A mixture of {1-ethyl-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol (134 mg, 0.475 mmol) and iodoethane (0.384 mL, 4.75 mmol) in THF (2 mL) was heated in a microwave reactor at normal absorption for 2 h at 100°C. LC/MS showed the reaction hasn't gone to completion yet. It was resubmitted back to microwave for 1 h at 100°C°. The crude product was purified by preparatory HPLC (neutral conditions). The product fractions were evaporated, combined and concentrated under a stream of nitrogen at 50°C, giving the title compound (143 mg, 0.326 mmol, 68.7 % yield). LC/MS: m/z 31 1 .0 (M)+, Rt 0.75 min.

Intermediate 32

-Ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol

A mixture of (5-bromo-1-ethyl-1 H-benzimidazol-2-yl)methanol (300 mg, 1.176 mmol), [4- (methyloxy)phenyl]boronic acid (268 mg, 1.764 mmol), sodium carbonate (374 mg, 3.53 mmol) and PdCI₂(dppf) (86 mg, 0.1 18 mmol) in 1 ,4-Dioxane (3 mL) and water (1 mL) in a microwave vial was degassed for 2 min and then heated in a microwave at reactor at

100°C for 10 min. Water (1 mL), ether (3 mL) and ethyl acetate (1 mL) were added to the reaction mixture. Solid was filtered and then washed with water (2 mL) followed by ether (5 mL). Solid was dried to give the title compound (281 mg, 0.995 mmol, 85 % yield) as solid. LC/MS: m/z 283.2 (M+H)⁺, Rt 0.74 min.

Intermediate 33

{1 -Ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl 3,5-diamino-6-

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (238 mg, 0.995 mmol) and {1-ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol (281 mg, 0.995 mmol) in DMF (3 mL) was added triethylamine (0.277 mL, 1.991 mmol). The reaction mixture was heated in a microwave reactor at normal absorption for 1 h at 120°C. Ether (5 mL) was added and the solid was filtered and washed with ether to give the title compound (386 mg, 0.852 mmol, 86 % yield) as a yellow solid. LC/MS: m/z 453.0 (M+H)⁺, Rt 0.92 min. Intermediate 34

1 -Ethyl -6-(3-thienyl)-1 H-benzimidazol-2-yl]methanol

A mixture of (6-bromo-1-ethyl-1 H-benzimidazol-2-yl)methanol (200 mg, 0.784 mmol), 3- thienylboronic acid (100 mg, 0.784 mmol), sodium carbonate (249 mg, 2.352 mmol) and PdCI₂(dppf) (57.4 mg, 0.078 mmol) in 1 ,4-Dioxane (2 mL) and water (0.667 mL) in a microwave vial was degassed for 2 min. It was then heated in microwave reactor at 100°C for 10 min. Water (1 mL) and ethyl acetate (4 mL) were added to the reaction mixture. Solid precipitated out from the solution. Solid was filtered and then washed with water (2 mL) followed by ethyl acetate (5 mL). Solid was dried to give the title compound (144 mg, 0.557 mmol, 71 .1 % yield) as solid. LC/MS: m/z 259.0 (M+H)⁺, Rt 0.71 min.

Intermediate 35

1 ,3-Dieth l-2-(hydroxymethyl)-6-(3-thienyl)-1 H-3,1 -benzimidazol-3-ium iodide

A mixture of [1 -ethyl-6-(3-thienyl)-1 H-benzimidazol-2-yl]methanol (130 mg, 0.503 mmol) and iodoethane (0.407 mL, 5.03 mmol) in THF (2 mL) was heated in a microwave reactor at normal absorption for 45 min at 100°C. Solid was filtered and then washed with ether to give the title compound (123 mg, 0.297 mmol, 59.0 % yield). LC/MS: m/z 287.0 (M)⁺, Rt 0.77 min.

Intermediate 36

1 -Ethyl-2-(hydroxymethyl)-3-propyl-1 H-benzimidazol-3-ium iodide

A mixture of (1 -ethyl-1 H-benzimidazol-2-yl)methanol (100 mg, 0.567 mmol) and 1- iodopropane (0.554 mL, 5.67 mmol) in THF (1 mL) was heated in a microwave reactor at normal absorption for 45 min at 100°C. Solid precipitated out from the reaction mixture. Solid was filtered and then washed with ether to obtain the title compound (61 mg, 0.176 mmol, 31.0 % yield) as white solid. LC/MS: m/z 219.2 (M)⁺, Rt 0.46 min. Intermediate 37

Ethyl(5-methyl-2-nitrophenyl)amine

To a mixture of 3-fluoro-4-nitrotuluene (2 g, 12.89 mmol) and potassium carbonate (3.56 g, 25.8 mmol) in Ethanol (8 mL) was added dropwise 70 % ethylamine in water (3.13 mL, 38.7 mmol). The resulting mixture was heated in a microwave reactor at normal absorption for 10 min at 100°C. Water (20 mL) was added and extracted with DCM (2 x 40 mL). The combined organic layer was washed with saturated NaCI (20 mL), dried over sodium sulfate, filtered and then concentrated under vacuum to give the title compound (2.19 g, 12.15 mmol, 94 % yield) as yellow solid. LC/MS: m/z 180.7 (M+H)⁺, Rt 1.10 min.

Intermediate 38

( -Amino-5-methylphenyl)ethylamine

10 % Pd/C (0.193 g, 0.181 mmol) was added to a solution of ethyl(5-methyl-2- nitrophenyl)amine (2.18 g, 12.10 mmol) in Ethanol (40 mL). Hydrogen in balloon was applied and stirred at room temperature over the weekend. The reaction mixture was filtered through celite, washed with ethyl acetate (40 mL). Solvents were evaporated under reduced pressure to give the title compound (1 .80 g, 1 1 .98 mmol, 99 % yield). LC/MS: m/z 151.0 (M+H)⁺, Rt 0.58 min.

Intermediate 39

(1 -Ethyl -6-meth l-1 H-benzimidazol-2-yl)methanol

To a solution of (2-amino-5-methylphenyl)ethylamine (1.82 g, 12.12 mmol) and hydroxyacetic acid (1.106 g, 14.54 mmol) in water (10 mL) was added 6 N HCI,aq (3.03 mL, 18.17 mmol). The resulting reaction mixture was heated in a microwave reactor at normal absorption for 30 minutes at 120°C. LC/MS showed the reaction wasn't going to completion. It was resubmitted to a microwave reactor at normal absorption for 15 min at 150°C. NaOH 6 N was added to the reaction mixture till pH=12-13. Solid was precipitated out from the reaction mixture. Solid was filtered and then washed with ether to give the title compound (2.78 g, 14.61 mmol, 121 % yield) as solid. LC/MS: m/z 191.1 (M+H)⁺, Rt 0.53 min.

Intermediate 40

1 ,3-Diethyl-2-(hydroxymethyl)-6-methyl-1 H-benzimidazol-3-ium iodide

A mixture of (1-ethyl-6-methyl-1 H-benzimidazol-2-yl) methanol (150 mg, 0.788 mmol) and iodoethane (0.637 mL, 7.88 mmol) in THF (1.5 mL) was heated in a microwave reactor at normal absorption for 45 min at 100°C. Solid was filtered and then washed with ether to give the title compound (149 mg, 0.430 mmol, 54.6 % yield). LC/MS: m/z 219.1 (M)+, Rt 0.51 min.

Intermediate 41

[ -Amino-5-(methyloxy)phenyl]ethylamine

To a solution of 10 % Pd/C (0.181 g, 0.170 mmol) in Ethanol (10 mL) was added solution of ethyl[5-(methyloxy)-2-nitrophenyl]amine (2.23 g, 1 1 .37 mmol) in Ethanol (30 mL). Hydrogen was applied and stirred at room temperature for 4 h. LC/MS showed the reaction was not complete yet. It was stirred at room temperature for another 17 h. The reaction mixture was filtered through celite, washed with ethyl acetate (40 mL). Solvents were evaporated under reduced pressure to give the title compound (1 .87 g, 1 1.25 mmol, 99 % yield) as oil. LC/MS: m/z 167.1 (M+H)⁺, Rt 0.55 min.

Intermediate 42

[1 -Eth l -6-(methyloxy)-1 H-benzimidazol-2-yl]methanol

To a solution of [2-amino-5-(methyloxy)phenyl]ethylamine (1.87 g, 1 1.25 mmol) and hydroxyacetic acid (1.027 g, 13.50 mmol) in water (10 mL) was added 6 N HCI,aq (2.81 mL, 16.88 mmol). The resulting reaction mixture was heated in a microwave reactor at normal absorption for 30 minutes at 150°C. NaOH 6 N was added to the reaction mixture until pH=12-13. Solid was precipitated out from the reaction mixture. Solid was filtered and then washed with ether to give the title compound (2.32 g, 1 1 .25 mmol, 100 % yield) as solid. LC/MS: m/z 207.1 (M+H)⁺, Rt 0.39 min.

Intermediate 43

1 ,3-Diethyl-2-(hydroxymethyl)-6-(methyloxy)-1 H-benzimidazol-3-ium iodide

A mixture of [1 -ethyl-6-(methyloxy)-1 H-benzimidazol-2-yl]methanol (150 mg, 0.727 mmol) and iodoethane (0.588 mL, 7.27 mmol) in THF (1 mL) was heated in a microwave reactor for 45 min at 100°C. Solid was filtered and then washed with ether to give the title compound (267 mg, 0.737 mmol, 101 % yield) as a solid. LC/MS: m/z 234.7 (M)⁺, Rt 0.57 min.

Intermediate 44

ethyl)amine

A mixture of [(5-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]amine (100 mg, 0.394 mmol), [4-(methyloxy)phenyl]boronic acid (78 mg, 0.512 mmol), Na₂C0₃ (125 mg, 1 .181 mmol) and PdCI₂(dppf) (28.8 mg, 0.039 mmol) in 4 mL of a 1 :3 ratio of water (1 mL):1 ,4-Dioxane (3.00 mL) was degassed for 5 min and then submitted to Microwave at 100°C for 3 h. The crude product was filtered and the solid residue was rinsed with ethyl acetate. The organic layer was concentrated under reduced pressure to afford the title compound (196.4 mg, 0.384 mmol, 98 % yield). LC/MS: m/z 282.1 (M+H)⁺, Rt 0.76 min.

Intermediate 45

3,5-Diamino-6-chloro-N-({1 -ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2- yl}methyl)-2-pyrazinecarboxamide

To a solution of ({1 -ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl)amine (196.4 mg, 0.384 mmol) in DMSO (2 mL) was added 3-chloro-5-(1 H-imidazol-1 - ylcarbonyl)-2,6-pyrazinediamine (60 mg, 0.251 mmol), followed by the addition of triethylamine (0.076 mL, 0.545 mmol). The reaction mixtures were stirred at room temperature for 19 h. LC-MS showed the major peak was the product but since there was still some starting material, the reaction mixture was heated at 50°C for 3 h but the reaction did not improve a lot. The crude product was purified by preparatory HPLC (with 0.1 % NH₄OH). The desired fractions were evaporated down over geneVac EZ-2 evaporator under vacuum. It was then combined and concentrated under a stream of nitrogen at 50°C to afford the title compound (20.3 mg, 0.045 mmol, 1 1 .70 % yield). LC/MS: m/z 452.0 (M+H)⁺, Rt 0.84 min.

Intermediate 46

4-bromo-N-ethyl-2-nitroaniline

A mixture of 4-bromo-1 -fluoro-2-nitrobenzene (40 g, 182 mmol), ethylamine hydrochloride (74.1 g, 909 mmol) and Et₃N (1 10 g, 1091 mmol) was stirred at 80°C for 2h before was added NaHC0₃ (150 mL, sat. aq.). The layers were separated, and the aqueous layer was extracted with EtOAc (2 x 150 mL). Isolute was added to the combined organic layer and the mixture was dried over Na₂S0₄ for 1 h before was concentrated under reduced pressure to afford the title compound (40 g, 163 mmol, 90 % yield). LC/MS: m/z 245 (M+H)⁺, Rt 1.84 min.

Intermediate 47

(2-amino-4-bromophenyl)ethyl

A suspension of (4-bromo-2-nitrophenyl)ethylamine (40 g, 163 mmol), and tin(ll) chloride (155 g, 816 mmol) in HCI(4M) (15 mL) was stirred at 85°C for 2h. The reaction mixture was diluted with EtOAc then was added NaOH (500 mL, sat. aq.) and EtOAc (150 mL). The organic layer was separated, and the aqueous layer was extracted with EtOAc (2 x 150mL). Isolute was added to the combined organic layer. This mixture was dried over Na₂S0₄ for 1 h and concentrated under reduced pressure. This crude product was purified by column chromatography to afford the title compound (20 g, 93 mmol, 57.0 % yield). LC/MS: m/z 215 (M+H)⁺, Rt 1.60 min.

Intermediate 48

1 1 -dimethylethyl [(5-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate

To a mixture of (2-amino-4-bromophenyl)ethylamine (15 g, 69.7 mmol), 1 , 1 -dimethylethyl (2-oxoethyl)carbamate (16.65 g, 105 mmol) in ethanol (300 ml.) and water (100 ml.) was added sodium bisulfite (29.0 g, 279 mmol). The result mixture was heated to 60°C for 3h before was filtrated, washed with cold ethanol. The organic layer was separated and concentrated under reduced pressure. This crude product was added NaHC0₃ (150 ml_, sat. aq.), extracted with EtOAc (3 x 50 ml_). The combined organic layer was dried over Na₂S0₄ for 1 h and concentrated under reduced pressure. This crude product was purified by column chromatography to afford the title compound (6.5 g, 18.35 mmol, 26.3 % yield). LC/MS: m/z 354 (M+H)⁺, Rt 1.19 min.

Intermediate 49

-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]amine

To a solution of 1 , 1 -dimethylethyl [(5-bromo-1 -ethyl-1 H-benzimidazol-2- yl)methyl]carbamate (6.5 g, 18.35 mmol) in DCM (100 ml.) was added trifluoroacetic acid (31.4 g, 275 mmol) slowly. The result mixture was stirred at room temperature for 1 h before was basified with Et₃N. Then was added silica gel (100-200μηη) and concentrated under reduced pressure. This crude mixture was purified by column chromatography to afford the title compound (4.3 g, 16.92 mmol, 92 % yield). LC/MS: m/z 254 (M+H)⁺, Rt 1.49 min.

Intermediate 50

5-bromo-N-ethyl-2-nitroaniline

A mixture of 4-bromo-2-fluoro-1 -nitrobenzene (40 g, 182 mmol), ethylamine hydrochloride (44.5 g, 545 mmol) in Et₃N (73.6 g, 727 mmol) was stirred at 80°C for 2h before was added NaHC0₃ (200 ml_, sat. aq.). The layers were separated, and the aqueous layer was extracted with EtOAc (2 x 200 ml_). Isolute was added into the combined organic layer. Then this mixture was dried over Na₂S0₄ for 1 h and concentrated under reduced pressure to afford the title compound (20 g, 82 mmol, 90 % yield). LC/MS: m/z 245 (M+H)⁺, Rt 1.84 min.

Intermediate 51

(2-amino-5-bromophenyl)ethylamine

To a suspension of (5-bromo-2-nitrophenyl)ethylamine 5-bromo-N-ethyl-2-nitroaniline (8g, 32.6 mmol) in 4N HCI (200 ml.) was added tin(ll) chloride (30.9 g, 163 mmol). The resulting mixture was heated to 85°C and stirred at this temperature for 2 h. After completion, the reaction solution was basified with NaOH solution, extracted with EtOAc (500ml x 3). The organic phase was combined and concentrated under vacuum to afford the title compound (6g, 27.9 mmol, 85 % yield). LC/MS: m/z 215 (M+H)⁺, Rt 1 .39 min.

Intermediate 52

1 1 -dimethylethyl [(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate

To a solution of (2-amino-5-bromophenyl)ethylamine (6 g, 27.9 mmol) in ethanol (18 ml.) and water (6.00 ml.) was added 1 ,1 -dimethylethyl (2-oxoethyl)carbamate (4.44 g, 27.9 mmol) and sodium bisulfite (10.60 g, 102 mmol). The reaction mixture was stirred at 60°C for 26 h, cooled down and the solid was filtered. The filtrate was concentrated under vacuum to afford the title compound (6 g, 16.94 mmol, 60.7 % yield). LC/MS: m/z 354 (M+H)⁺, Rt 1.40 min. Intermediate 53

-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]amine dihydrochloric salt

To 1 ,1 -dimethylethyl [(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (6 g, 16.94 mmol) was added to 4M HCI in methanol (250 mL) at room temperature and stirred for 20 hours. Then the solvent was removed under vacuum to afford the title compound (5.1 g, 14.99 mmol, 89 % yield). LC/MS: m/z 254 (M+H)⁺, Rt 1.37 min.

Intermediate 54

3, 5-diamino-N-[(6-bromo-1 -ethyl-1 H-benzimidazol-2 -yl)methyl]-6-chloro-2- razinecarboxamide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (239 mg, 1 mmol) in DMSO (5 mL) was added 1 -(6-bromo-1 -ethyl-1 H-benzimidazol-2 - yl)methanamine dihydrochloric salt (327 mg, 1 .000 mmol) then TEA (0.418 mL, 3.00 mmol). The resulting reaction mixture was stirred at room temperature for 17 h before was purified with preparatory HPLC (with 0.1 % NH40H) to afford the title compound (261 .6 mg, 0.616 mmol, 61 .6 % yield). LC/MS: m/z 424 (M+H)⁺, Rt 0.79 min.

Intermediate 55

3,5-diamino-N-[(5-bromo-1 -ethyl-1 H-benzimidazol-2 -yl)methyl]-6-chloro-2- razinecarboxamide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (239 mg, 1 mmol) in DMSO (5 mL) was added 1 -(5-bromo-1 -ethyl-1 H-benzimidazol-2 - yl)methanamine (254 mg, 1 .000 mmol) then TEA (0.139 mL, 1 .000 mmol). The resulting reaction mixture was stirred at room temperature for 17 h before was purified with preparatory HPLC (with 0.1 % NH40H) to afford the title compound (235.4 mg, 0.554 mmol, 55.4 % yield). LC/MS: m/z 424 (M+H)⁺, Rt 0.76 min. Intermediate 56

N-ethyl-4-(methylsulfonyl)-2-nitroaniline

To a mixture of 1-fluoro-4-(methylsulfonyl)-2-nitrobenzene (2 g, 9.12 mmol) and potassium carbonate (3.15 g, 22.81 mmol) in ethanol (10 ml) was added a solution of ethylamine (22.81 ml, 45.6 mmol) in THF. The resulting mixture was stirred at 80°C for 1.5 hour. After cooling to room temperature, the mixture was filtered and evaporated to give N-ethyl-4-(methylsulfonyl)-2-nitroaniline as a yellow solid (1 g, 4.09 mmol, 44.9 % yield). LC/MS: m/z 245.2 (M+H)⁺, Rt 0.71 min.

Intermediate 57

-ethyl -4-(methylsulfonyl)-1 ,2-benzenediamine

N-ethyl-4-(methylsulfonyl)-2-nitroaniline (1 g, 4.09 mmol) was dissolved in hot ethanol (300 mL) and 1 N HCI (4.1 ml). Attempts were made to hydrogenate the starting material using the H-Cube hydrogenation apparatus (20-35°C, 1 ml/mn, 10%Pd/C or 10%

Pd(OH)₂ cartridges), which resulted in little conversion of the nitro derivative to the aniline. All the solutions (collected product and remaining starting material) were combined and evaporated to give a brown yellow residue (900 mgs) containing mainly the starting material with ca 20% of the desired product. The mixture was dissolved in THF (30 ml) and methanol (10 ml), 10% Pd/C (1 15 mg) was added. The resulting mixture was hydrogenated using conventional methods by applying hydrogen in a balloon over 17 hours. The catalyst was filtered over celite. The filtrate was evaporated to give a residue which was dissolved in DCM (100 mL) and washed with 2M Na₂C0₃ (20 mL). The aq. layer was separated and extracted with DCM (50mL). The DCM layers were combined, dried over MgS0₄ and evaporated to give N1 -ethyl-4-(methylsulfonyl)-1 ,2- benzenediamine (0.75 g, 3.50 mmol, 85 % yield). LC/MS: m/z 215.1 (M+H)⁺, Rt 0.58 min. Intermediate 58

1 ,1 -dimethylethyl {[1 -ethyl -5-(methylsulfonyl)-1 H-benzimidazol-2- yl]methyl}carbamate

A mixture of N 1-ethyl-4-(methylsulfonyl)-1 ,2-benzenediamine (0.75 g, 3.50 mmol) , 1 , 1- dimethylethyl (2-oxoethyl)carbamate (0.557 g, 3.50 mmol) and sodium bisulfite (1 .165 g, 1 1 .20 mmol) in ethanol (21 ml) and water (7.00 ml) was heated at 60°C for 16 hours. The solvents were evaporated and the residue was partitioned between ethyl acetate (70 mL) and water (30 mL). The water layer was separated and extracted with ethyl acetate (70 mL). The organics were combined, dried over MgS0₄ and evaporated to give 1 ,1 - dimethylethyl {[1-ethyl-5-(methylsulfonyl)-1 H-benzimidazol-2-yl]methyl}carbamate (536.9 mg, 1 .519 mmol, 43.4 % yield). LC/MS: m/z 354.1 (M+H)⁺, Rt 0.71 min.

Intermediate 59

3,5-diamino-6-chloro-N-{[1 -ethyl-5-(methylsulfonyl)-1 H-benzimidazol-2-yl]methyl}-2- pyrazinecarboxamide

A mixture of 1 , 1 -dimethylethyl {[1-ethyl-5-(methylsulfonyl)-1 H-benzimidazol-2- yl]methyl}carbamate (557 mg, 1.576 mmol) and 4M HCI in dioxane (5 mL, 20.00 mmol) in DCM (5 mL) was stirred at room temperature. Removal of the solvent under vacuum gave a brown solid, which was redissolved in DMSO (5 mL) and 3-chloro-5-(1 H-imidazol- 1-ylcarbonyl)-2,6-pyrazinediamine (376 mg, 1 .576 mmol) was added. The resulting mixture was stirred at room temperature. The reaction mixture was allowed to stir at room temperature for 5 days and then purified by preparatory HPLC under neutral conditions to give :3,5-diamino-6-chloro-N-{[1 -ethyl-5-(methylsulfonyl)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (1 10.2 mg, 0.260 mmol, 16.50 % yield). LC/MS: m/z 423.9 (M)⁺, Rt 0.65 min. Intermediate 60

-ethyl -6-[2-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol

A mixture of (6-bromo-1-ethyl-1 H-benzimidazol-2-yl)methanol (200 mg, 0.784 mmol), [2- (methyloxy)phenyl]boronic acid (179 mg, 1.176 mmol), Na₂C0₃ (249 mg, 2.352 mmol) and PdCI₂(dppf) (57.4 mg, 0.078 mmol) in 4 mL of a 1 :3 ratio of water: 1 ,4-dioxane was degassed for 2 min and then treated in a microwave reactor at 100°C for 10 min. Water (1 mL), ether (3 mL) and ethyl acetate (1 mL) were added to the reaction mixture. The solid present in the reaction mixture was filtered and the resulting filtrate was concentrated down. 2 mL of water were added to the filtrate to dissolve the base. The product was extracted with DCM (2 x 3 mL). The combined organic layers were concentrated down to afford the title compound (282.3 mg). LC/MS: m/z 283.0 (M+H)⁺, Rt 0.77 min.

