WO2006076318A1 - Monopotassium salt of an ikb kinase inhibitor - Google Patents

Abstract

The present invention is directed to the compound of formula (II), or a solvate thereof. The invention is also directed to the pharmaceutical use of the compound as an IκB kinase inhibitor, crystalline anhydrous and hydrated forms of the compound, pharmaceutical compositions comprising the compounds of the invention, and stable pharmaceutical products comprising the compound. As an inhibitor of IκB kinase, the compound of the invention, functions via the selective inhibition of IKK, particularly an IKK-2 inhibitor; as well as exhibiting localized activity, as opposed to a systemic activity. Such an inhibitor is particularly useful for treating a patient suffering from or subject to IKK-2 mediated pathological diseases or conditions, e.g., asthma, rhinitis, chronic obstructive pulmonary disorder (COPD), or COPD exacerbations, that could be ameliorated by the targeted administering of the inhibitor.

Description

MONOPOTASSIUM SALT OF AN IκB KINASE INHIBITOR

This application claims the benefit of U.S. provisional application No. 60/643,300, filed January 12, 2005.

FIELD OF THE INVENTION

The present invention is directed to the compound of formula II

π or a solvate thereof.

The invention is also directed to the pharmaceutical use of the compound as an IKB kinase inhibitor, crystalline anhydrous and hydrated forms of the compound, pharmaceutical compositions comprising the compounds of the invention, and stable pharmaceutical products comprising the compound.

As an inhibitor of IKB kinase, the compound of the invention, functions via the selective inhibition of IKK, particularly an IKK-2 inhibitor; as well as exhibiting localized activity, as opposed to a systemic activity. Such an inhibitor is particularly useful for treating a patient suffering from or subject to IKK- 2 mediated pathological diseases or conditions, e.g., asthma, rhinitis, chronic obstructive pulmonary disorder (COPD), or COPD exacerbations, that could be ameliorated by the targeted administering of the inhibitor.

REPORTED DEVELOPMENTS NF-κB is a heterodimeric transcription factor that regulates the expression of multiple inflammatory genes. NF-κB has been implicated in many pathophysiologic processes including angiogenesis (Koch et al., Nature 376:517-519, 1995), atherosclerosis (Brand et al., J Clin Inv. 97:1715-1722, 1996), endotoxic shock and sepsis (Bohrer et al., J. Clin. Inv. 100:972-985, 1997), inflammatory bowel disease (Panes et al., Am J Physiol. 269:H1955-H1964, 1995), ischemia/reperfusion injury (Zwacka et al., Nature Medicine 4:698-704, 1998), and allergic lung inflammation (Gosset et al., Int Arch Allergy Immunol. 106:69-77, 1995). Thus the inhibition of NF-κB by targeting regulatory proteins in the NF- KB activation pathway represents an attractive strategy for generating anti-inflammatory therapeutics due to NF-κB's central role in inflammatory conditions.

The IKB kinases (IKKs) are key regulatory signaling molecules that coordinate the activation of NF- KB. Many immune and inflammatory mediators including TNFα, lipopolysaccharide (LPS), EL- lβ, CD3/CD28 (antigen presentation), CD40L, FasL, viral infection, and oxidative stress have been shown to lead to NF-κB activation. Although the receptor complexes that transduce these diverse stimuli appear very different in their protein components, it is understood that each of these stimulation events leads to activation of the IKKs and NF-κB.

The IKK complex appears to be the central integrator of diverse inflammatory signals leading to the phosphorylation of IKB. Cell and animal experiments indicate that IKK-2 is a central regulator of the pro-inflammatory role of NF-κB, wherein the IKK-2 is activated in response to immune and inflammatory stimuli and signaling pathways. Many of those immune and inflammatory mediators, including DL- lβ, LPS, TNFα, CD3/CD28 (antigen presentation), CD40L, FasL, viral infection, and oxidative stress, play an important role in respiratory diseases. Furthermore, the ubiquitous expression of NF-κB, along with its response to multiple stimuli means that almost all cell types present in the lung are potential targets for anti-NF-κB/IKK-2 therapy. This includes alveolar epithelium, mast cells, fibroblasts, vascular endothelium, and infiltrating leukocytes, including neutrophils, macrophages, lymphocytes, eosinophils and basophils.

Inhibitors of IKK-2 are believed to display broad anti-inflammatory activity by inhibiting the expression of genes such as cyclooxygenase-2 and 12-lipoxygenase (synthesis of inflammatory mediators), TAP-I peptide transporter (antigen processing), MHC class I H-2K and class II invariant chains (antigen presentation), E-selectin and vascular cell adhesion molecule (leukocyte recruitment), interleukins-1, 2, 6, 8 (cytokines), RANTES, eotaxin, GM-CSF (chemokines), and superoxide dismutase and NADPH quinone oxidoreductase (reactive oxygen species).

NF-κB is activated beyond its normal extent in diseases such as rheumatoid arthritis, osteoarthritis, asthma, chronic obstructive pulmonary disorder (COPD), rhinitis, multiple sclerosis, cardiac infarction, Alzheimer's diseases, diabetes Type II, inflammatory bowel disease or atherosclerosis.

Chronic obstructive pulmonary disease (COPD) is a debilitating inflammatory disease of the lungs characterized by the progressive development of airflow limitation that is not fully reversible (Pauwels et al., 2001). The airflow limitation is associated with an abnormal inflammatory response of the lungs to noxious particles or gases, primarily caused by cigarette smoking. Although COPD affects the lungs, it also produces significant systemic consequences. The term COPD encompasses chronic obstructive bronchitis, with obstruction of small airways, and emphysema, with enlargement of air spaces and destruction of lung parenchyma, loss of lung elasticity, and closure of small airways. Chronic bronchitis, by contrast, is defined by the presence of a productive cough (due to hypersecretion of mucus) of more than three months' duration for more than two successive years. There is some epidemiologic evidence that mucus hypersecretion is accompanied by airflow obstruction, perhaps as a result of obstruction of peripheral airways. Most patients with COPD have all three pathologic conditions (chronic obstructive bronchitis, emphysema, and mucus plugging), but the relative extent of emphysema and obstructive bronchitis within individual patients can vary, Vestbo et al., 1996; Barnes,

2004a, Barnes, 2004b; Hogg, 2004.

In industrialized countries, cigarette smoking accounts for most cases of COPD. but in developing countries other environmental pollutants (particularly with sulfur dioxide and particulates) and certain occupational chemicals (such as cadmium), are important causes. Passive smoking is also a risk factor.

COPD patients are predisposed to exacerbations, that is, an acute worsening of their respiratory symptoms. An exacerbation of COPD is an event in the natural course of the disease characterized by a change in the patient's baseline dyspnea, cough and/or sputum beyond day-to-day variability sufficient to warrant a change in management (Rodriguez-Roisin, 2000; Burge and Wedzicha, 2003).

Tracheobronchial infections are believed to be a common cause of exacerbation in COPD, although controversy exists regarding the nature of the infectious agent as well as its exact role (Wedzicha, -A-

2002; White et al., 2003). In addition, exacerbations of COPD are clearly associated with the levels of respirable particles and environmental air pollutants, and these have been linked to hospital admission rates (Rennard and Farmer, 2004).

