CN103874701A - Pyridine compounds and uses thereof - Google Patents

Abstract

The present invention relates to pyridine compounds of formula (I). Independent aspects of the present invention relate to pharmaceutical compositions containing said compounds and the use of said compounds as therapeutic agents for the treatment of neurological and psychiatric disorders.

Description

Pyridine compounds and uses thereof

field of invention

The present invention relates to compounds useful as therapeutic agents for the treatment of neurological and psychiatric disorders. Independent aspects of the invention relate to pharmaceutical compositions containing said compounds and their use.

Background technique

Cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) function as intracellular second messengers regulating a range of processes in neurons. Intracellular cAMP and cGMP are produced by adenylate and guanylate cyclases and degraded by cyclic nucleotide phosphodiesterases (PDEs). Intracellular levels of cAMP and cGMP are controlled by intracellular signals, and stimulation/inhibition of adenylate and guanylate cyclases is a well-characterized control of cyclic nucleotide concentrations in response to GPCR activation way (Antoni, Front. Neuroendocrinol. 2000, 21 , 103-132).

Phosphodiesterase 2A (PDE2A) is a dual-substrate enzyme with a higher affinity for cGMP, although it can metabolize either cAMP or cGMP, depending on tissue status. cAMP is derived from adenosine triphosphate (ATP) and is used in many different organisms for intracellular signal transduction, conducting cAMP-dependent pathways. Although PDE2A is expressed in the periphery, PDE2A has the highest expression level in the brain. Recent immunohistochemical studies have shown that the expression pattern of PDE2A in the brain is consistent across mammalian species, including humans (Stephenson et al. J. Histochem. Cytochem. 2009, 57, 933). Expression of the enzyme has been shown to be prominent in regions associated with cognitive function and emotional control, including the cerebral cortex, striatum, hippocampus, amygdala, and habenula pineal.

A selective PDE2A inhibitor, Bay 60-7550, preferentially increases cGMP in primary neuronal cultures and hippocampal slices. Bay 60-7550 also enhances long-term potentiation (LTP) induction in rat hippocampal slices. Consistent with the biochemical and electrophysiological effects of Bay 60-7550, it was also found to be active in novel object and social recognition tasks (Boess et al. Neuropharmacology 2004, 47 , 1081). In recent years, it was reported that Bay 60-7550 was able to reverse the defect in target recognition produced by tryptophan depletion (van Donkelaar et al . Eur. J. Pharmacol. 2008, 600 , 98). These results sparked interest due to the identification of PDE2-positive cells in the dorsal raphe nucleus, a region known to contain cell bodies of serotonergic neurons that project into the forebrain (Stephenson et al . J. Histochem. Cytochem. . 2009, 57 , 933). Similar studies in aged rats showed that the beneficial effects of Bay 60-7550 on target recognition could be reversed by neuronal nitric oxide synthase (nNOS) inhibitors, suggesting that PDE2A in the central nervous system (CNS) The inhibitory effect is due to altered cGMP levels (Domek-Lopacinska and Strosznajder Brain Res. 2008, 1216 , 68).

Recent studies have shown that PDE2A inhibition is also effective in the treatment of anxiety states (Masood et al . J. Pharmacol. Exp. Ther. 2008, 326 , 369; and Masood et al . J. Pharmacol. Exp. Ther. 2009, 331 , 699). In mice, induction of oxidative stress by depletion of central glutathione levels by butionine sulfoximine (BSO) increases a number of anxiety-like behaviors as assessed by open field time and elevated plus maze tests . Treatment with Bay 60-7550 reversed these effects. The PDE2 inhibitor Bay 60- 7550 or ND7001, or NO donor detanonoate to increase cGMP signal can antagonize the anxiety effect of restraint stress on behavior. These drugs also produced anxiolytic effects on the behavior of non-stressed mice in the elevated plus maze and hole board behavioral tests. In contrast, administration of NOS inhibitors, which reduce cGMP signaling, was able to produce similar anxiety effects to restraint stress.

Phosphodiesterase 10A (PDE10A) is another dual specificity enzyme that can convert cAMP to AMP and cGMP to GMP (Soderling et al. Proc. Natl. Acad. Sci. 1999, 96, 7071). PDE10A hydrolyzes cAMP and cGMP, and has a higher affinity for cAMP. PDE10A is expressed in neurons in the striatum, nucleus accumbens, and olfactory tubercle (Seeger et al., Brain Research, 2003, 985, 113-126), as well as in the thalamus, hippocampus, frontal cortex, and olfactory tubercle expressed in (Menniti et al., William Harvey Research Conference, Porto, December 2001). All these brain regions are involved in the pathology of schizophrenia (Lapiz et al., Neurosci Behav Physiol 2003, 33, 13), thus the location of the enzymes may indicate a dominant role in the pathology of psychosis. In the striatum, PDE10A is mainly found in medium spiny neurons and is mainly associated with the postsynaptic membrane of these neurons (Xie et al., Neuroscience 2006, 139, 597). In this position, PDE10A may be important for the signaling cascade induced by dopaminergic and glutamatergic inputs on the two neurotransmitter systems of medium spiny neurons that play a dominant role in the pathology of psychosis. Influence.

It has been shown that cGMP and cAMP levels and their downstream substrates are dysfunctional in psychopaths (MuIy, Psychopharmacol Bull 2002, 36, 92). Additionally, haloperidol treatment was associated with increased levels of cAMP and cGMP in rats and patients, respectively (Leveque et al., J. Neurosci. 2000, 20, 4011). Since PDE10A hydrolyzes cAMP and cGMP, inhibition of PDE10A also induces an increase in cAMP and cGMP, thereby producing a similar effect on cyclic nucleotide levels as haloperidol. The antipsychotic potential of PDE 10A inhibitors is further supported by studies by Kostowski et al. (Pharmacol Biochem Behav 1976, 5, 15), which showed that papaverine, a moderately selective PDE 10A inhibitor, In the model, it can reduce the stereotypy caused by apomorphine, increase the catalepsy caused by haloperidol in rats, and reduce the concentration of dopamine in the brain of rats. Typical antipsychotics also have this activity. In addition to typical antipsychotics that mainly improve the positive symptoms of psychosis, PDE10A also has the potential to improve the negative and cognitive symptoms of psychosis.

Focusing on dopaminergic input on medium spiny neurons, PDE10A inhibitors act as D1 agonists and D2 antagonists by upregulating cAMP and cGMP levels, because activation of Gs-proteins linked to dopamine D1 receptors can Increases intracellular cAMP, while activation of Gi-proteins linked to dopamine D2 receptors can reduce intracellular cAMP levels by inhibiting adenylyl cyclase activity. Elevation of intracellular cAMP levels mediated by D1 receptor signaling appears to regulate a range of neuronal processes responsible for working memory in the prefrontal cortex (Sawaguchi, Parkinsonism Relat. Disord. 2000, 7, 9), and D1 has been reported to be regulated by Physical activation can improve working memory deficits in patients with schizophrenia (Castner et al., Science 2000, 287, 2020).

Rodefer et al. (Eur. J Neurosci 2005, 21, 1070) gave additional evidence of the effect of PDE10A inhibition on the negative symptoms of psychosis, which could show that papaverine was able to reverse the subchronic administration of phenixidine in rats ( It is an attentional setting-switching deficit caused by NMDA antagonists). Attention deficits, including impaired switching of attention to novel stimuli, are among the negative symptoms of schizophrenia. In this study, administration of benzikidine for 7 days, followed by a washout time, was able to induce attention deficits. The PDE10A inhibitor papaverine reverses persistent deficits induced by subchronic treatment.

These pooled results suggest that inhibition of PDE2A and/or PDE10A may be a therapeutic target for the treatment of certain neurological and psychiatric disorders. Accordingly, the present invention relates to pyridines containing triazolopyrazines, processes for their preparation, their medical use and medicaments containing them.

SUMMARY OF THE INVENTION

The object of the present invention is to provide compounds capable of inhibiting PDE2A and/or PDE10A. Accordingly, the present invention relates to compounds of formula I:

wherein X ¹ , X ² , X ³ and X ⁴ are each independently N or CR ³ with the proviso that one X is N and the remaining X are each independently CR ³ ;

Wherein R ¹ is C ₁ -C ₆ alkyl, C ₃ -C ₆ cycloalkyl, tetrahydropyranyl, benzyl, phenyl and pyridyl, wherein benzyl, phenyl and pyridyl are optionally replaced by one or Multiple halogen, CN, C ₁ -C ₄ alkyl/fluoroalkyl or C ₁ -C ₄ alkoxy/fluoroalkoxy substitutions;

wherein R ² is C ₁ -C ₄ alkyl or C ₃ -C ₆ cycloalkyl; and

Wherein R ³ is hydrogen, halogen, CN, -CO ₂ H, -CON(H or C ₁ -C ₄ alkyl) ₂ , CHO, C ₁ -C ₄ alkyl/fluoroalkyl, heterocycle containing cyclic amino , C ₂ -C ₄ alkenyl, C ₂ -C ₄ alkenyl or C ₁ -C ₄ alkoxy/fluoroalkoxy; or a pharmaceutically acceptable salt thereof.