Intermediate 61

1 ,3-diethyl-2-(hydroxymethyl)-6-[2-(methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

A mixture of {1-ethyl-6-[2-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol (282.3 mg,

1.000 mmol) and iodoethane (0.808 mL, 10.00 mmol) in THF (2 mL) was heated in a microwave reactor for 40 min at 100°C. 1 mL of Et₂0 was added. The solid was filtered and then washed with ether to afford the title compound (141 .8 mg, 32.4 % yield). LC/MS: m/z 31 1 .1 (M)⁺, Rt 0.82 min.

Intermediate 62

1.1 -dimethylethyl ({1 -ethyl -5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2- yl}methyl)carbamate

A mixture of 1 , 1 -dimethylethyl [(5-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (50 mg, 0.141 mmol), [4-(methyloxy)phenyl]boronic acid (27.9 mg, 0.183 mmol), Na₂C0₃ (44.9 mg, 0.423 mmol) and PdCI₂(dppf) (10.33 mg, 0.014 mmol) in 4 ml. of a 1 :3 ratio of water: 1 ,4-dioxane was degassed for 5 min and treated in a microwave reactor at 100°C. It was passed through a PL-Thiol MP SPE+ cartridge. The cartridge and the vial were rinsed with EtOAc. 1 ml. of water was added to dissolve the base and the product was extracted with EtOAc. The organic layer was concentrated down to afford the title compound (61 .7 mg, 80 % yield). LC/MS: m/z 382.0 (M+H)⁺, Rt 0.90 min.

Intermediate 63

( -ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl)amine

To a solution of 1 , 1 -dimethylethyl ({1-ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2- yl}methyl)carbamate (61.7 mg, 0.1 13 mmol) in DCM (1 .5 mL) was added 4M HCI in dioxane (0.404 mL, 1.617 mmol). The reaction mixture was stirred at room temperature for 25.5 h. The reaction mixture was concentrated down to afford the title compound (40.1 mg, 100 % yield). LC/MS: m/z 282.0 (M+H)⁺, Rt 0.76 min.

Intermediate 64

3,5-diamino-6-chloro-N-({1 -ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2- l}methyl)-2-pyrazinecarboxamide

To a solution of ({1 -ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl)amine (40.1 mg, 0.1 13 mmol) in DMSO (2 mL) was added 3-chloro-5-(1 H-imidazol-1 - ylcarbonyl)-2,6-pyrazinediamine (27.0 mg, 0.1 13 mmol) and triethylamine (0.189 mL, 1.358 mmol). The reaction mixtures were stirred at room temperature for 17 h. Another equivalent of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (38.6 mg, 0.162 mmol) was added and the mixture was stirred at room temperature for 24 h. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (19 mg, 37.1 % yield). LC/MS: m/z 452.0 (M+H)⁺, Rt 0.86 min. Intermediate 65

1 ,1 -dimethylethyl ({1 -ethyl -6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2- yl}methyl)carbamate

A mixture of 1 , 1-dimethylethyl [(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (50 mg, 0.141 mmol), [4-(methyloxy)phenyl]boronic acid (27.9 mg, 0.183 mmol), Na₂C0₃ (44.9 mg, 0.423 mmol) and PdCI₂(dppf) (10.33 mg, 0.014 mmol) in 4 mL of a 1 :3 ratio of water: 1 ,4-dioxane was degassed for 5 min and heated in a microwave reactor at 100°C for 10 min. The mixture was passed through a PL-Thiol MP SPE+ cartridge. The cartridge and the vial were rinsed with EtOAc. 1 mL of water was added to dissolve the base and the product was extracted with EtOAc. The organic layer was concentrated down to afford the title compound (44 mg, 82 % yield). LC/MS: m/z 382.0 (M+H)⁺, Rt 0.90 min.

Intermediate 66

( 1 -ethyl-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl)amine

To a solution of 1 , 1 -dimethylethyl ({1-ethyl-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2- yl}methyl)carbamate (44 mg, 0.1 15 mmol) in DCM (1.5 mL) was added 4M HCI in dioxane (0.288 mL, 1.153 mmol). The reaction mixture was stirred at room temperature for 23 hours and concentrated down to afford the title compound (28 mg, 86 % yield). LC/MS: m/z 282.0 (M+H)⁺, Rt 0.75 min.

Intermediate 67

3,5-diamino-6-chloro-N-({1 -ethyl-6-[4-(methyloxy)phenyl]-1 H-benzi

yl}methyl)-2-pyrazinecarboxamide

o— To a solution of ({1-ethyl-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl)amine (28 mg, 0.100 mmol) in DMSO (1 .5 mL) was added 3-chloro-5-(1 H-imidazol-1-ylcarbonyl)- 2,6-pyrazinediamine (28.5 mg, 0.1 19 mmol) and triethylamine (0.160 mL, 1.144 mmol). The reaction mixture was stirred at room temperature for 70 h and purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (13 mg, 28.9 % yield). LC/MS: m/z 452.0 (M+H)⁺, Rt 1 .07 min.

Intermediate 68

1 ,1 -dimethylethyl {[1 -ethyl -6-(4-methylphenyl)-1 H-benzimidazol-2- yl]methyl}carbamate

A mixture of 1 , 1 -dimethylethyl [(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (120 mg, 0.339 mmol), (4-methylphenyl)boronic acid (59.9 mg, 0.440 mmol), Na₂C0₃ (108 mg, 1 .016 mmol) and PdCI₂(dppf) (24.79 mg, 0.034 mmol) in 4 mL of a 1 :3 ratio of water: 1 ,4-dioxane was degassed for 5 min and then heated in a microwave reactor at 100°C for 10 min. The mixture was passed through a PL-Thiol MP SPE+ cartridge. The cartridge and the vial were rinsed with EtOAc. 1 mL of water was added to dissolve the base and the product was extracted with EtOAc. The organic layer was concentrated down to afford the title compound (100 mg, 81 % yield). LC/MS: m/z 366.1 (M+H)⁺, Rt 1.01 min.

Intermediate 69

3,5-diamino-6-chloro-N-{[1 -ethyl-6-(4-methylphenyl)-1 H-benzimidazol-2-yl]methyl}- 2-pyrazinecarboxamide

To a solution of 1 , 1 -dimethylethyl {[1 -ethyl-6-(4-methylphenyl)-1 H-benzimidazol-2- yl]methyl}carbamate (100 mg, 0.274 mmol) in DCM (3 mL) was added 4 M HCI in dioxane (0.684 mL, 2.74 mmol). The reaction mixture was stirred at room temperature for 18 h and concentrated down to afford the intermediate. It was taken back in DMSO (2 mL). 3- chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (65.3 mg, 0.274 mmol) and triethylamine (0.496 mL, 3.56 mmol) were added to the reaction mixture which was stirred at room temperature for 72 h and purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (51 mg, 42.8 % yield). LC/MS: m/z 436.0 (M+H)⁺, Rt 0.95 min.

Intermediate 70

4- ethylamino)-3-nitrobenzoic acid

To a mixture of 4-fluoro-3-nitrobenzoic acid (2 g, 10.80 mmol) and potassium carbonate (2.99 g, 21.61 mmol) in ethanol (10 mL) was added dropwise 70 % ethylamine in water (2.62 mL, 32.4 mmol). The resulting mixture was heated in microwave for 10 min at 100°C. HCI 1 N was added until we observed the formation of a yellow precipitate. The precipitate was filtered out and dried to afford the title compound (2.1 1 g, 93 % yield). LC/MS: m/z 21 1 .0 (M+H)⁺, Rt 0.77 min.

Intermediate 71

-amino-4-(ethylamino)benzoic acid

10 % Pd/C (0.136 g, 0.128 mmol) was added to a solution of 4-(ethylamino)-3- nitrobenzoic acid (1.79 g, 8.52 mmol) in THF (40 mL). Hydrogen in balloon was applied and the mixture was stirred at room temperature for 66 h. The reaction mixture was filtered through celite and the celite was washed with ethyl acetate. The resulting filtrate was concentrated down and dried to afford the title compound (1 .475 g, 96 % yield) as 74-A1. LC/MS: m/z 181.1 (M+H)⁺, Rt 0.52 min.

Intermediate 72

2-[({[(1 ,1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1 -ethyl-1 H-benzimidazole-5- carboxylic acid

A mixture of 3-amino-4-(ethylamino)benzoic acid (500 mg, 2.77 mmol), 1 , 1-dimethylethyl (2-oxoethyl)carbamate (442 mg, 2.77 mmol) and sodium bisulfite (924 mg, 8.88 mmol) in ethanol (9 mL) and water (3 mL) was heated at 60°C for 48 h. The reaction mixture was filtered and the solid was rinsed with DCM. The resulting filtrate was concentrated down to afford the title compound (1250 mg). LC/MS: m/z 320.0 (M+H)⁺, Rt 0.68 min.

Intermediate 73

2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-5-carboxylic acid

To a solution of 2-[({[(1 , 1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1 -ethyl-1 H- benzimidazole-5-carboxylic acid (1 100 mg, 2.41 1 mmol) in DCM (12 ml.) was added 4 M HCI in dioxane (3.01 ml_, 12.06 mmol). The reaction mixture was stirred at room temperature for 22 h and was concentrated down to afford the deboc intermediate. It was taken back in DMSO (12 ml_). 3-chloro-5-(1 H-imidazol-1-ylcarbonyl)-2,6-pyrazinediamine (575 mg, 2.41 1 mmol) and triethylamine (5.5 ml_, 39.5 mmol) were added to the reaction mixture which was stirred at room temperature for 66h and heated at 50°C for 24 h. The salts present in the reaction mixture were filtered and rinsed with DMSO. The filtrate was concentrated down. The crude product was taken back in DCM and the product crashed out from the solution. Thus the solid was filtered and rinsed with DCM to afford the title compound (960 mg). LC/MS: m/z 390.0 (M+H)⁺, Rt 0.56 min.

Intermediate 74

Ethyl-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-5-carboxylate

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl- 1 H-benzimidazole-5-carboxylic acid (100 mg, 0.257 mmol) in EtOH (8 mL) was added H₂S0₄ (0.137 mL, 2.57 mmol). The reaction mixture was heated at reflux overnight and then concentrated down. 2 mL of water were added to the crude product and it was neutralized with NaOH 6N. It was then extracted with DCM and the resulting organic layer was washed with saturated NaHC0₃ and with brine. The organic layer was concentrated down to afford the title compound (35 mg, 32.7 % yield). LC/MS: m/z 418.0 (M+H)⁺, Rt 0.79 min. Intermediate 75

Methyl-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-5-carboxylate

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-5-carboxylic acid (250 mg, 0.641 mmol) in dry MeOH (5 mL) was added H₂S0₄ (0.068 mL, 1 .283 mmol). The reaction mixture was heated at reflux for 20 h and then concentrated down. The crude product was taken back in DCM, washed 3 times with saturated NaHC0₃ and once with brine. The organic layer was concentrated down to afford the title compound (94 mg, 36.3 % yield). LC/MS: m/z 404.0 (M+H)⁺, Rt 0.74 min.

Intermediate 76

Methyl 4-amino-3-(ethylamino)benzoate

10 % Pd/C (0.367 g, 0.345 mmol) was added to a solution of methyl 3-(ethylamino)-4- nitrobenzoate (5.15 g, 22.97 mmol) in THF (100 mL). Hydrogen in balloon was applied and it was stirred at room temperature overnight. The reaction mixture was filtered through celite and the celite was washed with ethyl acetate. The resulting filtrate was concentrated down and dried to afford the title compound (4.268 g, 96 % yield) as 8-A1. LC/MS: m/z 195.0 (M+H)⁺, Rt 0.61 min.

Intermediate 77

Methyl 2-[({[(1 ,1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1 -ethyl-1 H-

A mixture of methyl 4-amino-3-(ethylamino)benzoate (4.128 g, 21.25 mmol), 1 , 1- dimethylethyl (2-oxoethyl)carbamate (3.38 g, 21.25 mmol) and sodium bisulfite (7.08 g, 68.0 mmol) in MeOH (90 mL) and water (30 mL) was heated at 60°C for 70.5 h and then concentrated down. The product was extracted from the remaining water with DCM and the organic layer was then concentrated down. The crude product was purified by automated flash chromatography to afford the title compound (5.683 g, 80.2 % yield). LC/MS: m/z 334.0 (M+H)⁺, Rt 0.80 min.

Intermediate 78

Methyl-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H-

To a solution of methyl 2-[({[(1 , 1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1 -ethyl-1 H- benzimidazole-6-carboxylate (3.5175 g, 10.55 mmol) in DCM (100 mL) was added 4 M HCI in dioxane (14.55 mL, 58.2 mmol). The reaction mixture was stirred at room temperature for 24 hours and was then concentrated down. The intermediate was taken back in DMSO (30 mL). 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (2.52 g, 10.55 mmol) and triethylamine (14.71 mL, 106 mmol) were added to the reaction mixture which was stirred at 50C for 28 h. The salts present in the reaction mixture were filtered and washed with DCM. The reaction mixture was then concentrated down to get rid of the DCM. Since the solution was saturated we could observe some solid in the reaction mixture. This precipitate was filtered out and washed with ether. To encourage the precipitation of the remaining product, a large amount of ether was added to the filtrate and it was sonicated. The precipitate was filtered out and washed with ether. Both solid were combined and dried to afford the title compound (3.783 g, 88.8 % yield). LC/MS: m/z 403.9 (M+H)⁺, Rt 0.74 min.

Intermediate 79

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H-

A mixture of methyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 - ethyl-1 H-benzimidazole-6-carboxylate (1 g, 2.476 mmol), and potassium carbonate (1 .027 g, 7.43 mmol) in MeOH (10 mL) and water (5 mL) was heated in a microwave reactor for 1 hour at 100°C. 10 mL of water were added and the resulting aqueous layer was washed with 10 mL of DCM. HCI 1 N was then added until pH < 1 . The precipitate the reaction mixture was filtered, washed with ether, and dried to afford the title compound (514 mg, 53.2 % yield). LC/MS: m/z 389.9 (M+H)⁺, Rt 0.61 min.

Intermediate 80

2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-N,1 -diethyl-1 H-

A solution of 50 wt% T₃P in EtOAc (0.1 19 mL, 0.200 mmol) was added to a solution of 2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H-benzimidazole-6- carboxylic acid (60 mg, 0.154 mmol), ethylamine, 2M in THF (0.231 mL, 0.462 mmol), and triethylamine (0.085 mL, 0.616 mmol) in DMSO (2 mL). It was stirred at 50°C for 72 hours. More triethylamine (0.043 mL, 0.308 mmol) and ethylamine, 2M in THF (0.154 mL, 0.308 mmol) were added. It was heated at 80°C for 2 hours. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (18 mg, 28.1 % yield). LC/MS: m/z 416.9 (M+H)⁺, Rt 0.65 min.

Intermediate 81

Butyl-2 -({[(3, 5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylate

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylic acid (100 mg, 0.257 mmol) in n-butanol (2 mL) and DMSO

(0.5 mL) was added H₂S0₄ (0.041 mL, 0.770 mmol). The reaction mixture was heated at reflux for 6 hours. 0.5 mL of saturated NaHC0₃ was added to neutralize the reaction mixture, which was then concentrated down. The crude product was then purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (134 mg, 35.1 % yield).

LC/MS: m/z 446.2 (M+H)⁺, Rt 0.92 min. Intermediate 82

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-N-(2- oxotetrahydro-3-thienyl)-1 H-benzimidazole-6-carboxamide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylic acid (100 mg, 0.257 mmol) in DMSO (2 mL) at room temperature was added HATU (102 mg, 0.268 mmol) then triethylamine (0.072 mL, 0.513 mmol). After stirring for 5 mins, 3-aminodihydro-2(3H)-thiophenone (50 mg, 0.325 mmol) was added and the reaction mixture was stirred at room temperature for 5 hours.

The crude product was then purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (77 mg, 61 .4 % yield). LC/MS: m/z 489.2 (M+H)⁺, Rt 0.62 min.

Intermediate 83

Methyl N-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- -benzimidazol-6-yl]carbonyl}glycinate

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylic acid (100 mg, 0.257 mmol) in DMSO (2 mL) at room temperature was added HATU (98 mg, 0.257 mmol) then triethylamine (0.072 mL, 0.513 mmol). After stirring for 5 mins, methyl glycinate (97 mg, 0.770 mmol) was added and it was stirred at room temperature for 26 hours. The crude product was then purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (64 mg, 54.1 % yield). LC/MS: m/z 461 .2 (M+H)⁺, Rt 0.58 min.

Intermediate 84

Ethyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylate

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl- 1 H-benzimidazole-6-carboxylic acid (60 mg, 0.154 mmol) in ethanol (3 ml.) was added H₂S0₄ (0.041 ml_, 0.769 mmol). The mixture was heated at reflux for 7 hours then cooled down. 1 ml. of DMSO was added to dissolve the product which had partially crashed out and 0.4 ml. of a saturated solution of NaHC0₃ was added to neutralize the solution. The reaction mixture was concentrated down. The crude product was then purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (35.8 mg, 55.7 % yield). LC/MS: m/z 418.1 (M+H)⁺, Rt 0.75 min.

Intermediate 85

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-N-(2- oxotetrahydro-3-furanyl)-1 H-benzimidazole-6-carboxamide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylic acid (100 mg, 0.257 mmol) in DMSO (2 ml.) at room temperature was added HATU (102 mg, 0.268 mmol) and triethylamine (0.072 ml_, 0.513 mmol). After stirring for 5 mins, 3-aminodihydro-2(3H)-furanone (90 mg, 0.494 mmol) was added and it was stirred at room temperature for 5 hours. More HATU was added (48 mg, 0.126 mmol) and it was stirred at room temperature for 3 hours. The sample was concentrated down and the crude product was carried on to the next step. LC/MS: m/z 473.2 (M+H)⁺, Rt 0.57 min.

Intermediate 86

{1 -Ethyl -6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl 3,5-diamino-6-chloro- 2- razinecarboxylate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (250 mg, 1.048 mmol) and {1-ethyl-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methanol (296 mg, 1 .048 mmol) in DMF (3 ml.) was added triethylamine (0.292 ml_, 2.095 mmol). The reaction mixture was heated in a microwave reactor at normal absorption for 1 h at 120°C and then for another hour at 120°C. A solid precipitated out from the reaction mixture and was filtered then washed with ether to give the target compound (410 mg, 0.905 mmol, 86 % yield) as a solid. LC/MS: m/z 453.0 (M+H)⁺, Rt 0.88 min.

Intermediate 87

3-Butyl-1 -ethyl-2-(hydroxymethyl)-1 H-benzimidazol-3-ium iodide

A mixture of (1 -ethyl-1 H-benzimidazol-2-yl)methanol (100 mg, 0.567 mmol) and 1- iodobutane (1044 mg, 5.67 mmol) in THF (1.5 mL) was heated in a microwave reactor at normal absorption for 45 min at 100°C. Some solid precipitated out from the reaction mixture. It was stirred at room temperature over the weekend. More solid precipitated out from the reaction mixture. The solid was filtered and then washed with ether to obtain the title compound (77 mg, 0.214 mmol, 37.7 % yield) as a white solid. LC/MS: m/z 233.1 (M)⁺, Rt 0.57 min.

Intermediate 88

1 -Ethyl-3-(2-hydroxyethyl)-2-(hydroxymethyl)-1 H-benzimidazol-3-ium bromide

r

HO Br

A mixture of (1 -ethyl-1 H-benzimidazol-2-yl)methanol (100 mg, 0.567 mmol) and 2- bromoethanol (709 mg, 5.67 mmol) in THF (1.5 mL) was heated in a microwave reactor at normal absorption for 2 h at 100°C. A solid precipitated out from the reaction mixture and was filtered off to give the title compound (56 mg, 0.186 mmol, 32.8 % yield).

LC/MS: m/z 221.1 (M)⁺, Rt 0.49 min.

Intermediate 89

3-(cyclopropylmethyl)-1 -ethyl-2- hydroxymethyl)-1 H-benzimidazol-3-ium bromide

A mixture of (1 -ethyl-1 H-benzimidazol-2-yl)methanol (100 mg, 0.567 mmol) and

(bromomethyl)cyclopropane (766 mg, 5.67 mmol) in THF (1.5 mL) was heated in a microwave reactor at normal absorption for 1 h at 100°C. Some solid was precipitated out from the reaction mixture. It was re-heated in a microwave reactor at normal absorption for 1 h at 100 °. A solid was filtered off to give the title compound (42 mg, 0.135 mmol, 23.78 % yield). LC/MS: m/z 231 .2 (M)⁺, Rt 0.66 min.

Intermediate 90

1 ,1 -dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate

A mixture of 1 ,2-phenylenediamine (1 g, 9.25 mmol), 1 , 1 -dimethylethyl (2- oxoethyl)carbamate (1 .472 g, 9.25 mmol) and sodium bisulfite (3.51 g, 33.8 mmol) in ethanol (6 mL) and water (2 mL) was heated at 60°C for 17 h. It was cooled to room temperature and stirred at room temperature for 17 h. The solid from the reaction mixture was filtered and then washed with DCM (10 mL). The filtrate was concentrated to give the crude product. It was purified by automated flash chromatography to give the title compound (1.61 g, 6.51 mmol, 70.4 % yield). LC/MS: m/z 248.1 (M+H)+, Rt 0.63 min.

Intermediate 91

3,5-diamino-N-(1 -benzimidazol-2-ylmethyl)-6-chloro-2-pyrazinecarboxamide

H_{2 M}N A NAM NHH₂

To a solution of 1 , 1 -dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate (31 1 mg, 1.257 mmol) in DCM (4 mL) was added 4 M HCI in dioxane (3.14 mL, 12.57 mmol). The mixture was stirred at room temperature for 8 h. It was concentrated under a stream of nitrogen at 50°C to give the intermediate. 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6- pyrazinediamine (300 mg, 1 .257 mmol), triethylamine (3.50 mL, 25.1 mmol) and DMSO (4 mL) were added to the intermediate. The reaction mixture was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (245 mg, 0.771 mmol, 61.33 % yield). LC/MS: m/z 317.9 (M+H)⁺, Rt O.49 min. Intermediate 92

-dimethyleth l [(1 -ethyl -6-methyl-1 H-benzimidazol-2-yl)methyl]carbamate

A mixture of (2-amino-5-methylphenyl)ethylamine (250 mg, 1 .664 mmol), 1 , 1- dimethylethyl (2-oxoethyl)carbamate (265 mg, 1 .664 mmol) and sodium bisulfite (632 mg, 6.07 mmol) in ethanol (3 mL) and water (1 mL) was heated at 60°C for 17 h. The solid was filtered and then washed with methanol (10 mL). The filtrate was then concentrated to give the crude residue. The crude product was dissolved in DCM (5 mL). The solid precipitated out from the DCM solution. The solid was filtered and the filtrate was purified by automated flash chromatography to give the title compound (380 mg, 1 .313 mmol, 79 % yield). LC/MS: m/z 290.1 (M+H)+, Rt 0.73 min.