The frequency of exacerbations is linked to disease severity in COPD. Exacerbations, may adversely affect the natural history of these disorders, perhaps by contributing to increased rates of lung function decline, systemic effects, and premature mortality. Unfortunately, to date, there is no widely accepted definition of what constitutes an exacerbation of COPD (Rodriguez-Roisin, 2000). The intensity and duration of increased symptoms required to qualify as an "exacerbation" are difficult to define. Indeed, several definitions co-exist, and many clinical trials employ substantially different criteria or describe poorly the definition(s) used to diagnose exacerbation. The most widely quoted clinical criteria used in the characterization of acute exacerbation of COPD are those described by Anthonisen et al., (1987). In that study exacerbations were divided into three groups: type 1 exacerbations were characterized by increased breathlessness, increased sputum volume, and new or increased sputum purulence; type 2 included any two of these symptoms; and type 3 consisted of any one of the symptoms together with at least one additional feature, including sore throat or nasal discharge within the last five days; unexplained fever; increased wheeze; increased cough; or a 20% increase in respiratory or heart rate compared with baseline. These criteria have been used as a benchmark ever since, and all proposed etiologies of exacerbation need to establish their relationship to these key features.

The inhibition of NF-κB is also described as being useful for treating hypoproliferative diseases, e.g., solid tumor and leukemias, on its own or in addition to cytostatic therapy. Inhibition of the NF-κB~ activating signal chain at various points or by interfering directly with the transcription of the gene by glucocorticoids, salicylates or gold salts, has been shown as being useful for treating rheumatism.

Many inhibitors of IKB kinase are known to frequently suffer from the disadvantage of being nonspecific for inhibiting only one class of kinases. For example, most inhibitors of IKB kinase inhibit several different kinases at the same time because the structures of the catalytic domains of these kinases are similar. Consequently, the inhibitors act, in an undesirable manner on many enzymes, including those that possess the vital function.

Patent application WO2001/30774 discloses indole derivatives and patent application

WO2004/022553, discloses indole and benzimidazole derivatives, which derivatives are able to modulate NF-κB and which exhibit a strong inhibitory effect on IKB kinase. Particularly, patent application WO2004/022553 discloses indole and benκimidazole derivatives of formula (I), their preparation,

pharmaceutical compositions containing these compounds, and methods for the prophylaxis and therapy of a disease associated with an increased activity of IKB kinase comprising administering such compounds. Furthermore, patent application WO2004/022553 discloses the following compounds of formulae (B), (C), and (D):

(compound 43),

C (compound 32); and

D (compound 48).

Patent application WO2004/022553 does not specifically disclose the compound of Formula (I) wherein M is N; Rl is hydrogen, R2 is carboxyl (-COOH), R3 is methyl, R4 is pyridin-2-yl, Rl 1 is hydrogen, and X is CH. The structure and synthesis of this compound as the free carboxylic acid is disclosed, and claimed as being useful in the treatment of asthma and COPD, in patent application WO2005/113544, hereby incorporated by reference. While patent application WO2005/113544 generically discloses how to prepare a wide variety of base addition salts, however the working examples are limited to the preparation of the free acid, hereinafter referred to as "free form." Thus, patent application WO2005/113544 provides no specific disclosure or suggestion of the potassium salt hydrolyzed analogue of compound 32 or 43.

The large-scale manufacturing of a pharmaceutical composition poses many challenges to the chemist and chemical engineer. While many of these challenges relate to the handling of large quantities of reagents and control of large-scale reactions, the handling of the final product poses special challenges linked to the nature of the final active product itself. Not only must the product be prepared in high yield, be stable, and capable of ready isolation, the product must possess properties that are suitable for the types of pharmaceutical preparations in which they are likely to be ultimately used. The stability of the active ingredient of the pharmaceutical preparation must be considered during each step of the manufacturing process, including the synthesis, isolation, bulk storage, pharmaceutical formulation and long-term formulation. Each of these steps may be impacted by various environmental conditions of temperature and humidity.

The pharmaceutically active substance used to prepare the pharmaceutical compositions should be as pure as possible and its stability on long-term storage must be guaranteed under various environmental conditions. This is absolutely essential to prevent the appearance of unintended degradation products in pharmaceutical compositions, which degradation products may be potentially toxic or result simply in reducing the potency of the composition. A primary concern for the manufacture of large-scale pharmaceutical compounds is that the active substance should have a stable crystalline form to ensure consistent processing parameters and pharmaceutical quality. If an unstable crystalline form is used, that crystalline form may change during manufacture and/or storage resulting in quality control problems, and formulation irregularities. Such a change may affect the reproducibility of the manufacturing process and thus lead to final formulations which do not meet the high quality and stringent requirements imposed on formulations of pharmaceutical compositions. In this regard, it should be generally borne in mind that any change to the solid state of a pharmaceutical composition which can improve its physical and chemical stability gives a significant advantage over less stable forms of the same drug. No reference describes an inhibitor of IKB kinase that operates through the selective inhibition of IKK, particularly an IKK-2 inhibitor, as well as such an inhibitor that exhibits localized activity as opposed to a systemic activity as well as having properties suitable for large-scale manufacturing and formulation.

No reference discloses or teaches any particular crystalline potassium salt form of 2-{[2-(2- methylamino-pyrimidin-4-yl)- 1 H-indole-5-carbonyl]~amino} -3-(phenylpyridin-2-yl-amino)-propionic acid; or one that is particularly useful for large-scale manufacturing and pharmaceutical formulation.

SUMMARY OF THE INVENTION

The present invention is directed to a compound of formula II

π or a solvate form thereof; a pharmaceutical composition comprising a pharmaceutically effective amount of the compound of formula II, and a pharmaceutically acceptable carrier; and the use of a compound of formula II for treating a patient suffering from, or subject to, a pathological condition capable of being ameliorated by inhibiting IKK-2 comprising administering to said patient a pharmaceutically effective amount of the compound of formula II, and stable pharmaceutical products comprising the compound. The present invention is more fully discussed with the aid of the following figures and detailed description below.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGURE 1 is a powder X-ray diffractogram of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5- carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, as the monopotassium monohydrate salt.

FIGURE 2 is the corresponding Table of XRPD d-Spacings and Relative Intensities for the powder X- ray diffractogram in Figure 1 of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]- amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, as the monopotassium monohydrate salt.

FIGURE 3 is a differential scanning calorimeter-thermal gravimetric analyzer (DSC-TGA) thermogram of 2-{[2-(2-methylamino-pj'rimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-phenylpyridin-2-yl- amino)-propionic acid, as the monopotassium monohydrate salt. This DSC-TGA thermogram shows the dehydration of crystalline form α-1 at >100°C to about 140°C, to form anhydrous crystalline form β-1, which then recrystallizes at about 220°C to form a second anhydrous crystalline form β-2, having a melting point of about 293 °C.

FIGURE 4 is a DSC thermogram of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]- amino}-3-(phenylpyridin-2- yl-amino)-propionic acid, as the monopotassium monohydrate salt.

FIGURE 5 are photomicrographs post milling of the 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH- indole-5-carbonyl]-amino}-3-phenylpyridin-2-yl-amino)-propionic acid, as the monopotassium monohydrate salt, showing good reduction in particle size with no change in physicochemical properties and no degradation is observed using HPLC.

FIGURE 6 is a Dynamic Vapor Sorption Analyzer (DVS) Isothgrm plot of 2-{[2-(2-methylamino- pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, as the monopotassium monohydrate salt and the corresponding Table Water Sorption Profile.

FIGURE 7 is a Fourier Transform-Infrared (FT-IR) spectrum comparing free form 2-{[2- (2-methylamino-pyrhnidin-4-yl)- 1 H-indole-5-carbonyl]-amino} -3 -(phenylpyridin-2-yl-amino)- propionic acid and the crystalline monopotassium monohydrate salt and its corresponding Table of FT- ER peaks. FlGURE 8 are IKK cell-free assay results comparing free form 2-{[2-(2-methylamino-pyrimidin- 4- yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, S- and R- enantiomers; and 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3- (phenylpyridin-2- yl-amino)-propionic acid monopotassium monohydrate salts, S- and R-enantiomers.

