WO2009002746A1

WO2009002746A1 - Dosing schedules of leukotriene synthesis inhibitors for human therapy

Info

Publication number: WO2009002746A1
Application number: PCT/US2008/067103
Authority: WO
Inventors: David J. Hermann; Daniel L. Hartman; Peter James Van Ess; Wenping Wang
Original assignee: Decode Genetics Ehf.
Priority date: 2007-06-22
Filing date: 2008-06-16
Publication date: 2008-12-31

Abstract

The invention relates to improved materials and methods for therapy to inhibit production of leukotrienes, and all therapeutic applications thereof.

Description

DOSING SCHEDULES OF LEUKOTRIENE SYNTHESIS INHIBITORS FOR

HUMAN THERAPY

This application claims priority to U.S. Provisional Patent Application No. 60/945,871, filed June 22, 20007, which is incorporated herein by reference in its entirety.

FIELD OF INVENTION

The invention relates to improved dosing materials and methods for therapy to inhibit production of leukotrienes, and all therapeutic applications thereof.

BACKGROUND The present invention pertains to novel dose or dosing regimen to improve efficacy and/or safety of a class of drugs.

Typical concentration-response relationship

Most drugs exert a direct, reversible effect which is often mediated through binding with a specific cellular receptor. In theory, the number of drug molecule-occupied receptors is proportional to the magnitude of clinical effects. For these drugs, there typically is a direct relationship between the time course of drug concentrations and pharmacologic effect. An EMAX model (or its extension) proves to be a useful and concise description of the concentration-effect relationship for these classes of compounds. Assuming that drug binding to the receptor is reversible and the extent of pharmacologic response (E) is proportional to the concentration of receptors that are occupied, an EMAX model can be derived by applying the law of mass action, a principle widely recognized since the early twentieth century. An EMAX model takes the form as follows:

F C

E = - ^™ EC₅₀ + C

Where E_max is the maximal response, C is the concentration of a drug in the receptor "compartment," and EC50 is the concentration of a drug at which 50% of the maximum response is achieved.

If n drug molecules bind to each receptor site, an EMAX model can be modified to cope with the situation taking the form of a sigmoidal EMAX model: E. - C"

E =

EC₅"₀ + C"

A corollary of a drug that follows an EMAX or a sigmoidal EMAX is that a higher concentration of the drug leads to a higher clinical effect.

Chronobiology and Chronotherapy/Chronopharmacology Chronobiology is the study of biological rhythms and the biochemical mechanisms that underlie them. Circadian rhythms operate on a periodicity of about 24 hours (one day). A number of clinically relevant conditions are known to vary with time of day or with the daytime-active/nighttime-rest patterns of a typical diurnal schedule. For example, it has been known for hundreds of years that asthma is worse at night for many subjects. Blood pressure is highest during the day, known to exhibit a nocturnal decline, the extent of which may vary in different subjects, and then rise between 0400 h to 1200 h. The extent of the nocturnal decline has been correlated with cardiovascular injury and risk. Myocardial infarctions, on the other hand, occur most often in the early morning. There is emerging evidence on the molecular level of circadian rhythms too, e.g., involving hormones and other molecules. Kraft, Chronobiology Int'l, 16(5): 683-93 (1999) have reported that leukotrienes in nocturnal asthma patients, as measured in urine or bronchoalviolar lavage, exhibit circadian variation, and also hypothesized that the circadian pattern of Cortisol expression might contribute to the circadian pattern of asthma. Chronotherapy may be related to chronobiology and pertains to timing of therapeutic intervention to maximize efficacy and/or minimize toxicity. Skloot reported that leukotriene -modifying agents used for asthma are reportedly administered at bedtime with chronotherapeutic strategies in mind. See Skloot, "Nocturnal Asthma: Mechanisms and Management," Mount Sinai J. Med. 69(3): 140 (2002). However, many such agents are administered multiple times during the day. Bisgaard, Pediatrics 107(2): 381 (2001). Hermida et al, Hypertension, 46(part 2): 1060 (2005) reported that aspirin administered at bedtime caused a highly significant blood pressure reduction in patients.

Leukotriene Pathway Leukotrienes, which are formed in vivo from an arachidonic acid acid metabolism pathway, are potent inflammamtory lipid mediators and have been implicated in a variety of diseases processes, including but not limited to asthma, atherosclerosis, Chronic Obstructive Pulmonary Disease (COPD), and inflammatory bowel disease (IBD). They can potentially contribute to development of atherosclerosis and destabilization of atherosclerotic plaques through lipid oxidation or other pro-inflammatory effects. Leukotriene C4 (LTC4), leukotriene D4 (LTD4), and leukotriene E4 (LTE4), are known to induce vasoconstriction. Allen et al. , Circulation, 97:2406-2413 (1998) described a novel mechanism in which atherosclerosis is associated with the appearance of a leukotriene receptor(s) capable of inducing hyperactivity of human epicardial coronary arteries in response to LTC4 and LTD4. LTB4, on the other hand, is a strong pro-inflammatory agent.

Emerging evidence from genetic studies by deCODE genetics indicates that certain haplotypes for genes involved in leukotriene synthesis (FLAP and LT A4 Hydrolase) correlate with increased risk of myocardial infarction and/or stroke in humans, and the predictive haplotypes have been found to correlate with increased LTB4 production. Variants of the 5 -lipoxygenase (5-LO) Activating Protein (FLAP) gene and the leukotriene A4 hydrolase (LT A4H) gene have been associated with elevated risk for myocardial infarction in Icelandic, British and North American populations, as described in PCT Application Nos. PCT/US03/32805, PCT/US03/32556, PCT/US04/030582 and PCT/US05/03312. Leukotriene inhibitors and FLAP inhibitors specifically, have been tested for a variety of indications

(including those listed above. The results have been mixed, sometimes demonstrating an unfavorable risk (e.g., from liver function tests (LFT) elevation and/or skin lesions) and/or lack of or insufficient efficacy to continue development.

A need exists to develop improved anti-leukotriene therapies for treatment or prophylaxis of the inflammatory conditions to which leukotrienes contribute. Previous studies conducted by deCODE genetics with a leukotriene inhibitor known as DG-031 (or BAY x 1005) demonstrated a somewhat linear dose response when LTB4 production was measured at trough levels of DG-031. Data generated by deCODE has indicated that sustained-release or multi-dose regimens for DG-031 , to maintain a high steady state concentration of the drug (e.g., to maintain an elevated Cmin or a low Cmax/Cmin ratio, relative to previously studied regimen) was effective at reducing inflammatory markers. See International Application No. PCT/US2006/015542 (published as WO 2006/116349). SUMMARY OF INVENTION

The invention provides materials and methods to achieve steady state plasma concentration of a leukotriene synthesis inhibitor, such as DG-031, in a human, wherein these steady-state concentration ranges exhibit improved effects and reduce drug exposure, thereby achieving maximum efficiency and reducing the possibility of short term or long term drug side-effects. To achieve a steady-state plasma concentration pharmacokinetic profile of the leukotriene synthesis inhibitor, the invention provides for doses and dosing schedules that are effective to attain this effect. Numerous compounds are described below in the compound section, and each represents an embodiment of the invention. For brevity, the invention is described in the context of one of the preferred embodiments, DG-031.