Intermediate 93

3,5-diamino-6-chloro-N-[(1 -ethyl-6-methyl-1 H-benzimidazol-2-yl)methyl]-2- pyrazinecarboxamide

To a solution of 1 , 1 -dimethylethyl [(1 -ethyl-6-methyl-1 H-benzimidazol-2- yl)methyl]carbamate (196 mg, 0.677 mmol) in DCM (4 mL) was added 4 M HCI in dioxane (1 .693 mL, 6.77 mmol). The mixture was stirred at room temperature for 18.5 hours. It was concentrated under a stream of nitrogen at 50°C to give the intermediate. 3-chloro- 5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (162 mg, 0.677 mmol), triethylamine (0.944 mL, 6.77 mmol) and DMSO (4 mL) was added to the intermediate. The reaction mixture was stirred at room temperature for 3 h. Additional triethylamine (1 mL) and DMSO (2 mL) were added until all the solid was dissolved in the reaction mixture. It was stirred for 17 h. The reaction had not gone to completion yet. It was stirred at room temperature over the weekend. The crude product was purified by preparatory HPLC

(0.1 % NH₄OH) to give the title compound (167 mg, 0.464 mmol, 68.5 % yield) as a solid. LC/MS: m/z 360.0 (M+H)+, Rt 0.67 min. Intermediate 94

1 -Dimethylethyl {[1 -ethyl-6-(methyloxy)-1 H-benzimidazol-2-yl]methyl}carbamate

A mixture of N2-ethyl-4-(methyloxy)-1 ,2-benzenediamine [2-amino-5- (methyloxy)phenyl]ethylamine (360 mg, 2.166 mmol), 1 , 1-dimethylethyl (2- oxoethyl)carbamate (345 mg, 2.166 mmol) and sodium bisulfite (721 mg, 6.93 mmol) in ethanol (9 mL) and water (3 mL) was heated at 60°C for 17 h. It was cooled to RT. The solid from the reaction mixture was filtered and then washed with DCM (10 mL). The filtrate was concentrated to give the crude product. It was purified by automated flash chromatography to give the title compound (376 mg, 1 .231 mmol, 56.9 % yield). LC/MS: m/z 306.0 (M+H)+, Rt 0.73 min.

Intermediate 95

3,5-Diamino-6-chloro-N-{[1 -ethyl-6-(methyloxy)-1 H-benzimidazol-2-yl]methyl}-2- pyrazinecarboxamide

To a solution of 1 , 1 -dimethylethyl {[1 -ethyl-6-(methyloxy)-1 H-benzimidazol-2- yl]methyl}carbamate (200 mg, 0.655 mmol) in DCM (4 mL) was added 4 M HCI in dioxane

(1 .637 mL, 6.55 mmol). The mixture was stirred at room temperature for 8 h. It was concentrated under a stream of nitrogen at 50°C to give the intermediate. 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (156 mg, 0.655 mmol), triethylamine (1.826 mL, 13.10 mmol) and DMSO (4.00 mL) was added to the intermediate. The reaction mixture was stirred at room temperature over the weekend. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (145 mg, 0.386 mmol, 58.9 % yield) as a solid. LC/MS: m/z 376.0 (M+H)+, Rt 0.64 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.38 - 8.32 (m, 1 H), 7.46 (d, J = 8.8 Hz, 2 H), 7.1 1 - 7.01 (m, 4 H), 6.82 - 6.75 (m, 1 H), 4.69 - 4.63 (m, 2 H), 4.32 - 4.21 (m, 2 H), 3.81 (s, 3 H), 1 .29 (s, 3 H). Intermediate 96

1 -Ethyl-2-(hydroxymethyl)-3-{2-oxo-2-[(phenylmethyl)oxy]ethyl}-1 H-benzimidazol-3- ium bromide

A mixture of 1 -ethyl-1 H-benzimidazol-2-yl)methanol (100 mg, 0.567 mmol) and phenylmethyl bromoacetate (0.089 ml_, 0.567 mmol) in THF (1 .5 ml.) was heated in a microwave reactor at normal absorption for 1 h at 100°C. The solid was filtered and then washed with ether to obtain the title compound (82 mg, 0.202 mmol, 35.7 % yield) as a white solid. LC/MS: m/z 325.1 (M+H)+, Rt 0.73 min.

Intermediate 97

1 1 -Dimethylethyl [(1 -propyl-1 H-benzimidazol-2-yl)methyl]carbamate

A mixture of 1 , 1 -dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate (50 mg, 0.202 mmol), bromopropane (0.022 ml_, 0.243 mmol) and K₂C0₃ (33.5 mg, 0.243 mmol)in acetone (2 ml.) was stirred at room temperature for 17 h. It was heated at 50°C over the weekend. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (50 mg, 0.173 mmol, 85 % yield). LC/MS: m/z 290.2 (M+H)+, Rt 0.72 min.

Intermediate 98

3,5-Diamino-6-chloro-N-[(1 -propyl-1 H-benzimidazol-2-yl)methyl]-2- razinecarboxamide

To a solution of 1 , 1 -dimethylethyl [(1 -propyl-1 H-benzimidazol-2-yl)methyl]carbamate (48.5 mg, 0.168 mmol) in DCM (3 mL) was added 4 M HCI in dioxane (0.419 mL, 1.676 mmol). The mixture was stirred at room temperature for 8 h. It was concentrated under a stream of nitrogen at 50°C and then redissolved in DMSO (3 mL). 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (40 mg, 0.168 mmol) and triethylamine (0.234 mL, 1 .676 mmol) were added to the intermediate. The reaction mixture was stirred at room temperature for 17 h. The reaction had not gone to completion yet. It was stirred at room temperature over the weekend. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (39 mg, 0.108 mmol, 64.7 % yield) as a solid. LC/MS: m/z 360.0 (M+H)+, Rt 0.65 min.

Intermediate 99

4-[1 -Ethyl-2-(hydroxymethyl)-1 H-benzimidazol-6-yl]phenol

A mixture of (6-bromo-1-ethyl-1 H-benzimidazol-2-yl)methanol (200 mg, 0.784 mmol), (4- hydroxyphenyl)boronic acid (108 mg, 0.784 mmol), sodium carbonate (249 mg, 2.352 mmol) and PdCI₂(dppf) (57.4 mg, 0.078 mmol) in 1 ,4-dioxane (2 mL) and water (0.667 mL) in a microwave vial was degassed for 2 min and then heated in a microwave reactor at 100°C for 10 min. Water (1 mL) and ethyl acetate (4 mL) were added to the reaction mixture. A solid precipitated out from the solution. The solid was filtered and then washed with water (2 mL) followed by ethyl acetate (5 mL). The solid was dried to give the title compound (212 mg, 0.790 mmol, 101 % yield). LC/MS: m/z 269.1 (M+H)⁺, Rt 0.38 min.

Intermediate 100

1 ,3-Diethyl-2-(hydroxymethyl)-6-(4-hydroxyphenyl)-1 H-3,1 -benzimidazol-3-ium iodide

A mixture of 4-[1-ethyl-2-(hydroxymethyl)-1 H-benzimidazol-6-yl]phenol (120 mg, 0.447 mmol) and iodoethane (0.361 mL, 4.47 mmol) in THF (2 mL) was heated in a microwave reactor at normal absorption for 40 at 100°C. It was then stirred at 50°C for 17 h. The solid was filtered and then washed with ether to give the title compound (85 mg, 0.200 mmol, 44.8 % yield) as a solid. LC/MS: m/z 297.0 (M)+, Rt 0.65 min. Intermediate 101

1 ,1 -Dimethylethyl ({1 -[2-(methyloxy)ethyl]-1 H-benzimidazol-2-yl}methyl)carbamate

A mixture of 1 , 1-dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate (180 mg, 0.728 mmol), 2-bromoethyl methyl ether (0.123 mL, 1.310 mmol) and K₂C0₃ (181 mg, 1.310 mmol) in DMSO (2 mL) was stirred at 50°C for 17 h. The solid was filtered and the filtrate was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (127 mg, 0.416 mmol, 57.1 % yield). LC/MS: m/z 306.0 (M+H)+, Rt 0.71 min.

Intermediate 102

3,5-Diamino-6-chloro-N-({1 -[2-(methyloxy)ethyl]-1 H-benzimidazol-2-yl}methyl)-2- pyrazinecarboxamide

To a solution of 1 , 1 -dimethylethyl ({1-[2-(methyloxy)ethyl]-1 H-benzimidazol-2- yl}methyl)carbamate (122 mg, 0.398 mmol) in DCM (3 mL) was added 4 M HCI in dioxane (0.995 mL, 3.98 mmol). The mixture was stirred at room temperature for 17 h and then concentrated under a stream of nitrogen at 50°C to give the intermediate. 1 , 1- dimethylethyl ({1 -[2-(methyloxy)ethyl]-1 H-benzimidazol-2-yl}methyl)carbamate (122 mg, 0.398 mmol), triethylamine (0.555 mL, 3.98 mmol) and DMSO (3 mL) were added to the intermediate. The reaction mixture was stirred at room temperature for 17 h and then for another 24 h. The solid precipitated from the reaction mixture. The solid was filtered to give the title compound (51 mg, 0.136 mmol, 34.1 % yield). The filtrated was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (47 mg, 0.125 mmol, 31 .4 % yield) as a solid. LC/MS: m/z 376.0 (M+H)+, Rt 0.67 min.

Intermediate 103

1 1 -Dimethylethyl {[1 -(cyclobutylmethyl)-1 H-benzimidazol-2-yl]methyl}carbamate

A mixture of 1 , 1-dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate (150 mg, 0.607 mmol), (bromomethyl)cyclobutane (0.102 mL, 0.910 mmol) and K₂C0₃ (126 mg, 0.910 mmol) in DMSO (2 mL) was heated at 50°C initially for 17 h and then for another 48 h. The crude product was filtered and purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (83 mg, 0.263 mmol, 43.4 % yield). LC/MS: m/z 316.0 (M+H)+, Rt 0.84 min.

Intermediate 104

3,5-Diamino-6-chloro-N-{[1 -(cyclobutylmethyl)-1 H-benzimidazol-2-yl]methyl}-2- razinecarboxamide

To a solution of 1 , 1 -dimethylethyl {[1 -(cyclobutylmethyl)-1 H-benzimidazol-2- yl]methyl}carbamate (86 mg, 0.272 mmol) in DCM (3 mL) was added 4 M HCI in dioxane (0.681 mL, 2.72 mmol). The mixture was stirred at room temperature for 17 h. It was concentrated under a stream of nitrogen at 50°C to give the intermediate. 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (65 mg, 0.272 mmol), triethylamine (0.380 mL, 2.72 mmol) and DMSO (3 mL) were added to the intermediate. The reaction mixture was stirred at room temperature for 17 h and then for another 24 h. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (66 mg, 0.171 mmol, 62.8 % yield) as a solid. LC/MS: m/z 386.0 (M+H)+, Rt 0.81 min.

Intermediate 105

-Dimethylethyl ({5-[(trifluoromethyl)thio]-1 H-benzimidazol-2-yl}methyl)carbamate

A mixture of 4-[(trifluoromethyl)thio]-1 ,2-benzenediamine (300 mg, 0.847 mmol), 1 ,1 - dimethylethyl (2-oxoethyl)carbamate (135 mg, 0.847 mmol) and sodium bisulfite (441 mg, 4.24 mmol) in ethanol (3 mL) and water (1 mL) was heated at 60°C for 17 h. The solid was filtered and then washed with DCM (10 mL). The filtrate was concentrated to give the crude product. It was purified by automated flash chromatography to give the title compound (100 mg, 0.288 mmol, 34.0 % yield). LC/MS: m/z 348.0 (M+H)+, Rt 0.90 min. Intermediate 106

3,5-Diamino-6-chloro-N-({5-[(trifluorom

pyrazinecarboxamide

To a solution of 1 , 1 -dimethylethyl ({5-[(trifluoromethyl)thio]-1 H-benzimidazol-2- yl}methyl)carbamate (1 16 mg, 0.335 mmol) in DCM (4 mL) was added 4 M HCI in dioxane (0.838 mL, 3.35 mmol). The mixture was stirred at room temperature for 8 h. The solvent was concentrated under a stream of nitrogen at 50°C and then redissolved in DMSO (4 mL). 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (80 mg, 0.335 mmol) and triethylamine (0.467 mL, 3.35 mmol) were added. The reaction mixture was stirred at room temperature for 24 h. It was stirred for another 17 h. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (53 mg, 0.127 mmol, 37.8 % yield) as a solid. LC/MS: m/z 417.9 (M+H)+, Rt 0.84 min.

Intermediate 107

1 ,1 -Dimeth lethyl {[1 -(phenylmethyl)-l H-benzimidazol-2-yl]methyl}carbamate

A mixture of 1 , 1-dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate (150 mg, 0.607 mmol), benzyl bromide (104 mg, 0.607 mmol) and K₂C0₃ (151 mg, 1.092 mmol) in DMSO (2 mL) was stirred at 50°C for 17 h. It was heated at 50°C for 52 h. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (90 mg, 0.267 mmol, 44.0 % yield). LC/MS: m/z 338.0 (M+H)+, Rt 0.91 min.

Intermediate 108

3,5-Diamino-6-chloro-N-{[1 -(phenylmethyl)-1 H-benzimidazol-2-yl]methyl}-2- razinecarboxamide

To a solution of 1 , 1 -dimethylethyl {[1 -(phenylmethyl)-1 H-benzimidazol-2- yl]methyl}carbamate (92 mg, 0.272 mmol) in DCM (3 mL) was added 4 M HCI in dioxane (0.681 mL, 2.72 mmol). The mixture was stirred at room temperature for 17 h. It was concentrated under a stream of nitrogen at 50°C to give the intermediate. 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (65 mg, 0.272 mmol), triethylamine (0.380 mL, 2.72 mmol) and DMSO (3 mL) were added to the intermediate. The reaction mixture was stirred at room temperature for 17 h. It was then stirred at room temperature over the weekend. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (82 mg, 0.201 mmol, 73.8 % yield) as a solid. LC/MS: m/z 408.0 (M+H)+, Rt 0.81 min.

Intermediate 109

1 1 -Dimethylethyl {[1 -(2-methylpropyl)-1 H-benzimidazol-2-yl]methyl}carbamate

A mixture of 1 , 1-dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate (150 mg, 0.607 mmol), 1-bromo-2-methylpropane (0.066 mL, 0.607 mmol) and K₂C0₃ (151 mg, 1.092 mmol) in DMSO (2 mL) was heated at 50°C for 52 h. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (108 mg, 0.356 mmol, 58.7 % yield). LC/MS: m/z 304.0 (M+H)+, Rt 0.85 min.

Intermediate 110

3,5-Diamino-6-chloro-N-{[1 -(2-methylpropyl)-1 H-benzimidazol-2-yl]methyl}-2- razinecarboxamide

To a solution of 1 , 1 -dimethylethyl {[1-(2-methylpropyl)-1 H-benzimidazol-2- yl]methyl}carbamate (108 mg, 0.356 mmol) in DCM (3 mL) was added 4 M HCI in dioxane (0.890 mL, 3.56 mmol). The mixture was stirred at room temperature for 17 h. It was concentrated under a stream of nitrogen at 50°C to give the intermediate. 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (85 mg, 0.356 mmol), triethylamine (0.496 mL, 3.56 mmol) and DMSO (3 mL) were added to the intermediate. The reaction mixture was stirred at room temperature for 17 h. It was then stirred at room temperature over the weekend. The crude product was purified by preparatory HPLC (0.1 % NH₄OH) to give the title compound (102 mg, 0.273 mmol, 77 % yield) as a solid. LC/MS: mlz 374.0 (M+H)+, Rt 0.77 min. Intermediate 111

1 ,1 -dimethylethyl{[1 -ethyl-6-(4 ,4,5,5-tetramethyM ,3,2-dioxaborolan-2-yl)-1 H- nzimidazol-2-yl]methyl}carbamate

solution of 1 ,1 -dimethylethyl [(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl] carbamate (0.886 g, 2.5 mmol) in DMF (25 mL) were added 4,4,4',4',5,5,5',5'-octamethyl-2,2'-bi- 1 ,3,2-dioxaborolane (0.952 g, 3.75 mmol), potassium acetate (0.736 g, 7.50 mmol) and PdCI₂(dppf) (0.091 g, 0.125 mmol). The resulting reaction mixture was stirred and heated to 1 10°C in a microwave reactor for 40 minutes. The organic layer was filtered, concentrated and purified by automated flash chromatography to give 0.446 g (44 %) of the title compound. LC/MS : m/z 401 .0 (M+H)⁺ ; Rt 0.99 min.

Intermediate 112

1 ,1 -dimethyleth l [(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl)methyl]carbamate

To a solution of 1 , 1-dimethylethyl {[1 -ethyl-6-(4,4,5,5-tetramethyl-1 ,3,2-dioxaborolan-2-yl)- 1 H-benzimidazol-2-yl]methyl}carbamate (1.83 g, 4.56 mmol) in DMF (40 mL) was added hydrogen peroxide (aqueous) (1.165 mL, 1 1 .4 mmol). The resulting reaction mixture was stirred at room temperature for 48 hours. The remaining oxidation agent was quenched with 5% aq. Na₂S0₃ 5% and the mixture was filtered. Water was added to the solution and the organic layer was extracted with EtOAc. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated to afford 0.7644 g (58%) of the title compound. LC/MS: m/z 292.0 (M+H)⁺ ; Rt 0.66 min.

Intermediate 113

3,5-diamino-6-chloro-N-[(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl)methyl]-2- pyrazinecarboxamide

To a solution of 1 , 1 -dimethylethyl [(1-ethyl-6-hydroxy-1 H-benzimidazol-2- yl)methyl]carbamate (750 mg, 2.57 mmol) in DCM (15 mL) was added HCI (4N) in dioxane (3.22 mL, 12.87 mmol). The reaction mixture was stirred at room temperature for 22 hours. The reaction mixture was concentrated and the intermediate was taken back in DMSO (7.50 mL). 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (614 mg, 2.57 mmol) and triethylamine (3.59 mL, 25.7 mmol) were added to the reaction mixture which was stirred at 50°C overnight. After removal of the solvent, the crude residue was purified by preparatory HPLC (NH₄OH). The relevant fractions were combined and concentrated under vacuum to afford the title compound (742 mg, 80%). LC/MS: m/z 361.9 (M+H)⁺ ; Rt 0.64 min.

Intermediate 114

3,5-diamino-6-chloro-N-({1 -ethyl-6-[(phenylmethyl)oxy]-1 H-benzimidazol-2- yl}methyl)-2-pyrazinecarboxamide

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl)methyl]- 2-pyrazinecarboxamide (100 mg, 0.276 mmol) in DMSO (2 mL) were added K₂C0₃ (95 mg, 0.692 mmol) and (bromomethyl)benzene (0.065 mL, 0.553 mmol). The resulting reaction mixture was stirred at room temperature for 24 hours. The crude residue was filtered and purified by preparatory HPLC (NH₄OH). The relevant fractions were combined and concentrated under vacuum to afford the title compound (33 mg, 27%). LC/MS: m/z 452.2 (M+H)⁺ ; Rt 0.89 min.

Intermediate 115

Methyl {[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazol-6-yl]oxy}acetate

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl)methyl]- 2-pyrazinecarboxamide (100 mg, 0.276 mmol) in DMSO (2 mL) were added Cs₂C0₃ (180 mg, 0.553 mmol) and methyl bromoacetate (0.039 mL, 0.415 mmol). The resulting reaction mixture was stirred at room temperature overnight. The crude residue was filtered and purified by preparatory HPLC (NH₄OH). The relevant fractions were combined and concentrated under vacuum to afford the title compound (28 mg, 23 %). LC/MS: m/z 434.1 (M+H)⁺; Rt 0.64 min.

Intermediate 116

3,5-diamino-6-chloro-N-({1 -ethyl-6-[(2-oxotetrahydro-3-furanyl)oxy]-1 H- benzimidazol-2-yl}methyl)-2-pyrazinecarboxamide

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl) methyl]-2-pyrazinecarboxamide (100 mg, 0.276 mmol) in DMSO (2 mL) were added K₂C0₃ (95 mg, 0.692 mmol) and 3-bromodihydro-2(3H)-furanone (91 .2 mg, 0.553 mmol). The resulting reaction mixture was stirred at room temperature for 24 hours. The crude residue was filtered and purified by preparatory HPLC (NH₄OH). The relevant fractions were combined and concentrated under vacuum to afford the title compound (35 mg, 28 %). LC/MS: m/z 446.1 (M+H)⁺; Rt 0.64 min.

Intermediate 117

1 ,1 -dimethylet -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate

In a flask, 1 , 1 -dimethylethyl [(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (0.496 g, 1.4 mmol) was dissolved in methanol (70 mL). The starting material was hydrogenated using an hydrogenation apparatus under the following conditions: flow rate: 1 mL/min; cartridge: 20 % Pd(OH)₂/C; temperature: 20°C; pressure: 1 bar. The reaction crude was concentrated down to afford the title compound (0.385 g, 100 %). LC/MS: m/z 276.0 (M+H)⁺; Rt 0.72 min.

Intermediate 118

3, 5-diamino-6-chloro-N-[(1 -ethyl-1 H-benzimidazol-2-yl)methyl]-2- pyrazinecarboxamide

To a solution of 1 , 1 -dimethylethyl [(1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (385 mg, 1 .398 mmol) in DCM (15 mL) was added HCI (4N in dioxane) (1 .748 mL, 6.99 mmol). The reaction mixture was stirred at room temperature for 18 hours. The reaction product was filtered, washed with DCM and dried. The intermediate was taken back in DMSO (4 mL). 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (417 mg, 1 .747 mmol) and triethylamine (1.949 mL, 13.98 mmol) were added to the reaction mixture which was stirred at 50°C for 72 hours. The excess of triethylamine was evaporated. The crude residue was filtered and purified by preparatory HPLC (NH₄OH). The relevant fractions were combined and concentrated under vacuum to afford the title compound (276.8 mg, 60 %). LC/MS: m/z 346.1 (M+H)⁺; Rt 0.60 min.