FIGURE 9 represents the through inhaler life emitted weight performance of dry powder inhalers (Ultrahaler®) that are filled with a dry powder blend of monopotassium monohydrate salt of 2- {[2 -(2- methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid (active) and are mixed with α-lactose monohydrate (excipient). The proportion of active to excipient is chosen to give an approximate single actuation dose of 0.5 mg active. The graph shows consistent emitted weight performance through device life with 227 of 230 emitted weights within ±25% of mean value.

FIGURE 10 represents the through inhaler life emitted dose performance of dry powder inhalers (Ultrahaler®) that are filled with a dry powder blend of monopotassium monohydrate salt of 2-{[2-(2- methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid (active) and are mixed with α-lactose monohydrate (excipient). The proportion of active to excipient is chosen to give an approximate single actuation dose of 0.5 mg active. The graph shows consistent emitted dose (μg) performance through device life with 48 of 50 emitted doses within ±35% of mean value (0.76 mg/actuation).

FIGURE 11 represents the through inhaler life emitted weight performance of dry powder inhalers (Ultrahaler®) that are filled with a dry powder blend of monopotassium monohydrate salt of 2-{[2-(2- methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid (active) and are mixed with α-lactose monohydrate (excipient). The proportion of active to excipient is chosen to give an approximate single actuation dose of 5 mg active. The graph shows consistent emitted weight performance through device life with 209 of 217 emitted weights within ±25% of mean value.

FIGURE 12 represents the through inhaler life emitted dose performance of dry powder inhalers (Ultrahaler®) that are filled with a dry powder blend of monopotassium monohydrate salt of 2-{[2-(2- methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid (active) and that are mixed with α-lactose monohydrate (excipient). The proportion of active to excipient is chosen to give an approximate single actuation dose of 5 mg active. The graph shows consistent emitted dose (μg) performance through device life with 40 of 40 emitted doses within ±35% of mean value (4.7 mg/actuation).

FIGURE 13 represents the fine particle performance of dry powder inhalers (Ultrahaler®) that are filled with a dry powder blend of monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4- yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid (active) and that are mixed with α-lactose monohydrate (excipient). The proportion of active to excipient in each blend is chosen to give approximate single actuation doses of 1 mg and 5 mg active. The graph shows that both dry powder inhaler formulations are able to deliver a significant proportion (ca. 40-60%) of the emitted dose (μg) as fine particles.

DETAILED DESCRIPTION OF THE INVENTION

With reference to the present invention described herein, below are particular embodiments related thereto.

Embodiments

One particular embodiment of the invention is the compound of formula II as a monohydrate; more particularly to the crystalline form α-1.

Another particular embodiment of the invention is the compound of formula II as anhydrous; more particularly to the crystalline form β-1.

Another particular embodiment of the invention is the compound of formula II as anhydrous; more particularly to the crystalline form β-2.

Another particular embodiment of the invention is the compound of formula II as a solvate or hydrate.

Another particular embodiment of the invention is the compound of formula π as the monopotassium monohydrate salt, i.e., Form α-1, having unexpected properties that are useful for large-scale manufacturing and pharmaceutical formulation.

Another particular embodiment of the invention is the use of a compound of formula π for treating a patient suffering from, or subject to, a pathological condition capable of being ameliorated by inhibiting EKK-2, such as, asthma, rhinitis, chronic obstructive pulmonary disorder or chronic obstructive pulmonary disorder exacerbations. Another particular embodiment of the invention is the compound of formula II wherein the administration is intratracheal, intranasal, inhalational, or by aerosolization.

Another particular embodiment of the invention is a stable pharmaceutical product comprising a metered dose inhaler and respirable crystalline form α-1, respirable anhydrous crystalline form β-1, or respirable anhydrous crystalline form β-2.

Another particular embodiment of the invention is a stable pharmaceutical product comprising a dry powder inhaler and respirable crystalline form α-1, respirable anhydrous crystalline form β-1, or respirable anhydrous crystalline form β-2.

Another particular embodiment of the invention is directed to respirable crystalline form α-1, respirable anhydrous crystalline form β-1, or respirable anhydrous crystalline form β-2, wherein about 90% of the crystalline particles are not more than about 7 microns in size; or more particularly, from about 2 to about 6 microns in size.

Another particular embodiment of the invention is directed to respirable crystalline form α-1, respirable anhydrous crystalline form β-1, or respirable anhydrous crystalline form β-2, wherein the median diameter of the crystalline particles is from about 1 to 3 microns in size; or more particularly about 1.5 microns in size.

Another particular embodiment of the invention is directed to crystalline form α-1 comprising less than about 10% of amorphous product upon micronization; more particularly less than about 7% of amorphous product upon micronization; or further more particularly less than about 5% of amorphous product upon micronization.

It is to be understood that this invention covers all appropriate combinations of the particular embodiments herein.

Definitions and Abbreviations

As used above, and throughout the description of the invention, the following abbreviations, unless otherwise indicated, shall be understood to have the following meanings:

HFA 227 1,1,1,2,3,3,3-heptafluoropropane

HFA 134a 1,1,1,2-tetrafluoroethane HPLC high performance liquid chromatography i.n. intranasally p.o. oral i.p. intraperitoneal i.t. intratracheally mpk mg/kg

SD standard deviation

EtOH ethanol

PG propylene glycol

THP tetrahydrofuran

MDI metered-dose inhaler

DPI dry powder inhaler rpm revolutions per minute

RT room temperature

As used above, and throughout the description of the invention, the following terms, unless otherwise indicated, shall be understood to have the following meanings.

"Treating" or "treatment" means prevention, partial alleviation, or cure of the disease. The compound and compositions of this invention are useful in treating conditions that are characterized by the activation of NF-κB and/or enhanced levels of cytokines and mediators that are regulated by NF-κB including, but not limited to TNFα and EL-I β. Inhibition or suppression of NF-κB and/or NF-κB- regulated genes such as TNFα may occur locally, for example, within certain tissues of the subject, or more extensively throughout the subject being treated for such a disease. Inhibition or suppression of NF-κB and/or NF-κB-regulated genes such as TNFα may occur by one or more mechanisms, e.g., by inhibiting or suppressing any step of the pathway(s) such as inhibition of HCK.

The term "NF-κB-associated condition" refers to diseases that are characterized by activation of NF- KB in the cytoplasm (e.g., upon phosphorylation of IKB).

The term "TNFα-associated condition" is a condition characterized by enhanced levels of TNFα. In the instant specification, the term NF-κB-associated condition will include a TNFα-associated condition but is not limited thereto as NF-κB is involved in the activity and up regulation of other proinflammatory proteins and genes.

The term "inflammatory or immune diseases or disorders" is used herein to encompass both NF~κB- associated conditions and TNFα-associated conditions, e.g., any condition, disease, or disorder that is associated with release of NF-κB and/or enhanced levels of TNFα, including conditions as described herein.

"Patient" includes both human and other mammals.

"Pharmaceutically effective amount" is meant to describe an amount of a compound, composition, medicament or other active ingredient effective in producing the desired therapeutic effect.

"Form α-1" is meant to describe a crystalline form of the compound of formula II as a monohydrate that may be characterized using distinguishing data. Exemplary data is found in Figures 1, 2, 3, 4, 5, 6 and/or 7.

"Form β-1" is meant to describe the anhydrous crystalline form of a compound of formula π, derived from dehydration of the monohydrate crystalline form that may be characterized using distinguishing data. Exemplary data is found in Figure 3.