In an independent aspect of the invention, the compound is selected from one of the illustrated compounds of formula I.

The present invention provides a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula I and a pharmaceutically acceptable carrier.

The present invention also provides a method of treating an individual suffering from anxiety, cognitive impairment or schizophrenia, the method comprising: administering a therapeutically effective amount of a compound of formula I.

The present invention further provides the use of the compound of formula I in the preparation of a medicament for treating anxiety, cognitive impairment or schizophrenia.

Another aspect of the invention provides compounds for use in the treatment of anxiety, cognitive impairment or schizophrenia.

Detailed Description of the Invention

The present invention is based on the discovery that compounds of formula I are capable of inhibiting PDE2A and/or PDE10A and are therefore useful in the treatment of certain neurological and psychiatric disorders. Specific aspects of the invention are explained in more detail below, but this description is not to be considered an exhaustive catalog of all the various ways in which the invention may be implemented, or of all features that may be incorporated into the invention. Therefore, the following descriptions only illustrate some embodiments of the present invention, rather than exhaustively specifying all permutations, combinations and changes thereof.

Those practicing the art will appreciate that the compounds may exist in tautomeric forms. When the application refers to a specific tautomer, it should be understood that its tautomeric form and its mixture are included.

The present invention relates to compounds of formula I as defined in the Summary of the Invention, their pharmaceutical compositions and uses.

In one embodiment, R ² is C ₁ -C ₄ alkyl. In one embodiment, R ² is methyl.

In one embodiment, R ² is C ₃ -C ₆ cycloalkyl.

In one embodiment, R ¹ is C ₁ -C ₄ alkyl.

In one embodiment, R ¹ is C ₃ -C ₆ cycloalkyl.

In one embodiment, R ¹ is tetrahydropyranyl.

In one embodiment, R ¹ is benzyl optionally substituted with one or two F, Cl or C ₁ -C ₃ alkyl.

In one embodiment, R ¹ is phenyl optionally substituted with one or two F, Cl or C ₁ -C ₃ alkyl.

In one embodiment, R ¹ is pyridyl optionally substituted with one or two F, Cl or C ₁ -C ₃ alkyl.

In one embodiment, R ¹ is phenyl substituted with one or two F, Cl or C ₁ -C ₃ alkyl.

In one embodiment, ^Xi is N. In one embodiment, ^X2 is N. In one embodiment, ^X3 is N. In one embodiment, ^X4 is N.

In one embodiment, ^R3 is hydrogen. In one embodiment, ^R3 is halo or CHO.

In one embodiment, R ³ is C ₁ -C ₄ alkyl or C ₁ -C ₄ alkoxy.

In one embodiment, R ³ is C ₂ -C ₄ alkenyl or C ₂ -C ₄ alkenyl.

In one embodiment, ^R3 is a heterocyclic ring containing a cyclic amino group.

In one embodiment, R ³ is CN, —CO ₂ H, or —CON(H or C ₁ -C ₄ alkyl) ₂ .

The racemic forms can be resolved into the optical antipodes by known methods, for example, separation of the diastereomeric salts with an optically active acid and treatment with a base to liberate the optically active amine compound. Such diastereomeric salts can be isolated, for example, by fractional crystallization. Optically active acids suitable for this purpose may include, but are not limited to, d- or l-tartaric acid, mandelic acid, or camphorsulfonic acid. Another method for the resolution of racemates into optical antipodes is a chromatography-based method (on an optically active matrix). Compounds of the invention can also be resolved by forming diastereomeric derivatives from chiral derivatizing reagents (e.g., chiral alkylating or acylating reagents) and chromatographically separating them followed by cleavage of the chiral auxiliary . Any of the above methods can be used to resolve the optical antipodes of the compound of the present invention itself, or to resolve the optical antiantiomers of the synthetic intermediates, which can then be converted into optically resolved final products using the methods described herein, such The final product is a compound of the invention.

Other methods of resolution of optical isomers known to those skilled in the art may be used. Such methods include those discussed in J. Jaques, A. Collet and S. Wilen, Enantiomers, Racemates, and Resolutions, John Wiley and Sons, New York, 1981. Optically active compounds can also be prepared from optically active starting materials.

definition

As used herein, the term "C ₁ -C ₆ alkyl" refers to a straight or branched chain saturated hydrocarbon having one to six carbon atoms, inclusive. Examples include, but are not limited to: methyl, ethyl, 1-propyl, 2-propyl, 1-butyl, 2-butyl, 2-methyl-2-propyl, 2-methyl-1- Propyl, n-pentyl and n-hexyl. Similarly, the term "straight or branched C ₁ -C ₄ alkyl" refers to a saturated hydrocarbon having one to four carbon atoms. Examples include methyl, ethyl and n-propyl.

Likewise, the term "C ₁ -C ₄ alkoxy" refers to a straight or branched chain saturated oxygen-containing hydrocarbon group having one to four carbon atoms, with an open valence on the oxygen. Examples include, but are not limited to, methoxy, ethoxy, n-butoxy, and tert-butoxy.

The term "C ₁ -C ₆ fluoroalkyl" refers to a linear or branched saturated hydrocarbon having one to six carbon atoms, inclusive, substituted with one or more fluorine atoms. Examples include trifluoromethyl, pentafluoroethyl, 1-fluoroethyl, monofluoromethyl, difluoromethyl, 1,2-difluoroethyl and 3,4-difluorohexyl. Similarly, the term "straight chain or branched _C1 - _C4 fluoroalkoxy" refers to a saturated hydrocarbon having one to four carbon atoms, substituted by one or more fluorine atoms, with an open valence on the oxygen.

The term "halogen" refers to fluorine, chlorine, bromine and iodine.

The term "C ₂ -C ₄ -alkenyl" refers to a branched or straight chain alkenyl group having two to four carbon atoms and a double bond, including ethenyl, propenyl and butenyl. The term " _C2 - _C4 -alkynyl" refers to branched or straight chain alkynyl groups having two to four carbon atoms and a triple bond, including ethynyl, propynyl and butynyl.

For the purposes of the present invention, the term "heterocycle containing a cyclic amino group" refers to azetidine, pyrrolidine, piperidine, piperazine and morpholine. The "heterocyclic ring containing cyclic amino group" may be optionally substituted by one or more linear or branched C ₁ -C ₄ alkyl groups.

The term "CON(H or C ₁ -C ₄ alkyl) ₂ " refers to an amido group in which the substituents of the amido moiety are each independently selected from H or C ₁ -C ₄ alkyl. Examples include _-CONH2 , _-CONHCH3 , -CON( _CH3 ) ₂ and -CON( _CH3 ) _{CH2CH3 .}

As used herein, the term "treating" means ameliorating or reversing the course or severity of a disease or disorder, or ameliorating or reversing one or more symptoms or side effects of such a disease or disorder. As used herein, "treating" also refers to inhibiting or blocking, such as delaying, retarding, limiting, hindering or impeding the progression of a system, symptom or state of a disease or disorder. For the purposes of the present invention, "treatment" further refers to a method of obtaining a beneficial or objective clinical outcome, wherein "beneficial or objective clinical outcome" includes, but is not limited to: alleviating symptoms, reducing the extent of a disorder or disease, stabilizing a disease or disorder state ( That is, without worsening), delaying or slowing a disease or disorder state, ameliorating or alleviating a disease or disorder state, and alleviating a disease or disorder, whether partial or total, detectable or undetectable.

As used herein, the phrase "effective amount", when applied to a compound of the present invention, means an amount sufficient to produce the desired biological effect.

The phrase "therapeutically effective amount", when applied to a compound of the present invention, means an amount of the compound sufficient to ameliorate, alleviate, stabilize, reverse, slow or delay the progression of a disorder or disease state or the progression of symptoms of a disorder or disease. In one embodiment, the methods of the invention provide for administering a combination of compounds. In this context, an "effective amount" is an amount of the composition sufficient to produce the desired biological effect.

medicinal salt

The present invention also includes salts of the compounds of the present invention, generally referring to pharmaceutically acceptable salts. Such salts include pharmaceutically acceptable acid addition salts. Acid addition salts include salts of inorganic acids as well as organic acids.