Intermediate 119

1 -ethyl-2-methyl-1 H-benzimidazole

00 of-

To a solution of 2-methyl-1 H-benzimidazole (6.61 g, 50 mmol) in MeCN (250 mL) was added KOH (5.61 g, 100 mmol). The mixture was then stirred for 10 min and then iodoethane (4.24 mL, 52.5 mmol) was added. Stirring was continued to at room temperature stirred for 21 h. The reaction mixture was evaporated down under vacuum and H₂0 (50 mL) was added. The mixture was extracted with EtOAc (100 + 50 mL). The combined organic layers were washed with brine (30 mL), dried over Na₂S0₄, filtered, evaporated down under vacuum and dried under high vacuum to afford the title compound (8.1771 g, 102 % yield). LC/MS: m/z 161 (M+H)⁺, Rt 0.44 min.

Intermediate 120

1 3-diethyl-2-methyl-1 H-3, -benzimidazol-3-ium iodide

To a solution of 1-ethyl-2-methyl-1 H-benzimidazole (1 .602 g, 10 mmol) in MeCN (20 mL) was each added iodoethane (1.616 mL, 20.00 mmol). The mixture was heated in a microwave reactor at 120°C for 1 h. The reaction mixture was evaporated down under vacuum, washed with acetone (2 x 5 mL) and then dried under high vacuum to afford the title compound (2.6884 g, 85 % yield). LC/MS: m/z 189 (M)⁺, Rt 0.56 min. Intermediate 121

1 ,3-diethyl-2-(iodomethyl)-1 H-3,1 -benzimidazol-3-ium iodide

To a solution of 1 ,3-diethyl-2-methyl-1 H-3, 1 -benzimidazol-3-ium iodide (632 mg, 2 mmol) in MeCN (10 mL) were added l₂ (558 mg, 2.200 mmol) and TEA (0.335 mL, 2.400 mmol). The resulting mixture was stirred at room temperature for 20.5 h before more TEA (0.139 mL, 1 .000 mmol) was added. Stirring was then continued at room temperature for 4 h. The precipitate in the reaction mixture was filtered out, washed with acetone (2 + 1 mL) and then dried under high vacuum to afford the title compound (656.2 mg, 74.2 % yield). LC/MS: m/z 315 (M)⁺, Rt 0.50 min.

Intermediate 122

2- (azidomethyl)-1 ,3-diethyl-1 H-3,1 -benzimidazol-3-ium iodide

To a mixture of sodium azide (102 mg, 1.575 mmol) in EtOH (7.5 mL) was added 1 ,3- diethyl-2-(iodomethyl)-1 H-3, 1 -benzimidazol-3-ium iodide (663 mg, 1 .5 mmol) and then stirred at room temperature for 20 h before addition of more MeOH (5 mL). Stirring was continued at room temperature for 2 h. The precipitate in the reaction mixture was filtered out, washed with EtOH (1 mL) and then dried under high vacuum to afford the title compound (231.3 mg, 43.2 % yield). LC/MS: m/z 230 (M)⁺, Rt 0.58 min.

Intermediate 123

-(aminomethyl)-1 ,3-diethyl-1 H-3,1 -benzimidazol-3-ium iodide

To a solution of 2-(azidomethyl)-1 ,3-diethyl-1 H-3,1 -benzimidazol-3-ium iodide (146 mg, 0.409 mmol) in MeOH (20 mL) was hydrogenated in an H-Cube hydrogenation apparatus (1 atm H₂, 1 mL/min, 20% Pd(OH)₂/C cartridge, 20°C). The reaction mixture was evaporated down under vacuum to afford the title compound (134.5 mg, 99 % yield). LC/MS: m/z 204 (M)⁺, Rt 0.42 min.

Intermediate 124

methyl (2E)-3-{2-[({[(1 ,1 -dimethylethyl)oxy]carbonyl}amino) methyl]-1 -ethyl-1 H- benzimidazol-6- l -2- ro enoate

To a solution of 1 , 1 -dimethylethyl [(6-bromo-1-ethyl-1 H-benzimidazol-2- yl)methyl]carbamate (708 mg, 2 mmol) in DMF (20 mL) were added methyl 2-propenoate (0.360 mL, 4.00 mmol), PdCI₂(dppf) (1 17 mg, 0.160 mmol) and K₃P0₄ (849 mg, 4.00 mmol). The resulting reaction mixture was flushed with N₂ for 5 min and then heated in a microwave reactor at 100°C for 2 h. The reaction mixture was evaporated down under vacuum, purified by automated flash chromatography, evaporated down under vacuum and dried under high vacuum to afford the title compound (504.4 mg, 70.2 % yield). LC/MS: m/z 360 (M+H)⁺, Rt 0.83 min.

Intermediate 125

methyl 3-{2-[({[(1 ,1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1 -ethyl-1 H- benzimidazol-6- l}propanoate

A solution of methyl (2E)-3-{2-[({[(1 , 1-dimethylethyl)oxy]carbonyl}amino)methyl]-1-ethyl- 1 H-benzimidazol-6-yl}-2-propenoate (500 mg, 1 .391 mmol) in MeOH (140 mL) and acetone (50 mL) was hydrogenated in an H-Cube hydrogenation apparatus (1 atm H₂, 1 mL/min, 20% Pd(OH)₂/C cartridge, 20°C, 2 runs). The reaction mixture was evaporated down under vacuum, purified with automated flash chromatography, evaporated down under vacuum, dried under high vacuum to afford the title compound (427.6 mg, 85 % yield). LC/MS: m/z 362 (M+H)⁺, Rt 0.74 min. Intermediate 126

{1 -ethyl -6-[3-(methyloxy)-3-oxopropyl]-1 H-benzimidazol-2-yl}methanaminium trifluoroacetate

To a solution of methyl 3-{2-[({[(1 , 1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1-ethyl-1 H- benzimidazol-6-yl}propanoate (425 mg, 1.176 mmol) in DCM (1 mL) was added TFA (0.453 mL, 5.88 mmol). The mixture was stirred at room temperature for 18 h and then heated at 30°C for 25 h. The reaction mixture was evaporated down under vacuum to afford the title compound (654.2 mg, 149 % yield) as a crude residue which was used for the next reaction step without further purification. LC/MS: m/z 262 (M+H)⁺, Rt 0.64 min.

Intermediate 127

methyl 3-[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino} methyl)-1 -ethyl- 1 H- nzimidazol-6-yl]propanoate

To a solution of methyl {1-ethyl-6-[3-(methyloxy)-3-oxopropyl]-1 H-benzimidazol-2- yl}methanaminium trifluoroacetate (438 mg, 1 .17 mmol) in DMF (5ml_) was added 3- chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (279 mg, 1 .170 mmol) and TEA (0.815 mL, 5.85 mmol). The resulting reaction mixture was stirred at room temperature for 5 h and then heated at 60°C for 17 h. The reaction mixture was evaporated down under vacuum and then DMSO (2 mL) and MeOH (4 mL) were added. The solution was filtered and the solid was washed with MeOH (1 mL). The combined filtrate was purified by preparatory HPLC (with 0.1 % NH₄OH) to afford the title compound (314.9 mg, 62.3 % yield). LC/MS: m/z 432 (M+H)⁺, Rt 0.76 min. Intermediate 128

Methyl 3-{[2-(aminomethyl)-1 -ethyl-1 H-benzimidazol-6-yl]thio}propanoate

A mixture of 1 -(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methanamine (150 mg, 0.459 mmol), methyl 3-mercaptopropionate (0.061 ml_, 0.550 mmol), Pd₂(dba)₃ (21.00 mg,

0.023 mmol), N,N-diisopropylethylamine (0.160 ml_, 0.917 mmol) and Xanphos (26.5 mg, 0.046 mmol) in 1 ,4-dioxane (1 .5 ml.) was heated in a microwave reactor at normal absorption for 40 minutes at 120°C. Some solid precipitated out from the reaction mixture. It was filtered through a PL-Thiol MP SPE+ and was then washed with ethyl acetate. LC/MS indicated the solid was not the desired product. The desired product was in the mother liquor layer. The mother liquor was concentrated and carried to the next step without further purification.

Intermediate 129

Methyl 3-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 1 H-benzimidazol-6-yl]thio}propanoate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (35 mg, 0.147 mmol) in DMSO (4 mL) was added methyl 3-{[2-(aminomethyl)-1 -ethyl-1 H-benzimidazol- 6-yl]thio}propanoate (43.0 mg, 0.147 mmol) and triethylamine (0.204 mL, 1.467 mmol). The mixture was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (0.1 % NH₄OH), giving the title compound (23 mg, 0.050 mmol, 33.8 % yield). LC/MS: m/z 463.9 (M+H)+, Rt 0.72 min.

Intermediate 130

-Dimethylethyl [(6-chloro-1 -ethyl-1 H-benzimidazol-2- l)methyl]carbamate

A mixture of 4-chloro-N2-ethyl-1 ,2-benzenediamine (450 mg, 2.64 mmol), 1 ,1 - dimethylethyl (2-oxoethyl)carbamate (420 mg, 2.64 mmol) and sodium bisulfite (823 mg, 7.91 mmol) in ethanol (18 mL) and water (6 mL) was heated at 60°C for 17 h then cooled to room temperature. The solid from the reaction mixture was filtered and then washed with DCM (15 mL). The filtrate was concentrated to give the crude product. It was purified by automated flash chromatography. The desired fractions were concentrated under reduced pressure then triturated with ether to give the title compound (406 mg, 1.31 1 mmol, 49.7 % yield). LC/MS: m/z 310.1 (M+H)+, Rt 0.81 min.

Intermediate 131

3,5-Diamino-6-chloro-N-[(6-chloro-1 -ethyl-1 H^enzimidazol-2-yl)methyl]-2- pyrazinecarboxamide

A mixture of 1 , 1-dimethylethyl [(6-chloro-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (325 mg, 1 .048 mmol) and 4 N HCI in dioxane (1 .310 mL, 5.24 mmol) in DCM (5.00 mL) was stirred at room temperature over the weekend. It was concentrated under a stream of nitrogen at 50°C to dryness. It was redissolved in DMSO (5 mL). 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (250 mg, 1.048 mmol) and triethylamine

(1 .168 mL, 8.38 mmol) were added and stirred at room temperature for 18 h then at room temperature for another 19 h. Some solid precipitated out from the reaction mixture and was filtered. The filtrate was recovered and purified by preparatory HPLC (0.1 % NH₄OH), giving the title compound (198 mg, 0.521 mmol, 49.7 % yield) as a solid. LC/MS: m/z 380.0 (M+H)+, Rt 0.68 min.

Intermediate 132

-Bromo-N-ethyl-6-nitroaniline

To a mixture of 1-bromo-2-fluoro-3-nitrobenzene (653 mg, 2.97 mmol) and potassium carbonate (1366 mg, 9.89 mmol) in ethanol (3 mL) was added dropwise 70 % ethylamine in water (1 .2 mL, 14.83 mmol). The resulting mixture was heated in a microwave reactor at normal absorption for 10 min at 100°C. Water (5 mL) was added and extracted with DCM (2 x 20 mL). The combined organic layer was washed with saturated NaCI (10 mL), dried over sodium sulfate, filtered and then concentrated under vacuum to give the title compound (613 mg, 2.501 mmol, 84 % yield) as a solid. LC/MS: m/z 244.9 (M+H)⁺, Rt 1.15 min. Intermediate 133

-Bromo-N2-ethyl-1 ,2-benzenediami

A mixture of 2-bromo-N-ethyl-6-nitroaniline (613 mg, 2.501 mmol), tin(ll) chloride dihydrate (2822 mg, 12.51 mmol) in ethanol (30 mL) was heated at reflux for 17 h.

Saturated NaHC0₃ was added until the pH of the solution was around 7. A solid precipitated out from the solution and was filtered then washed with ethyl acetate (300 mL). The organic layer was concentrated under vacuum to give the title compound (540 mg, 2.51 mmol, 100 % yield) as a solid. LC/MS: m/z 215.0 (M+H)+, Rt 0.71 min.

Intermediate 134

1 -Dimethylethyl [(7-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate

A mixture of 3-bromo-N2-ethyl-1 ,2-benzenediamine (540 mg, 2.51 mmol), 1 ,1 - dimethylethyl (2-oxoethyl)carbamate (400 mg, 2.51 mmol) and sodium bisulfite (784 mg, 7.53 mmol) in ethanol (18 mL) and water (6 mL) was heated at 60°C for 17 h. It was cooled to room temperature. Solid from the reaction mixture was filtered and then washed with DCM (15 mL). The filtrate was concentrated to give the crude product. It was purified by automated flash chromatography eluting with ethyl acetate to obtain the title compound (506 mg, 1.428 mmol, 56.9 % yield). LC/MS: m/z 353.9 (M+H)+, Rt 0.94 min.

Intermediate 135

3, 5-Diamino-A -[(7-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- pyrazinecarboxamide

A mixture of 1 , 1-dimethylethyl [(7-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]carbamate (341 mg, 0.964 mmol) and 4 N HCI in dioxane (1 .205 mL, 4.82 mmol) in DCM (5.00 mL) was stirred at room temperature overnight. It was concentrated under a stream of nitrogen at 50°C to dryness. It was redissolved in DMSO (5 mL). 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (230 mg, 0.964 mmol) and triethylamine (1 .343 mL, 9.64 mmol) were added and stirred at room temperature for 18 h. Some solid was precipitated out from the reaction mixture and was filtered. The filtrate was purified by preparatory HPLC (with basic conditions) then triturated with ether to give the target compound (185 mg, 0.436 mmol, 45.2 % yield) as a solid. LC/MS: m/z 423.8 (M+H)+, Rt 0.78 min.

Intermediate 136

-Chloro-A -ethyl-2-nitroaniline

To a mixture of 4-chloro-2-fluoro-1 -nitrobenzene (868 mg, 4.94 mmol) and potassium carbonate (1366 mg, 9.89 mmol) in ethanol (3 mL) was added dropwise 70 % ethylamine in water (1 .2 mL, 14.83 mmol). The resulting mixture was heated in a microwave reactor at normal absorption for 10 min at 100°C. Water (5 mL) was added and extracted with DCM (2 x 20 mL). The combined organic layer was washed with saturated NaCI (10 mL), dried over sodium sulfate, filtered and then concentrated under vacuum to give the target compound (982 mg, 4.89 mmol, 99 % yield) as a solid. LC/MS: m/z 201 .0 (M+H)⁺, Rt 1.15 min.

Intermediate 137

-Chloro-A ²-ethyl-1 ,2-benzenediamine

A mixture of 5-chloro-N-ethyl-2-nitroaniline (982 mg, 4.89 mmol), tin(ll) chloride dihydrate (5522 mg, 24.47 mmol) in ethanol (30 mL) was heated at reflux for 17 h. Saturated NaHC0₃ was added until the pH of the solution was around 7. Solid was precipitated out from the solution and was filtered then washed with ethyl acetate (300 mL). The combined organic layer was concentrated to give the target compound (906 mg, 5.31 mmol, 108 % yield) as a solid. LC/MS: m/z 171 .0 (M+H)+, Rt 0.70 min. Intermediate 138

6-Chloro-1 -eth l-1H-benzimidazol-2-yl)methanol

A mixture of 4-chloro-N2-ethyl-1 ,2-benzenediamine (450 mg, 2.64 mmol), hydroxyacetic acid (201 mg, 2.64 mmol) and 6 N HCI (0.044 ml_, 0.267 mmol) in water (1 ml.) was heated in a microwave reactor at normal absorption for 30 minutes at 150°C. It was then heated again in a microwave reactor at normal absorption for 15 minutes at 150°C. 6 N NaOH was added until the pH of the mixture was around 9. A solid precipitated out from the reaction mixture and was filtered. It was purified by preparatory HPLC (with basic conditions). The product fractions were dried to give the target compound (301 mg, 1 .429 mmol, 54.2 % yield). LC/MS: m/z 21 1 .0 (M+H)+, Rt 0.65 min.

Intermediate 139

6-Chloro-1 ,3-diethyl-2-(hydroxymethyl)-1 H-benzimidazol-3-ium iodide

A mixture of (6-chloro-1 -ethyl-1 H-benzimidazol-2-yl)methanol (100 mg, 0.475 mmol) and iodoethane (0.384 ml_, 4.75 mmol) in THF (2 ml.) was heated in a microwave reactor at normal absorption for 45 min at 100°C then at 50°C for 35 h. A solid precipitated out from the reaction mixture and was filtered then washed with ether to give the target compound (51 mg, 0.139 mmol, 29.3 % yield). LC/MS: m/z 238.9 (M)+, Rt 0.57 min.

Intermediate 140

{5-[(Trifluoromethyl)thio]-1 H-benzimidazol-2-yl}methanol

CIH

A mixture of {2-amino-4-[(trifluoromethyl)thio]phenyl}amine dihydrochloride (500 mg, 1.779 mmol), hydroxyacetic acid (176 mg, 2.312 mmol) and 6 N HCI (0.044 ml_, 0.267 mmol) in water (3 ml.) was heated in a microwave reactor at normal absorption for 1 h at 150°C. 40 mg of hydroxyacetic acid was added and the reaction mixture was heated in a microwave reactor at normal absorption for 1 h at 150°C. A solid precipitated out from the reaction mixture and was filtered to give the target compound (520 mg, 2.095 mmol, 1 18 % yield). LC/MS: m/z 249.0 (M+H)+, Rt 0.65 min.

Intermediate 141

1 -Ethyl -6-[(trifluoromethyl)thio]-1 H-benzimidazol-2-yl}methanol

A mixture of {5-[(trifluoromethyl)thio]-1 H-benzimidazol-2-yl}methanol (520 mg, 2.095 mmol), bromoethane (0.313 ml_, 4.19 mmol) and K₂C0₃ (290 mg, 2.095 mmol) in DMSO (4 ml.) was heated at 50°C for 50 h. 0.1 ml. bromoethane and 100 mg of K₂C0₃ were added and it was heated at 50°C for 24 h. A solid was filtered and the filtrate was purified by preparatory HPLC (with basic conditions), giving the title compound (389 mg, 1.408 mmol, 67.2 % yield). LC/MS: m/z 211 A (M+H)+, Rt 0.69 min.

Intermediate 142

1 ,3-Diethyl-2-(hydroxymethyl)-6-[(trifluoromethyl)thio]-1 H-3,1 -benzimidazol-3-ium iodide

A mixture of {1-ethyl-6-[(trifluoromethyl)thio]-1 H-benzimidazol-2-yl}methanol (100 mg, 0.362 mmol) and iodoethane (0.292 ml_, 3.62 mmol) in THF (3 ml.) was heated in a microwave reactor at normal absorption for 45 min at 100°C. It was then heated at 50°C for 5 days. The crude product was concentrated and carried to the next step without purification.

Intermediate 143

3-[2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazol-6-yl]propanoic acid

To a solution of methyl 3-[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)- 1-ethyl-1 H-benzimidazol-6-yl]propanoate (250 mg, 0.579 mmol) in THF (10 mL) was added NaOH (1 N) (2.89 mL, 2.89 mmol). The resulting reaction mixture was stirred at room temperature for 17 h. The reaction was quenched with HCI (0.5 mL, 6 N). The layers were separated. The aqueous layer was extracted with THF (3 mL). The combined organic layers were dried over Na₂S0₄, filtered, evaporated down under vacuum, dried under high vacuum to afford the title compound (224.3 mg, 93 % yield). LC/MS: m/z 418(M+H)⁺, Rt 0.57 min.

Intermediate 144

3,5-diamino-6-chloro-N-[(1 -ethyl-6-{3-[(3-hydroxypropyl)amino]-3-oxopropyl}-1 H- b

To a solution of 3-[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 1 H-benzimidazol-6-yl]propanoic acid (44 mg, 0.105 mmol) in DMSO (1 mL) was added 3- amino-1 -propanol (0.012 mL, 0.158 mmol) followed by TEA (0.029 mL, 0.21 1 mmol) and HBTU (59.9 mg, 0.158 mmol). The resulting reaction mixture was stirred at room temperature for 66 h. The reaction mixture was purified by preparatory HPLC (with 0.1 % NH₄OH). The relevant fractions were evaporated down under vacuum to afford the title compound (27.0 mg, 54.0 % yield). LC/MS: m/z 475 (M+H)⁺, Rt 0.52 min. Intermediate 145

3,5-diamino-6-chloro-N-({1 -ethyl-6-[3-(4-morpholinyl)-3-oxopropyl]-1 H- ben

To a solution of 3-[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 1 H-benzimidazol-6-yl]propanoic acid (44 mg, 0.105 mmol) in DMSO (1 mL) was added morpholine (0.014 mL, 0.158 mmol) followed by TEA (0.029 mL, 0.21 1 mmol) and HBTU (59.9 mg, 0.158 mmol). The resulting reaction mixture was stirred at room temperature for 66 h. The reaction mixture was purified by preparatory HPLC (with 0.1 % NH₄OH). The relevant fractions were evaporated down under vacuum to afford the title compound (32.4 mg, 63.2 % yield). LC/MS: m/z 487 (M+H)⁺, Rt 0.57 min.

Intermediate 146

3,5-diamino-6-chloro-N-({1 -ethyl-6-[3-(ethylamino)-3-oxopropyl]-1 H-benzimidazol-2- yl}methyl)-2-pyrazinecarboxamide

To a solution of 3-[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 1 H-benzimidazol-6-yl]propanoic acid (44 mg, 0.105 mmol) in DMSO (1 mL) was added ethylamine (2.0 M in THF) (0.079 mL, 0.158 mmol) followed by TEA (0.029 mL, 0.21 1 mmol) and HBTU (59.9 mg, 0.158 mmol). The resulting reaction mixture was stirred at room temperature for 2.5 h. The reaction mixture was purified by preparatory HPLC (with 0.1 % NH₄OH). The relevant fractions were evaporated down under vacuum to afford the title compound (28.1 mg, 60.0 % yield). LC/MS: m/z 445 (M+H)⁺, Rt 0.56 min. Intermediate 147

3,5-diamino-6-chloro-N-[(1 -ethyl-6-{3-[(2-hydroxyethyl)amino]-3-oxopropyl}-1 H- ben

To a solution of 3-[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 1 H-benzimidazol-6-yl]propanoic acid (44 mg, 0.105 mmol) in DMSO (1 mL) was added ethanolamine (9.51 μΙ_, 0.158 mmol) followed by TEA (0.029 mL, 0.21 1 mmol) and HBTU (59.9 mg, 0.158 mmol). The resulting reaction mixture was each stirred at room temperature for 2.5 h. The reaction mixture was purified by preparatory HPLC (with 0.1 % NH₄OH). The relevant fractions were evaporated down under vacuum to afford the title compound (36.1 mg, 74.4 % yield). LC/MS: m/z 461 (M+H)⁺, Rt 0.52 min.