"Form β-2" is meant to describe the anhydrous crystalline form of a compound of formula π, derived from recrystallizing Form β-1, that may be characterized using distinguishing data. Exemplary data is found in Figure 3.

The term "solvate or solvated" means a physical association of a compound of this invention with one or more solvent molecules. This physical association includes hydrogen bonding. In certain instances the solvate will be capable of isolation, for example when one or more solvent molecules are incorporated within a crystalline solid. "Solvate or solvated" encompasses both solution-phase and isolable solvates. Representative solvates include, for example, a hydrate, ethanolates or a methanolate.

The term "hydrate"' is a solvate wherein the solvent molecule is H₂O that is present in a defined stoichiometric amount, and may for example, include hemihydrate, monohydrate, dihydrate, or trihydrate.

The term "monohydrate" is meant to describe a compound associated with one equivalent of water.

The term "respirable" is meant to describe a substance, e.g., crystalline form α-1, capable of being taken in by breathing. Combination therapies may improve efficacy and decrease the risk of side effects compared with increasing the dose of a single agent. DCK inhibitors can be combined with bronchodilators including but not limited to short-acting β2-agonists; long-acting B2-agonists such as salmeterol and foπnoterol; anticholinergic agents such as ipratropium bromide and tiotropium bromide. IKK inhibitors can also be combined with methylxanthines such as theophylline.

Inhibitors of IKK2 can be combined with several anti-inflammatory therapies including but not limited to immunomodulators directed at various stages of the inflammatory cascade and directed to ameliorating inflammatory processes. Such therapies include, but are not limited to:

(A) Inhibitors of cellular recruitment and toxic inflammatory mediators including but not limited to phosphodiesterase-4 inhibitors; inhibitors of p38 mitogen-activated protein kinase; biopharmaceuticals such as anti-tumor necrosis factor-alpha, anti-interleukin-8, and anti-monocyte chemoattractant protein- 1; inhibitors of adhesion molecules and chemotactic factors; and molecules that interfere with cell survival and clearance/ apoptosis;

(B) Inhibitors of proteolytic enzymes including but not limited to inhibitors of neutrophil-derived serine proteases such as neutrophil elastase; and inhibitors of matrix metalloproteinases (MMPs) such as MMP-2, MMP-9 and MMP- 12;

(C) Antioxidants including but not limited to N-acetylcysteine and inhibitors or scavengers of reactive oxygen species; and toxic peptides such as defensins that can directly cause cell injury;

(D) Inhibitors of mucus production including but not limited to inhibitors of mucous genes; and also mucus clearing agents such as expectorants, mucolytics, and mucokinetics; and

(E) Antibiotic therapy such as with a ketolide, for example Ketek®.

The drug combinations of the present invention can be provided to a cell or cells, or to a human patient, either in separate pharmaceutically acceptable formulations administered simultaneously or sequentially, formulations containing more than one therapeutic agent, or by an assortment of single agent and multiple agent formulations. However administered, these drug combinations form a pharmaceutically effective amount of components. The treatment regimen/dosing schedule can be rationally modified over the course of therapy so that the lowest amounts of each of the pharmaceutically effective amount of compounds used in combination which together exhibit satisfactory pharmaceutical effectiveness are administered, and so that administration of such pharmaceutically effective amount of compounds in combination is continued only so long as is necessary to successfully treat the patient.

The compound of the present invention may be administered with any suitable metered-dose inhaler, for example, as described herein, or any suitable dry powder inhaler, such as the Eclipse, Spinhaler®, or Ultrahaler® as described in patent application WO2004/026380, or US Patent No. 5,176,132.

By the suitable choice of manufacturing conditions, crystalline form α-1 can be obtained in its monohydrate form, which is particularly suitable for large-scale manufacturing processes.

Compounds of the present invention may be prepared, or formed during the process of the invention, as solvates, e.g. hydrates. Solvates of compounds of the present invention may be prepared for example, by recrystallization or evaporation from water, an aqueous/organic solvent mixture, or organic solvent, using for example, dioxane, tetrahydrofuran or methanol.

The preparation and properties of the compounds of the invention are described in the following experimental section.

EXAMPLES

Example 1, Step 1

Synthesis of 2-{[2-(2-Methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]amino}-3-(phenyl-pyridin-

2-yl-amino)-propionic acid

6.04 mmol of the 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenyl- ρyridin-2-yl-amino)-propionic acid, methyl ester prepared essentially as described in patent application WO2005/113544, is dissolved in 70 mL of ethanol. 24.2 mL of 0.5 N aqueous ΝaOΗ is added and the mixture is stirred at room temperature for 2 h. After the reaction is complete, the pH is adjusted to ~5 using 1 N HCl. Water is added slowly and the resulting precipitate is filtered off and washed with water. After drying under reduced pressure of about 1 mbar at 40⁰C, 2.49 g of 2-{[2-(2-methylamino- pyrimidin-4-yl)-lH-indole-5-carbonyl]-arnino}-3-(phenyl-pyridin-2-yl-amino)-propionic acid is isolated. Empirical formula C₂₈H₂₅N₇O₃; M. W. = 507.56; MS (M+H) 508.3. ¹H NMR (DMSO-^₆) 2.95 (s, 3 H), 4.32-4.50 (m, 2 H), 4.65-4.72 (m, 1 H), 6.29-6.36 (d_? 1 H), 6.70- 6.79 (m, 1 H), 6.90-7.10 (sb, 1 H), 7.13-7.19 (m, 1 H), 7.22-7.38 (m, 4 H), 7.40-7.48 (m, 3 H), 7.50-7.55 (m, 1 H), 7.57-7.60 (m, 1 H), 7.96 (bs, 1 H), 8.34-8.40(m, 2 H), 8.80-8.90 (d, 1 H), 11.80 (s, 1 H) 12.8 (bs, IH). Chiral HPLC shows 94% ee.

Example 1, Step 2

Enantiomeric Purification of 2-{[2-(2-Methylaminopyrimidin-4-yl)-lH-indole-5-carbonyl]amino}-3-

(phenylpyridin-2-yl-amino)-propionic acid

2- { [2-(2-methylaminopyrimidin-4-yl)- lH-indole-5-carbonyl]amino} -3-(phenylpyridin-2-yl-amino)- propionic acid, prepared essentially according to Example 1, Step 1 above, is heated under reflux for 15 minutes. The insoluble racemic compound is removed by hot filtration. The TΗF of the resulting filtrate is removed by distillation and the residue is precipitated by the addition of isopropanol. After drying under reduced pressure of about 1 mbar at 40⁰C, the desired 2-{[2-(2-methylaminopyrimidin-4- yl)-lH-indole-5-carbonyl]amino}-3-(phenylpyridin-2-yl-amino)-propionic acid is isolated with an ee = 98.5%.