One embodiment of the invention is a method of reducing leukotriene production in a human, the method comprising administering to a human a composition that comprises a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof, wherein the composition is administered according to a dosing regimen that achieves a single daily peak plasma concentration of the inhibitor between the hours of 6 A.M. (0600 h) and 2 P.M (1400 h). These methods include administering the composition once daily and may be administered in the morning.

Adminstration in the "morning" refers to administering a composition and dose between the the before twelve noon. For example, administration in the morning is between the hours of 1A.M. and and twelve noon, such as between 1 A.M. and 2 A.M., between 1 A.M. and 3 A.M., between 1 A.M. and 4 A.M., between 1

A.M. and 5 A.M., between 1 A.M. and 6 A.M., between 1 A.M. and 7 A.M., between 1 A.M. and 8 A.M., between 1 A.M. and 9 A.M., between 1 A.M. and 10 A.M., between 1 A.M. and 11 A.M., 2 A.M and 3 A.M., between 2 A.M. and 4 A.M., between 2 A.M. and 5 A.M., between 2 A.M. and 6 A.M., between 2 A.M. and 7, between 2 A.M. and 8 A.M., between 2 A.M. and 9 A.M., between 2 A.M. and 10 A.M., between 2 A.M. and 11 A.M., 2 A.M. and twelve noon, between 3 A.M. and 4 A.M., between 3 A.M. and 5 A.M., between 3 A.M. and 6 A.M., between 3 A.M. and 7, between 3 A.M. and 8 A.M., between 3 A.M. and 9 A.M., between 3 A.M. and 10 A.M., between 3 A.M. and 11 A.M., 3 A.M. and 12 noon, between 4 A.M. and 5 A.M., between 4 A.M. and 6 A.M., between 4 A.M. and7, between 4 A.M. and 8 A.M., between 4 A.M. and 9 A.M., between 4 A.M. and 10 A.M., between 4 A.M. andl 1 6 A.M., between 4 A.M. and twelve noon, between 5 A.M. and 6 A.M., between 5 A.M. and 7 A.M., between 5 A.M. and 8 A.M., between 5 A.M. and 9 A.M., between 5 A.M. andlO A.M., between 5 A.M. and 11 A.M., between 5 A.M. and 12 noon, between 6 A.M. and 7 A.M., between 6 A.M. and 8 A.M., between 6 A.M. and 9 A.M., between 6 A.M. and 10 A.M., between 6 A.M. and 11 A.M., between 6 A.M. and twelve noon, between7 A.M. and 8 A.M., between7 A.M. and 9 A.M., between7 A.M. and 10 A.M., between7 A.M. and 11 A.M., between7 A.M. and twelve noon, between 8 A.M. and 9 A.M., between 10 A.M. and 11 A.M., between 10 A.M. and twelve noon, 11 A.M. and twelve noon.

Another embodiment of the invention is a use of a leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, in the manufacture of a medicament for administration to a human for reducing leukotriene synthesis production in said human, wherein the medicament is formulated into a dose that is administered according to a dosing regimen that is effective to achieve a single daily peak plasma concentration of the inhibitor between the hours of 6 A.M.(0600 h) and 2 P.M. (1400 h). These methods include administering the composition once daily and may be administered in the morning.

In a further embodiment, the invention provides for a leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, for reducing leukotriene synthesis in a human, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration according to a dosing regimen that is effective to achieve a single daily peak plasma concentration of the inhibitor between the hours of 6 A.M. (0600 h) and 2 P.M. (1400 h). The does of this leukotriene synthesis inhibitor may be administered in the morning.

More preferably, the peak plasma concentration occurs during the day during a time range that begins at 6:30 A.M., 7, 7:30, 8, 8:30, 9, 9:30, or 10 A.M.; and that ends at a later time during the day selected from 10 A.M., 10:30, 11, 11 :30 A.M., 12 noon, 12:30 P.M., 1, 1 :30, and 2 P.M. To provide just a few examples of the many permutations encompassed by these values, in one variation, the peak plasma concentration occurs during the day between the hours of 8 A.M. and 2 P.M.; or between the hours of 9 A.M. and 2 P.M.

A single daily peak refers to a single maxima on a plot of concentration versus time in the course of a day. A single daily peak is readily achieved with a single daily dose.

In other variations of the invention, the dosing regimen is chosen to achieve a peak plasma concentration within a certain range of time after administration of the composition that comprises the leukotriene synthesis inhibitor. For example, in some variations of the invention, the composition is formulated to cause a peak plasma concentration of the leukotriene synthesis inhibitor to occur within 7 hours of administering the composition; or within 6.5 or 6 hours of administering the composition; or within 5.5 or 5 hours of administering the composition; or within 4.5 or 4 hours of administering the composition; or within 3.5 or 3 hours of administering the composition. The preferred time delay for achieving peak concentration also can be expressed as a range, e.g., at least 1 or 1.5 or 2 or 2.5 hours after dosing, up until one of the aforementioned time limits. To provide just one example of the many permutations contemplated by these values, the composition is formulated to cause a peak plasma concentration 2-4 hours after administration.

In the context of the invention, the term "dose" refers to a quantity of a therapeutic agent to be administered at one time, and the terms "dosing schedule" or "dosing regimen" describe the time course and frequency during which doses of a therapeutic agent are administered to a human or animal subject for therapeutic or prophylactic purposes. For example, doses of 1000 mg of therapeutic agent might be administered on a two times per day dosing schedule, which would normally be administered approximately every twelve hours. Alternatively, doses of 500 mg of therapeutic agent might be administered on a three times per day dosing schedule, which would normally be administered approximately every eight hours. The invention provides dosing schedules which effectively achieve and maintain a steady state concentration of a leukotriene inhibitor such as DG-031 , wherein the steady state concentration exhibits a desired pharmacokinetic profile and attains a desired therapeutic effect in a human and reduces the potential for adverse events. The invention also provides for methods of administering doses of DG-031 to a human according to a dosing schedule of the invention in order to attain the desired therapeutic effect in said human. Preferred embodiments involve doses and dosing schedules that are convenient to patients, e.g., with fewer daily doses.