Intermediate 148

3,5-diamino-6-chloro-N-{[1 -ethyl-6-(ethyloxy)-1 H-benzimidazol-2-yl]methyl}-2- pyrazine carboxamide

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl)methyl]- 2-pyrazinecarboxamide (90 mg, 0.250 mmol) in DMF (2 mL) were added K₂C0₃ (121 mg, 0.875 mmol) and iodoethane (0.160 mL, 2 mmol). The resulting reaction mixture was stirred at 50°C for 5 days. The crude residue was filtered and purified by preparatory HPLC (basic conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (9.5 mg, 10%). LC/MS: m/z 390.1 (M+H)⁺: Rt 0.76 min. Intermediate 149

3,5-diamino-6-chloro-N-{[1 -ethyl-6-(propyloxy)-1 H-benzimidazol-2-yl]methyl}-2- pyrazine carboxamide

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl)methyl]- 2-pyrazinecarboxamide (90 mg, 0.250 mmol) in DMF (2 mL) were added K₂C0₃ (121 mg, 0.875 mmol) and iodopropane (0.156 mL, 2 mmol). The resulting reaction mixture was stirred at 50°C for 5 days. The crude residue was filtered and purified by preparatory HPLC (basic conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (1 1 .2 mg, 1 1 %). LC/MS: m/z 404.1 (M+H)⁺: Rt 0.86 min.

Intermediate 150

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H-

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-hydroxy-1 H-benzimidazol-2-yl)methyl]- 2-pyrazinecarboxamide (1 10 mg, 0.304 mmol) in dry pyridine (3 mL) cooled to 0°C was added acetyl chloride (0.043 mL, 0.608 mmol). The resulting reaction mixture was stirred at room temperature for 24 hours. The mixture was then concentrated under a stream of nitrogen at 50°C to afford the title compound. LC/MS: m/z 404.2 (M+H)⁺ ; Rt 0.62 min.

Intermediate 151

3,5-diamino-6-chloro-N-{[1 -ethyl-6-(hydroxymethyl)-1 H-benzimidazol-2-yl]methyl}-2- pyrazinecarboxamide

A solution of methyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 - ethyl-1 H-benzimidazole-6-carboxylate (1300 mg, 3.22 mmol) in dry THF (30 mL) was added dropwise to a slurry of LiAIH₄ (244 mg, 6.44 mmol) in THF (30 mL). The reaction mixture was stirred at rt for 70 h, then cooled in ice-bath and 1 .2 mL of saturated Na₂S0₄ was added. The reaction mixture was stirred at rt for 1 h. The solid was filtered and then washed with ethyl acetate (20 mL), DCM (20 mL) and MeOH (15 mL). Part of the crude product obtained at this stage was not purified. The rest of it was dissolved in 10 mL of DMSO and purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (300 mg, 0.798 mmol, 24.80 % yield). LC/MS: m/z 376.1 (M+H)⁺, Rt 0.52 min.

Intermediate 152

methyl ({[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazol-6-yl]methyl}oxy)acetate

To a solution of 3,5-diamino-6-chloro-N-{[1 -ethyl-6-(hydroxymethyl)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (50 mg, 0.133 mmol) in DMF (2 mL) was added NaH as a 60 % suspension in mineral oil (7.3 mg, 0.183 mmol). The reaction mixture was stirred at rt for 1 hour. Methyl bromoacetate (0.019 mL, 0.200 mmol) was added and the reaction mixture was stirred at rt for 18 hours and then heated at 50°C for 4 hours. 2 mL of water were added to quench the reaction mixture which was then concentrated under vacuum. This experiment was repeated a second time with the same quantities of starting materials and both batch were combined and purified by preparatory HPLC (0.1 % NH₄OH) to afford the title compound (25 mg). LC/MS: m/z 448.0 (M+H)⁺, Rt 0.67 min.

Intermediate 153

Methyl 3-{2-[({[(1 ,1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1 H-benzimidazol-1 - yl}propanoate

A mixture of 1 , 1-dimethylethyl (1 H-benzimidazol-2-ylmethyl)carbamate (150 mg, 0.607 mmol), methyl 3-bromopropanoate (0.066 mL, 0.607 mmol) and K₂C0₃ (151 mg, 1.092 mmol) in DMSO (2 mL) was heated at 50°C for 17 h. The reaction mixture was purified by preparatory HPLC (0.1 % NH₄OH). The product fractions were dried under vacuum, combined and concentrated under a stream of nitrogen at 50°C, giving the title compound (157 mg, 0.471 mmol, 78 % yield). LC/MS: m/z 334.2 (M+H)+, Rt 0.64 min.

Intermediate 154

Methyl 3-[2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-1 H- benzimidazol-1 - l ro anoate\

To a solution of methyl 3-{2-[({[(1 , 1 -dimethylethyl)oxy]carbonyl}amino)methyl]-1 H- benzimidazol-1-yl}propanoate (154 mg, 0.461 mmol) in DCM(3 mL) was added 4 M HCI in dioxane (0.890 mL, 3.56 mmol). The mixture was stirred at rt for 6 h then concentrated under a stream of nitrogen at 50°C to give the deprotected amine. 3-chloro-5-(1 H- imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (1 10 mg, 0.461 mmol), triethylamine (0.642 mL, 4.61 mmol) and DMSO (3 mL) were added to the amine. The reaction mixture was stirred at rt for 17 h then stirred for another 18 h. The crude product was by preparatory HPLC (0.1 % NH₄OH). The product fractions were dried under EZ2 Genevac evaporator, combined and concentrated under a stream of nitrogen at 50°C, giving the title compound (108 mg, 0.267 mmol, 58.0 % yield) as a solid. LC/MS: m/z 404.3 (M+H)+, Rt 0.58 min.

Intermediate 155

N-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazol-6-yl]carbonyl}glycine

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylic acid (1 .3 g, 3.34 mmol) in DMSO (20 mL) at room

temperature was added HATU (1.268 g, 3.34 mmol) then triethylamine (1 .395 mL, 10.01 mmol). After stirring for 1 hour, methyl glycinate (0.419 g, 3.34 mmol) was added and the resulting solution was stirred at room temperature over the week-end (65 hours). More HATU (0.6 g, 1 .578 mmol) was added and the reaction mixture was stirred at rt for one hour before adding an excess of methyl glycinate (0.198 g, 1.577 mmol). The reaction mixture was stirred at rt overnight (22 hours). Diethyl ether (40 mL) was added and the resulting salts were filtered off. The filtrate was concentrated under reduced pressure to give a DMSO solution, which was purified by preparatory HPLC under basic conditions to give 1 197 mg of a mixture containing the title acid products and the corresponding methyl ester precursor. The material was re-dissolved in 20 mL of DMSO and further purified by preparatory HPLC under neutral conditions. The fractions containing the carboxylic acid were evaporated down under vacuum, combined, re-concentrated under reduced pressure and dried overnight under high vacuum to afford N-{[2-({[(3,5-diamino-6-chloro- 2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H-benzimidazol-6-yl]carbonyl}glycine (419.6 mg, 0.939 mmol, 28.2 % yield). LC/MS 70-B1 : m/z 447.2 (M+H)⁺, Rt 0.57 min.

Intermediate 156

Ethyl ({[2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazol-6-yl]carbonyl}thio)acetate

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylic acid (0.164 g, 0.421 mmol) in DMSO (3 mL) at room temperature was added HATU (0.160 g, 0.421 mmol) then triethylamine (0.070 mL, 0.505 mmol). After stirring for 15 mins, ethyl mercaptoacetate (0.046 mL, 0.421 mmol) was added and the resulting mixture was stirred at room temperature for 16 hours. Purification was done by injecting the reaction mixture (after filtration) straight onto a preparatory HPLC system and eluting with a acetonitrile/water gradient (neutral conditions). The relevant fractions were combined and evaporated to give ethyl ({[2-({[(3,5-diamino-6- chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H-benzimidazol-6- yl]carbonyl}thio)acetate (134 mg, 0.272 mmol, 64.7 % yield) as a pale yellow solid.

LC/MS: m/z 492.3 (M+H)+, Rt 0.82 min

EXAMPLES

Example 1

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 W-

To a suspension of1 H-benzimidazol-2-ylmethyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (50 mg, 0.157 mmol) and potassium carbonate (54.2 mg, 0.392 mmol) in DMF (1 ml) was added iodomethane (0.05 ml, 0.800 mmol). The resulting mixture was allowed to stir for 14 hours at room temperature. The potassium carbonate was filtered off. To the filtrate was added 5ml of ether. A precipitate was formed. The solid was filtered off and washed with 2 ml of MeOH, 2ml of DCM and then 2 ml of ether, to give 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H- benzimidazol-3-ium iodide (19.7 mg, 0.042 mmol, 26.5 % yield). LC/MS: m/z 348.8 (M+H)⁺, Rt 0.71 min.

Example 2

6^romo-2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl- 1 H-benzimidazol-3-ium iodide

A solution of (5-bromo-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (150 mg, 0.377 mmol) in DMF (1 ml) was added to a suspension of sodium hydride. (15.84 mg, 0.396 mmol) in DMF (1 ml). The reaction mixture was stirred for 1 hour at room temperature, and then iodomethane (0.08 ml, 1 .279 mmol) was added drop wise. The reaction mixture was stirred 5 hours at 50°C. The formed solid was filtered, washed with methanol (5 ml) and ether(10 ml), then dried in air to give 6-bromo- 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H- benzimidazol-3-ium iodide (75 mg, 0.135 mmol, 35.9 % yield). LC/MS: m/z 429.9 (M)⁺, Rt 0.70 min.

Example 3

6-chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl- 1 H-benzimidazol-3-ium iodide

A solution of (5-chloro-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (150 mg, 0.425 mmol) in DMF (1 ml) was added to a suspension of sodium hydride (16.99 mg, 0.425 mmol) in DMF (1 ml). The mixture was stirred for 1 hour at room temperature, and then iodomethane (0.09 ml, 1.439 mmol) was added. The reaction mixture was stirred 10 hours at 50°C then cooled down. To the solution was added 5 ml of ether. The formed solid was filtered, washed with methanol (5 ml) and ether (5 ml), then dried in air to give 6-chloro-2-({[(3,5-diamino-6-chloro-2- pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H-benzimidazol-3-ium iodide (106 mg, 0.208 mmol, 49.0 % yield). LC/MS: m/z 381 .0 (M)⁺, Rt 0.65 min.

Example 4

6-cyano-2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl- -benzimidazol-3-ium iodide

To a solution of NaH (25.6 mg, 0.640 mmol) in DMF (2 ml) was added a solution of (5- cyano-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate (200 mg, 0.582 mmol) in DMF (2ml). The mixture was stirred for 1 hour at room temperature, and then iodomethane (0.15 mL, 2.399 mmol) was added. The reaction mixture was stirred 4 hours at 50°C. To the reaction mixture were added 20 ml of ether. The formed solid was filtered, washed with 5 ml of methanol and 10ml of ether giving 65 mg of a crude product which was dissolved in DMSO (3 mL) and purified by preparatory HPLC (neutral conditions). The desired fractions were concentrated under a stream of nitrogen at 50°C, to give 6-cyano-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl- 1 H-benzimidazol-3-ium iodide (23 mg, 0.046 mmol, 7.91 % yield). LC/MS: m/z 371 .9 (M)⁺, Rt 0.55 min.

Example 5

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-6- (methyloxy)-1 H-benzimidazol-3-ium iodide

r

To a suspension of sodium hydride (12.24 mg, 0.306 mmol) in DMF(1 ml) was added a solution [5-(methyloxy)-1 H-benzimidazol-2-yl]methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (97 mg, 0.278 mmol) in DMF(1 ml). The resulting reaction mixture was stirred for 1 hour at room temperature, then iodomethane (0.061 ml, 0.973 mmol) was added drop-wise. The reaction mixture was stirred 6 hours at 50°C. To the solution were added 5 ml of ether. The formed solid was filtered, washed with methanol (5 ml), water (2 ml) and ether (10 ml) and dried to give 83 mg of a product which was dissolved in DMSO (3 ml_), and purified by preparatory HPLC (neutral conditions). The desired fractions were concentrated to give 2-({[(3,5-diamino-6-chloro-2- pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-6-(methyloxy)-1 H-benzimidazol-3-ium iodide (60 mg, 0.1 19 mmol, 42.7 % yield). LC/MS: m/z 376.9 (M)⁺, Rt 0.63 min. Example 6

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3,6-trimethyl-1 H-

To a suspension of sodium hydride (14.15 mg, 0.354 mmol) in DMF (1 ml) was added a solution (5-methyl-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (107 mg, 0.322 mmol) in DMF(1 ml). The resulting reaction mixture was stirred for 1 hour at room temperature, and then iodomethane (0.070 ml, 1 .125 mmol) was added. The reaction mixture was stirred 6hours at 50°C. To the solution were added 5 ml of ether. The formed solid was filtered, washed with methanol (5 ml), water (2 ml) and ether (10 ml) and dried in air giving 72 mg of a product which was dissolved in DMSO (3 ml_), and purified by preparatory HPLC (neutral conditions). The desired fractions were concentrated to give 2-({[(3,5-diamino-6-chloro-2- pyrazinyl)carbonyl]oxy}methyl)-1 ,3,6-trimethyl-1 H-benzimidazol-3-ium iodide (43.9 mg, 0.090 mmol, 27.9 % yield). LC/MS: m/z 361 .0 (M)+, Rt 0.66 min.

Example 7

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide

Method A

To a suspension of 1 H-benzimidazol-2-ylmethyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (100 mg, 0.314 mmol) in DMF(1 ml) was added 1 H-benzimidazol-2- ylmethyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate (100 mg, 0.314 mmol) in DMF (1 ml). The resulting mixture was stirred for 1 hour at room temperature, and then iodoethane (0.1 ml, 1.237 mmol) were added. The reaction mixture was allowed to stir 7 hours at

50°C. To the reaction mixture were added 10 ml of ether. A precipitate was formed. The solid was filtered off and washed with methanol (5 ml), water (2 ml) and ether (10 ml). The solid was dried to give 35mg of a product, which was dissolved in DMSO (2 mL), and purified by preparatory HPLC (neutral conditions). The desired fractions were

concentrated to give 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3- diethyl-1 H-benzimidazol-3-ium iodide (18 mg, 0.036 mmol, 1 1 .41 % yield). LC/MS: m/z 374.8 (M)+, Rt 0.65 min.

Method B

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (150 mg, 0.629 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-1 H-benzimidazol-3-ium iodide (251 mg, 0.754 mmol) in DMSO (2.5 mL) was added triethylamine (0.131 mL, 0.943 mmol). The reaction mixture was stirred at room temperature for 16 h. The crude reaction mixture was concentrated and then purified by preparatory HPLC (neutral conditions) to give the title compound (207.2 mg, 0.412 mmol, 65.58 % yield). LC/MS: m/z 375.0 (M)⁺, Rt 0.64 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.16 - 8.10 (m, 2 H), 7.77 - 7.70 (m, 2 H), 7.18 (br s., 4 H), 5.89 (s, 2 H), 4.72 (q, J = 7.2 Hz, 4 H), 1 .48 (t, J = 7.2 Hz, 6 H). Example 8

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-6-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

A mixture of {5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl 3,5-diamino-6-chloro- 2-pyrazinecarboxylate (150 mg, 0.353 mmol) and sodium hydride (15.53 mg, 0.388 mmol) in DMSO (1.5 mL) in a microwave vial was stirred at room temperature for 1 hour then iodomethane (0.077 mL, 1 .236 mmol) was added to the mixture. The resulting mixture was stirred at room temperature for 3 h. The crude product was purified by automated flash chromatography to give the title compound (18.7 mg, 0.032 mmol, 9.12 % yield). LC/MS: m/z 453.1 (M)⁺, Rt 0.77 min.

Example 9

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium trifluoroacetate

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (40 mg, 0.168 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-1 H-benzimidazol-3-ium iodide (55.7 mg, 0.168 mmol) in DMSO (1 mL) was added triethylamine (0.047 mL, 0.335 mmol). The reaction mixture was stirred at room temperature over the weekend and then for another day. The crude product was concentrated under vacuum and then purified by preparatory HPLC (0.1 % TFA) to give the title compound (15.9 mg, 0.033 mmol, 19.40 % yield). LC/MS: m/z 375.0 (M)+, Rt 0.65 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.27 - 8.00 (m, 2 H), 7.83 - 7.67 (m, 2 H), 7.19 (br. s., 4 H), 5.89 (s, 2 H), 4.72 (d, J = 7.3 Hz, 4 H), 1 .48 (t, J = 7.2 Hz, 6 H) Example 10

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-1 H- benzimidazol-3-ium iodide

To a suspension of (1 -ethyl-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (50 mg, 0.144 mmol) in DMSO (1 mL) was added iodomethane (0.036 mL, 0.577 mmol). The resulting mixture was stirred at room temperature for 17 h. LC/MS showed 50 % conversion. It was stirred at room temperature for 8 days. The crude product was purified by preparatory HPLC (neutral conditions). The product fractions were evaporated, combined (using acetronitrile) and concentrated under a stream of nitrogen at 50°C, followed by high vacuum for 4 h to give the title compound (16 mg, 0.033 mmol, 22.71 % yield) as a solid. LC/MS: m/z 361 .0 (M)⁺, Rt 0.59 min.

Example 11

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3- henylmethyl)-l H-benzimidazol-3-ium bromide

To a suspension of (1 -ethyl-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (100 mg, 0.288 mmol) in DMF (2 mL) was added. Benzyl bromide (0.137 mL, 1.153 mmol). The resulting mixture was stirred at room temperature for 17 h. LC/MS showed 29 % conversion of the desired product. It was stirred at 50°C for 1 day. The crude product was purified by preparatory HPLC (neutral conditions). The product fractions were evaporated, combined (using acetronitrile) and concentrated under a stream of nitrogen at 50°C to give the title compound (45 mg, 0.087 mmol, 30.1 % yield) as solid. LC/MS: m/z 437.1 (M)⁺, Rt 0.78 min. Example 12

6-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl- 1 H-benzimidazol-3-ium iodide

To a suspension of NaH (8.85 mg, 0.221 mmol) in DMF (1 .5 ml) was added (5-bromo-1 H- benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate (80 mg, 0.201 mmol). The resulting mixture was stirred at room temperature for 1 h, then iodoethane (0.065 ml, 0.805 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The reaction mixture was heated at 50°C for 17 h and then at room temperature for 10 days. The crude product was purified by preparatory HPLC (neutral conditions). The product fractions were evaporated, combined and concentrated under a stream of nitrogen at 50°C, giving the title compound (20 mg, 0.034 mmol, 17.09 % yield) as a solid. LC/MS: m/z 453.0 (M+)⁺, Rt 0.78 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.51 (d, J = 1.8 Hz, 1 H), 8.1 1 (d, J = 8.8 Hz, 1 H), 7.90 (dd, J = 1.8, 9.0 Hz, 1 H), 7.18 (br. s., 4 H), 5.87 (s, 2 H), 4.70 (dd, J = 2.8, 7.3 Hz, 4 H), 1 .46 (td, J = 1.6, 7.2 Hz, 6 H).

Example 13

6-Cyano-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide

To a suspension of NaH (10.24 mg, 0.256 mmol) in DMF (1.5 ml) was added (5-cyano- 1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2-pyrazinecarboxylate. The resulting mixture was stirred at room temperature for 1 h, and then iodoethane (0.075 ml, 0.931 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The reaction mixture was heated at 50°C for 17 h. and then stirred at room temperature for 10 days. The crude product was purified by preparatory HPLC (neutral conditions). The product fractions were evaporated, combined and concentrated under a stream of nitrogen at 50°C, giving the title compound (3 mg, 5.68 μηηοΙ, 2.442 % yield) as solid. LC/MS: m/z 400.0 (M)⁺, Rt 0.66 min. Example 14

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3,6-dimethyl- 1 H-benzimidazol-3-ium iodide

Γ

A mixture of (1-ethyl-6-methyl-1 H-benzimidazol-2-yl)methyl 3,5-diamino-6-chloro-2- pyrazinecarboxylate (20 mg, 0.055 mmol) and iodomethane (10.40 μΙ_, 0.166 mmol) in

DMF (1 mL) was stirred at 50°C overnight. The crude product was purified by

preparatory HPLC (neutral conditions). The product fractions were evaporated, combined and concentrated under a stream of nitrogen at 50°C, giving the title compound (3 mg,

5.97 μπΊθΙ, 10.77 % yield) as solid. LC/MS: m/z 374.8 (M)⁺, Rt 0.70 min.

Example 15

3,5-diamino-N-[(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- razinecarboxamide iodide

To a solution of 3,5-diamino-N-[(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro- 2-pyrazinecarboxamide (42.5 mg, 0.1 mmol) in ethanol (0.5 mL) and tetrahydrofuran (1 mL) was added Mel (0.013 mL, 0.200 mmol). The resulting reaction mixture was stirred at room temperature for 67 h and then was heated at 50°C for 165 h. The precipitate in the reaction mixture was filtered out, washed with acetone (0.3 mL), dried under high vacuum to afford the title compound (20.6 mg, 0.036 mmol, 36.4 % yield). LC/MS: m/z 438 (M)⁺, Rt 0.74 min. ¹ H NMR (400 MHz, DMSO-d6) δ ppm 8.92 (1 H, t, J=5.27 Hz), 8.43 (1 H, d, J=1.51 Hz), 8.01 (1 H, d, J=8.78 Hz), 7.87 (1 H, dd, J=8.78, 1 .76 Hz),7.18 (2 H, br. s.), 4.94 (2 H, d, J=5.52 Hz), 4.64 (2 H, q, J=7.03 Hz), 4.1 1 (3 H, s), 1 .36 (3 H, t, J=7.15 Hz) Example 16

3,5-diamino-N-[(5-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- pyrazinecarboxamide iodide

To a solution of 3,5-diamino-N-[(5-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro- 2-pyrazinecarboxamide (42.5 mg, 0.1 mmol) in ethanol (0.5 mL) and tetrahydrofuran (1 mL) was added Mel (0.013 mL, 0.200 mmol). The resulting reaction mixture was stirred at room temperature for 67 h and then was heated at 50°C for 48 h. The precipitate in the reaction mixture was filtered out, washed with acetone (0.3 mL), dried under high vacuum to afford the title compound (12.1 mg, 0.021 mmol, 21 .35 % yield). LC/MS: m/z 438 (M)⁺, Rt 0.73 min. ¹H NMR (400 MHz, DMSO-d6) δ ppm 8.92 (1 H, t, J=5.27 Hz), 8.41 (1 H, d, J=1.51 Hz), 8.03 (1 H, d, J=9.03 Hz), 7.86 (1 H, dd, J=8.78, 1 .76 Hz),7.18 (2 H, br. s.), 4.94 (2 H, d, J=5.52 Hz), 4.65 (2 H, q, J=7.19 Hz), 4.10 (3 H, s), 1.36 (3 H, t, J=7.15 Hz) Example 17

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6- (methyloxy)-l H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (150 mg, 0.629 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-6-(methyloxy)-1 /-/-benzimidazol-3-ium iodide (250 mg, 0.691 mmol) in DMSO (2.5 mL) was added triethylamine (0.131 mL, 0.943 mmol). The reaction mixture was stirred at room temperature for 16 h. The crude reaction mixture was purified by preparatory HPLC (neutral conditions), giving the title compound (127.07 mg, 0.238 mmol, 37.93 % yield). LC/MS: m/z 405.3 (M)+, Rt 0.69 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.02 (d, J = 9.0 Hz, 1 H), 7.63 (d, J = 2.3 Hz, 1 H), 7.33 (dd, J = 2.4, 9.2 Hz, 1 H), 7.19 (br. s., 2 H), 5.84 (s, 2 H), 4.73 - 4.62 (m, 4 H), 3.93 (s, 3 H), 1.46 (td, J = 3.5, 7.2 Hz, 6 H). Example 18

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-methyl- 1 H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (100 mg, 0.419 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-6-methyl-1 H-benzimidazol-3-ium iodide (145 mg, 0.419 mmol) in DMSO (2 mL) was added triethylamine (0.088 mL, 0.629 mmol). The reaction mixture was stirred at room temperature for 16 h. The crude reaction mixture was purified by preparatory HPLC (neutral conditions), giving the title compound (73.53 mg, 0.143 mmol, 33.1 1 % yield). LC/MS: m/z 389.2 (M)+, Rt 0.60 min. ¹ H NMR (400MHz ,DMSO-d₆) δ = 8.00 (d, J = 8.5 Hz, 1 H), 7.93 (s, 1 H), 7.57 (d, J = 8.5 Hz, 1 H), 7.18 (br. s., 4 H), 5.86 (s, 2 H), 4.73 - 4.62 (m, 4 H), 2.55 (s, 3 H), 1.52 - 1.43 (m, 6 H).