Example 1, Step 3

Synthesis of 2-{[2-(2-Methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenyl-pjτidin- 2-yl-amino)-propionic acid monopotassium monohydrate salt

To a slurry of 2-{[2-(2-methylaminopyrimidin-4-yl)-lH-indole-5-carbonyl]amino}-3-(phenylpyridin- 2-yl-amino)-propionic acid (50.8 mmol from Example 1, Step 2 above) in H₂O and EtOH is added 1.02 M KOH (2.00 equiv) with vigorous swirling. The mixture is heated to 67⁰C with swirling on a steam bath to dissolve the starting material, while braking up any remaining clumps. After several minutes the clear orange solution is filtered and the flask containing the filtrate is wrapped in aluminum foil and allowed to cool slowly to room temperature in the hot water remaining in the steam bath. After 17 hours, the mixture is cooled in an ice-bath and the salt is collected by filtration and washed 4 times with ice-cold H₂O. The last two washes have a pH of 8. The salt is dried in a vacuum oven at 45 ⁰C with an N₂ bleed to yield the desired compound as fine needles: ¹H NMR (DMSO-«k) 2.95 (s,3 H)₅ 3.95-4.05 (m, 1 H), 4.35-4.40 (m, IH), 4.55-4.62 (m, 1 H), 6.35-6.39 (d, 1 H), 6.58-6.60 (m, IH), 6.90-7.10 (sb, 1 H), 7.13-7.19 (m, 1 H), 7.22-7.38 (m, 6 H), 7.40-7.48 (m, 3 H), 7.57-7.60 (m,l H), 7.70 (s, 1 H), 8.10-8.15(d, 1 H), 8.30 (bs, 1 H), 11.80 (s, 1 H); LC-MS m/z 509 (M⁺ + 2), 508 (M⁺ H- I), 275, 254 (100). Anal. Calcd for C₂₈H₂₄KN₇O₃-H₂O (563.66): C, 59.67; H, 4.65; N, 17.39; K. 6.94; H₂O (Karl Fischer), 3.20. Found: C, 59.59; H, 4.66; N, 17.39; K₅ 6.44; H₂O (Karl Fischer), 3.16. Chiral HPLC showed 99.5% S-enantiomer.

Example 2 Synthesis of 2-{[2-(2-Methylammo-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenyl-pyridin-

2-yl-amino)-propionic acid monopotassium monohydrate salt

As an alternative procedure for preparing the compound of formula Ha₃ (3.8 mmol) of methyl ester 1 is dissolved in ethanol and water and 2 N aqueous KOH is added and the mixture is stirred at room temperature for 4 h. The product starts to crystallize and the mixture is diluted with additional water. The resulting crystalline precipitate is filtered off and washed with water. After drying under reduced pressure of about 1 mbar at 40⁰C, the monopotassium monohydrate salt π is isolated. Empirical formula C₂₈H₂₄KN₇O₃-H₂O M.W. = 563.65; MS (free acid, M+H) 508.3. ¹H ΝMR (DMSO-J₆) 2.95 (s, 3 H), 3.95-4.05 (m, 1 H), 4.35-4.40 (m, IH), 4.55-4.62 (m, 1 H), 6.35-6.39 (d, 1 H), 6.58-6.60 (m, 1 H), 6.90-7.10 (sb, 1 H), 7.13-7.19 (m, 1 H), 7.22-7.38 (m, 6 H), 7.40-7.48 (m, 3 H), 7.57-7.60 (m, 1 H), 7.70 (s, 1 H), 8.10-8.15(d, 1 H), 8.30 (bs, 1 H), 11.80 (s, 1 H). Water (Karl-Fischer): 3.2% (Monohydrate). XRPD (2 theta): 5.28, 6.45, 7.97, 9.46, 10.18, 10.93, 13.23, 13.66, 14.94, 15.94, 16.71, 18.15, 19.49, 20.38, 21.04, 21.42, 23.76, 24.38, 25.36, 25.71, 26.19, 27.13, 27.67, 28.13, 28.61, 29.12, 29.75, 30.95, 31.37, 32.94. ee: 99.8% (Chiralpak AD-H, 250 x 4.6mm, Heptane : EtOH : MeOH 5 : 1 : 1, RT).

IN VITRO TEST PROCEDURE IKK-EΝZYME ELISA

The assay buffer has the following composition (50 mM HEPES, 10 mM MgCl₂, 10 mM β- Glycerophosphate, 2 μM Microcystin-LR, 0,01% ΝP-40, 5 mM DTT).

The IKK enzyme preparation is diluted 1 :50 (in-house-made preparation) plus test compound in DMSO (final concentration in well: 2 %).

The assay procedures are as follows:

Incubation of enzyme and compound for 30 min; Addition of 1 mM ATP; pSer36-DcB Peptide (Substrate): 40 μM;

Incubation for 45 min and addition of anti- pSer32-pSer36-DcB Peptid-antibody;

Incubation for 45 min and transfer to protein-G-coated plate;

Incubation for 90 min followed by 3x washing; Addition of streptavidin-HRP, then incubation 45 min followed by 6x washing;

Addition of TMB and incubation for 15 min; and

Stop solution and read using photometer.

The results from the in vitro profiling are shown in Table I below. Table I

In the IKK-enzyme ELISA described above, at a 1 mM ATP, the compound 2-{[2-(2-methylamino- pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino} -3-(phenyl-pyridin-2-yl-amino)-propionic acid, free form (S-enantiomer), exhibits a 4 times greater binding affinity than 2-{[2-(2-methylamino-pyrimidin- 4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenyl-pyridin-2-yl-amino)-propionic acid, free form (R- enantiomer); and 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenyl- pyridin-2-yl-amino)-propionic acid monopotassium monohydrate salt (S-enantiomer) exhibits a 250 times greater binding affinity than 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]- amino}-3-(phenyl-pyridin-2-yl-amino)-propionic acid monopotassium monohydrate salt (R- enantiomer).

Physico-Chemical Characterization

The compound of formula Da is non-hygroscopic. A particular aspect of the compound of IIa is where it has an enantiomeric purity of greater than about 99% ee S-enantiomer. It has considerably increased aqueous solubility over the free form (See Table II below, which lists the solubilities in different solvents of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lΗ-indole-5-carbonyl]-amino}-3-(phenylpyridin-2- yl-amino)-propionic acid and its monopotassium monohydrate salt). It is a crystalline solid that is chemically and physically stable when exposed to heat (2 weeks, 80⁰C), light (500 watts/m2, 6 hrs), and humidity (2 weeks, 40°C/75% RH). It is chemically unstable under acidic conditions due to amide hydrolysis, and solutions are unstable to light (accelerated conditions, 500 watts/m2, 6 hrs) but under normal lab light conditions, solutions are stable for at least 2 weeks. The crystalline solid is chemically stable upon micronization with good reduction in particle size, which is a particularly useful component for inhalation dosage form development (suspension MDI, DPI). Respirable crystalline solid comprises about 90% of crystalline form α-1 wherein the particles are not more than about 7 microns in size. In particular, the particles are from about 2 to about 6 microns in size. Additionally, the median diameter of the particles is from about 1 to about 3 microns in size. In particular, the median diameter of the crystalline particles is about 1.5 microns in size. Respirable crystalline solid may form amorphous material upon micronization. That amorphous material may thereafter recrystallize to original crystalline form α-1.

Table π

K salt pH Free Form pH

Solubility (mg/ml) Solubility (mg/ml)

Water 5.9 8.4 0.2 6.1

10% Ethanol 8.7 8.2 0.2 6.4

20% Ethanol 10.0 8.0 0.3 6.0

Ethanol 1 .5 2.8

2-Propanol 0.4 1 .6

Acetone 0.2 4.7

Acetonitrile 0.2 2.2

Ethyl acetate 0.3 1 .9

0.2% Tween 80 /water 9.9 8.4 0.08 5.3

0.2% Tween 80 / 0.5%Methylcellulose 8.5 8.4 0.07 5.3

20% P G /water 12.2 8.0 0.16 4.8

0.9% Sodium Chloride 1 1.9 8.0

Furthermore, the crystalline solid has low solubility in propellants HFA227 and HFA 134a with and without 5% ethanol. Low solubility of a suspendant is a particularly useful factor in formulating MDI suspensions. Low solubility of the compounds of this invention in particular pharmaceutically acceptable propellants indicate that suspension MDI may be a feasible inhalation dosage form.