A "steady state concentration" in a human subject receiving treatment is a concentration of therapeutic agent that is at a dynamic equilibrium, fluctuating periodically within a reasonably predictable and periodic range with the fluctuation determined by the dosing schedule. The invention provides for dosing schedules of DG-031 that attain a dynamic equilibrium of DG-031 within a desired range in the plasma of the individual receiving the doses according to the dosing schedule. Of particular importance to the present invention is the timing during the day, and the magnitude, of the steady state peak concentration that is achieved in the patient after repeated dosing for several days, according to dosing regimens of the invention.

The term "peak concentration," also referred to as "Cmax," refers to the maximum concentration achieved in the steady state dynamic equilibrium, which can be visualized as the top of a peak or maxima on a graph of plasma concentration plotted against time. With oral or other bolus dosing, the peak concentration usually occurs some time between doses, with the time depending on the route of administration and formulation. The "trough concentration," also referred to as "Cmin," refers to the minimum concentration achieved in the steady state dynamic equilibrium, which can be visualized as the minima on a graph of plasma concentration plotted over time. With oral or other bolus dosing, the trough concentration following a dose usually is observed at a time corresponding to immediately before administration of a new/next dose.

In the context of treatment of a single individual, a steady state or peak concentration may be expected to fluctuate from day-to-day with variations in the individual's diet, level activity, state of health, co-administration of other medications, and the like. For example, when the leukotriene synthesis inhibitor DG-031 is administered with food, adsorption is increased. For instance, administration with a high fat meal caused a 150% increase in C_max, 30% increase in AUC, a shorter half- life and no effect on T_max. The present invention can be practiced by administering DG-031 with food or, alternatively, DG-031 is administered in the absence of food. The decision whether or not to administer with food may be based on the Cmax and Cmin observed following administration of a novel formulation where the Cmax and Cmin fall within the desired range. Many indications for the therapies described herein, such as prophylaxis for myocardial infarction, benefit from repeat dosing for weeks or months or years. In this context, the steady state concentration for an individual refers to an average concentration taken at multiple time points, to adjust for such fluctuation. In the context of a treatment regimen, a steady state concentration refers to an average or mean (preferably, a geometric mean) dynamic equilibrium obtained from observations of a statistically representative number of individuals, taking into account factors such as sex, weight, race and age. Likewise, in the context of evaluating the properties of a particular formulation or dose or dosing regimen, a steady state concentration refers to an average or mean (preferably, a geometric mean) dynamic equilibrium obtained from observations of a statistically representative number of individuals, taking into account the same aforementioned factors.

In a preferred embodiment, the doses of the methods and uses of the invention are formulated for oral administration and are administered orally. Preferably, the invention provides for dosing schedules that are no more than three times a day and more preferably no more than two times per day. Single daily dosing is highly preferred. Morning dosing is a preferred dosing time, and conventionally occurs a short time after awaking in diurnal individuals that are active during the daytime and rest during nighttime hours. However, other dosing times are suitable, depending on formulation and pharmacokinetics, to achieve the desired peak concentration during the desired time window during the day.

Measurements of plasma DG-031 can be carried out by methods and uses that are standard in the art. In a preferred method of measuring plasma DG-031 , plasma is separated from the blood samples from the treated patients and the protein in the plasma is precipitated by an organic solvent such as acetonitrile. The DG-031 is then measured by liquid chromatography and/or mass spectrometry and the measured concentration is compared to a standard curve.

The term "treating" refers to providing any measure of therapeutic benefit, such as reduction of symptoms, measurable improvement in therapeutically meaningful biological molecules, slowing of deterioration, or curing. The term "preventing" refers to any measurable preventative/prophylactic benefit. For example, effective prevention can be measured in an individual by a slowing or elimination of deterioration or a delay in an expected adverse event. Prevention is often more readily demonstrated in a clinical setting or population study that demonstrates that a population of individuals that receive a therapy suffer fewer adverse events, or survive longer, or suffer less severe adverse events, or enjoy any other benefit as a group that a physician would characterize as prophylactic or beneficial.

Preferred embodiments of the invention exhibit a treatment or preventive effect with respect to an inflammatory condition or disease such as asthma, IBD, COPD, MI, or stroke. Highly preferred embodiments exhibit a greater therapeutic or prophylactic effect than conventional dosing regimes for the same drug. Similarly, reduction in leukotriene production, which is reasonably expected to correlate with a therapeutic benefit for numerous inflammatory conditions, is readily demonstrated in a clinical setting or population study that demonstrates that a population of individuals that receive a therapy exhibit a statistically significant reduction in leukotriene production (p < 0.05) from the dosing according to the invention.

The effect of a treatment regimen on leukotriene production is preferably assessed by taking a biological sample from the human subject at the time when leukotriene production is at a daily peak. In some variations of the invention, a leukotriene such as LTB4 is measured directly in a biological sample such as serum or plasma. In other variations, whole blood (or at least blood containing leukocytes) is first stimulated with a calcium ionophore or other agent to stimulate release of leukotrienes, and then a leukotriene such as LTB4 is measured in the serum or plasma. An evening peak has been observed for LTB4 production when measured in this manner in human subjects. A preferred time point at which to assess efficacy is a time point approximating an evening peak of LTB4 production.

In some variations of methods and uses and leukotriene synthesis inhibitors of the invention, the doses and dosing schedule are effective to achieve a reduction of leukotriene B4 (LTB4) or leukotriene A4 (LTA4)) of at least 20% within one week of commencing administration, and maintain said reduction with continued administration of doses according to the dosing schedule. Greater reductions (e.g., 25%, 30%, 35%, 40%, 45%, 50%, or more, and faster reductions, e.g., within 5 days or 4 days or 3 days, is preferred. Another criteria for measuring successful reduction, in human subjects with an inflammatory condition, is achievement of a reduction to the baseline LTB4 level in control patients that do not have the inflammatory condition, e.g., 30% to 35% reduction. In some variations, measurements of stimulated LTB4 production obtained by treating a whole blood sample (or sample containing leukocytes) with a calcium ionophore, such as ionomycin, are utilized for monitoring efficacy. In addition, LTB4 may be measured in serum or plasma.

In another embodiment, the invention provides for any of the preceding methods or uses to further comprise a step of measuring at least one inflammatory marker in a sample from the human to monitor efficacy of the therapy, wherein a reduction in the inflammatory marker compared to pre-treatment levels is indicative of efficacy. The invention provides for measuring the at least one inflammatory marker at least annually during treatment. The invention also provides for measuring the at least one inflammatory marker within 45 days of beginning the administering. The invention contemplates that the at least one inflammatory marker is a MPO or a leukotriene, such as LTB4.