Representative examples of suitable inorganic acids include hydrochloric, hydrobromic, hydroiodic, phosphoric, sulfuric, sulfamic, nitric, and the like. Representative examples of suitable organic acids include: formic acid, acetic acid, trichloroacetic acid, trifluoroacetic acid, propionic acid, benzoic acid, cinnamic acid, citric acid, fumaric acid, glycolic acid, itaconic acid, lactic acid, Methanesulfonic acid, maleic acid, malic acid, malonic acid, mandelic acid, oxalic acid, picric acid, pyrogluconic acid, salicylic acid, succinic acid, methanesulfonic acid, ethanesulfonic acid, tartaric acid, ascorbic acid, pamoic acid, Bismethylene salicylic acid, ethanedisulfonic acid, gluconic acid, citraconic acid, aspartic acid, stearic acid, palmitic acid, EDTA, glycolic acid, p-aminobenzoic acid, glutamic acid, benzene Sulfonic acid, p-toluenesulfonic acid, theophylline acetic acid, and 8-halophylline (for example, 8-bromophylline, etc.). Other examples of pharmaceutically acceptable inorganic or organic acid addition salts include the pharmaceutically acceptable salts listed in S. M. Berge et al. J. Pharm. Sci., 1977, 66, 2.

Furthermore, the compounds of the present invention may exist in unsolvated forms as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like.

pharmaceutical composition

The present invention further provides a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula I and a pharmaceutically acceptable carrier. The present invention also provides a pharmaceutical composition comprising a therapeutically effective amount of a specific compound disclosed in the Experimental Section and a pharmaceutically acceptable carrier.

The compounds of the present invention may be administered alone, or together with a pharmaceutically acceptable carrier or excipient in single or multiple doses. The pharmaceutical compositions according to the present invention may be formulated according to conventional techniques together with pharmaceutically acceptable carriers or diluents and any other known adjuvants and excipients, for example, those methods disclosed in: Remington: The Science and Practice of Pharmacy, ^19th Edition, Gennaro, Ed., Mack Publishing Co., Easton, PA, 1995.

Pharmaceutical compositions for oral administration include solid dosage forms such as capsules, tablets, troches, pills, lozenges, powders and granules. The compositions may, if appropriate, be prepared with coatings such as enteric coatings, or they may be formulated so as to provide controlled, eg sustained or prolonged release of the active ingredient, according to methods well known in the art. Liquid dosage forms for oral administration include solutions, emulsions, suspensions, syrups and elixirs.

Pharmaceutical compositions for parenteral administration include sterile aqueous and non-aqueous injection solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions before use.

Oral doses are usually administered in one or more doses, generally one to three doses per day. The exact dosage will depend on the frequency and mode of administration, the sex, age, weight and general condition of the individual being treated, the nature and severity of the condition being treated and any concomitant diseases being treated, and other factors as will be apparent to those skilled in the art. The formulations can also be presented in unit dosage form by methods known to those skilled in the art. For example, unit dosage forms for oral administration may contain from about 0.01 to about 1000 mg, from about 0.05 to about 500 mg, or from about 0.5 to about 200 mg.

The compounds of the invention are generally employed as free material or as a pharmaceutically acceptable salt thereof. An example is the acid addition salts of compounds with free base availability. When a compound of formula I contains a free base, such salts are prepared in conventional manner by treating a solution or suspension of the free base of formula I with a molar equivalent of a pharmaceutically acceptable acid. Representative examples of suitable organic and inorganic acids are described above.

For parenteral administration, solutions of a compound of formula I in sterile aqueous solution, aqueous propylene glycol, aqueous vitamin E or sesame or peanut oil may be used. Such aqueous solutions should be suitably buffered, if necessary, first with sufficient saline or glucose to render the liquid diluent isotonic. Aqueous solutions are especially suitable for intravenous, intramuscular, subcutaneous and intraperitoneal administration. Compounds of formula I can be readily incorporated into known sterile aqueous media using standard techniques known to those skilled in the art.

Suitable pharmaceutical carriers include: inert solid diluents or fillers, sterile aqueous solutions and various organic solvents. Examples of solid carriers include lactose, terra alba, sucrose, cyclodextrin, talc, gelatin, agar, pectin, acacia, magnesium stearate, stearic acid and lower alkyl ethers of cellulose. Examples of liquid carriers include, but are not limited to, syrup, peanut oil, olive oil, phospholipids, fatty acids, fatty acid amines, polyoxyethylene and water. Similarly, the carrier or diluent may include any sustained release material known in the art, such as glyceryl monostearate or glyceryl distearate, alone or mixed with a wax. Then, the pharmaceutical composition formed by combining the compound of formula I and a pharmaceutically acceptable carrier can be easily administered in various dosage forms suitable for the disclosed administration routes. The formulations may conveniently be presented in unit dosage form by methods known in the art of pharmacy.

If a solid carrier is used for oral administration, the preparation may be tabletted, placed in a hard gelatin capsule in powder or granule form, or may be in the form of a troche or lozenge. The amount of solid carrier may vary widely but will range from about 25 mg to about 1 g per dosage unit. If a liquid carrier is used, the preparation can be in the form of a syrup, emulsion, soft gelatin capsule, or sterile injectable solution, such as an aqueous or non-aqueous liquid suspension or solution.

therapeutic use

The present invention provides a method of treating an individual suffering from anxiety, cognitive impairment or schizophrenia, the method comprising: administering a therapeutically effective amount of a compound of formula I.

The present invention further provides the use of the compound of formula I in the preparation of a medicament for treating anxiety, cognitive impairment or schizophrenia. Another aspect of the invention provides compounds for use in the treatment of anxiety, cognitive impairment or schizophrenia. The present invention provides a method for treating anxiety, cognitive impairment or schizophrenia, the method comprising: administering a therapeutically effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof.

The invention provides a method for treating anxiety disorders selected from the group consisting of: anxiety, panic disorder, agoraphobia, specific phobia, social phobia, obsessive-compulsive disorder, post-traumatic stress disorder, acute stress disorder disorder and generalized anxiety disorder.

The invention further provides a method of treating an individual suffering from cognitive impairment, the method comprising: administering to the individual a therapeutically effective amount of a compound of formula I. Examples of cognitive disorders that may be treated in accordance with the present invention include, but are not limited to: Alzheimer's disease, multi-infarct dementia, alcoholic dementia or other drug-related dementia, dementia associated with intracranial tumors or brain trauma , dementia associated with Huntington's disease or Parkinson's disease, or AIDS-related dementia and age-related cognitive decline.

The present invention also provides a method of treating movement disorders, the method comprising: administering to a subject a therapeutically effective amount of a compound of formula I. Examples of movement disorders that may be treated in accordance with the present invention include, but are not limited to, Huntington's disease and movement disorders associated with dopamine agonist treatment. The present invention further provides a method of treating a movement disorder selected from Parkinson's disease and restless legs syndrome, the method comprising: administering to a subject a therapeutically effective amount of a compound of formula I.

The present invention provides methods for the treatment of schizophrenia, e.g., paranoid, disorganized, catatonic, undifferentiated, or other types of schizophrenia; schizophrenia-like disorders; schizoaffective disorders, For example, delusional or depressive type; delusional disorder; substance-induced psychosis, eg, psychosis induced by alcohol, amphetamines, marijuana, cocaine, hallucinogens, inhalants, opioids, or phenixidine; paranoid type and personality disorder of the schizophrenic type; where the drug addiction is alcohol, amphetamine, cocaine, or opiate addiction.

Experimental part

Compounds of formula I can be prepared using the methods outlined below and in the examples. In the following methods, variants or modifications, which are known per se to a chemist in the art, or are apparent to a person of ordinary skill in the art, may be used. Additionally, other methods for preparing compounds of the present invention will be apparent to those skilled in the art with reference to the following schemes and examples. For example, these methods describe the use of selective protecting groups during the synthesis of compounds of the invention. For a particular reaction, those skilled in the art will be able to select an appropriate protecting group. Methods for the protection and deprotection of such groups are well known in the art and can be found in: T. Green et al. Protective Groups in Organic Synthesis, 1991, ^2nd Edition, John Wiley & Sons, New York.