The results of a first solubility study of monopotassium monohydrate salt of 2-{[2-(2-methylamino- pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, in inhaler propellants is depicted in Table m below. The poorest solubility is found in the propellant composition of HFA 134a and 5% ethanol. Ethanol appears to act as a solvation inhibitor when combined with HFA 134a. This data supports the use of the monopotassium salt in a suspension MDI an inhalation dosage form.

Table m

The results of a second solubility study of monopotassium monohydrate salt of 2-{[2-(2-methylamino- pyrirnidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, in inhaler propellants is depicted in Table IV below, wherein the poorest solubility is now found in the propellant HFA227, wherein no ethanol is present. Such data also supports the use of the monopotassiura salt in a suspension MDI inhalation dosage form.

Table IV

Table V below shows the results of particle size distribution measurements (change in median diameter) following 14 days temperature cycling (0 to 40⁰C) storage. No dissolution and particle growth is evident in the HFA formulations following temperature cycling storage. Such data also supports the use of the monopotassium salt in a suspension MDI inhalation dosage form.

Table V

Run Target PEG600 PVP K30 HFA D₅₀

Product Propellant difference Strength between t₀ & (mg/act) t, (μm)

2 3 NO NO 227 +0.5

3 0.5 NO NO 134a +0.5

5 0.5 NO YES 227 +0.3

8 3 NO YES 134a +0.1

9 0.5 YES NO 227 +1

12 3 YES NO 134a -0.5

14 3 YES YES 227 -0.1

15 0.5 YES YES 134a 0

In addition to the meter dose inhaler methods described herein, Table VI shows results of the visual assessment of 2-{[2-(2-methylamino-pyrLtnidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-

(phenylpyridin-2-yl-amino)-propionic acid monopotassium monohydrate salt HFA suspension formulations in glass bottles. The results show both HFA227 and HFA134a form suspensions that are visually stable for periods between 7 and 89 seconds. The more stable suspensions are associated with HFA227 propellant indicating a better density match between the active and the propellant. All HFA suspensions are easily resuspended with HFA227 suspensions requiring only a single inversion. Table VI: Formulations and Experimental Results

to to

Feasibility data for delivering monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4- yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, in an Ultrahaler® DPI inhalation dosage form is confirmed. Such DPI dosage form is capable for delivery up to 5 mg API per actuation and the powder dispersion from lactose is demonstrated (Figures 9, 10, 11, 12 and 13).

Experimental

The following materials are useful for each of the appropriate formulations in the Experimentals below.

Monopotassium monohydrate salt of 2-{[2-(2-methylamino-pvrimidin-4-yl)-lH-indole-5-carbonyl]- amino}-3-(phenylpyridin-2-yl-amino)-propionic acid, micronized (Form α-1); α-lactose monohydrate;

Aerosol propellant HFA 227; Aerosol propellant HFA134a;

Polyethylene glycol 600, i.e., PEG600;

Povidone (PVP);

Lactose Monohydrate Respitose MLOOl, obtained from DMV International;

Lactose Monohydrate Respitose SV003, obtained from DMV International; Dry Powder Inhalers, as described herein;

Standard metered dose inhaler cans 19ml unlined, obtained from Presspart;

Plastic coated glass bottles; and

Metered dose inhaler valves (50μl and lOOμl), obtained from Bespak PIc.

Solvent Solubility Method (Results listed in Table II)

The solubilities of monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH- indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid in various aqueous and nonaqueous solvents are determined at 25 ⁰C. An excess amount of the drug (~15mg) is added to 1 mL of the various solvents. The material is shaken at 500 rpm for 3 days. The samples are centrifuged at 10000 rpm for 20 minutes, the pH of the solutions is recorded, and the drug concentrations are determined by HPLC.

Propellant Solubility Method (Results listed in Table m)

The solubilities of monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH- indole-5-carbonyl]-amino}-3-(phenylp3τidin-2-yl-amino)-propionic acid in HFA 227 and HFA 134a are measured at 25°C. Twenty milligrams of monopotassium monohydrate salt of 2-{[2-(2- methylamino-pyrimidin-4-yl)- 1 H-indole-5-carbonyl]-amino} -3 -(phenylpyridin-2-yl-amino)-propionic acid and ~0.8g of ethanol are added to polymer coated glass vials. A Valois DF 10/50 continuous valve is crimped to the container and ~15g of HFA propellant is added through the valve. The bottles are shaken at 200 rpm on a planetary mixer for 4 days. The suspensions are filtered (0.45 μm filter) into empty crimped bottles, and the propellants are evacuated by depressing the valve. The valves are removed using a pipe cutter and the bottles are allowed to dry in the hood overnight. The drug remaining in the bottles are dissolved in 1.5 mL mobile phase (70:30, acetonitrile : water) and the samples are assayed by HPLC.

Meter Dose Inhaler Method

Pre-weighed amounts of monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4-yl)- lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid and excipients (PEG600 & PVP as required) are added to cans or glass bottles. A metering valve is crimped to the container and a specified amount of HFA propellant is added through a metering valve. The quantities of monopotassium monohydrate salt of 2-{[2-(2-methylamino-p)τimidin-4-yl)-lH-indole-5-carbonyl]- amino}-3-(phenylpyridin-2-yl-amino)-propionic acid and excipients used in each formulation are shown in Table VI herein. Following addition of all the formulation ingredients the filled metered dose inhalers are mixed by shaking and sonication. The suspension characteristics of the bottles are visually assessed. The cans are assessed for single actuation particle size distribution by laser diffraction (Sympatec) before and after storage. Solubility in propellant is assessed by filtration of the suspension followed by quantitative analysis of dissolved monopotassium monohydrate salt of 2-{[2- (2-memylammo-pyrimidim-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)- propionic acid.

Dry Powder Inhaler Method

Two dry powder blends are manufactured with the formulations shown in Table VTl.

Table VH

Monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]- amino}-3-(phenylpyridin-2-yl-amino)-propionic acid Dry Powder Inhaler Formulations

For each formulation, the monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4-yl)- lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid is deaggregated by sieving and mixing with the specified amount of lactose to form a powder blend. Following confirmation of homogeneity of mix, the powder blend is filled into inhalers using Bespak inhaler filling equipment. The filled devices are tested for single actuation emitted weight (total weight of powder emitted), single actuation emitted dose (mass of active emitted) and single actuation aerodynamic particle size by cascade impaction. The weight of powder blend used, the weight of powder emitted per actuation and the mass of monopotassium monohydrate salt of 2-{[2-(2- methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-propionic acid emitted for each formulation is shown in Table VDI.

Table VTJI

Monopotassium monohydrate salt of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]- amino}-3-(phenylpyridin-2-yl-amino)-propionic acid Dry Powder Inhaler Formulations

Characterization Methods

Provided herein is an assortment of characterizing information to describe monopotassium salt forms of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl- amino)-propionic acid. It should be understood, however, that not all such information is required for one skilled in the art to determine that such particular form is present in a given composition, but that the determination of a particular form can be achieved using any portion of the characterizing information that one skilled in the art would recognize as sufficient for establishing the presence of a particular form, e.g., even a single distinguishing peak can be sufficient for one skilled in the art to appreciate that such particular form is present.