The term "prodrug" refers to a chemical entity that is metabolized in vivo into an active drug such as DG-031 , such entities being designable and identifiable by pharmaceutical chemists.

In a related embodiment, the invention is a method of reducing leukotriene production in a human comprising administering to the human a composition that comprises a leukotriene synthesis inhibitor, wherein the composition is administered once daily in the morning, and wherein the leukotriene synthesis inhibitor is present in the composition in an amount effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 20%. In addition, the invention provides for use of a leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, in the manufacture of a medicament for administration to a human for reducing leukotriene production in said human, wherein the medicament is formulated into a dose that is administered once daily in the morning, and the dose is effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 20%. In a further embodiment, the invention provides for a leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, for reducing leukotriene production in a human, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration once daily in the morning, and the dose is effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 20%.

Greater reductions, e.g., of 25%, 30%, 35%, 40%, or more are preferred. Exemplary reduction goals also can be expressed as a range, e.g., with a reduction of at least 20, 25, 30, or 35% up to a higher value of 35, 40, 45, 50, 55, 60, or 65%. To provide just a few examples of the permutations contemplated using these numbers, reductions of 20-60%, or 25-55% or 30-50% are contemplated. Although the therapeutic agent is a leukotriene inhibitor, the efficacy of the invention alternatively can be demonstrated by measuring reduction in other markers of inflammation, as described herein.

The invention also provides for methods of reducing leukotriene production in a human comprising administering a composition to the human in single daily dose in the morning, wherein the composition comprises 500-2500 mg of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof. In another embodiment, the invention provides for uses of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof, in the manufacture of a medicament for administration to a human for reducing leukotriene production in said human, wherein the medicament is formulated into a does that is administered once daily in the morning, and wherein the dose is in a range of 500-2500 mg of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof. In addition, the invention provides for leukotriene synthesis inhibitors or a pharmaceutically acceptable salt, ester, or pro-drug thereof, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration once daily in the morning, and wherein the dose is in a range of 500- 2500 mg of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof.

The invention described herein may be practiced with compounds that inhibit any one or more of the enzymes involved in leukotriene biosynthesis. Exemplary agents are inhibitors of at least one enzyme selected from the group consisting of 5 -lipoxygenase (5-LO), 5 -lipoxygenase activating protein (FLAP), Leukotriene A4 hydrolase (LTA4H), Leukotriene C4 synthase, gamma- glutamyltranspeptidase, and leukotriene D4 dipeptidase. Compounds that inhibit 5- LO, FLAP, or LTA4H are highly preferred. Preferred compounds include any of the leukotriene synthesis compounds that are in clinical trials and/or have been approved for one or therapeutic indications, such as asthma. Some preferred compounds are listed in Tables 1, 2and 3 in the detailed description below.

A genus of highly preferred compounds for practice of the invention include compounds of the formula:

wherein Ar is selected from the group consisting of: aryl; heteroaryl; aryl substituted with from one to three substituents independently selected from the group consisting of halogen, loweralkyl, loweracyl, loweralkoxy, fluoro loweralkyl, fluoroloweralkoxy, hydroxy, hydroxy(Ci-C4) alkyl, formyl, formyl(Ci-C4) alkyl, cyano, cyano(Ci-C4) alkyl, benzyl, benzyloxy, phenyl, substituted phenyl, heteroaryl, heterocyclylalkyl, substituted heteroaryl, and nitro; and heteroaryl substituted with from one to three substituents independently selected from the group consisting of halogen, loweralkyl, loweracyl, loweralkoxy, fluoroloweralkyl, fluoroloweralkoxy, formyl, cyano, benzyl, benzyloxy, phenyl, heteroaryl, heterocyclylalkyl and nitro; wherein X is selected from the group consisting of direct bond, O, SO,

S(O₂), NR¹, CH₂, CF₂, CH₂CH₂, CH₂NR¹, NR¹CH₂, CH=CH, C=O, CH₂C=O, _CR ^ia _R ^ib oCR^laR^lb , CR^laR^lbO; SO₂NR¹^R¹SO₂, C(O)NR¹ and NR¹C(O); wherein R¹ is selected separately in each occurrence from the group consisting of H and lower alkyl; R^la is selected from the group consisting of H, OH and lower alkyl; R^lb is selected from the group consisting of H and lower alkyl, or R^la and R^lb taken together may form a 3-6 membered ring, which may optionally contain a heteroatom chosen from O, S, and N; wherein HetAr is an aryl or heteroaryl ring attached via a ring carbon to Q, further characterized in that Q and X cannot be on adjacent positions in said aryl or heteroaryl ring;

Q is chosen from -O-, -NR¹- and S(O)_P;

Q and X cannot be on adjacent positions in said benzene or pyridine ring; p is zero, 1 or 2; n is an integer selected from 1-5;

HET is selected from the group consisting of

4-7-membered saturated nitrogenous heterocycle and

4-7-membered saturated nitrogenous heterocycle substituted with one or two substituents independently selected from the group consisting of halogen, hydroxyl, amino, carboxy, loweralkyl, loweracyl, loweralkoxy, N-oxide, fluoroloweralkyl, fluoroloweralkoxy, formyl, cyano, benzyl, benzyloxy, phenyl, heteroaryl and nitro; and taken together ZW is H or Z is (CH₂)I-Io, in which one or two (CH₂) may optionally be replaced by -O-, -NR¹-, -SO-, -S(O)₂-, -C(O)- or -C=O(NH)-, provided that said -0-, -NR¹-, -SO-, -S(O)₂-, -C(=0)- or -C=O(NH)- are not at the point of attachment to HET and are separated by at least one -(CH₂)-;

W is selected from the group consisting of acyl, hydroxyl, carboxyl, amino, -C(O)NHR⁴, aminoacyl, -COOalkyl, -CHO, heterocyclyl, substituted aryl, substituted heterocyclyl, sulfonamide, -C(O)fiuoroalkyl, -C(O)CH₂C(O)Oalkyl, -C(O)CH₂C(O)Ofluoroalkyl, -SH, -C(O)NH(OH), -C(O)N(OH)R⁴, -N(OH)C(O)OH, - N(OH)C(O)R⁴; and

R⁴ is selected from the group consisting of H, (C₁-C₄) alkyl, and phenyl(Ci-C₄) alkyl; with the provisos that;

(a) when Q is -O-, HET is (^-pyrrolidine, rac-pyrrolidine or piperidine, Ar is phenyl or halo-substituted phenyl, and HetAr is /?-phenylene, then the Z-W combination is other than H;

(b) when Q is NR¹, HET is thiazolidine, Ar is phenyl or substituted phenyl and HetAr is meta-phenylene, then the ZW combination is other than H; and

(c) when Q is -O-, HET is azetidine, Ar is phenyl, n is 1 and HetAr is a 2,5-substituted pyridine, then the Z-W combination is other than H; or a pharmaceutically acceptable salt, ester, or prodrug of said compound.