Abbreviations and chemical reagents used

AcOH = acetic acid (eg, Sigma-Aldrich 242853). Acetonitrile (eg, Aldrich 271004). APPI = Atmospheric Pressure Photoionization. Aq = aqueous solution. Saline = saturated aqueous sodium chloride (eg, Aldrich S7653). Boc ₂ O=di-tert-butyl dicarbonate (eg, Aldrich 361941 ). 2-Chlorobenzoic acid (eg, Aldrich 135577). 3-Chlorobenzoic acid (eg, Fluka 23530). 3-Chlorobenzoyl chloride (eg, Aldrich C26801). Chloroform (eg, Sigma-Aldrich C2432). 2-Chloro-6-methyl-benzoic acid (eg Lancaster X18348 or Matrix 002794). 2-Chloro-6-methyl-benzoyl chloride (eg, Fluorochem 38160 or Betapharm 15-47106). DBU = 1,8-diazabicyclo[5.4.0]undec-7-ene (eg, Aldrich 139009). DCM=methylene chloride/dichloromethane (eg, Aldrich 270997). 2,6-Dichloro-benzoic acid (eg, Aldrich D57450). 2,3-Diaminopyridine (eg, Aldrich 125857). 3,4-Diaminopyridine (eg, Aldrich D7148). Diethyl ether (eg, Sigma-Aldrich 346136). DMAP = 4-(dimethylamino)pyridine (eg, Aldrich 522805). 2,6-Dimethylbenzoyl chloride (eg, Fluorochem 017526 or ABCR AB173115). DMF = Dimethylformamide (eg, Sigma-Aldrich 227056). DIPEA = Diisopropylethylamine (eg, Aldrich 387649). ELS = evaporative light scattering. Ethanol (eg, Sigma-Aldrich 459844). Ethyl pyruvate (eg, Fluka 15960). EtOAc = ethyl acetate (eg, Fluka 34972). 2-Fluoro-3-nitro-pyridine (eg, Fluorochem 03250 or Matrix 018339). h = hours. 4M HCl/1,4-dioxane (eg, Sigma-Aldrich 345547). Heptane (eg, Sigma-Aldrich 730491). HPLC = high performance liquid chromatography. 30% hydrogen peroxide in water (for example, Sigma-Aldrich 216763). Hydrazine hydrate (eg, Sigma-Aldrich 225819). Iron powder (eg, Aldrich 12310). LC = liquid chromatography. LC/MS = liquid chromatography/mass spectrometry. 4M = 4 molar solution. Methanol (eg, Sigma-Aldrich 34860). MTBE = methyl tert-butyl ether (eg, Sigma-Aldrich 306975). MW = microwave. MW conditions = reactions in sealed tubes using Biotage Initiator instrument or CEM Explorer-48 instrument. _Na2CO3 (eg, Sigma _- Aldrich S7795). _NaHCO3 (eg, Sigma-Aldrich S6014). NaOH (eg, Sigma-Aldrich S5881). _{Na2SO4 (} eg, Sigma-Aldrich 238597). 3-Nitro-pyridin-4-ylamine (eg, Aldrich 646962). 10% palladium on charcoal (eg, Aldrich 75990). PDA = photodiode array. Pd(DPPF)Cl ₂ =[1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (eg, Aldrich 697230), Pd(DPPF)Cl ₂ -DCM complex = [ 1,1'-Bis(diphenylphosphino)ferrocene]dichloropalladium(II) in complex with dichloromethane (for example, Aldrich 379670). Pentane (eg, Sigma-Aldrich 236705). _PhPOCl2 = phenylphosphine dichloride (eg, Aldrich 389560). POCl ₃ = phosphorus oxychloride (eg, Aldrich 262099). PyBroP = benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate (eg, Fluka 12809). Racemic alanine (eg, Sigma A7502). RT = retention time. Sat = saturated. T = time. _Tf2O =trifluoromethanesulfonic anhydride (eg Aldrich 176176). TLC = thin layer chromatography. Et ₃ N=triethylamine (eg, Sigma-Aldrich T0886).

LC/MS Method 131: LC/MS was performed on a Sciex API 150EX equipped with an APPI source (operated in positive ion mode). The HPLC consisted of a Shimadzu LC10-ADvp LC pump, a SPD-M20A PDA detector (operating at 254 nM), and a SCL-10A system controller. The autosampler was a Gilson 215, the column oven was a Jones Chromatography 7990R, and the ELS detector was a Sedere Sedex 85. LC conditions: The column is Waters Symmetry C-18, 4.6 x 30 mm, 3.5 microns, run at 60°C, binary gradient consisting of water+0.05% TFA(A) and methanol+0.05% TFA, 3.0 mL/min. Gradient: 0.01 min, 17% B; 0.27 min, 28% B; 0.53 min, 39% B; 0.80 min, 50% B; 1.07 min, 59% B; 1.34 min, 68% B; 1.60 min, 78% B ; 1.87 min, 86% B; 2.14 min, 93% B; 2.38 min, 100% B; 2.40 min, 17% B; 2.80 min, 7% B; total running time: 2.8 min.

LC/MS Method 132: The hardware is the same as LC/MS Method 131. LC conditions: Column was Waters Symmetry C-18, 4.6 x 30 mm, 3.5 microns, run at 60°C, binary gradient consisting of water + 0.05% TFA(A) and methanol + 0.05% TFA, 2.5 mL/min. Gradient: 0.01 min, 5% B; 2.38 min, 100% B; 2.40 min, 5% B; 2.80 min, 5% B. Total running time: 2.8 min.

Method 550: LC-MS on a Waters Aquity UPLC-MS composed of Waters Aquity including column manager, binary solvent manager, sample organizer, PDA detector (operating at 254 nM), ELS detection detector and a TQ-MS equipped with an APPI source (running in positive ion mode). LC conditions: column is Acquity UPLC BEH C18 1.7μm, 2.1x50mm, run at 60°C, binary gradient consists of water + 0.05% trifluoroacetic acid (A) and acetonitrile + 5% water + 0.05% trifluoroacetic acid, 1.2 mL/min. Gradient: 0.00 min, 10% B; 1.00 min, 100% B; 1.01 min, 10% B; 1.15 min, 10% B. Total run time: 1.15 min.

general method

Briefly, compound I of the present invention can be prepared from hydrazine II under the conditions described in method 1 or method 2, respectively. Hydrazine II can be prepared from compound III under the conditions described in Method 3. In some cases, it is also possible to convert III directly to I using Method 4. Precursor III can be obtained from lactam IV using method 5 or method 6.

Method 1 involves treatment of hydrazine II with the appropriate acid chloride _R1COCl in a suitable solvent such as acetonitrile at elevated temperature. Sometimes it may be advantageous to add POCl ₃ or PhPOCl ₂ . Acid chlorides can be prepared in situ from the corresponding acids by adding _POCl3 . Alternatively, _reaction of hydrazine II with _R1CO2H and PyBroP affords the corresponding hydrazide, followed by two-step treatment of the hydrazide with _PhPOCl2 and base to give compounds of the invention as described in Example Ia1.

Method 2 is an alternative to method 1 in which hydrazine II is condensed with a suitable aldehyde _R1CHO in a suitable solvent such as dichloromethane (DCM) to form the corresponding hydrazone. Subsequently, a suitable oxidizing agent is added to obtain the compounds of the invention, for example, under the conditions reported by Sadana et al. (AK Sadana, Y. Mirza, KR Aneja, O. Prakash European Journal of Medicinal Chemistry 2003, 38, 533), wherein The oxidizing agent is iodobenzene diacetate (PhI(OAc) ₂ ). Alternatively, the method reported by Mogilaiah et al. (K. Mogilaiah, T. Kumara Swamy, K. Shiva Kumar J. Heterocyclic Chem. 2009, 46, 124) can be used, in which by treatment with chloramine-T, Oxidative cyclization of the intermediate hydrazone.

Method 3 involves: replacing X in compound III with hydrazine, where X is a chlorine atom or other leaving group, eg, phosphonium as depicted in the reaction scheme. The reaction is generally carried out with hydrazine hydrate in a suitable solvent such as ethanol at elevated temperature. Preparation IIc and IId are examples of Method 3.

Method 4 is: in a suitable solvent (such as acetonitrile) at high temperature, compound III is reacted with a suitable acylhydrazine _R1CONHNH2 to directly convert to compound I of the present invention, as described in Example Id1.

Method 5 is conversion of lactam IV to compound III, where X is a chlorine atom, by heating the substrate in excess phosphorus oxychloride as described for IId; sometimes, addition of a suitable base (e.g. triethylamine or diiso Propylethylamine (DIPEA)) is advantageous.

Method 6 is: treatment with benzotriazol-1-yl-oxytripyrrolidino-phosphonium hexafluorophosphate (PyBroP) or similar peptide coupler in the presence of a suitable base such as DIPEA Lactam IV, provides compound III, wherein X is phosphonium, as depicted in the reaction scheme. For other lactams, this method is known in the literature. (TD Ashton, PJ Scammells Australian Journal of Chemistry 2008, 61 , 49). Preparation Ha is an example of Method 6.

Preparation of intermediates

(3-Methyl-pyrido[2,3-b]pyrazin-2-yl)-hydrazine (IIa)

A mixture of 2-fluoro-3-nitro-pyridine (12 g), _Et3N (30 mL) and racemic alanine (11.87 g) was refluxed in methanol (200 mL) overnight. The mixture was cooled to ambient temperature, and the filtrate was concentrated in vacuo. The residue was partitioned between DCM and water. The organic layer was dried over _Na2CO3 , filtered and concentrated _in vacuo to give 2-(3-nitro-pyridin-2-ylamino)-propionic acid (7.8 g). This material was dissolved in AcOH (50 mL) and iron (8.2 g) was added. The mixture was refluxed for 1.5 hours. After cooling to ambient temperature, the mixture was filtered, and the filtrate was concentrated in vacuo. The residue was washed with water and dried to give 3-methyl-3,4-dihydro-1H-pyrido[2,3-b]pyrazin-2-one (1.4 g). This material was mixed with 5% aqueous NaOH (92 mL) and water (18 mL), followed by the addition of 30% aqueous hydrogen peroxide (9.2 mL). The mixture was stirred at 60°C for 10 hours. After cooling to ambient temperature, the pH was adjusted to neutral and 3-methyl-1H-pyrido[2,3-b]pyrazin-2-one (1.2 g) was precipitated. This material was dissolved in DMF (10 mL) and treated with PyBroP (4.6 g) and DIPEA (1.6 mL) at ambient temperature for 16 hours. The precipitated white solid was filtered off, washed with ethanol and dried to give 2-(benzotriazol-1-yloxy)-3-methyl-pyrido[2,3-b]pyrazine (0.4 g). This material (0.4 g) and hydrazine hydrate (0.5 mL) were refluxed in ethanol (5 mL) for 10 minutes. After cooling to ambient temperature, the precipitated white solid was filtered off, washed with ethanol and dried to afford Ila (0.2 g) sufficiently pure for use in the next step.