X-Ray Powder Diffractometry (XRPD) (Figures 1 and 2)

X-ray powder diffractometry is performed on a Siemens-Bruker D5000 diffractometer, using the parafocusing Bragg-Brentano (theta-two-theta)-type geometry. 2-{[2-(2-Methylamino-pyrimidin-4- yl)-lH-indole-5-carbonyl]-amino}-3-(phenyl-pyridin-2-yl-amino)-propionic acid monopotassium monohydrate salt, as a powder, is deposited on a single-crystal silicon wafer, cut according to the (510) crystallographic orientation. Copper K-alpha radiation (1.54056 angstroms), emitted from a copper anticathode tube (45kV/40mA) is used as the x-ray source, with Cu K-beta radiation filtered out using a reflected beam monochromator. A scintillation counter is used for detection. A divergence slit of 0.6mm, an anti-scatter slit of 0.6 mm, a monochromator slit of 0.1mm, and detector slit of 0.6 mm are used. The diffraction pattern is obtained using the following conditions: 2.0 to 40.0 degree scan in angle 2-tbeta, 1.0 second count time per step, 0.02 degree step size, under ambient conditions of pressure, temperature, and relative humidity.

Solvation/Hydration state by Thermal Gravimetric Analysis (Figure 3)

Thermal analysis of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3- (phenylpyridin-2-yI-amino)-propionic acid monopotassium monohydrate salt of formula H is performed using a TA Instruments Model Q-600 Simultaneous Differential Scanning

Calorimeter/Thermal Gravimetric Analyzer (DSC/TGA) under a dry nitrogen atmosphere. The TGA temperature is calibrated using an indium standard. The compound of formula II powder is transferred to an aluminum pan (TA Instruments part number 900793.901). The thermogram is acquired at a linear heating rate of 1O⁰C per minute. Dehydration of the compound of formula II begins at a temperature of approximately 100⁰C under these experimental conditions, with a total weight loss of approximately

3.0% from ambient room temperature to 200⁰C (theoretical 1 :1 hydrate would be 3.3%). Upon dehydration, an anhydrous crystalline form is produced, i.e., Form β-1, which then upon heating recrystallizes at about 22O⁰C into a second anhydrous crystalline form, i.e., Form β-2. The recrystallized material subsequently melts with decomposition at about 293⁰C. The XRPD of the two anhydrous crystalline forms are specific to the two forms.

Differential Scanning Calorimetry (Figure 4)

Differential Scanning Calorimetry (DSC) is performed using a TA Instruments Model Q- 1000 DSC equipped with a refrigerated cooling system under a dry nitrogen atmosphere. The DSC is calibrated using an indium standard. The compound of formula II powder is transferred to an aluminum pan, and a lid with laser-drilled pinhole (TA Instruments part numbers 900793.901 and 900860.901, respectively) is cold welded to the pan. The DSC thermogram is acquired at a linear heating rate of 1O⁰C per minute. Dehydration of the compound of formula II begins at a temperature of approximately 142⁰C under these experimental conditions. The thermograms show a retarded dehydration (because of the encapsulation) compared to the DSC/TGA, with the dehydration and recrystallization occurring as overlapping events. The melt of the anhydrous phase is observed at an onset temperature of approximately 291⁰C under these conditions. Micronization Photomicrographs (Figure 5)

Photomicrographs post milling, labeled as Milled; d50^~ 1.8 μm, show a well-micronized, crystalline material with no particles greater than 10 microns in length during a search of ~30 fields at 20Ox magnification. The particle size distribution is measured using a Sympatec HELOS-BF laser diffraction particle size analyzer with the R3 measurement lens, RODOS dry disperser, and laser tuned to 632.8 run. The system is calibrated using silicon carbide standards. The powder is dispersed using the RODOS dry dispersion attachment with a primary pressure of 3.0 bar and the depression is maximized. The volume based particle size distribution is calculated using the Fraunhofer method by the Sympatec Windox (Version 4.0) software. The particle size distribution of the micronized API is determined to be a single mode curve. The median [x(50)] is 1.8 microns and 90% of the particles are 3.6 microns or less. The micronization process reduces both the median (3.9 microns before) and the x(90) size (~9 microns before). Comparison of the XRPD, i.e., peak intensities, location (d-spacing) and resolution, of 2-{[2-(2-methylamino-pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenyl- pyridin-2-yl-amino)-propionic acid, monopotassium monohydrate salt, before and after micronization, are the same.

Dynamic Water Vapor Sorption (Figure 6)

The water sorption profile of the compound of formula II is determined using a SMS Instruments Dynamic Vapor Sorption Analyzer (DVS) Model DVS-I. Relative humidity (RH) and weight are calibrated using standards. The compound of formula II powder is loaded and dried at 0% RH for 4 hours prior to starting the experiment. The RH is stepped from 0.1 to 94.4% in 10 steps. The specimen weight is considered constant at each step when percent mass change is less than 0.001% over a 5- minute interval with a minimum absolute equilibration time of 15 minutes.

FT-IR Spectroscopy (Figure 7)

Fourier Transform-Infrared (FT-IR) spectra of the free form and the crystalline monopotassium monohydrate salt are obtained using a Nicolet Magna-IR Spectrometer 55 attached to Nicolet Nic-Plan FT-IR Microscope. A solid sample is analyzed on a KBr disk. Spectrum is obtained after 32 scans from 4000-400 cm^"1 with 4 cm^"1 resolution. A comparison of the ER. spectra of 2-{[2-(2-methylamino- pyrimidin-4-yl)-lH-indole-5-carbonyl]-amino}-3-(phenylpyridin-2-yl-amino)-ρropionic acid (free form) and 2- { [2-(2-methylamino-pyrimidin-4-yl)- 1 H-indole-5-carbonyl]-amino} -3-(phenylpyridin-2- yl-amino)-propionic acid monopotassium monohydrate salt H show two distinct patterns. Notable is the peak corresponding to the carbonyl group of the carboxylic acid which shifts to a lower wave number upon formation of the salt. Pharmaceutical Compositions and Methods

The pharmacological properties of the compound II are such that it is suitable for use in the treatment of all those patients suffering from or subject to conditions that can be ameliorated by the targeted administration of an inhibitor of IKB kinase to a site where the treatment is better effected by localized versus systemic activity, e.g., asthma, or chronic obstructive pulmonary disorder (COPD), among others noted above.

Topical application by inhalation, e.g., intratracheal, or intranasal, is a preferred mode for administering the compound according to the present invention.

A pharmaceutical composition according to the invention is preferably produced and administered in dosage units, with each unit containing, as the active constituent, a particular dose of the compound.

In practice, the compound of the present invention is administered in a suitable formulation to patients such that its activity is particularly localized. It will be appreciated that the preferred route can be varied depending on the site of the condition for which administration is directed.

Pharmaceutically acceptable dosage forms refers to dosage forms of the compound of the invention, and includes, for example, powders, suspensions, sprays, inhalants, tablets, emulsions, and solutions, particularly suitable for inhalation. Techniques and formulations generally may be found in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA, 20th edition.

If desired, and for more effective distribution, the compound can be microencapsulated in, or attached to, a slow release or targeted delivery systems such as biocompatible, biodegradable polymer matrices, e.g., poly (d^-lactide-co-glycolide), liposomes, and microspheres and subcutaneously or intramuscularly injected by a technique called subcutaneous or intramuscular depot to provide continuous slow release of the compound(s) for a period of 2 weeks or longer.

The compound may also be sterilized, for example, by filtration through a bacteria-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions, which can be dissolved in sterile water, or some other sterile medium immediately before use.

Formulations suitable for nasal or tracheal administration means formulations that are in a form suitable to be administered nasally or by inhalation to a patient. The formulation may contain a carrier, in a powder form, having a particle size for example in the range 1 to 500 microns (including particle sizes in a range between 20 and 500 microns in increments of 5 microns such as 30 microns, 35 microns, etc.). Suitable formulations wherein the carrier is a liquid, for administration as for example a nasal spray or as nasal drops, include aqueous or oily solutions of the active ingredient. Formulations suitable for aerosol administration may be prepared according to conventional methods and may be delivered with other therapeutic agents. Metered dose inhalers readily administer inhalation therapy.