A very highly preferred compound is

referred to as DG-051 by deCODE genetics ehf, or a pharmaceutically acceptable salt, ester, or prodrug thereof. With respect to DG-051, doses of 10, 15, 20, 25, 30 mg...120, 130, 140, 150 or 160 mg, and any interger value within the range of 10 mg to 160 mg, and any integer sub-range, are specifically contemplated for practicing methods of the invention. Another preferred genus of compounds for practice of the invention are leukotriene synthesis inhibitor compounds represented by the formula:

or pharmaceutically acceptable salt thereof , wherein R¹ represents a group of the formula:

FT

/

-OFT or -N

\

R°

R² and R³ are identical or different and represent hydrogen, lower alkyl, phenyl, benzyl or a group of the formula:

R represents hydrogen, lower alkyl, phenyl or benzyl, which can optionally be substituted by hydroxyl, carboxyl, lower alkoxycarbonyl, lower alkylthio, heteroaryl or carbamoyl, R⁵ represents hydrogen, lower alkyl, phenyl or benzyl, R⁶ represents a group of the formula -COR⁵ or -CO² R⁵, R⁷ represents hydrogen, lower alkyl or phenyl, Y represents a group of the formula:

wherein R⁸ represents hydrogen, lower alkyl or phenyl and n denotes a number of 0 to 5, Z represents norbornyl, or represents a group of the formula:

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or a pharmaceutically acceptable salt thereof. A highly preferred group of leukotriene synthesis inhibitors are 2-[4-

(quinolin-2-yl-methoxy)phenyl]-2-cyclopentylacetic acid, 2-[4-(quinolin-2-yl- methoxy)phenyl]-2-cyclohexylacetic acid, and 2-[4-(quinolin-2-yl-methoxy)phenyl]- 2-cycloheptylacetic acid, (+)-enantiomer of 2-[4-(quinolin-2-yl-methoxy)phenyl]-2- cyclopentylacetic acid, (-)-enantiomer of 2-[4-(quinolin-2-yl-methoxy)phenyl]-2- cyclopentylacetic acid, and pharmaceutically acceptable salts thereof.

A very highly preferred leukotriene synthesis inhibitor comprises BAY-X- 1005 (DG-031) or a physiologically acceptable salt, formulation, or pro-drug thereof. In preferred variations, the composition is administered according to a dosing regimen that achieves a single daily peak plasma concentration of the inhibitor of at least 11.2 micrograms per milliliter (μg/ml), with the upper limit being the maximum safely tolerated dose.

In a related variation, the invention is a method or use of inhibitor for reducing leukotriene production in a human comprises administering to the human a composition that comprises a leukotriene synthesis inhibitor of the formula:

wherein R¹ represents a group of the formula:

R^

/

-OR^ or -N

\

R°

R⁴ R⁴ R⁴

-CH CO_?R^S -CH- -CH; -OR^ϋ -CH- -C-

R⁴ represents hydrogen, lower alkyl, phenyl or benzyl, which can optionally be substituted by hydroxyl, carboxyl, lower alkoxycarbonyl, lower alkylthio, heteroaryl or carbamoyl, R⁵ represents hydrogen, lower alkyl, phenyl or benzyl, R⁶ represents a group of the formula -COR⁵ or -CO² R⁵, R⁷ represents hydrogen, lower alkyl or phenyl, Y represents a group of the formula:

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or pharmaceutically acceptable salt, ester, or pro-drug thereof, and wherein the composition is administered to the human in the morning without food, in an amount effective to achieve a peak plasma concentration (Cmax) of the inhibitor greater than 11.2 μg/ml. Preferably, the peak plasma concentration occurs during the day between the hours of 6 A.M. and 2 P.M.

As indicated above, one preferred leukotriene synthesis inhibitor is DG-031 , and the concentrations in the following paragraphs are particularly relevant to DG-031.

In some variations of the invention, the dosing regimen achieves a daily peak plasma concentration of at least 11.2, 11.3, 11.4, 11.5, 11.7, 11.9, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16, 16.5, 17, 17.5, 18, 18.5, 19, 19.5, or 20 micrograms of DG-031 per milliliter. Preferred daily peak plasma concentrations also can be expressed as any value falling within a range, where the minimum concentration for the range is any value in the preceding sentence, and the maximum value is any higher value than the minimum, selected from 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 2, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, or 40 micrograms per milliliter. To provide just a few examples, in one variation, the dosing regimen is effective to achieve a peak plasma concentration within a range of 12 to 30 micrograms per milliliter. In another variation, the peak plasma concentration is within a range of 11.2 to 25 micrograms per milliliter. In yet another variation, the peak plasma concentration is within a range of 11.2 to 20 micrograms per milliliter; or 12 to 15 micrograms per milliliter.

For compounds that are structurally similar to DG-031 , the same exemplary minimums and maximums are contemplated, adjusting for differences in molecular weight, so as to achieve micromolar equivalence.

Methods of the invention can be further refined to achieve the desired therapeutic effect and peak plasma concentration while minimizing unnecessary excess drug exposure, a parameter which is reflected by AUC measurements or reflected by determining an average plasma concentration of the compound during the day (from a plot or curve obtained from multiple measurements). In some variations using DG-031 , for example, the average plasma concentration is preferably less than 5.5 μg/ml; or less than 5.25 μg/ml or less that 5.0 μg/ml; or less than 4.75 μg/ml, or less than 4.5 μg/ml; or less that 4.0 μg/ml; or less than 3.5 μg/ml. In some variations of the invention, the composition is formulated for oral administration and administered orally. As explained herein, DG-031 can be administered with or without food, and adsorption is increased when taken with food.