(3-Methyl-pyrido[2,3-b]pyrazin-2-yl)-hydrazine (IIc)

A mixture of 3,4-diaminopyridine (10 g) and ethyl pyruvate (53 g) in chloroform (100 mL) was stirred overnight at ambient temperature. The precipitated solid was filtered off, washed with DCM and dried to give 2-methyl-4H-pyrido[3,4-b]pyrazin-3-one (14 g) as a pale yellow solid. 2 g of this material were suspended in DMF (10 mL) and treated with PyBroP (6 g) and DIPEA (3.3 mL) overnight at ambient temperature. The precipitated solid was filtered off, washed with ethanol, and dried to give 3-(benzotriazol-1-yloxy)-2-methyl-pyrido[3,4-b]pyrazine (1.7 g) as a white solid . This material was suspended in ethanol (50 mL), hydrazine hydrate was added, and the resulting mixture was heated at 85 °C for 15 min. The solid was filtered off, washed with ethanol and dried to afford Ilc (0.95 g) as a yellow solid sufficiently pure to be used in the next step.

(3-Methyl-pyrido[3,4-b]pyrazin-2-yl)-hydrazine (IIb)

To a suspension of 3-nitro-pyridin-4-ylamine (15 g) in DCM (250 mL) was added triethylamine (30 mL), Boc ₂ O (23.5 g) and DMAP (1.31 g). The mixture was stirred at ambient temperature for 2 days. The solid was filtered off, and the filtrate was concentrated in vacuo. The residue was washed with MTBE. Yellow crystals were collected to give (3-nitro-pyridin-4-yl)-carbamic acid tert-butyl ester (17 g). This material and ethyl pyruvate (100 mL) were dissolved in ethanol (150 mL) and treated with hydrogen (50 psi) at 50 °C for two days in the presence of 10% palladium on carbon (3 g). The catalyst was filtered off, and the filtrate was concentrated in vacuo. The residue was purified by silica gel chromatography (eluent: pentane/EtOAc, 10:1 → 2:1) to give 2-(4-tert-butoxycarbonylamino-pyridin-3-ylamino)-propionic acid ethyl ester ( 5 g) Yellow solid. A larger sample (9 g) of this compound prepared in a similar manner was stirred in 4M HCl/1,4-dioxane (40 mL) at ambient temperature for 5 hours. The precipitated solid was filtered off, washed with MTBE, and dried. The filtrate was concentrated to dryness, washed with MTBE and dried to give a second crop of product. In total 3-methyl-3,4-dihydro-1H-pyrido[3,4-b]pyrazin-2-one hydrochloride (5 g) was obtained as a white solid. 2.2 g of this material were suspended in water (20 mL) and treated with 30% aqueous hydrogen peroxide (1.2 mL) and NaOH (adjust pH to 7-8). The mixture was stirred at 75°C for 2 days. More 30% aqueous hydrogen peroxide (0.15 mL) was added and stirring was continued for another 2 days. Volatiles were removed in vacuo. The residue was washed with EtOAc to give 3-methyl-1H-pyrido[3,4-b]pyrazin-2-one (0.8 g) as a pale yellow solid. 0.7 g of this material and PyBroP (2.11 g) were suspended in DMF (3 mL). DIPEA (1.16 mL) was added and the mixture was stirred at ambient temperature overnight. The precipitated solid was filtered off, washed with ethanol, and dried to give 2-(benzotriazol-1-yloxy)-3-methyl-pyrido[3,4-b]pyrazine (0.7 g) as a white solid . 430 mg of this material was suspended in ethanol (13 mL) and treated with hydrazine hydrate (0.5 mL) at 85°C for 20 minutes. The solid was filtered off, washed with ethanol, and dried to afford IId (180 mg) as a pale yellow solid sufficiently pure to be used in the next step.

3-Chloro-2-methyl-pyrido[2,3-b]pyrazine (IIId)

Ethyl pyruvate (1.22 mL) was dissolved in methanol (10 mL) and added to a cold solution of 2,3-diaminopyridine (1.09 g)/methanol MeOH (20 mL), and the mixture was heated at ambient temperature Stir down. The solid was filtered off and washed with cold methanol to give 2-methyl-4H-pyrido[2,3-b]pyrazin-3-one (1.1 g) as a gray solid. 1.0 g of this material was suspended in acetonitrile (10 mL), and phosphorus oxychloride (1.16 mL) was added. The mixture was heated at 120 °C for 0.5 h under MW conditions. The crude mixture was partitioned between water and EtOAc. The organic layer was washed with brine, dried over _Na2SO4 , filtered and concentrated _in vacuo to afford IIId (460 mg) as a red solid sufficiently pure for the next step.

(2-Methyl-pyrido[2,3-b]pyrazin-3-yl)-hydrazine (IId)

A mixture of IIId (0.61 g) and hydrazine hydrate (0.5 mL) in ethanol (5 mL) was stirred at room temperature for 10 min. The precipitated white solid was filtered off, washed with ethanol, and dried to afford IId (0.43 g) pure enough to be used in the next step.

(8-Methoxy-2-methyl-pyrido[2,3-b]pyrazin-3-yl)-hydrazine (IIe)

To 4-chloro-pyridin-2-ylamine (5 g) in 96% _H2SO4 (20 mL) in water was added dropwise 70% _HNO3 in _water (2.5 mL) and 96% _H2SO at 0 °C A mixed solution of ₄ aqueous solutions (10 mL). After the addition was complete, the reaction mixture was stirred at room temperature for 2 hours. The solution was poured into ice/water and the pH was adjusted to 9 by adding 6M aqueous NaOH dropwise. The solid was then filtered off, washed with water, dried and purified by alumina chromatography (eluent: pentane:EtOAc, 2:1) to give 4-chloro-3-nitro-pyridin-2-ylamine (1.2 g ). 200 mg of this material was dissolved in methanol (3 mL), NaOMe (125 mg) was added, and the mixture was stirred at 60° C. for 16 hr. 37% aqueous HCl was added dropwise to adjust the pH to 6. The mixture was cooled to 0 °C and the precipitated solid was filtered off, washed with water and dried to give 4-methoxy-3-nitro-pyridin-2-ylamine (180 mg). This material was dissolved in methanol (20 mL) and treated with hydrogen (1 bar) in the presence of 10% palladium on charcoal at room temperature for 2 hours. The mixture was filtered and the filtrate was concentrated in vacuo to give 4-methoxy-pyridine-2,3-diamine (50 mg). Repeat the process for more material. In the next step, 4-methoxy-pyridine-2,3-diamine (6.8 g) was dissolved in ethanol (200 mL). To this solution was added methyl pyruvate (4.9 mL) slowly at room temperature, and the mixture was stirred at ambient temperature for 3 hours, after which time the volatiles were removed in vacuo. The residue was purified by silica gel chromatography (eluent: pentane:EtOAc, 5:1 to 0:1) to afford 8-methoxy-2-methyl-4H-pyrido[2,3-b]pyrazine -3-one (6.1 g). To a suspension of 3 g of this material in DMF (12 mL) was added PyBroP (7.65 g) followed by DBU (3.58 g). The mixture was stirred overnight, then the precipitated solid was filtered off, washed with ethanol and dried to give 3-(benzotriazol-1-yloxy)-8-methoxy-2-methyl-pyrido[2 ,3-b]pyrazine (3.5 g). 1.5 g of this material were dissolved in a mixture of ethanol (10 ml) and DCM (60 ml). Hydrazine monohydrate (3.6 g) was added and the mixture was stirred at ambient temperature overnight before the volatiles were removed in vacuo. The residual solid was triturated with ethanol, filtered off, washed with ethanol and dried to give (8-methoxy-2-methyl-pyrido[2,3-b]pyrazin-3-yl)-hydrazine IIe ( 0.9 g).