Actual dosage levels of active ingredient(s) in the compositions of the invention may be varied so as to obtain an amount of active ingredient(s) that is (are) effective to obtain a desired therapeutic response for a particular composition and method of administration for a patient. A selected dosage level for any particular patient therefore depends upon a variety of factors including the desired therapeutic effect, on the route of administration, on the desired duration of treatment, the etiology and severity of the disease, the patient's condition, weight, sex, diet and age, the type and potency of each active ingredient, rates of absorption, metabolism and/or excretion and other factors.

Total daily dose of the compounds of this invention administered to a patient in single or divided doses to about 1000 mg, more particularly from about 50 mg to 300 mg, and, further particularly from about 10 mg to 100 mg. However, higher or lower daily doses can also be appropriate. The daily dose can be administered either by means of a once-only administration in the form of a single dosage unit, or of several smaller dosage units, or by means of the multiple administration of subdivided doses at predetermined intervals. The percentage of active ingredient in a composition may be varied, though it should constitute a proportion such that a suitable dosage shall be obtained. Obviously, several unit dosage forms can be administered at about the same time. A dosage may be administered as frequently as necessary in order to obtain the desired therapeutic effect. Some patients may respond rapidly to a higher or lower dose and may find much weaker maintenance doses adequate. For other patients, it may be necessary to have long-term treatments at the rate of 1 to 4 doses per day, in accordance with the physiological requirements of each particular patient. It goes without saying that, for other patients, it will be necessary to prescribe not more than one or two doses per day.

The formulations can be prepared in unit dosage form by any of the methods well known in the art of pharmacy. Such methods include the step of bringing into association the active ingredient with the carrier that constitutes one or more accessory ingredients. In general the formulations are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product.

Claims

We Claim:

1. A compound of formula II

π or a solvate thereof.

2. The compound of formula II according to claim 1 as anhydrous crystalline form β-1.

3. The compound of formula II according to claim 1 as anhydrous crystalline form β-2.

4. The compound of formula II according to claim 1 in the solvate form.

5. The compound of formula II according to claim 4 wherein the solvate form is a hydrate.

6. The compound of formula II according to claim 5 wherein the hydrate is a monohydrate.

7. The compound of formula II according to claim 6 as crystalline form α-1.

8. A pharmaceutical composition comprising a pharmaceutically effective amount of a compound according to any one of claims 1, 2, 3, 4, 5, 6 or 7 in admixture with a pharmaceutically acceptable carrier.

9. A method for treating a patient suffering from, or subject to, a pathological condition capable of being ameliorated by inhibiting DCK-2 comprising administering to said patient a pharmaceutically effective amount of the compound according to any one of claims 1, 2, 3, 4, 5, 6 or 7.

10. The method according to claim 9 wherein the pathological condition is asthma, rhinitis, chronic obstructive pulmonary disorder or chronic obstructive pulmonary disorder exacerbations.

11. A method for treating a patient suffering from asthma, comprising administering to the patient a pharmaceutically effective amount of the compound according to any one of claims 1, 2, 3, 4, 5, 6 or 7.

12. A method for treating a patient suffering from rhinitis, comprising administering to the patient a pharmaceutically effective amount of the compound according to any one of claims 1, 2, 3, 4, 5, 6 or 7.

13. A method for treating a patient suffering from chronic obstructive pulmonary disorder, comprising administering to the patient a pharmaceutically effective amount of the compound according to any one of claims 1, 2, 3, 4, 5, 6 or 7.

14. A method for treating a patient suffering from chronic obstructive pulmonary disorder exacerbations, comprising administering to the patient a pharmaceutically effective amount of the compound according to any one of claims 1, 2, 3, 4, 5, 6 or 7.

15. The method according to claim 9 wherein the administering is local.

16. The method according to claim 9 wherein the administering is intratracheal.

17. The method according to claim 9 wherein the administering is intratracheal, intranasal, inhalational, or by aerosolization administration.

18. The pharmaceutical composition according to claim 8 further comprising a pharmaceutically effective amount of a compound selected from the group consisting of a bronchodilator, a long-acting beta-2 agonist, an anticholinergic agents, a methylxanthine and an anti-inflammatory therapy, in admixture with a pharmaceutically acceptable carrier.

19. The pharmaceutical composition according to claim 18, wherein the bronchodilator is a short- acting beta 2-agonist; the long-acting beta 2-agonist is selected from salmeterol and formoterol; the anticholinergic agent is selected from ipratropium bromide and tiotropium bromide; the methylxanthine is theophylline; and the anti-inflammatory therapy is selected from inhibitors of cellular recruitment and toxic inflammatory mediators, inhibitors of proteolytic enzymes, antioxidants, inhibitors of mucus production and antibiotic therapy.

20. The method for treating according to claim 9 further comprising administering a pharmaceutically effective amount of a compound selected from the group consisting of a bronchodilator, a long-acting beta-2 agonist, an anticholinergic agents, a methylxanthine and an antiinflammatory therapy, in admixture with a pharmaceutically acceptable carrier.

21. The method for treating according to claim 20 wherein the bronchodilator is a short-acting beta 2-agonist; the long-acting beta 2-agonist is selected from salmeterol and formoterol; the anticholinergic agent is selected from ipratropium bromide and tiotropium bromide; the methylxanthine is theophylline; and the anti-inflammatory therapy is selected from inhibitors of cellular recrutiment and toxic inflammatory mediators, inhibitors of proteolytic enzymes, antioxidants, inhibitors of mucus production and antibiotic therapy.

22. A stable pharmaceutical product comprising a dry powder inhaler and respirable crystalline form α-1.

23. A stable pharmaceutical product comprising a dry powder inhaler and respirable crystalline form β-1.

24. A stable pharmaceutical product comprising a dry powder inhaler and respirable crystalline form β-2.

25. A stable pharmaceutical product comprising a metered dose inhaler and respirable crystalline form α-1.

26. A stable pharmaceutical product comprising a metered dose inhaler and respirable crystalline form β-1.

27. A stable pharmaceutical product comprising a metered dose inhaler and respirable crystalline form β-2.

28. The stable pharmaceutical product of claim 22, wherein particles of crystalline form α-1 have a median diameter from about 1 to about 3 microns.

29. The stable pharmaceutical product of claim 23, wherein particles of crystalline form β-1 have a median diameter from about 1 to about 3 microns.

30. The stable pharmaceutical product of claim 24, wherein particles of crystalline form β-2 have a median diameter from about 1 to about 3 microns.

31. The stable pharmaceutical product of claim 28, wherein particles of crystalline form α-1 have a median diameter of about 1.5 microns.

32. The stable pharmaceutical product of claim 29, wherein particles of crystalline form β-1 have a median diameter of about 1.5 microns.

33. The stable pharmaceutical product of claim 30, wherein particles of crystalline form β-2 have a median diameter of about 1.5 microns.

34. The product according to any one of claims 25, 26 or 27, wherein about 90% of the particles are not more than about 7 microns in size.

35. The product according to claim 34 wherein about 90% of the particles are from about 2 to about 6 microns in size.

36. The crystalline form according to claim 7 which upon micronization contains less than about 10% of amorphous product upon micronization.

37. The crystalline form according to claim 7 which upon micronization contains less than about 7% of amorphous product upon micronization.

38. The crystalline form according to claim 7 which upon micronization contains less than about 5% of amorphous product upon micronization.

39. A preparation of a medicament used for treating a patient suffering from, or subject to, a pathological condition capable of being ameliorated by inhibiting DCK-2 comprising administering to the patient a pharmaceutically effective amount of a compound according to any one of claims 1 to 7.