In some embodiments, the composition is administered with food. For the example, the method of the invention is practiced wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, the composition is administered orally with food in the morning, and the dose is within the range of 250- 1500 mg, inclusive. More preferably, the dose with food falls within a range having a minimum of at least 300, 350, 400, 450, 500, 550, 600, 650, 700, or 750 mg, and having a maximum that is higher than the minimum and at least 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 1000, 1050, 1100, 1200, 1250, 1300, 1350, 1400, 1450, 1500mg. For example, the dose may be in the range of 750-2000 mg, 1000-1500 mg, 750-1000 mg, 250-1500 mg, 750-1000 mg, 1000-1500 mg, inclusive. To provide another example of the many permutations contemplated by these values, the method is practiced wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, the composition is administered orally with food in the morning, and the dose is within the range of 750-1000 mg, inclusive. In other embodiments, the composition is administered on an empty stomach. For the purposes of the invention, empty stomach constitutes fasting for at least 4 hours before administration (but preferably at least 5, 6, 7, or 8 hours) and continuing to fast for at least 30 minutes (but preferably at least 60, 90, or 120 minutes) after administration. For example, in one variation, the method of the invention is practiced wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, the composition is administered orally on an empty stomach, and the dose is within the range of 500-2000 mg, inclusive. More preferably, the dose with food falls within a range having a minimum of at least 500, 550, 600, 650, 700, 750, 800, 850, 900, 950 mg, or 1000 mg, and having a maximum that is higher than the minimum and at least 800, 850, 900, 950, 1000, 1050, 1100, 1200, 1250, 1300, 1350, 1400, 1450, 1500, 1600, 1700, 1750, 1800, 1900, 2000, 2100, 2200, 2300, 2400, and 2500mg. To provide another example of the many permutations contemplated by these values, the method of a invention is practiced wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or prodrug thereof, the composition is administered orally on an empty stomach, and the dose is within the range of 1000-1500 mg, inclusive. To provide one example of a specific embodiment, the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, and the composition contains a 1000 mg dose of the inhibitor.

A preferred human subject for treatment according to the invention is an adult human, particularly an adult humans suffering from an inflammatory disease and/or an adult human identified as being at risk for developing an inflammatory disease or condition. Treatment of adult humans identified as having or at risk for developing cardiovascular disease is specifically contemplated. An exemplary human subject of the invention is an individual who is at risk for suffering a myocardial infarction (MI) as indicated by elevated levels of a leukotriene (e.g., LTB4) or an inflammatory marker such as C-reactive protein (CRP) or myeloperoxidose (MPO). For example, thresholds for identifying "elevated" may be 20% or more higher than median for age and sex-matched controls, or more preferably 25% or 30% higher; or 1, 1.5, 2, or 2.5 standard deviations above mean values for age and sex matched controls; or top quartile or quintile of a population. Other "conventional" risk factors for cardiovascular events may be used to select the subject for treatment according to the invention. For example, exemplary human subjects for practice of the invention have been diagnosed with at least one risk factor for myocardial infarction or stroke selected from the group consisting of advanced age, gender, smoking, physical activity, waist-to-hip circumference ratio, family history of cardiovascular disease or myocardial infarction, previously diagnosed cardiovascular disease or MI, obesity, diabetes, hypertriglyceridemia, low HDL cholesterol, hypertension, elevated blood pressure, cholesterol levels (total cholesterol >200mg/dL), HDL cholesterol, LDL cholesterol, triglycerides, apolipoprotein AI and B levels, fibrinogen, ferritin, C-reactive protein, and leukotriene levels.

Another exemplary human subject of the invention is an individual who has suffered at least one myocardial infarction in the past. Another exemplary human subject is an individual identified as at-risk for MI due to a genetic predisposition, such as a predisposing single nucleotide polymorphism (SNP) or haplotype in a gene such as FLAP, LTA4H, or 5-LO. See PCT Application No. PCT/US03/32805, filed October 16, 2003, PCT/US03/32556 filed October 16, 2003, PCT/US04/030582 filed September 17, 2004, PCT/US05/03312 filed January 31, 2005 and PCT Application No. PCT/US06/12073 filed March 30, 2006 (attorney docket no. 30847/40807A), which are incorporated by reference herein in their entirety. Also see U.S. Publication No. US-2006-0019269-A1 filed March 30, 2005 and U.S. Patent Application No. 11/270,804 (Publication No. ) filed

November 9, 2006, which are incorporated by reference herein in their entirety.

An exemplary FLAP haplotype that is associated with risk for MI is HapA, which is defined by allele G at marker SG13S25, allele T at marker

SG13S114, allele G at marker SG13S89, and allele A at marker SG13S32 within the FLAP gene.

Another example of a FLAP haplotype that is associated with risk for MI is HapC, which is defined by the T allele of marker SG13S375, allele G of marker SG12S25, allele G of marker SG12S106 and allele A of marker SG12S32 within the FLAP gene. There are 4 additional variations of the HapC haplotype which comprise SNPs in addition to the T allele of SG13S375. HapC2 is defined by allele T of the SNPs SG13S375 and allele G of the SNP SG13S25. HapC3 is defined by allele T of the SNPs SG13S375 and allele G of the SNP SG13S25 and allele A of SNP SG13S32. HapC4-A is defined by allele G of the SNP SG13S106 in addition to allele T of the SNPs SG13S375, allele G of the SNP SG13S25 and allele A of SNP SG13S32. HapC4-B is defined by allele A of the SNP SG13S106 in addition to allele T of the SNPs SG13S375, allele G of the SNP SG13S25 and allele A of SNP SG13S32. HapC4-A correlates with HapA and HapB.

An exemplary LTA4H haplotype that is associated with risk for MI is HapK. HapK is defined by allele C of the SNP SG 12S16, allele G of the SNPs SG12S21, allele T of the SNP SG12S23, allele A of the SNP SG12S25, allele T of the SNP SG12S26, allele T of the SNP SG12S100, allele T of the SNP SG12S28, allele C of the SNP SG12S143, allele G of the SNP SG12S144, and allele G of the SNP SG12S221. Numerous haplotypes with apparent perfect or near perfect correlation with HapK are described in the aforementioned patent documents.

Cardiovascular patients are not the only patients expected to benefit from the invention. Other exemplary patients for treatment according to the invention have other inflammatory conditions, such as asthma or arthritis.

In preferred variations of the invention, measurable reductions in one or more inflammatory markers in achieved, preferably an inflammatory marker that correlates with a disease state or is predictive of a likelihood of a disease or condition. Thus, in one variation of methods of the invention, the doses and dosing schedule are effective to cause a reduction of serum C-reactive protein (CRP) of at least 20% within two weeks of commencing administration and maintain said reduction with continued administration of doses according to the dosing schedule. More significant reduction, e.g., at least 25%, 28%, 30%, 32%, 35%, 40%, 45%, 50%, or more, is preferred. Faster reduction, e.g., within 10 days or one week of commencing therapy, is preferred. With respect to a single human, percent reduction is measured relative to pre-treatment levels measurable in the human. Pre -treatment levels can be measured at any time before administration of the drug, although inflammatory markers, such as CRP will vary with diet, activity, infection, other medications, and the like. For greater accuracy, two or more pre-treatment measurements from different times can be used to establish the pre-treatment baseline. With respect to evaluating a dose and dosing schedule or a particular sustained or controlled release formulation in a clinical setting or in a population, mean reductions are used.