Compounds of the invention

Example Ia1

1-(3-Chloro-phenyl)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta[a]naphthalene

A mixture of IIa (100 mg), PyBroP (277 mg), 3-chlorobenzoic acid (88.9 mg) and DIPEA (0.3 mL) in DMF (10 mL) was stirred at ambient temperature for 16 hours. Volatiles were removed in vacuo. The residue was stirred in DCM (5 mL) and the corresponding hydrazide (150 mg) precipitated. This material was suspended in a mixture of acetonitrile (10 mL) and DIPEA (0.4 mL) at ambient temperature for 5 min. _PhPOCl2 (0.13 mL) was added and the mixture was stirred at ambient temperature until it became homogeneous. Stirring was continued until a white solid precipitated. The material was filtered off, washed with acetonitrile, and dried. This material (140 mg) was dissolved in methanol (10 mL) and _treated with _Na2CO3 (10 mg) at ambient temperature for 0.5 h. Then, reduce the volume by approximately 50% in a flask open to air. The precipitated solid was filtered off and dried to afford Example Ia1 (60 mg). LC/MS (Method 132): RT (PDA) = 1.59 min; PDA/ELS - Purity 100%/100%; Mass observed: 296.1.

Example Ia2

1-(2-Chloro-6-methyl-phenyl)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta[a]naphthalene

A mixture of IIa (90 mg), PyBroP (264 mg), 2-chloro-6-methylbenzoyl chloride (87 mg) and DIPEA (0.7 mL) was stirred in DMF (2 mL) at ambient temperature for 2 sky. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20/1) to give the corresponding hydrazide (80 mg). More of this material (100 mg), prepared in a similar manner, was dissolved in a mixture of acetonitrile (5 mL) and DIPEA (0.42 mL) and treated with _PhPOCl2 (0.08 mL) at ambient temperature for 5 minutes. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20/1) to afford Example Ia2 (85 mg). LC/MS (Method 132): RT (PDA) = 1.55 min; PDA/ELS-purity: 97.8%/100%; Mass observed: 310.1.

Example Ia3

1-(2,6-Dichloro-phenyl)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta[a]naphthalene

A mixture of IIa (80 mg), PyBroP (221 mg), 2,6-dichloro-benzoic acid (87 mg) and DIPEA (0.2 mL) was stirred in DMF at ambient temperature for 2 days. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20/1) to give the corresponding hydrazide (100 mg). This material was suspended in acetonitrile (3 mL) and treated with DIPEA (0.5 mL) and _PhPOCl2 (0.08 mL) at ambient temperature for 5 min. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20/1) to afford Example Ia3 (15 mg). LC/MS (Method 131): RT (PDA) = 1.07 min; PDA/ELS-purity: 96.1%/100%; mass observed: 330.1.

Example Ia4

1-(2,6-Dimethyl-phenyl)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta[a]naphthalene

To a mixture of IIa (100 mg), DIPEA (0.2 mL) in DMF (2 mL) was added 2,6-dimethylbenzoyl chloride (96 mg), and the mixture was stirred at ambient temperature for 5 minutes. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20/1) to give the corresponding hydrazide (114 mg). 45 mg of this material was suspended in acetonitrile (2 mL) and treated with DIPEA (0.2 mL) for 5 minutes followed by _PhPOCl2 (0.11 mL). The mixture was directly purified by preparative TLC (eluent: pentane/EtOAc, 1/1) to afford Example Ia4 (7.2 mg). LC/MS (Method 132): RT (PDA) = 1.54 min; PDA/ELS-purity: 98.0%/100%; mass observed: 290.0.

Example Ib1

1-(2-Chloro-phenyl)-4-methyl-2,3,5,7,9b-pentaaza-cyclopenta[a]naphthalene

A mixture of lib (480 mg), 2-chlorobenzoic acid (430 mg) and PyBroP (1.33 g) was suspended in DMF (12 mL), and DIPEA (0.97 mL) was added. The mixture was stirred overnight. Volatiles were removed in vacuo. The residue was suspended in DCM. The solid was filtered off, washed with DCM and dried to give 2-chlorobenzoic acid N'-(3-methyl-pyrido[3,4-b]pyrazin-2-yl)-hydrazide (400 mg) as a yellow solid . This material was suspended in acetonitrile (30 mL) at 0 °C. DIPEA (1.7 mL) and _PhPOCl2 (0.35 mL) were added. The mixture was stirred at 0 °C for 0.5 h, then allowed to warm to ambient temperature and stirred for 10 min. Most of the volatiles were removed in vacuo. The solid was filtered off, washed with acetonitrile and dried to give a solid (0.3 g). This material was dissolved in methanol (30 mL) and treated with _Na2CO3 (100 mg) at ambient temperature for 1 _h . The solid was filtered off, and the residue was concentrated in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 10:1) to give Example Ib1 (180 mg) as a white solid. LC/MS (Method 132): RT (PDA) = 1.54 min; PDA/ELS-purity: 99.0%/100%; mass observed: 296.1.

Example Ic1

1-(2-Chloro-phenyl)-4-methyl-2,3,5,8,9b-pentaaza-cyclopenta[a]naphthalene

A mixture of IIc (200 mg), 2-chlorobenzoic acid (179 mg) and PyBroP (554 mg) was suspended in DMF (7 mL). DIPEA (0.41 mL) was added, and the mixture was stirred at ambient temperature. Volatiles were removed in vacuo. The residue was suspended in DCM. The solid was filtered off, washed with DCM and dried to give 2-chlorobenzoic acid N'-(2-methyl-pyrido[3,4-b]pyrazin-3-yl)-hydrazide (313 mg) as a yellow solid . 250 mg of this material was suspended in acetonitrile (200 mL) and treated with DIPEA (1.1 mL) and _PhPOCl2 (0.22 mL) at 0°C. After 10 minutes, the mixture was warmed to ambient temperature and stirred for an additional 5 minutes. Most of the volatiles were removed in vacuo. The solid was filtered off, washed with acetonitrile, and dried. This material (0.32 g) was dissolved in a mixture of methanol (100 mL) and acetonitrile (20 mL). Na ₂ CO ₃ (30 mg) and a few drops of water were added, and the mixture was stirred at ambient temperature for 0.5 h. The solid was filtered off. The filtrate was concentrated in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 10:1) to give Example Ic1 (162 mg) as a white solid. LC/MS (Method 132): RT (PDA) = 1.58 min; PDA/ELS-purity: 94.0%/100%; mass observed: 296.1.

Example Id1

1-(2-Chloro-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

IIId (450 mg) was dissolved in acetonitrile (10 mL), and the solution was purged with nitrogen, followed by the addition of 2-chlorobenzohydrazide (0.44 g). The mixture was heated at 150 °C for 0.5 h under MW conditions. The crude mixture was partitioned between water and EtOAc and basified with NaHCO ₃ . The solid was filtered off, and the organic portion of the filtrate was washed with brine, dried over _MgSO4 , filtered, and concentrated in vacuo. The residue was purified by silica gel chromatography (eluent: 1:1 heptane/EtOAc→EtOAc) to give a sticky solid. This material was suspended in a mixture of diethyl ether and EtOAc (10:1) and the solid was filtered off and dried to give Example Id1 (82 mg). LC/MS (Method 131): RT(PDA) = 1.33 min; PDA/ELS-purity: 97.1%/100%; mass observed: 296.2.

Example Id2

1-(2,6-Dichloro-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

Preparation was carried out as described in Example Id3, using IId (100 mg) and 2,6-dichlorobenzoyl chloride (118 mg), to obtain the corresponding hydrazide (170 mg), which was converted to Example Id2 (18 mg ). LC/MS (Method 132): RT(PDA) = 1.77 min; PDA/ELS-purity: 99.0%/100%; mass observed: 330.1.

Example Id3

1-(2,6-Dimethyl-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

To a mixture of IId (50 mg), DIPEA (0.2 mL) in DMF (2 mL) was added 2,6-dimethylbenzoyl chloride (48 mg), and the mixture was stirred at ambient temperature for 5 minutes. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20/1) to give the corresponding hydrazide (45 mg). This material was mixed with acetonitrile (2 mL) and DIPEA (0.2 mL) and treated with _PhPOCl2 (0.04 mL) at ambient temperature for 3 minutes. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 30/1) to afford Example Id3 (12 mg). LC/MS (Method 132): RT (PDA) = 1.78 min; PDA purity: 100%; Mass observed: 290.0.

Example Id4

1-(2-Chloro-6-methyl-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

IId (90 mg) and DIPEA (0.4 mL) were dissolved in DMF, and 2-chloro-6-methylbenzoyl chloride (106 mg) was added. The mixture was stirred at ambient temperature for 5 minutes. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20:1) to give 2-chloro-6-methyl-benzoic acid N'-(2-methyl-pyrido[2,3-b] pyrazin-3-yl)-hydrazide (110 mg). This material was mixed with DIPEA (0.24 mL) in acetonitrile (2 mL) and treated with _PhPOCl2 (0.1 mL) at ambient temperature for 5 minutes. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: pentane/EtOAc, 1:3) to afford Example Id4 (60 mg). LC/MS (Method 132): RT (PDA) = 1.79 min; PDA/ELS-purity: 98.4%/100%; Mass observed: 310.3.