Example 19

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-propyl-1 H- benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (42 mg, 0.176 mmol) and 1 -ethyl-2-(hydroxymethyl)-3-propyl-1 H-benzimidazol-3-ium iodide (60.9 mg, 0.176 mmol) in DMSO (2 mL) was added triethylamine (0.037 mL, 0.264 mmol). The reaction mixture was stirred at room temperature for 16 h. The crude reaction mixture was purified by preparatory HPLC (neutral conditions), giving the title compound (38 mg, 0.074 mmol, 41.8 % yield). LC/MS: m/z 389.1 (M)+, Rt 0.65 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.19 - 8.09 (m, 2 H), 7.78 - 7.70 (m, 2 H), 7.22 - 7.14 (m, 4 H), 5.89 (s, 2 H), 4.77 - 4.67 (m, 2 H), 4.66 - 4.58 (m, 2 H), 1.94 - 1.83 (m, 2 H), 1.48 (t, 3 H), 0.97 (t, 3 H). Example 20

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-(3- thien l)-1 H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (70 mg, 0.293 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-6-(3-thienyl)-1 H-3, 1-benzimidazol-3-ium iodide (122 mg, 0.293 mmol) in DMSO (2 ml.) was added triethylamine (0.061 ml_, 0.440 mmol). The reaction mixture was stirred at room temperature for 16 h. The reaction mixture was stirred at room temperature for 16 h. The crude reaction mixture was purified by preparatory HPLC (neutral conditions), giving the title compound (39.37 mg, 10.317 mmol, 22.95 %). LC/MS: m/z 457.0 (M)+, Rt 0.82 min.

Example 21

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-[4- methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (75 mg, 0.314 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide (138 mg, 0.314 mmol) in DMSO (2 ml.) was added triethylamine (47.7 mg, 0.471 mmol). The reaction mixture was stirred at room temperature for 17 h. The crude reaction mixture was purified by preparatory HPLC (neutral conditions), giving the title compound (73 mg, 0.12 mmol, 38.1 % yield). LC/MS: m/z 481 .1 (M)+, Rt 0.89 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.34 (d, J = 1 .3 Hz, 1 H), 8.16 (d, J = 8.8 Hz, 1 H), 8.00 (dd, J = 1.8, 8.8 Hz, 1 H), 7.85 - 7.77 (m, 2 H), 7.21 (br. s., 4 H), 7.14 - 7.06 (m, 2 H), 5.90 (s, 2 H), 4.76 (dd, J = 7.3, 17.6 Hz, 4 H), 3.84 (s, 3 H), 1.57 - 1 .42 (m, 6 H). Example 22

6-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3- meth l-1H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (130 mg, 0.545 mmol) and 6-bromo-1 -ethyl-2-(hydroxymethyl)-3-methyl-1 /-/-benzimidazol-3-ium iodide (216 mg, 0.545 mmol) in DMSO (2 mL) was added triethylamine (0.1 14 mL, 0.817 mmol). The reaction mixture was stirred at room temperature for 16 h. The crude reaction mixture purified by preparatory HPLC (neutral conditions), giving the title compound (50.5 mg, 0.089 mmol, 16.33 % yield). LC/MS: m/z 438.8 (M)+, Rt 0.72 min.

Example 23

5-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3- meth l-1H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (130 mg, 0.545 mmol) and 5-bromo-1 -ethyl-2-(hydroxymethyl)-3-methyl-1 H-benzimidazol-3-ium iodide (216 mg, 0.545 mmol) in DMSO (2 mL) was added triethylamine (0.1 14 mL, 0.817 mmol). The reaction mixture was stirred at room temperature for 16 h. The crude product was purified by preparatory HPLC (neutral conditions). The product fractions were evaporated, combined and concentrated under a stream of nitrogen at 50°C, giving the title compound (37.21 mg, 0.066 mmol, 12.03 % yield). LC/MS: m/z 439.0 (M)+, Rt 0.72 min. Example 24

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-5- -(methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-({1-ethyl-5-[4-(methyloxy)phenyl]-1 H- benzimidazol-2-yl}methyl)-2-pyrazinecarboxamide (20.3 mg, 0.045 mmol) in a 1 :1 ratio of Acetone (0.5 ml_):DMSO (0.500 mL) was added Mel (5.62 μΙ_, 0.090 mmol). The reaction mixtures were stirred at room temperature for 48 h. The crude product was purified by preparatory HPLC (neutral conditions), giving the title compound (3.9 mg, 6.57 μηηοΙ, 14.62 % yield). LC/MS: m/z 466.0 (M)⁺, Rt 0.83 min. ¹H NMR (400MHz

,ACETONITRILE-d₃) δ = 8.29 - 8.21 (m, 1 H), 8.03 - 7.99 (m, 1 H), 7.95 - 7.86 (m, 2 H), 7.72 (d, J = 8.8 Hz, 2 H), 7.09 (d, J = 8.8 Hz, 2 H), 5.86 (br. s., 4 H), 4.92 (d, J = 5.5 Hz, 2 H), 4.64 (dd, 2 H), 4.13 (s, 3 H), 3.86 (s, 3 H), 1.50 (t, J = 7.3 Hz, 3 H)

Example 25

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-5- (methylsulfonyl)-l H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-{[1 -ethyl-5-(methylsulfonyl)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (80 mg, 0.189 mmol) in DMSO (2 mL) was added iodomethane (50 μΙ, 0.800 mmol). The resulting mixture was allowed to stir at room temperature. The solution in DMSO was purified by preparatory HPLC under neutral conditions to give 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 3-methyl-5-(methylsulfonyl)-1 H-benzimidazol-3-ium iodide (38.9 mg, 0.069 mmol, 36.4 % yield). LC/MS: m/z 437.9 (M)⁺, Rt 0.61 min. Example 26

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-5-[2- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (77 mg, 0.324 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-5-[2-(methyloxy)phenyl]-1 H-benzimidazol-3-ium (141 .8 mg, 0.324 mmol) in DMSO (2.5 ml.) was added triethylamine (0.090 ml_, 0.647 mmol). The reaction mixture was stirred at room temperature for 2 days, concentrated and then purified by preparatory HPLC (neutral conditions) to afford the title compound (13 mg, 6.60 % yield). LC/MS: m/z 480.9 (M)⁺, Rt 0.85 min.

Example 27

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-5- 4-(methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-({1-ethyl-5-[4-(methyloxy)phenyl]-1 H- benzimidazol-2-yl}methyl)-2-pyrazinecarboxamide (19 mg, 0.042 mmol) in DMSO (1.5 ml.) was added Mel (5.26 μΙ_, 0.084 mmol). The reaction mixture was stirred at room temperature for 70 h and then purified by preparatory HPLC (neutral conditions) to afford the title compound (9 mg, 36.0 % yield). LC/MS: m/z 466.2 (M)⁺, Rt 0.81 min. 1 H NMR (400 MHz, ACETONITRILE-ds) δ ppm 1 .50 (3 H, t, J=7.28 Hz) 3.87 (2 H, s) 4.14 (2 H, s) 4.64 (2 H, q, J=7.19 Hz) 4.91 (2 H, d, J=5.52 Hz) 5.86 (2 H, br. s.) 7.08 - 7.12 (2 H, m) 7.70 - 7.74 (2 H, m) 7.90 (2 H, qd) 8.01 - 8.02 (1 H, m) 8.23 - 8.28 (1 H, m) Example 28

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- (4-methylphenyl)-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-{[1-ethyl-6-(4-methylphenyl)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (51 mg, 0.1 17 mmol) in DMSO (1 mL) was added Mel (0.015 mL, 0.234 mmol). The reaction mixture was stirred at room temperature for 48 h, concentrated down and purified by preparatory HPLC (neutral conditions) to afford the title compound (28.9 mg, 42.7 % yield). LC/MS: m/z 450.0 (M)⁺, Rt 0.88 min. ¹H NMR (400 MHz, DMSO-de) δ ppm 1.42 (3 H, t, J=7.15 Hz) 2.39 (3 H, s) 4.14 (3 H, s) 4.72 (2 H, q) 4.97 (2 H, d, J=5.27 Hz) 7.21 (2 H, br. s.) 7.34 - 7.38 (2 H, m) 7.74 - 7.78 (2 H, m) 7.97 - 8.01 (1 H, m) 8.08 (1 H, d, J=8.53 Hz) 8.29 - 8.32 (1 H, m) 8.97 (1 H, t, J=5.40 Hz)

Example 29

5-carboxy-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- methyl-1 H-benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-5-carboxylic acid (87 mg, 0.223 mmol) in DMSO (2 mL) was added Mel (0.028 mL, 0.446 mmol). The reaction mixture was stirred at room temperature for 23 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (6.5 mg, 5.48 % yield). LC/MS: m/z 404.0 (M)⁺, Rt 0.51 min. Example 30

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-5- [(ethyloxy)carbonyl]-3-methyl-1 H-benzimidazol-3-ium iodide

To a solution of ethyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 - ethyl-1 H-benzimidazole-5-carboxylate (35 mg, 0.084 mmol) in DMSO (1 mL) was added Mel (10.47 μί, 0.168 mmol). The reaction mixture was heated at 50°C for 19 h. 2 other equivalents of Mel (10.47 μί, 0.168 mmol) were added and the reaction mixture was heated at 50°C for 48 h. The crude product was purified by preparatory HPLC (neutral conditions) to afford the title compound (14.5 mg, 30.9 % yield). LC/MS: m/z 432.0 (M)⁺, Rt 0.76 min. ¹H NMR (400 MHz, DMSO-d₆) δ ppm 1.35 - 1.42 (6 H, m) 4.19 (3 H, s) 4.41 (2 H, q, J=7.19 Hz) 4.69 (2 H, q) 4.98 (2 H, d, J=5.27 Hz) 7.18 (2 H, br. s.) 8.15 - 8.25 (2 H, m) 8.61 (1 H, s) 8.94 (1 H, t, J=5.52 Hz)

Example 31

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-5- [(methyloxy)carbonyl]-1 H-benzimidazol-3-ium iodide

To a solution of methyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1- ethyl-1 H-benzimidazole-5-carboxylate (94 mg, 0.233 mmol) in DMSO (2 mL) was added Mel (0.029 mL, 0.466 mmol). The reaction mixture was stirred at room temperature for 17 h, and then heated at 50°C for 4 h. More Mel (0.029 mL, 0.446 mmol) was added and the reaction was put back to heat at 50°C for 72 h. The mixture was purified by preparatory HPLC (neutral conditions) to afford the title compound (7 mg, 5.51 % yield). LC/MS: m/z 417.9 (M)⁺, Rt 0.66 min. ¹ H NMR (400 MHz, ACETONITRILE-c/₃) δ ppm 1 .48 (3 H, t, J=7.40 Hz) 3.96 (3 H, s) 4.15 (3 H, s) 4.65 (2 H, q, J=7.28 Hz) 4.92 (2 H, d, J=5.77 Hz) 5.85 (2 H, br. s.) 7.94 (1 H, dd, J=8.78, 0.75 Hz) 8.23 - 8.30 (2 H, m) 8.50 (1 H, dd, J=1.38, 0.63 Hz). Example 32

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- [(methyloxy)carbonyl]-1 H-benzimidazol-3-ium iodide

To a solution of methyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1- ethyl-1 H-benzimidazole-6-carboxylate (200 mg, 0.495 mmol) in DMSO (2.5 mL) was added Mel (0.062 mL, 0.991 mmol). The reaction mixture was stirred at room

temperature for 40 hours. Another equivalent of Mel (0.031 mL, 0.495 mmol) was added and the reaction mixture was stirred at room temperature for 96 h. The crude product was purified by preparatory HPLC (neutral conditions) and dried to afford the title compound (13.6 mg, 5.03 % yield). LC/MS: m/z 418.0 (M)⁺, Rt 0.66 min.

Example 33

6-carboxy-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- methyl-1 H-benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazole-6-carboxylic acid (100 mg, 0.257 mmol) in DMSO (2 mL) was added Mel (0.032 mL, 0.513 mmol). The reaction mixture was stirred at room temperature for 23 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (25 mg, 18.33 % yield). LC/MS: m/z 403.9 (M)⁺, Rt 0.63 min. Example 34

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-6- [(ethylamino)carbonyl]-3-methyl-1 H-benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-N, 1- diethyl-1 H-benzimidazole-6-carboxamide (18 mg, 0.043 mmol) in DMSO (0.5 mL) was added Mel (5.40 μΙ_, 0.086 mmol). The reaction mixture was stirred at room temperature for 69 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (4.9 mg, 20.31 % yield). LC/MS: m/z 431 .1 (M)\ Rt 0.65 min.

Example 35

6-[(butyloxy)carbonyl]-2-({[(3,5-diamino-6-chloro-2- pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -3-methyl-1 H-benzimidazol-3-ium iodide

To a solution of butyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1- ethyl-1 H-benzimidazole-6-carboxylate (31 mg, 0.070 mmol) in DMSO (700 μΙ_) was added Mel (8.69 μΙ_, 0.139 mmol). The reaction mixture was stirred at room temperature for 48 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (17.1 mg, 41 .8 % yield). LC/MS: m/z 460.2 (M)⁺, Rt 0.86 min.

Example 36

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- {[(2-oxotetrahydro-3-thienyl)amino]carbonyl}-1 H-3,1 -benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-N- (2-oxotetrahydro-3-thienyl)-1 H-benzimidazole-6-carboxamide (77 mg, 0.157 mmol) in DMSO (1 .5 ml.) was added Mel (0.020 ml_, 0.315 mmol). The reaction mixture was stirred at room temperature for 27 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (13.3 mg, 13.39 % yield). LC/MS: m/z 503.2 (M)⁺, Rt 0.62 min. ¹ H NMR (400MHz, DMSO-d₆) δ = 9.1 1 - 9.00 (m, 1 H), 8.95 (t, J = 5.3 Hz, 1 H), 8.49 (s, 1 H), 8.16 (s, 2 H), 7.19 (br. s., 2 H), 5.06 - 4.87 (m, 3 H), 4.77 - 4.65 (m, 2 H), 4.15 (s, 3 H), 3.60 - 3.46 (m, 1 H), 3.44 - 3.35 (m, 1 H), 2.64 - 2.53 (m, 1 H), 2.43 - 2.26 (m, J = 7.0, 12.2, 12.4, 12.4 Hz, 1 H), 1.43 (t, J = 7.2 Hz, 3 H).

Example 37

Methyl N-{[2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- methyl-1 H-3,1 -benzimidazol-3-ium-6-yl]carbonyl}glycinate iodide

To a solution of methyl N-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-

1- ethyl-1 H-benzimidazol-6-yl]carbonyl}glycinate (64 mg, 0.139 mmol) in DMSO (1 .2 mL) was added Mel (0.017 mL, 0.278 mmol). The reaction mixture was stirred at room temperature for 48 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (17.1 mg, 20.43 % yield). LC/MS: m/z 475.2 (M)⁺, Rt 0.57 min. ¹H NMR (400MHz, DMSO-d₆) δ = 9.26 (t, J = 5.8 Hz, 1 H), 8.95 (t, 1 H), 8.52 (s, 1 H), 8.23 - 8.10 (m, 2 H), 7.21 (br. s., 1 H), 4.99 (d, J = 5.3 Hz, 2 H), 4.76 - 4.61 (m, 2 H), 4.23 - 4.05 (m, 5 H), 3.69 (s, 3 H), 1.44 (t, 3 H).

Example 38

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-6- [(ethyloxy)carbonyl]-3-methyl-1 H-benzimidazol-3-ium iodide

To a solution of ethyl 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 - ethyl-1 H-benzimidazole-6-carboxylate (35.8 mg, 0.086 mmol) in DMSO (700 μΙ_) was added Mel (10.71 μΙ_, 0.171 mmol). The reaction mixture was stirred at room

temperature for 24 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (1 1 mg, 22.94 % yield). LC/MS: m/z 432.0 (M)⁺, Rt 0.76 min.

Example 39

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- {[(2-oxotetrahydro-3-furanyl)amino]carbonyl}-1 H-3,1 -benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-N- (2-oxotetrahydro-3-furanyl)-1 H-benzimidazole-6-carboxamide (91 mg, 0.192 mmol) in DMSO (2 mL) was added Mel (0.024 mL, 0.385 mmol). The reaction mixture was stirred at room temperature for 97 hours and purified by preparatory HPLC (neutral conditions) to afford the title compound (13.0 mg, 1 1 % yield). LC/MS: m/z 486.8 (M)⁺, Rt 0.57 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 9.29 (d, J = 7.8 Hz, 1 H), 8.95 (t, J = 5.3 Hz, 1 H), 8.50 (s, 1 H), 8.16 (s, 2 H), 7.21 (br. s., 2 H), 4.99 (d, J = 5.3 Hz, 2 H), 4.93 - 4.80 (m, 1 H), 4.71 (q, J = 7.1 Hz, 2 H), 4.53 - 4.42 (m, 1 H), 4.39 - 4.26 (m, 1 H), 4.15 (s, 3 H), 2.59 - 2.54 (m, 1 H), 2.46 - 2.31 (m, 1 H), 1.44 (t, J = 7.2 Hz, 3 H).

Example 40

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- [4-(methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-({1-ethyl-6-[4-(methyloxy)phenyl]-1 H- benzimidazol-2-yl}methyl)-2-pyrazinecarboxamide (13 mg, 0.029 mmol) in DMSO (1.5 mL) was added Mel (3.60 μί, 0.058 mmol). The reaction mixture was stirred at room temperature for 5 days, concentrated down and then purified by preparatory HPLC (neutral conditions) to afford the title compound (2.52 mg, 14.75 % yield) as 86-A1.

LC/MS: m/z 466.1 (M)⁺, Rt 0.82 min. ¹ H NMR (400 MHz, ACETONITRILE-d₃) δ ppm 1 .50 (3 H, t, J=7.28 Hz) 3.86 (2 H, s) 4.10 (2 H, s) 4.67 (2 H, q, J=7.28 Hz) 4.91 (2 H, d, J=5.77 Hz) 5.86 (2 H, br. s.) 7.06 - 7.10 (2 H, m) 7.69 - 7.74 (2 H, m) 7.83 - 7.93 (2 H, m) 8.01 - 8.02 (1 H, m) 8.27 (1 H, t, J=5.52 Hz).

Example 41

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-5-[4- (methyloxy)phenyl]-1 H-3,1 -benzimidazol-3-ium iodide

A mixture of {1-ethyl-5-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl 3,5-diamino-6- chloro-2-pyrazinecarboxylate (100 mg, 0.221 mmol) and iodomethane (0.138 mL, 2.208 mmol) in DMSO (1.5 mL) was stirred at room temperature for 17 h. The solid from the reaction mixture was filtered. The filtrate was purified by preparatory HPLC (neutral conditions) to give the title compound (13 mg, 0.022 mmol, 9.90 % yield). LC/MS: m/z

467.0 (M)⁺, Rt 0.85 min.

Example 42

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl -3 -methyl -6-[4- methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide

I

A mixture of {1-ethyl-6-[4-(methyloxy)phenyl]-1 H-benzimidazol-2-yl}methyl 3,5-diamino-6- chloro-2-pyrazinecarboxylate (150 mg, 0.331 mmol) and iodomethane (0.062 mL, 0.994 mmol) in DMSO (1.5 mL) was stirred at room temperature over the weekend. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (5 mg, 8.41 mrnol, 2.54 %). LC/MS: m/z 467.0 (M)⁺, Rt 0.83 min.

Example 43

3-Butyl-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-1 H- benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (50 mg, 0.210 mmol) and 3-butyl-1 -ethyl-2-(hydroxymethyl)-1 H-benzimidazol-3-ium iodide in DMSO (1 mL) was added triethylamine (0.044 mL, 0.314 mmol). The reaction mixture was stirred at room temperature for 16 h. The reaction mixture was concentrated under vacuum. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (5.38 mg, 10.14 μπιοΙ, 4.84 %). LC/MS: m/z 403.0 (M)+, Rt 0.79 min. ¹ H

NMR (400MHz, DMSO-d₆) δ = 8.17 - 8.09 (m, 2 H), 7.79 - 7.68 (m, 2 H), 7.19 (br. s., 2 H), 5.89 (s, 2 H), 4.80 - 4.56 (m, 4 H), 1 .89 - 1 .76 (m, 2 H), 1 .53 - 1 .35 (m, 5 H), 0.93 - 0.84 (m, 3 H). Example 44

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-(2- h droxyethyl)-1 H-benzimidazol-3-ium bromide

A mixture of 3-chloro-5-(1 H-imidazol-1-ylcarbonyl)-2,6-pyrazinediamine (45 mg, 0.189 mmol), 1-ethyl-3-(2-hydroxyethyl)-2-(hydroxymethyl)-1 H-benzimidazol-3-ium bromide (56.8 mg, 0.189 mmol) and Triethylamine (28.6 mg, 0.283 mmol) in DMSO (1 mL) was stirred at room temperature for 17 h. The reaction mixture was concentrated under vacuum. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (24.29 mg, 0.051 mmol, 27.3 % yield). LC/MS: m/z 391.0 (M)+, Rt 0.63 min. ¹H NMR (400MHz, DMSO-d₆) δ = 8.16 - 8.10 (m, 2 H), 7.76 - 7.70 (m, 2 H), 7.18 (br. s., 2 H), 5.89 (s, 2 H), 5.17 (t, 1 H), 4.85 - 4.68 (m, 4 H), 3.86 - 3.77 (m, 2 H), 1.55 - 1.48 (m, 3 H).