In another variation, the inflammatory marker is used as a primary measure of the method of the invention. Thus, in another embodiment, the invention is a method of treating or preventing an inflammatory condition or disease in a human comprising administering doses of DG-031, or a pharmaceutically acceptable salt or ester or prodrug thereof, according to a dosing schedule that is effective to cause a reduction of serum C-reactive protein (CRP) of at least 20% within two weeks of commencing administration, and maintain said reduction with continued administration of doses according to the dosing schedule. As described above, more significant reduction and faster reduction is preferred.

Myeloperoxidase is another preferred inflammatory marker for use as a primary or secondary measure of a method of the invention. Thus, in another variation of methods of the invention, the doses and dosing schedule are effective to achieve a reduction of serum myeloperoxidase (MPO) of at least 30% within one week of commencing administration and maintain said reduction with continued administration of doses according to the dosing schedule. More significant reductions, e.g., of 35% or 40%, are preferred. As described below, measurement of stimulated MPO production, obtained by treating a whole blood sample with a calcium ionophore, such as ionomycin, are contemplated for monitoring drug effects.

In some variations, the materials and methods/uses of the invention are selected to minimize undesirable side-effects while still meeting therapeutic objectives set forth herein. Undesirable side effects include elevations of serum LDL (e.g., LDL-C), fatigue, dizziness, hyperhidosis, adverse indicators of liver function, increases in serum creatinine (or other potential indicators of adverse affects on renal function), and increases in creatine kinase.

The invention provides methods and uses of the invention wherein the human subject has or is at elevated risk for an inflammatory disease or condition such as a cardiovascular disease or condition. Exemplary disease or conditions for therapeutic or prophylactic therapy include cardiovascular disease, or more particularly, atherosclerosis, arteriosclerosis, or PAOD; and patients at increased risk for myocardial infarction or stroke due to family history, medical history, behavior (e.g., smoking), or genetic predisposition. Such family or medical history risk factor include diabetes; hypertension; hypercholesterolemia; elevated triglycerides; elevated lp(a); obesity; ankle/brachial index (ABI) less than 0.9; a past or current smoker; transient ischemic attack; transient monocular blindness; carotid endarterectomy; asymptomatic carotid stenosis; claudicatioin; limb ischemia leading to gangrene, ulceration or amputation; a vascular or peripheral artery revascularization graft; increased serum LDL cholesterol and/or decreased HDL cholesterol; serum total cholesterol >200 mg/dl, increased leukotriene synthesis; and/or at least one previous myocardial infarction, ACS, stable angina, previous transient ischemic attack , transient monocular blindness, or stroke, asymptomatic carotid stenosis or carotid endarterectomy, atherosclerosis, requires treatment for restoration of coronary artery blood flow (e.g., angioplasty, stent, revascularization procedure).

It will be understood that a molar quantity of DG-031 will not necessarily weigh the same as the same molar quantity of one of its salts, esters, or prodrugs. The molecular weight of DG-031 is 361 gram/mole. DG-031 concentration values herein expressed as mass/volume units can be converted into molarity concentrations or other units that are more readily transferable to salts, esters, prodrugs, or chemical variants.

In one preferred embodiment of this dosing schedule, the doses are pills or capsules containing 693 or 1385 micromoles of the DG-031 (250 or 500 mg) or the salt or ester thereof, (where 750, 1000, or larger doses are achieved by consuming multiple pills or capsules). In one variation, the doses are formulated for oral administration and administered orally. An exemplary formulation for use in the method of the invention is a solid tablet that is orally administered and that consists essentially of 250 mg of DG-031, 40 mg of corn starch, 96.24 mg of microcrystalline cellulose, 1.24 mg, 10 mg providone 25 [poly(l-vinyl-2-pyrrolidinone 25], 2.52 mg magnesium stearate and purified water and further comprising a film coating consisting essentially of 6 mg methylhydroxypropylcellulose, 1.5 mg polyethylene glycol 4000, 2.5 mg titanium oxide and purified water. For a 500 mg dose, two tablets are consumed.

In another embodiment, the invention is a method of treating or preventing an inflammatory condition or disease in a human comprising administering an initial dose of DG-031, or a pharmaceutically acceptable salt or ester or prodrug thereof according to any of the variation of the methods already described, continuing administering doses of DG-031 according to the initial dosing schedule for a time effective to cause a reduction of leukotriene B4 (LTB4) of at least 30% or at least 35%, and administering a maintenance dose of DG-031, or a pharmaceutically acceptable salt or ester or prodrug thereof, that is less than the initial dose of DG-031 administered. The LTB4 levels may be measured using the ionophore-stimulation assay. A maintenance dose of DG-031 is a dose that effectively maintains a therapeutic benefit such as a measurable reduction in LTB4. A maintenance dosing schedule is a dosing schedule that effectively maintains a therapeutic benefit of the initial dosing schedule that has a reduced quantity of DG-031 per dose and/or a reduced frequency of administration. Preferably, administration of the initial dose is continued for 2 weeks. An exemplary maintenance dose is a total daily administered according to the maintenance doses and dosing schedule is at least 25% less than the total daily administration according to the initial doses and dosing schedule. In another embodiment, the invention is a method of treating or preventing an inflammatory condition or disease in a human comprising administering an initial dose of DG-031, or a pharmaceutically acceptable salt or ester or prodrug thereof according to any of the variation of the methods already described, continuing administering an initial doses of DG-031 according to the initial dosing schedule for a time effective to cause a reduction of serum C-reactive protein (CRP) of at least 20%, and administering a maintenance dose of DG-031, or a pharmaceutically acceptable salt or ester or prodrug thereof, that is less than the initial dose of DG-031 administered. A maintenance dose of DG-031 is a dose that effectively maintains a therapeutic benefit such as a measurable reduction in serum CRP. The invention also provides for any of the preceding methods or uses comprising a step, prior to the administering step, of selecting a human at risk for myocardial infarction to receive the doses of DG-031. The selecting step comprises determining a level of an inflammatory marker in a human subject and selecting a subject with an elevated measurement of the marker. The inflammatory markers for selection include CRP, MPO and leukotriene, preferably LTB4.

In one variation, the selecting step comprises selecting a human who has suffered at least one myocardial infarction. The invention also provides for selecting for administration a human with cardiovascular disease, such as atherosclerosis, PAOD, myocardial infraction or stroke.

In one variation, the selecting step comprises selecting a human female. In another variation, the selecting step comprises selecting a human that is at least 40 years old , or at least 50 years old, or at least 60 years old, or at least 65 years old.