Example Id5

1-(3-Chloro-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

Preparation as described in Example Id4 using IId (200 mg) and 3-chlorobenzoyl chloride (200 mg) afforded the corresponding hydrazine (130 mg). This material was suspended in acetonitrile (10 mL) and treated with DIPEA (0.4 mL) and _PhPOCl2 (0.11 mL) until the mixture became homogeneous, after which a white solid precipitated. The material was filtered off, washed with acetonitrile, and dried. The dry material (170 mg) _was dissolved in methanol (20 mL) and treated with _Na2CO3 (10 mg) at ambient temperature for 2 hours. Volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM/methanol, 20:1) to afford Example Id5 (31.2 mg). LC/MS (Method 132): RT (PDA) = 1.88 min; PDA/ELS-purity: 99.6%/100%; mass observed: 296.2.

Example Id6

1-(2,6-Dichloro-4-iodo-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

4-Amino-2,6-dichloro-phenol (50 g) was treated with _Boc2O (69 g) in 1,4-dioxane (0.8 L) at reflux for 18 hours and removed in vacuo The volatiles gave (3,5-dichloro-4-hydroxy-phenyl)-carbamic acid tert-butyl ester (70 g), which was used in the next step without further purification. This process is repeated to obtain more of the substance. 86 g of this compound and 2,6-lutidine (49 g) were dissolved in DCM (0.9 L). At -78°C, _Tf2O (104 g) was added dropwise. The mixture was warmed to room temperature, then stirred for 2 hours. The crude mixture was partitioned between water and DCM. _The organic layer was dried over _Na2SO4 , filtered and concentrated in vacuo. The residue was purified by column chromatography on silica gel (eluent: pentane: EtOAc 30:1) to give 4-tert-butoxycarbonylamino-2,6-dichloro-phenyl trifluoro-methanesulfonate (73 g) . This material was mixed with Pd(DPPF) _Cl2 (4 g), triethylamine (102 mL) in a mixture of methanol (580 mL) and DMF (384 mL). The mixture was refluxed overnight under an atmosphere of carbon monoxide, then cooled and concentrated in vacuo. The residue was partitioned between water and EtOAc. The organic layer was _washed with brine, dried over _Na2SO4 , filtered and concentrated in vacuo. The residue was purified by silica gel column chromatography (eluent: pentane:EtOAc, 80:1) to give methyl 4-tert-butoxycarbonylamino-2,6-dichloro-benzoate (12 g). 7 g of this material was dissolved in 37% aqueous HCl (70 mL), and aqueous sodium nitrite (3.75 g) (100 mL) was added dropwise at 0°C. The mixture was stirred at 0°C for 30 minutes, filtered, and the filtrate was added to a solution of potassium iodide (24 g) previously cooled at 0°C. The mixture was warmed to room temperature and stirred overnight. The mixture was extracted with EtOAc. The organic layer was washed with saturated aqueous NaHSO ₃ , then dried over Na ₂ SO ₄ , filtered, and concentrated in vacuo. The residue was purified by silica gel column chromatography (eluent: pentane:EtOAc, 50:1) to give methyl 2,6-dichloro-4-iodobenzoate (7.9 g). This material was dissolved in a mixture of pyridine (40 mL) and water (7 mL) and treated with lithium iodide (3.2 g) at 130 °C for 30 h, after which time the volatiles were removed in vacuo. The residue was partitioned between 2M aqueous HCl and EtOAc. The organic layer was concentrated in vacuo to give 2,6-dichloro-4-iodobenzoic acid (3 g). 0.5 g of this material was stirred in thionyl chloride (8 mL) at 60 °C for 3 hours, after which time the excess thionyl chloride was removed in vacuo. The residue was washed with ether and dried to give 2,6-dichloro-4-iodo-benzoyl chloride (0.53 g), which was used directly in the next step where it was dissolved in DMF (20 mL ) and DIPEA (0.57 mL). To this solution was added IId (277 mg). The mixture was stirred at room temperature for 1 hour. The volatiles were removed in vacuo and the residue was purified by silica gel chromatography (eluent: DCM:MeOH, 100:1 to 30:1) to give 2,6-dichloro-4-iodobenzoic acid N'-(2-methyl -pyrido[2,3-b]pyrazin-3-yl)-hydrazide (250 mg). Repeat the process for more material. 380 mg of this compound was dissolved in 1,4-dioxane (5 mL). Phosphorus oxychloride (4 mL) was added, and the mixture was stirred at 90°C for 1.5 hr. Volatiles were removed in vacuo. The residue was partitioned between DCM and water. The organic layer was washed with saturated aqueous NaHCO ₃ , dried over Na ₂ SO ₄ , filtered, and concentrated in vacuo. The residue was purified by preparative TLC (eluent: pentane:EtOAc, 2:1) to afford Example Id6 (45.5 mg). LC/MS (method WXE-AB10): RT (PDA) = 2.39 min; PDA/ELS purity: 97.1%/98.3%; mass observed: 456.0.

Example Id7

1-(2-Chloro-4-iodo-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

A mixture of 2-chloro-4-iodobenzoic acid (470 mg) and thionyl chloride (6 mL) was stirred at 60°C for 3 hours. Excess thionyl chloride was removed in vacuo. The residue was washed with ether and dried to give 2-chloro-4-iodo-benzoyl chloride (0.5 g), which was used directly in the next step where it was dissolved in 1,4-dioxane ( 10 mL). Phosphorus oxychloride (5 mL) and IId (246 mg) were added, and the mixture was stirred at 90°C for 1.5 hr. Volatiles were removed in vacuo. The residue was partitioned between water and DCM. The organic layer was washed with saturated aqueous NaHCO ₃ , dried over Na ₂ SO ₄ , filtered, and concentrated in vacuo. The residue was purified by column chromatography on silica gel (eluent: pentane:EtOAc, 10:1 to 5:1) to afford Example Id7 (250 mg). LC/MS (method WXE-AB10): RT (PDA) = 2.26 min; PDA/ELS purity: 99%/99%; mass observed: 422.0.

Example Id8

1-(2-Methyl-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

IId (400 mg) was dissolved in a mixture of DIPEA (0.8 mL) and DMF (8 mL). 2-Methyl-benzoyl chloride (368 mg) was added and the mixture was stirred at ambient temperature for 0.5 h before the volatiles were removed in vacuo. The residue was purified by preparative TLC (eluent: DCM:MeOH, 25:1) to give 2-methyl-benzoic acid N'-(2-methyl-pyrido[2,3-b]pyrazine-3 -yl)-hydrazide (310 mg). 100 mg of this material was dissolved in acetonitrile (2 ml), and DIPEA (0.24 mL) and PhP( ₀ )Cl2 (0.07 mL) were added. The mixture was stirred at ambient temperature for 15 minutes before the crude mixture was purified by preparative TLC (eluent: EtOAc:pentane, 3:2) to afford Example Id8 (5 mg). LC/MS (method WXE-AB01): RT (PDA) = 2.05 min; PDA/ELS purity: 95.3%/95.7%; mass observed: 276.2.

Example Ie1

1-(2-Chloro-phenyl)-6-methoxy-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

To a suspension of IIe (500 mg) in DCM (80 mL) was added compound 2-chloro-benzaldehyde (377 mg). The mixture was stirred at 40°C for two days, then the volatiles were removed in vacuo. The residue was purified by silica gel chromatography (eluent: pentane/EtOAc, 1:0 to 0:1) to afford N-[1-(2-chloro-phenyl)-methylene]-N'-(8 -Methoxy-2-methyl-pyrido[2,3-b]pyrazin-3-yl)-hydrazine (0.5 g). This material was dissolved in DCM (60 mL), (bisacethoxy)iodobenzene (540 mg) was added, and the reaction solution was stirred at room temperature overnight. The volatiles were removed in vacuo and the residue was purified by silica gel chromatography (eluent: pentane/EtOAc, 1:0 to 0:1) to afford Example Ie1 (100 mg). LC/MS (Method 550): RT(PDA) = 0.55 min; PDA/ELS-purity: 90%/100%; mass observed: 326.0.

Example Ie2

1-(2-Chloro-6-methyl-phenyl)-6-methoxy-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

To a suspension of IIe (0.5 g) in DCM (80 mL) was added 2-chloro-6-methyl-benzaldehyde compound (377 mg). The mixture was stirred at 40 °C for two days, then (diacetoxy)iodobenzene (864 mg) was added and stirring was continued at ambient temperature overnight. The volatiles were removed in vacuo and the residue was purified by preparative TLC (eluent: EtOAc) to afford Example 1e2 (185 mg). LC/MS (method WXE-AB01): RT (PDA) = 1.9 min; PDA/ELS purity: 100%/100%; mass observed: 340.1.