Example 45

3-(Cyclopropylmethyl)-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)- 1 -eth l -1 H-benzimidazol-3-ium bromide

Br

A mixture of 3-chloro-5-(1 H-imidazol-1-ylcarbonyl)-2,6-pyrazinediamine (30 mg, 0.126 mmol), [1 -ethyl-2-(hydroxymethyl)-3-pentyl-1 H-benzimidazol-3-ium bromide (41 .1 mg, 0.126 mmol) and Et₃N (0.026 mL, 0.189 mmol) in DMSO (1 mL) was stirred at room temperature over the weekend. The reaction mixture was concentrated under vacuum. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (23.88 mg, 0.050 mmol, 39.4 % yield). LC/MS: m/z 401.0 (M)⁺, Rt 0.74 min. ¹H NMR (400MHz, DMSO-d₆) δ = 8.23 - 8.10 (m, 2 H), 7.80 - 7.70 (m, 2 H), 7.21 (br. s., 2 H), 5.90 (s, 2 H), 4.78 - 4.57 (m, 4 H), 1 .56 - 1 .45 (m, 4 H), 0.63 - 0.51 (m, 4 H). Example 46

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide

Method a

^***To a solution of 3,5-diamino-N-(1 H-benzimidazol-2-ylmethyl)-6-chloro-2- pyrazinecarboxamide (80 mg, 0.252 mmol) in DMF (3 ml.) was added NaH (10.57 mg, 0.264 mmol). The resulting mixture was stirred at room temperature for 1 h, then iodoethane (0.102 ml_, 1 .259 mmol) was added. The reaction mixture was stirred 50°C for 17 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (23 mg, 0.046 mmol, 18.21 % yield). LC/MS: m/z 374.0 (M)⁺, Rt 0.63 min. ¹H NMR (400MHz, DMSO-d₆) δ = 9.06 - 8.93 (m, 1 H), 8.06 (dd, J = 3.1 , 6.1 Hz, 2 H), 7.68 (dd, J = 3.0, 6.3 Hz, 2 H), 7.20 (br. s., 2 H), 4.99 (d, J = 5.0 Hz, 2 H), 4.73 - 4.60 (m, 4 H), 1.40 (t, J = 7.2 Hz, 6 H).

Method b:

To a solution of 2-(aminomethyl)-1 ,3-diethyl-1 H-3, 1 -benzimidazol-3-ium iodide (134 mg, 0.405 mmol) in DMSO (2m L) was added 3-chloro-5-(1 H-imidazol-1-ylcarbonyl)-2,6- pyrazinediamine (97 mg, 0.405 mmol). The mixture was then stirred at 60°C for 65 h. and then purified by preparatory HPLC (neutral) to afford the desired product which was further triturated with MeOH (0.5 mL) and dried under high vacuum to afford the title compound (17.2 mg, 8.47 % yield). LC/MS: m/z 374 (M)⁺, Rt 0.63 min. ¹H NMR (400 MHz, DMSO-d6) δ ppm 7.93 - 8.01 (2 H, m), 7.66 - 7.74 (2 H, m), 5.04 (2 H, s), 4.76 (4 H, q, J=7.28 Hz), 1 .55 (6 H, t, J=7.28 Hz). Example 47

2-({[(3,5-Diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-1 -ethyl-3,6-dimethyl- 1 H-benzimidazol-3-ium iodide

A mixture of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-methyl-1 H-benzimidazol-2-yl)methyl]-2- pyrazinecarboxamide (157 mg, 0.436 mmol) and iodomethane (0.082 mL, 1.309 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (90 mg, 0.179 mmol, 41 .1 % yield). LC/MS: m/z 374.0 (M)+, Rt 0.69 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.94 (t, 1 H), 7.90 (d, J = 8.5 Hz, 1 H), 7.85 (s, 1 H), 7.54 - 7.48 (m, 1 H), 7.20 (br. s., 2 H), 4.93 (d, J = 0.3 Hz, 2 H), 4.60 (q, 2 H), 4.09 (s, 3 H), 2.53 (s, 3 H), 1.37 (t, 2 H).

Example 48

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- methyloxy)-l H-benzimidazol-3-ium iodide

A mixture of 3,5-diamino-6-chloro-N-{[1 -ethyl-6-(methyloxy)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (130 mg, 0.346 mmol) and iodomethane (0.065 mL, 1.038 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound

(52 mg, 0.100 mmol, 29.0 % yield) and (19 mg, 0.037 mmol, 10.61 % yield). LC/MS: m/z 390.2 (M)⁺, Rt 0.55 min. ¹ H NMR (400MHz ,DMSO-d₆) δ = 8.95 - 8.90 (m, 1 H), 7.91 (d, J = 9.0 Hz, 1 H), 7.55 (d, J = 2.3 Hz, 1 H), 7.28 (dd, 1 H), 7.19 (br. s., 2 H), 4.91 (d, J = 5.3 Hz, 2 H), 4.60 (q, 2 H), 4.07 (s, 3 H), 3.90 (s, 3 H), 1.36 (t, 3 H). Example 49

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-{2-oxo-2- [(phenylmethyl)oxy]ethyl}-1 H-benzimidazol-3-ium

A mixture of 3-chloro-5-(1 H-imidazol-1-ylcarbonyl)-2,6-pyrazinediamine (47.1 mg, 0.197 mmol), 1-ethyl-2-(hydroxymethyl)-3-{2-oxo-2-[(phenylmethyl)oxy]ethyl}-1 H-benzimidazol- 3-ium bromide (80 mg, 0.197 mmol) and triethylamine (0.041 mL, 0.296 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. The reaction mixture was concentrated under vacuum. The crude product was purified by preparatory HPLC (with acidic conditions) to give the title compound (20.6 mg, 0.034 mmol, 17.1 1 %). LC/MS: mlz 494.9 (M)+, Rt 0.83 min. ¹H NMR (400MHz, DMSO-d₆) δ = 8.20 - 8.14 (m, 1 H), 8.1 1 - 8.04 (m, 1 H), 7.78 - 7.68 (m, 2 H), 7.41 - 7.28 (m, 3 H), 7.14 (br. s., 2 H), 5.93 (s, 2 H), 5.87 (s, 2 H), 5.04 (s, 2 H), 4.87 - 4.76 (m, 2 H), 1 .53 (t, 3 H).

Example 50

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-dimethyl-1 H-

To a solution of 3,5-diamino-N-(1 H-benzimidazol-2-ylmethyl)-6-chloro-2- pyrazinecarboxamide (80 mg, 0.252 mmol) in DMF (3 mL) was added NaH (1 1.08 mg, 0.277 mmol). The resulting mixture was stirred at room temperature for 1 h, and then iodomethane (0.079 mL, 1 .259 mmol) was added. The reaction mixture was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (17 mg, 0.049 mmol, 19.47 % yield). LC/MS: mlz 346.0 (M)⁺, Rt 0.64 min. ¹ H NMR (400MHz, DMSO-d₆) δ = 8.94 - 8.87 (m, 1 H), 8.07 - 7.97 (m, 2 H), 7.69 (dd, J = 3.1 , 6.4 Hz, 2 H), 7.18 (br. s., 2 H), 4.95 (d, J = 5.3 Hz, 2 H), 4.1 1 (s, 6 H).

Example 51

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1 -propyl- 1 H-benzimidazol-3-ium iodide

I

A mixture of iodomethane (0.018 mL, 0.292 mmol) and 3,5-diamino-6-chloro-N-[(1-propyl- 1 H-benzimidazol-2-yl)methyl]-2-pyrazinecarboxamide (35 mg, 0.097 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (20 mg, 0.040 mmol, 41 .0 % yield). LC/MS: m/z 374.0 (M)+, Rt 0.72 min. ¹H NMR (400MHz, DMSO-d₆) δ = 8.99 - 8.88 (m, 1 H), 8.1 1 - 7.97 (m, 2 H), 7.74 - 7.60 (m, 2 H), 7.17 (br. s., 2 H), 5.02 - 4.92 (m, 2 H), 4.63 - 4.49 (m, 2 H), 4.1 1 (s, 3 H), 1 .86 - 1 .71 (m, 2 H), 0.94 (t, 3 H).

Example 52

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3 -diethyl -5 -(4- h droxyphenyl)-1 H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (50 mg, 0.210 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-5-(4-hydroxyphenyl)-1 H-benzimidazol-3-ium iodide (89 mg, 0.210 mmol) in DMSO (2 mL) was added triethylamine (0.044 mL, 0.314 mmol). The reaction mixture was stirred at room temperature for 40 h. The reaction mixture was concentrated under vacuum. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (7 mg, 0.012 mmol, 5.62 %).

LC/MS: m/z 467.0 (M)⁺, Rt 0.79 min. ¹H NMR (400MHz, DMSO-d₆) δ = 9.71 (s, 1 H), 8.27 (s, 1 H), 8.12 (d, J = 8.8 Hz, 1 H), 7.99 - 7.90 (m, 1 H), 7.73 - 7.65 (m, 2 H), 7.18 (br. s., 2 H), 6.91 (d, J = 8.5 Hz, 2 H), 5.88 (s, 2 H), 4.83 - 4.66 (m, 4 H), 1.58 - 1 .43 (m, 6 H).

Example 53

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1 -[2- (methyloxy)ethyl]-1 H-benzimidazol-3-ium iodide

A mixture of iodomethane (0.025 mL, 0.407 mmol) and 3,5-diamino-6-chloro-N-({1 -[2- (methyloxy)ethyl]-1 H-benzimidazol-2-yl}methyl)-2-pyrazinecarboxamide (51 mg, 0.136 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (30.74 mg, 0.059 mmol, 43.8 %). LC/MS: m/z 390.1 (M)⁺, Rt 0.60 min. ¹H NMR (400MHz ,DMSO- d₆) δ = 8.83 - 8.73 (m, 1 H), 8.09 - 7.98 (m, 2 H), 7.74 - 7.62 (m, 2 H), 7.17 (br. s., 2 H), 5.00 (d, J = 5.5 Hz, 2 H), 4.89 (t, J = 4.8 Hz, 2 H), 4.12 (s, 3 H), 3.73 (t, J = 4.9 Hz, 2 H), 3.24 (s, 3 H).

Example 54

1 -(Cyclobutylmethyl)-2-({[(3,5-diamino-6-chloro-2- pyrazinyl)carbonyl]amino}methyl)-3-methyl-1 H-benzimidazol-3-ium iodide

A mixture of iodomethane (0.029 mL, 0.467 mmol) and 3,5-diamino-6-chloro-N-{[1- (cyclobutylmethyl)-1 H-benzimidazol-2-yl]methyl}-2-pyrazinecarboxamide (60 mg, 0.156 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (33.19 mg, 0.063 mmol, 40.4 %). LC/MS: m/z 400.1 (M)⁺, Rt 0.70 min. ¹H NMR (400MHz, DMSO- d₆) δ = 8.98 - 8.91 (m, 1 H), 8.12 - 7.97 (m, 2 H), 7.71 - 7.64 (m, 2 H), 7.18 (br. s., 2 H), 4.96 (d, J = 5.3 Hz, 2 H), 4.67 (d, J = 7.0 Hz, 2 H), 4.1 1 (s, 3 H), 2.88 - 2.76 (m, 1 H), 1 .96 - 1.74 (m, 6 H). Example 55

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-diethyl-6- trifluoromethyl)thio]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-({5-[(trifluoromethyl)thio]-1 H-benzimidazol-2- yl}methyl)-2-pyrazinecarboxamide (48 mg, 0.1 15 mmol) in DMF (3 mL) was added NaH (4.82 mg, 0.121 mmol). The resulting mixture was stirred at room temperature for 1 h, and then iodoethane (0.046 mL, 0.574 mmol) was added. The reaction mixture was stirred at room temperature for 17 h. It was then heated at 50°C for 9 days. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (9.61 mg, 0.016 mmol, 13.90 % yield). LC/MS: m/z 473.8 (M)⁺, Rt 0.84 min. ¹H NMR (400MHz, DMSO-d₆) δ = 8.97 (t, J = 5.1 Hz, 1 H), 8.63 (s, 1 H), 8.24 (d, J = 8.5 Hz, 1 H), 8.02 (s, 1 H), 7.20 (br. s., 2 H), 5.02 (d, J = 5.0 Hz, 2 H), 4.71 (dd, J = 7.5, 15.8 Hz, 4 H), 1.45 - 1.37 (m, 6 H). Example 56

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-dimethyl-6- [(trifluoromethyl)thio]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-({5-[(trifluoromethyl)thio]-1 H-benzimidazol-2- yl}methyl)-2-pyrazinecarboxamide (50 mg, 0.120 mmol) in DMF (3 mL) was added NaH (5.03 mg, 0.126 mmol). The resulting mixture was stirred at room temperature for 1 h, and then iodomethane (0.037 mL, 0.598 mmol) was added. The reaction mixture was stirred at room temperature for 24 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (6.81 mg, 0.012 mmol, 9.92 % yield). LC/MS: m/z 446.0 (M)⁺, Rt 0.67 min. Example 57

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1 - (phenylmethyl)-l H-benzimidazol-3-ium iodide

A mixture of iodomethane (0.034 mL, 0.537 mmol) and 3,5-diamino-6-chloro-N-{[1- (phenylmethyl)-1 H-benzimidazol-2-yl]methyl}-2-pyrazinecarboxamide (73 mg, 0.179 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. It was stirred at room temperature for 67 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (37.66 mg, 0.068 mmol, 38.3 %). LC/MS: mlz 422.0 (M)⁺, Rt 0.72 min.

Example 58

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1 -(2- methylpropyl)-1 H-benzimidazol-3-ium iodide

A mixture of iodomethane (0.046 mL, 0.738 mmol) and 3,5-diamino-6-chloro-N-{[1-(2- methylpropyl)-1 H-benzimidazol-2-yl]methyl}-2-pyrazinecarboxamide (92 mg, 0.246 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. It was stirred at room temperature for 67 h. The crude product was purified by preparatory HPLC (neutral conditions) to give the title compound (38.23 mg, 0.074 mmol, 30.1 % yield). LC/MS: mlz 388.0 (M)⁺, Rt 0.77 min. Example 59

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- [(phenylmethyl)oxy]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-({1 -ethyl-6-[(phenylmethyl)oxy]-1 H-benzimidazol - 2-yl}methyl)-2-pyrazinecarboxamide (33 mg, 0.073 mmol) in DMSO (0.7 ml.) was added Mel (6.85 μΙ_, 0.1 10 mmol). The resulting reaction mixture was stirred at room

temperature for 17 hours. The crude residue was filtered and purified by preparatory HPLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (14.4 mg, 33 %). LC/MS: m/z 465.9 (M)⁺, Rt 0.94 min. ¹H NMR (400 MHz, DMSO-d) δ ppm: 7.93 (1 H, d, J=9.3 Hz), 7.70 (1 H, d, J=2.3 Hz), 7.51 (2H, d, J=7.0 Hz), 7.42 (2H, t, J=7.0 Hz), 7.37 (2H, dd, J=9.0, 2.0 Hz), 5.28 (2H, s), 4.92 (2H, d, J=5.5 Hz), 4.60 (2H, q, J=7.0 Hz), 4.08 (3H, s), 1 .35 (3H, t, J=7.2 Hz).

Example 60

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- -(methyloxy)-2-oxoethyl]oxy}-1 H-benzimidazol-3-ium iodide

To a solution of methyl {[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 - ethyl-1 H-benzimidazol-6-yl]oxy}acetate (27 mg, 0.062 mmol) in DMSO (0.6 ml.) was added Mel (5.84 μΙ_, 0.093 mmol). The resulting reaction mixture was stirred at room temperature for 20 hours. The crude residue was filtered and purified by preparatory HPLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (13.2 mg, 37%). LC/MS: m/z 448.0 M⁺, Rt 0.78 min. ¹H NMR (400MHz, DMSO-d) δ ppm: 7.73 (1 H, d, J=9.0 Hz), 7.43 (1 H, d, J=2.3 Hz), 7.13 (1 H, dd, J=2.4, 9.2 Hz), 4.79 (2 H, s), 4.71 (2H, d, J=5.3 Hz), 4.37 (2H, q, J=7.0 Hz), 3.87 (3 H, s), 3.51 (3 H, s), 1 .14 (3H, t, J=7.2 Hz). Example 61

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-[2- (methyloxy)-2-oxoethyl]-1 H-benzimidazol-3-ium bromide

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-1 H-benzimidazol-2-yl)methyl]-2- pyrazinecarboxamide (69.2 mg, 0.2 mmol) in DMSO (2 mL) was added methyl bromoacetate (0.076 mL, 0.8 mmol). The resulting reaction mixture was stirred at room temperature for 40 hours. The crude residue was filtered and purified by preparatory HPLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (10.2 mg, 10 %). LC/MS: m/z 418.0 (M)⁺, Rt 0.75 min. ¹H NMR (400MHz, DMSO-d) δ ppm: 8.16-8.08 (1 H, m), 8.06-7.98 (1 H, m), 7.76-7.64 (2 H, m), 5.66 (2 H, s), 4.99 (2 H, d, J=5.5 Hz), 4.79 (2 H, q, J=7.0 Hz), 3.53 (3 H, s), 1 .48 (3 H, t, J=7.2 Hz).

Example 62

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-[2- (ethyloxy)-2-oxoethyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-1 H-benzimidazol-2-yl)methyl]-2- pyrazinecarboxamide (69.2 mg, 0.2 mmol) in DMSO (2 mL) was added ethyl iodoacetate (0.096 mL, 0.8 mmol). The resulting reaction mixture was stirred at room temperature for 48 hours. The crude residue was filtered and purified by preparatory HPLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (25.5 mg, 23 %). LC/MS: m/z 431.9 (M)⁺, Rt 0.78 min. ¹H NMR (400MHz, DMSO-d) δ ppm: 8.12 (1 H, dd), 8.03 - 7.97 (1 H, m), 7.75 - 7.65 (2 H, m), 5.63 (2 H, s), 4.99 (2 H, d, J=5.3 Hz), 4.79 (2 H, q, J=7.0 Hz), 3.95 (2 H, q, J=7.0 Hz), 1.48 (3 H, t, J=7.3 Hz), 1.13 (3 H, t, J=7.2 Hz). Example 63

[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazol-3-ium-3-yl]acetate

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-1 H-benzimidazol-2-yl)methyl]-2- pyrazinecarboxamide (69.2 mg, 0.2 mmol) in DMSO (2 mL) was added methyl bromoacetate (0.048 mL, 0.5 mmol). The resulting reaction mixture was stirred at room temperature for 24 hours. The crude residue was filtered and purified by preparatory HPLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum. The intermediate was taken back in a mixture of DMSO / 1 % aqueous solution of NH₄OH and stirred at room temperature for 18 hours. The desired product crashed out of the solution. It was filtered, washed with acetone and dried under high vacuum to afford the title compound (8.6 mg, 9 %). LC/MS: m/z 403.9 (M)⁺, Rt 0.65 min. ¹H NMR (400MHz, DMSO-d) δ ppm: 8.05 - 7.99 (2 H, m), 7.68 - 7.60 (2 H, m), 4.95 (2 H, s), 4.89 (2 H, d, J=5.5 Hz), 4.63 (2 H, q, J=7.0 Hz), 1.36 (3 H, t, J=7.2 Hz).

Example 64

6-bromo-2-({[(3,5-diamino-6-chloro-2^yrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-[2- (ethyloxy)-2-oxoethyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-N-[(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro- 2-pyrazinecarboxamide (42.5 mg, 0.1 mmol) in acetone (0.5 mL) and THF (0.500 mL) was added ethyl iodoacetate (0.013 mL, 0.1 10 mmol). The resulting reaction mixture was stirred at 50°C for 42 h. The precipitate in the reaction mixture was filtered out, washed with acetone (0.3 mL) and dried under high vacuum to afford the title compound (7.7 mg, 12.06 % yield). LC/MS: m/z 510 (M)⁺, Rt 0.80 min. Example 65

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- -(methyloxy)-3-oxopropyl]-1 H-3,1 -benzimidazol-3-ium iodide

To a solution of methyl 3-[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-

1- ethyl-1 H-benzimidazol-6-yl]propanoate (21 .59 mg, 0.05 mmol) in acetone (0.25 mL) and THF (0.250 mL) was added Mel (6.25 μί, 0.100 mmol). The resulting mixture was then stirred at room temperature for 239 h. The precipitate in the reaction mixture was filtered out, washed with acetone (0.3 mL) and dried under high vacuum to afford the title compound (14.1 mg, 49.1 % yield). LC/MS: m/z 446 (M)⁺, Rt 0.68 min.

Example 66

2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -3 -methyl -6- {[3-(methyloxy)-3-oxopropyl]thio}-1 H-benzimidazol-3-ium iodide

A mixture of iodomethane (9.30 μί, 0.149 mmol) and methyl 3-{[2-({[(3,5-diamino-6- chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-1 H-benzimidazol-6-yl]thio}propanoate (23 mg, 0.050 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. It was stirred at room temperature for 5 days. The crude product was purified by preparatory HPLC (neutral conditions), giving the title compound (5.31 mg, 8.76 μηηοΙ, 17.68 % yield) LC/MS: m/z 477.8 (M)+, Rt 0.72 min.

Example 67

6-Chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- meth l-1 H-benzimidazol-3-ium iodide

I

A mixture of iodomethane (0.049 mL, 0.789 mmol) and 3,5-diamino-6-chloro-/\/-[(6-chloro- 1-ethyl-1 H-benzimidazol-2-yl)methyl]-2-pyrazinecarboxamide (100 mg, 0.263 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions), giving the title compound (14.58 mg, 0.028 mmol, 10.62 % yield). LC/MS: m/z 393.7 (M)+, Rt 0.65 min.

Example 68

7-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- meth l-1 H-benzimidazol-3-ium iodide

I

A mixture of iodomethane (0.044 mL, 0.706 mmol) and 3,5-diamino-N-[(7-bromo-1-ethyl- 1 H-benzimidazol-2-yl)methyl]-6-chloro-2-pyrazinecarboxamide (100 mg, 0.235 mmol) in DMSO (2 mL) was stirred at room temperature for 17 h. It was stirred furthermore at room temperature for 2 days. The crude product was purified by preparatory HPLC (neutral conditions), giving the title compound (41 .48 mg, 0.073 mmol, 31.1 % yield). LC/MS: m/z 437.9 (M)+, Rt 0.62 min.

Example 69

5-Chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl- 1 H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (35 mg, 0.147 mmol) and 5-chloro-1 ,3-diethyl-2-(hydroxymethyl)-1 H-3, 1-benzimidazol-3-ium iodide (53.8 mg, 0.147 mmol) in DMSO (2 mL) was added triethylamine (0.031 mL, 0.220 mmol). The reaction mixture was stirred at room temperature for 17 h. The crude product was purified by preparatory HPLC (neutral conditions), giving the title compound (15.15 mg, 0.028 mmol, 19.23 % yield). LC/MS: m/z 409.1 (M)+, Rt 0.68 min. ¹ H NMR (400MHz ,DMSO-d₆) δ = 8.40 (d, J = 1.8 Hz, 1 H), 8.18 (d, J = 8.8 Hz, 1 H), 7.80 (dd, J = 1.9, 8.9 Hz, 1 H), 7.18 (br. s., 2 H), 5.88 (s, 2 H), 4.71 (dd, J = 5.1 , 7.2 Hz, 4 H), 1 .46 (td, J = 1 .4, 7.1 Hz, 6 H)

Example 70

6-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- propyl-1 H-benzimidazol-3-ium iodide

A mixture of 3,5-diamino-N-[(6-bromo-1 -ethyl-1 H-benzimidazol-2-yl)methyl]-6-chloro-2- pyrazinecarboxamide (100 mg, 0.235 mmol) and 1-iodopropane (0.230 mL, 2.355 mmol) in THF (1 mL) and Acetone (1.000 mL) was heated at 50°C for 17 h. It was heated at 50°C for 75 h. Some solid precipitated out from the reaction mixture and was filtered to give a crude material, which was purified by preparatory HPLC (neutral conditions), giving the title compound (18.32 mg, 0.031 mmol, 13.08 % yield). LC/MS: m/z 465.8 (M)+, Rt 0.80 min. ¹H NMR (400MHz ,DMSO-d₆) δ = 8.95 (t, 1 H), 8.44 (d, J = 1.8 Hz, 1 H), 8.07 (d, J = 8.8 Hz, 2 H), 7.85 (dd, J = 1.8, 8.8 Hz, 2 H), 7.19 (br. s., 2 H), 4.98 (d, J = 5.3 Hz, 2 H), 4.70 - 4.58 (m, 2 H), 4.58 - 4.47 (m, 2 H), 1 .85 - 1.70 (m, 2 H), 1 .37 (t, J = 7.3 Hz, 3 H), 0.93 (t, J = 7.3 Hz, 3 H). Example 71

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6- trifluoromethyl)thio]-1H-benzimidazol-3-ium iodide

To a solution of 3-chloro-5-(1 H-imidazol-1 -ylcarbonyl)-2,6-pyrazinediamine (60 mg, 0.251 mmol) and 1 ,3-diethyl-2-(hydroxymethyl)-6-[(trifluoromethyl)thio]-1 H-3, 1-benzimidazol-3- ium iodide (109 mg, 0.251 mmol) in DMSO (2 ml.) was added triethylamine (0.053 ml_, 0.377 mmol). The reaction mixture was stirred at room temperature for 16 h. The crude product was purified by preparatory HPLC (neutral conditions), giving the title compound (20.22 mg, 0.034 mmol, 13.34 % yield. LC/MS: m/z 474.9 (M)+, Rt 0.82 min.

Example 72

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -6-{3-[(3- hydroxypropyl)amino]-3-oxopropyl}-3-methyl-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-[(1 -ethyl-6-{3-[(3-hydroxypropyl)amino]-3- oxopropyl}-1 H-benzimidazol-2-yl)methyl]-2-pyrazinecarboxamide (27 mg, 0.057 mmol) in DMSO (0.8 ml.) was added Mel (7.1 1 μΙ_, 0.1 14 mmol). The resulting reaction mixture was stirred at room temperature for 17 h. The reaction mixture was purified by

preparatory HPLC (neutral). The relevant fractions were evaporated down under vacuum, dried under high vacuum to afford the title compound (14.6 mg, 41 .6 % yield). LC/MS: m/z 489 (M)⁺, Rt 0.54 min.

Example 73

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- -(4-morpholinyl)-3-oxopropyl]-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-({1-ethyl-6-[3-(4-morpholinyl)-3-oxopropyl]-1 H- benzimidazol-2-yl}methyl)-2-pyrazinecarboxamide (31 mg, 0.064 mmol) in DMSO (0.8 mL) was added Mel (7.96 μί, 0.127 mmol). The resulting reaction mixture was stirred at room temperature for 17 h. The reaction mixture was purified by preparatory HPLC (neutral). The relevant fractions were evaporated down under vacuum, dried under high vacuum to afford the title compound (21.3 mg, 53.2 % yield). LC/MS: m/z 501 (M)⁺, Rt 0.59 min.

Example 74

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -6-[3- ethylamino)-3-oxopropyl]-3-methyl-1 H-be

To a solution of 3,5-diamino-6-chloro-N-({1-ethyl-6-[3-(ethylamino)-3-oxopropyl]-1 H- benzimidazol-2-yl}methyl)-2-pyrazinecarboxamide (28 mg, 0.063 mmol) in DMSO (0.8 mL) was added Mel (7.87 μί, 0.126 mmol). The resulting reaction mixture was stirred at room temperature for 22 h. The reaction mixture was purified by preparatory HPLC (neutral). The relevant fractions were evaporated down under vacuum, dried under high vacuum to afford the title compound (25.4 mg, 68.8 % yield). LC/MS: m/z 459 (M)⁺, Rt 0.68 min.

Example 75

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -6-{3-[(2- hydroxyethyl)amino]-3-oxopropyl}-3-methyl- -benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-[(1-ethyl-6-{3-[(2-hydroxyethyl)amino]-3- oxopropyl}-1 H-benzimidazol-2-yl)methyl]-2-pyrazinecarboxamide (36 mg, 0.078 mmol) in DMSO (0.8 mL) was added Mel (9.77 μί, 0.156 mmol). The resulting reaction mixture was stirred at room temperature for 22 h. The reaction mixture was purified by

preparatory HPLC (neutral). The relevant fractions were evaporated down under vacuum, dried under high vacuum to afford the title compound (31 .7 mg, 67.3 % yield). LC/MS: m/z 475 (M)⁺, Rt 1 .1 1 min.

Example 76

6-[(carboxymethyl)oxy]-2-({[(3,5-diamino-6-chloro-2- pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -3-methyl-1 H-benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- methyl-6-{[2-(methyloxy)-2-oxoethyl]oxy}-1 H-benzimidazol-3-ium iodide (52 mg, 0.090 mmol) in a mixture of methanol (0.6 mL) and water (0.3 mL) was added LiOH (18.95 mg, 0.452 mmol). The reaction mixture was stirred at room temperature for 48 hours. The reaction mixture was added HI (0.096 mL, 0.723 mmol). The reaction mixture was filtered, washed with acetone and dried under high vacuum to afford the title compound (29 mg, 57 %). LC/MS: m/z 434.2 M⁺ Rt 0.52 min.

Example 77

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -6-(ethyloxy)- - methyl-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-{[1 -ethyl-6-(ethyloxy)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (9.4 mg, 0.024 mmol) in DMSO (0.4 mL) was added Mel (9.04 L, 0.142 mmol). The reaction mixture was stirred at room temperature for 48 hours. The crude residue was filtered and purified by preparatory H PLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (3 mg, 23%). LC/MS: m/z 404.3 M⁺, Rt 0.69 min. Example 78

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- (propyloxy)-l H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-{[1 -ethyl-6-(propyloxy)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (1 1 mg, 0.027 mmol) in DMSO (0.4 mL) was added Mel (10.22 μί, 0.164 mmol). The reaction mixture was stirred at room temperature for 48 hours. The crude residue was filtered and purified by preparatory HPLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (5 mg, 37%). LC/MS: m/z 418.3 M⁺, Rt 0.78 min.

Example 79

6-(acetyloxy)-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- -methyl-1 H-benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-1 H- benzimidazol-6-yl acetate (100 mg, 0.248 mmol) in DMF (2 mL) was added methyl iodide (0.184 mL, 2.972 mmol). The reaction mixture was stirred at room temperature for 6 days. The crude residue was concentrated, dissolved in DMSO, filtered and purified by preparatory HPLC (neutral conditions). The relevant fractions were combined and concentrated under vacuum to afford the title compound (17.9 mg, 13 %). LC/MS: m/z 418.1 M⁺, Rt 0.69 min. Example 80

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-6- (hydroxymethyl)-3-methyl-1 H-benzimidazol-3-ium iodide

To a solution of 3,5-diamino-6-chloro-N-{[1 -ethyl-6-(hydroxymethyl)-1 H-benzimidazol-2- yl]methyl}-2-pyrazinecarboxamide (30 mg, 0.080 mmol) in DMSO (1 mL) was added methyl iodide (10 μΙ_, 0.160 mmol). The reaction mixture was stirred at rt for 2 days and purified by preparatory HPLC (neutral conditions) to afford the title compound (9.9 mg, 23.95 % yield). LC/MS: m/z 390.2 (M)⁺, Rt 0.50 min.

Example 81

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- ({[2-(methyloxy)-2-oxoethyl]oxy}methyl)-1 H-benzimidazol-3-ium iodide

To a solution of methyl ({[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)- 1-ethyl-1 H-benzimidazol-6-yl]methyl}oxy)acetate (25 mg, 0.056 mmol) in DMSO (1 .5 mL) was added methyl iodide (10.47 μί, 0.167 mmol). The reaction mixture was stirred at rt overnight and then purified by preparatory HPLC (neutral conditions) to afford the title compound (3.78 mg, 1 1 .48 % yield). LC/MS: m/z 462.3 (M)⁺, Rt 0.62 min. Example 82

N-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl- 1 H-3,1 -benzimidazol-3-ium-6-yl]carbonyl}homoserine iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3- methyl-6-{[(2-oxotetrahydro-3-furanyl)amino]carbonyl}-1 H-3,1 -benzimidazol-3-ium iodide (25 mg, 0.041 mmol) in DMSO (200 μΙ) was added NH₄OH, 0.5 % (500 μΙ_, 0.071 mmol). The reaction mixture was heated at 50°C overnight. The solid present in the reaction mixture was filtered, washed with ether, and dried to afford the title compound (10 mg, 38.9 % yield). LC/MS: m/z 505.4 (M)⁺, Rt 0.50 min.

Example 83

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1 -[3- (methyloxy)-3-oxopropyl]-1 H-benzimidazol-3-ium iodide

A mixture of iodomethane (0.050 mL, 0.802 mmol) and methyl 3-[2-({[(3,5-diamino-6- chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 H-benzimidazol-1-yl]propanoate (108 mg, 0.267 mmol) in DMSO (2 mL) was stirred at rt for 37 h. The crude product was purified by preparatory HPLC (neutral conditions). The desired fractions were concentrated to afford the title compound (44.8 mg, 0.082 mmol, 30.7 % yield). LC/MS: m/z 417.9 (M)⁺, Rt 0.66 min. Example 84

6-((carboxymethyl)carbamoyl)-2-((3,5-diamino-6-chloropyrazine-2- carboxamido)methyl)-1 -ethyl-3-methyl-1 H-benzo[d]imidazol-3-ium

To a solution of N-{[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 1 H-benzimidazol-6-yl]carbonyl}glycine (419.6 mg, 0.939 mmol) in DMSO (12 ml.) was added Mel (0.1 17 ml_, 1.878 mmol). The reaction mixture was stirred at room

temperature over the week-end (85 hours). Another equivalent of Mel (60 μΙ_) was added and the reaction mixture was stirred at rt for another 48 hours then another equivalent of Mel (60 μΙ_) was added and the reaction mixture was stirred at rt overnight. Two other equivalents of Mel were added and the reaction mixture was stirred over the week-end (73 hours). The reaction mixture was heated at 50°C for 6 hours. The crude product was purified by preparatory HPLC under neutral conditions to afford N-{[2-({[(3,5-diamino-6- chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-1 H-3, 1-benzimidazol-3-ium-6- yl]carbonyl}glycine iodide (81 mg, 0.138 mmol, 14.65 % yield). LC/MS: m/z 461.2 (M)+, Rt 0.52 min.

Example 85

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl -6-({[2- (ethyloxy)-2-oxoethyl]thio}carbonyl)-3-methyl-1 H-3 1 -benzimidazol-3-ium iodide

To a solution of ethyl ({[2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 - ethyl-1 H-benzimidazol-6-yl]carbonyl}thio)acetate (100 mg, 0.203 mmol) in DMSO (2 ml.) was added iodomethane (0.064 ml_, 1.016 mmol). The resulting solution was allowed to stir at room temperature. More iodomethane (0.064 ml_, 1.016 mmol) was added. The reaction mixture was filtered, then injected and purified by preparatory HPLC eluting with a gradient of acetonitrile in water (20 to 90%). The fractions containing the desired product were combined and evaporated to give 2-({[(3,5-diamino-6-chloro-2- pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-6-({[2-(ethyloxy)-2-oxoethyl]thi methyl-1 H-3,1 -benzimidazol-3-ium iodide (54 mg, 0.085 mmol, 41.9 % yield) as a yellow solid. LC/MS: m/z 506.2 (M)+, Rt 0.84 min.

Example 86

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6- ({[(3S)-2-oxotetrahydro-3-furanyl]amino}carbonyl)-1 H-3,1 -benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-1 H- benzimidazole-6-carboxylic acid (2.0 g, 5.13 mmol) in DMSO (30 mL) at room temperature was added HATU (3.90 g, 10.26 mmol) then triethylamine (2.503 mL, 17.96 mmol). After stirring for one hour, (3S)-3-aminodihydro-2(3H)-furanone (1.868 g, 10.26 mmol) was added and the resulting solution was allowed to stir at room temperature overnight. The reaction mixture was concentrated under a stream of nitrogen at 50°C in 8 different vials. A large excess of DCM was then added to the different vials to make the desired product crash out. The solid contents of the vials were combined, filtered off and washed with DCM to give the amide intermediate. The material was taken back in DMSO (50 mL) and Mel (0.625 mL, 10.00 mmol) was added. The reaction mixture was stirred at room temperature over the week end. The crude product was purified by two successive preparatory HPLC under neutral conditions. The title compound was recovered in three separate batches totaling 339 mgs with purity of 95-97%. LC/MS: m/z 487.1 (M)⁺, Rt 0.54 min. 1H NMR (400 MHz, DMSO-d₆) δ 9.32 (d, J = 8.03 Hz, 1H), 8.96 (t, J = 5.27 Hz,

1H), 8.52 (s, 1H), 8.17 (s, 2H), 7.20 (br. s., 4H), 5.00 (d, J = 5.02 Hz, 2H), 4.84 - 4.88 (m, 1H), 4.71 (q, J= 6.94 Hz, 2H), 4.43 - 4.51 (m, 1H), 4.29 - 4.37 (m, 1H), 4.16 (s, 3H), 2.53 - 2.60 (m, 1H), 2.37 - 2.44 (m, 1H), 1.44 (t, J= 7.15 Hz, 3H) Example 87

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl-6-

({[(3R)-2-oxotetrahydro-3-furanyl]amino}carbonyl)-1 H-3,1 -benzimidazol-3-ium iodide

To a solution of 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl- 1 H-benzimidazole-6-carboxylic acid (1 .949 g, 5 mmol) in DMSO (30 mL) at room temperature was added Ν,Ν-diethylethanamine triethylamine (2.426 mL, 17.50 mmol) then HATU (2.281 g, 6.00 mmol). After stirring for 1 hour, (3R)-3-aminodihydro-2(3H)- furanone (0.740 g, 5.50 mmol) was added and the resulting solution was allowed to stir at room temperature overnight (14 hours). The mixture was concentrated under a stream of nitrogen at 50°C in 8 different vials. A large excess of DCM was then added to the different batches and the desired product crashed out. The solids were filtered off each vial, combined and washed with DCM to the amide intermediate. The material was taken back in DMSO (50 mL) and Mel (0.625 mL, 10.00 mmol) was added. The reaction mixture was stirred at room temperature overnight. The crude product was purified by two successive preparatory HPLC under neutral conditions. The title compound was recovered in three separate batches totaling 214 mgs with purity of 97-100%. LC/MS: m/z 487.1 (M)⁺, Rt 0.56 min. 1H NMR (400 MHz, DMSO-d₆) δ 9.37 (d, J= 7.53 Hz, 1H), 8.96 (br. s., 1H), 8.54 (br. s., 1H), 8.17 (br. s., 2H), 7.20 (br. s., 4H), 5.01 (br. s., 2H), 4.82 - 4.90 (m, 1H), 4.72 (d, J = 6.27 Hz, 2H), 4.44 - 4.51 (m, 1H), 4.30 - 4.37 (m, 1H), 4.16 (br. s., 3H), 2.55 (br. s., 1H), 2.38 - 2.45 (m, 1H), 1.40 - 1.48 (m, 3H)

Claims

What is claimed is:

1. A compound of Formula (I):

wherein:

Ri is Ci_-6alkyl; wherein the Ci_-6alkyl, may be further substituted by phenyl; Ci_-6alkyl-OH, or (CH₂)_m-C₃-₆cycloalkyl;

R₂ is Ci-6 alkyl; wherein the Ci_-6 alkyl, may be further substituted by phenyl;

R₃ is hydrogen, C_1-6alkyl, halo, CN, OC_1-6alkyl, C_1-6alkyl-OH, or phenyl; wherein the phenyl may be further substituted by -OC_1-6alkyl, -C_1-6alkyl or -OH;

R₄ is independently Ci_-6alkyl;

R₅ is independently hydrogen, Ci_-6alkyl, or Ci_-6alkyl-OH;

X is oxygen or NH;

Y is CI or Br;

P is O or S;

Q is NH or S;

m is independently 1 , 2, or 3;

n is independently 0, 1 , 2, or 3; and

2. A compound of claim 1 wherein:

R- is C_1-6 alkyl; wherein the C_1-6 alkyl, may be further substituted by phenyl;

R₃ is hydrogen, Ci_-6 alkyl, halo, CN, OCi_-6 alkyl, or phenyl; wherein the phenyl may be further substituted by OCi_-6 alkyl;

X is oxygen or NH;

Y is CI or Br; and

3. A compound if claim 1 selected from the group consisting of:

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H- benzimidazol-3-ium iodide;

6-bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 /-/- benzimidazol-3-ium iodide;

6-chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H- benzimidazol-3-ium iodide;

6-cyano-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-1 H- benzimidazol-3-ium iodide;

3- ium iodide;

2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-dimethyl-6-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H-benzimidazol-

3- ium trifluoroacetate;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-1 H- benzimidazol-3-ium iodide; 2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1-ethyl-3-(phenylmethyl)-1 H benzimidazol-3-ium bromide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-methyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-propyl-1 /-/- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6-(3-thienyl)-1 ^ benzimidazol-3-ium iodide;

6-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-1 H^ benzimidazol-3-ium iodide;

5-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-1 H- benzimidazol-3-ium iodide;2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-

1- ethyl-3-methyl-5-[4-(methyloxy)phenyl]-1 /-/-benzimidazol-3-ium iodide;

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-5- (methylsulfonyl)-l H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-5-[4 (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide; 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbon

methylphenyl)-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-6- [(ethylamino)carbonyl]-3-methyl-1 H-benzimidazol-3-ium iodide;

6-[(butyloxy)carbonyl]-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1- ethyl-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-5-[4- (methyloxy)phenyl]-1 H-3, 1 -benzimidazol-3-ium iodide;

2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-3-methyl-6-[4- (methyloxy)phenyl]-1 H-benzimidazol-3-ium iodide;

3- Butyl-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 -ethyl-1 H- benzimidazol-3-ium iodide; 2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1-ethyl-3-(2-hydroxyethyl)- 1 H-benzimidazol-3-ium bromide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1-propyl-1 H- benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-5-(4- hydroxyphenyl)-1 H-benzimidazol-3-ium iodide;

1- (Cyclobutylmethyl)-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3- methyl-1 H-benzimidazol-3-ium;

2- ({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 ,3-diethyl-6- [(trifluoromethyl)thio]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-{[2- (methyloxy)-2-oxoethyl]oxy}-1 H-benzimidazol-3-ium iodide; 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbon

oxoethyl]-1 H-benzimidazol-3-ium bromide;

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-[2-(ethyloxy oxoethyl]-1 H-benzimidazol-3-ium iodide;

3- ium-3-yl]acetate;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-[3 (methyloxy)-3-oxopropyl]-1 H-3, 1-benzimidazol-3-ium iodide;

6-Chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-meth 1 H-benzimidazol-3-ium iodide;

7-Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1 -ethyl-3-methyl- 1 H-benzimidazol-3-ium iodide;

5- Chloro-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-1 H- benzimidazol-3-ium iodide;

6- Bromo-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-propyl- 1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]oxy}methyl)-1 ,3-diethyl-6- [(trifluoromethyl)thio]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]^

morpholinyl)-3-oxopropyl]-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-[3-(ethylam oxopropyl]-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-{3-[(2- hydroxyethyl)amino]-3-oxopropyl}-3-methyl-1 H-benzimidazol-3-ium; 6-[(carboxymethyl)oxy]-2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methy ethyl-3-methyl-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6- (propyloxy)-l H-benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-(hydroxymethyl)- 3-methyl-1 H-benzimidazol-3-ium iodide;

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-({[2- (methyloxy)-2-oxoethyl]oxy}methyl)-1 H-benzimidazol-3-ium iodide;

2-({[(3,5-Diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-3-methyl-1-[3-(methyloxy)-

3- oxopropyl]-1 /-/-benzimidazol-3-ium iodide;

1- ethyl-3-methyl-1 H-benzo[d]imidazol-3-ium iodide;

2- ({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-6-({[2-(ethyloxy)-2- oxoethyl]thio}carbonyl)-3-methyl-1 H-3,1 -benzimidazol-3-ium iodide;

2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-({[(3S)- 2-oxotetrahydro-3-furanyl]amino}carbonyl)-1 H-3, 1 -benzimidazol-3-ium iodide; and 2-({[(3,5-diamino-6-chloro-2-pyrazinyl)carbonyl]amino}methyl)-1-ethyl-3-methyl-6-({[(3R)- 2-oxotetrahydro-3-furanyl]amino}carbonyl)-1 H-3, 1 -benzimidazol-3-ium iodide.

3. A pharmaceutical composition comprising a compound of claim 1 and a pharmaceutically acceptable carrier or excipient.

4. A method of treating cystic fibrosis, COPD, (non-cystic fibrosis) and bronchiectasis, which comprises administering to a human in need thereof, a compound of claim 1.

5. A method according to claim 4 wherein the compound is administered orally.

6. A method according to claim 4 wherein the compound is inhaled.

7. A method according to claim 4 wherein the compound of claim 1 is administered in conjunction with one or more other therapeutic agents selected from the group consisting of: advair, tiotropium, tobi, pulmozyme, azithromycin, hypertonic saline, albuterol, pancreatic enzymes, oral antibiotics, and oral steroids.