Example Ie3

1-(2,6-Dimethyl-phenyl)-6-methoxy-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta[a]naphthalene

To a suspension of IIe (0.5 g) in DCM (80 mL) was added 2,6-dimethyl-benzaldehyde (377 mg), and the mixture was stirred at 40 °C for two days before adding (diacetoxy ) iodobenzene (864 mg), and the mixture was stirred overnight at ambient temperature. The volatiles were removed in vacuo and the residue was purified by silica gel chromatography (eluent: EtOAc) to afford Example Ie3 (175 mg). LC/MS (method WXE-AB10): RT (PDA) = 1.88 min; PDA/ELS purity: 100%/100%; mass observed: 320.1.

PDE in vitro test

In conjunction with the following methods, the inhibitory activity of the compounds of the present invention is determined:

PDE10A enzyme

Active PDE10A enzymes have been prepared in a number of ways for use in PDE assays (Loughney, K. et al. Gene 1999, 234, 109-117; Fujishige, K. et al. Eur J Biochem. 1999, 266, 1118-1127 and Soderling, S. et al. Proc. Natl. Acad. Sci. 1999, 96, 7071-7076). PDE10A can be expressed as a full-length protein or a truncated protein as long as they express the catalytic domain. PDE10A can be produced in various cell types, eg, insect cells or E. coli. An example of a method for obtaining catalytically active PDE10A is as follows: The catalytic domain of human PDE10A (amino acid sequence 440-779, accession number NP 006652) was amplified from total human whole brain RNA using standard RT-PCR and cloned into the pET28a vector (Novagen) BamH1 and Xho1 sites. Expression was performed in E. coli following standard protocols. Briefly, expression plasmids were transformed into the BL21(DE3) E. coli strain, 50 mL cultures were inoculated with cells, allowed to grow to an OD600 of 0.4-0.6, and then protein expression was induced with 0.5 mM IPTG. After induction, cells were incubated overnight at room temperature and harvested by centrifugation. Resuspend the PDE10A-expressing cells in 12 mL (50 mM TRIS-HCl-pH 8.0, 1 mM MgCl ₂ and protease inhibitors). Cells were lysed by sonication and after all cells were lysed, TritonX100 was added according to the Novagen protocol. PDE10A was partially purified on Q Sepharose and most of the active fractions were pooled.

PDE10A inhibition test

A typical PDE10A assay is performed as follows: 60 μL containing a fixed amount of PDE2A enzyme (enough to convert 20-25% of the cyclic nucleotide substrate), buffer (50mM HEPES, pH7.6; 10mM _MgCl2 ; 0.02% Tween20), The assay was performed with samples of 10 nM tritium-labeled cAMP and different amounts of inhibitor. Reactions were initiated by addition of cyclic nucleotide substrate, allowed to proceed for 1 hour at room temperature, and then terminated by mixing with 20 [mu]L (0.2 mg) yttrium silicate SPA microparticles (Amersham). The microparticles were allowed to settle for 1 hour in the dark before counting the plates in a Wallac 1450 Microbeta counter. The measured signal was converted to activity (relative to uninhibited control (100%)) and _IC50 values were calculated using XlFit (model 205, IDBS).

PDE2A enzyme

Likewise, active human PDE2A enzyme (ATCC 68585) can be prepared for use in PDE assays in a number of ways, methods well known to those skilled in the art.

PDE2A inhibition assay

A typical PDE2A assay is carried out as follows: 60 μL containing a fixed amount of PDE2A enzyme (enough to convert 20-25% of the cyclic nucleotide substrate), buffer (50mM HEPES, pH7.6; 10mM MgCl ₂ ; 0.02% Tween20), The assay was performed on samples of 0.1 mg/ml BSA, 15 nM tritiated cAMP and different amounts of inhibitors. Reactions were initiated by addition of cyclic nucleotide substrate, allowed to proceed for 1 hour at room temperature, and then terminated by mixing with 20 [mu]L (0.2 mg) yttrium silicate SPA microparticles (Amersham). The microparticles were allowed to settle for 1 hour in the dark before counting the plates in a Wallac 1450 Microbeta counter. The measured signal was converted to activity (relative to uninhibited control (100%)) and _IC50 values were calculated using XlFit (model 205, IDBS).

The data obtained for selected examples are listed in the table below.

Claims (26)

1. the compound of formula I:

Wherein X ¹, X ², X ³and X ⁴n or CR independently of one another ³, condition is, and an X is N, and all the other X are CR independently of one another ³;

Wherein R ¹c ₁-C ₆alkyl, C ₃-C ₆cycloalkyl, THP trtrahydropyranyl, benzyl, phenyl and pyridyl, wherein benzyl, phenyl and pyridyl are optionally by one or more halogens, CN, C ₁-C ₄alkyl/fluoroalkyl or C ₁-C ₄alkoxyl group/Fluoroalkyloxy replaces;

Wherein R ²c ₁-C ₄alkyl or C _3-c ₆cycloalkyl;

Wherein R ³hydrogen, halogen, CN ,-CO ₂h ,-CON (H or C _1-c ₄alkyl) ₂, CHO, C ₁-C ₄alkyl/fluoroalkyl, contain amino heterocycle, the C of ring ₂-C ₄thiazolinyl, C ₂-C ₄thiazolinyl or C ₁-C ₄alkoxyl group/Fluoroalkyloxy; Or its pharmacologically acceptable salt.

2. the compound of claim 1, wherein R ²c ₁-C ₄alkyl.

3. the compound of claim 1 or 2, wherein R ²it is methyl.

4. the compound of claim 1, wherein R ²c ₃-C ₆cycloalkyl.

5. the compound of any one of claim 1-4, wherein R ¹c ₁-C ₄alkyl.

6. the compound of any one of claim 1-4, wherein R ¹c ₃-C ₆cycloalkyl.

7. the compound of any one of claim 1-4, wherein R ¹it is THP trtrahydropyranyl.

8. the compound of any one of claim 1-4, wherein R ¹optionally by one or two F, Cl or C ₁-C ₃the benzyl that alkyl replaces.

9. the compound of any one of claim 1-4, wherein R ¹optionally by one or two F, Cl or C ₁-C ₃the phenyl that alkyl replaces.

10. the compound of any one of claim 1-4, wherein R ¹optionally by one or two F, Cl or C ₁-C ₃the pyridyl that alkyl replaces.

The compound of any one of 11. claim 1-4, wherein R ¹by one or two F, Cl or C ₁-C ₃the phenyl that alkyl replaces.

The compound of any one of 12. claim 1-11, wherein X ¹n.

The compound of any one of 13. claim 1-11, wherein X ²n.

The compound of any one of 14. claim 1-11, wherein X ³n.

The compound of any one of 15. claim 1-11, wherein X ⁴n.

The compound of any one of 16. claim 1-15, wherein R ³hydrogen.

The compound of any one of 17. claim 1-15, wherein R ³halogen or CHO.

The compound of any one of 18. claim 1-15, wherein R ³c ₁-C ₄alkyl or C ₁-C ₄alkoxyl group.

The compound of any one of 19. claim 1-15, wherein R ³c ₂-C ₄thiazolinyl or C ₂-C ₄thiazolinyl.

The compound of any one of 20. claim 1-15, wherein R ³to contain the amino heterocycle of ring.

The compound of 21. claims 1, wherein this compound is selected from: 1-(the chloro-phenyl of 3-)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta [a] naphthalene; 1-(the chloro-6-methyl-phenyl of 2-)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta [a] naphthalene; 1-(the chloro-phenyl of 2,6-bis-)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta [a] naphthalene; 1-(2,6-dimethyl-phenyl)-4-methyl-2,3,5,6,9b-pentaaza-cyclopenta [a] naphthalene; 1-(the chloro-phenyl of 2-)-4-methyl-2,3,5,7,9b-pentaaza-cyclopenta [a] naphthalene; 1-(the chloro-phenyl of 2-)-4-methyl-2,3,5,8,9b-pentaaza-cyclopenta [a] naphthalene; 1-(the chloro-phenyl of 2-)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta [a] naphthalene; 1-(the chloro-phenyl of 2,6-bis-)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta [a] naphthalene; 1-(2,6-dimethyl-phenyl)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta [a] naphthalene; 1-(the chloro-6-methyl-phenyl of 2-)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta [a] naphthalene; And 1-(the chloro-phenyl of 3-)-4-methyl-2,3,5,9,9b-pentaaza-cyclopenta [a] naphthalene.

The pharmaceutical composition of 22. compound or pharmaceutically acceptable salt thereofs that contain claim 1 and pharmaceutically acceptable carrier.

The method of 23. treatment anxiety disorders, the method comprises: the compound for the treatment of any one of the claim 1-21 of significant quantity.

The method of 24. treatment cognitive disorders, the method comprises: the compound for the treatment of any one of the claim 1-21 of significant quantity.

The schizoid method of 25. treatment, the method comprises: the compound for the treatment of any one of the claim 1-21 of significant quantity.

The compound of any one of 26. claim 1-21 is used for the treatment of.