In another variation, the selecting step comprises selecting a human with a genetic predisposition to increased risk for myocardial infarction. For example, the genetic predisposition comprises presence of a polymorphism or haplotype in the human that correlates with increased risk for MI, wherein the polymorphism or haplotype is in a gene selected from the group consisting of FLAP, LTA4-H, and 5-LO.

The invention also provides for the selecting step further comprising determining if a human has a race that includes black African ancestry, and selecting for dosing with DG-031 a human with a race that includes black African ancestry. Further, the selecting step comprises determining if a human has a race that includes European ancestry, and selecting a human with a race that includes European and African ancestry.

Methods of the invention can be practiced by any mode/route of drug administration, including but not limited to oral, transdermal, transmucosal (e.g., sublingual, buccal), intradermal, subcutaneous, intramuscular, intravenous, pulmonary (e.g., nebulizer, metered-dose inhaler) anal, rectal, vaginal, inhalation and intranasal administration. Oral administration, e.g., by a tablet/pill or capsule, is preferred.

Because conditions for which the treatment is indicated may be chronic or progressive, and also because the treatment may be prophylactic, repeated dosing according to the dosing schedule is specifically contemplated, For example, the methods of the invention may be practiced where the administering is performed for at least 15 days, 30 days, 60 days, 90 days, 120 days, 180 days, 1 year, 2 years, 3 years, or longer, e.g., for the duration of a person's life. As described above with respect to methods of the invention, all integer and half- integer subranges of the maximum recited above are specifically contemplated as embodiments of the invention. It should be understood that all aspects of the invention that have been described as methods of treating a human subject also should be interpreted as a description of medical uses of a compound. For example, the invention includes a leukotriene synthesis inhibitor for use as a medicament for once daily administration in the morning; or use of a leukotriene synthesis inhibitor for the manufacture of a medicament for once daily morning administration; according to the teachings herein.

In a further embodiment, the invention provides for a use of a leukotriene synthesis inhibitor such as DG-031, or a pharmaceutically acceptable salt or ester or prodrug thereof, for the preparation of a medicament for human administration for the treatment or prevention of an inflammatory condition or disease, wherein the medicament is to be administered at an initial dose according to an initial dosing schedule that is effective in said human as described above, wherein the initial dose and dosing schedule are continued for a time effective to cause a reduction of LTB4 of at least 30%, and wherein the medicament is then administered at a maintenance dose of DG-031, or a pharmaceutically acceptable salt or ester or prodrug thereof, according to a maintenance dosing schedule after the reduction in LTB4, wherein the maintenance dose of DG-031 and the maintenance dosing schedule are effective to maintain a reduction of serum LTB4 of at least 30%.

The foregoing summary is not intended to define every aspect of the invention, and additional aspects are described in other sections, such as the Detailed Description. The entire document is intended to be related as a unified disclosure, and it should be understood that all combinations of features described herein are contemplated, even if the combination of features are not found together in the same sentence, or paragraph, or section of this document. In addition to the foregoing, the invention includes, as an additional aspect, all embodiments of the invention narrower in scope in any way than the variations specifically mentioned above. With respect to aspects of the invention described as a genus, all individual species are individually considered separate aspects of the invention. With respect to aspects described as a range, all subranges and individual values are specifically contemplated.

Although the applicant(s) invented the full scope of the claims appended hereto, the claims appended hereto are not intended to encompass within their scope the prior art work of others. Therefore, in the event that statutory prior art within the scope of a claim is brought to the attention of the applicants by a Patent Office or other entity or individual, the applicant(s) reserve the right to exercise amendment rights under applicable patent laws to redefine the subject matter of such a claim to specifically exclude such statutory prior art or obvious variations of statutory prior art from the scope of such a claim. Variations of the invention defined by such amended claims also are intended as aspects of the invention. Additional features and variations of the invention will be apparent to those skilled in the art from the entirety of this application, and all such features are intended as aspects of the invention.

BRIEF DESCRIPTION OF THE DRAWING

Figure 1 shows a summary of PK (upper panel; micrograms per mL) and PD (lower panel; percentage LTB4 lowering) for 1000 mg QD dosing (solid lines) and 500 mg BID dosing (dashed line), based on data for 12 subjects for each dosing regimen. The average daily DG-031 concentration (area under curves in upper panel) is greater for 500 mg BID than for 1000 mg QD. However, LTB4 lowering effect (lower panel) is more efficient for 1000 mg QD than for 500 mg BID. X-axis denotes time ofadministration of DG-031, wherein TO is 8 AM.

Figure 2 is a scatter plot depicting a comparison of 500 mg BID and 1000 mg QD effects for all study subjects. The plot shows paired, rank-ordered data for DG-031 concentration (open circles) and LTB4 lowering effect (closed circles). Coordinates [x,y] on the graph are based on the actual data points after rank-ordering the data. For example, for DG-031 concentration for 500 mg BID and 1000 mg QD dosing on Day 7 (highest open circle on right half of figure), the coordinates are [x,y] given by the DG-031 concentration achieved by the 500 mg BID (y) and 1000 mg QD (x) doses for the individual with the highest DG-031 plasma concentration on that day. The solid line represents the line around which all the dots would fall on to or close to (or randomly scattered around) in absence of any differences achieved by the 1000 mg QD vs. 500 mg BID dosing regimens. DETAILED DESCRIPTION

The use of leukotriene inhibitors to treat cardiovascular diseases is described in International Patent Application Nos. PCT/US03/32805, PCT/US04/30582 and PCT/US05/00312, incorporated herein by reference in their entirety. One preferred class of compounds for use in such materials and methods are FLAP inhibitors described in U.S. Patent Nos. 4,970,215 and 5,693,650, also incorporated herein by reference in their entirety. A preferred compound for use in such therapeutic materials and methods is DG-031 (also known as Bay x-1005), an orally active inhibitor of the synthesis of leukotrienes B4 and C4 through inactivation of FLAP. DG-031 is a substituted 4-(quinolin-2-YL-methoxy)phenyl-acetic acid derivative. Clinical studies have demonstrated DG-031 was safe and well tolerated in healthy volunteers at a total daily dose of 100 mg, 200 mg, 250 mg, 300 mg, 500 mg, 750 mg, and 1000 mg DG-031 administered for less than 14 days. In addition, total daily doses of 500 mg and 1000 mg DG-031 administered for 14-42 days were safe and well tolerated in healthy volunteers. (See, Dahlen et al. Thorax 523: 348-354, 1997; Hamilton et al, Thorax 52: 348-54, 1997).

In clinical studies in asthma patients, total daily dose of 250 mg, 500 mg, 750 mg and 1000 mg were safe and well tolerated when administered for 14 days. Total daily doses of 100 mg, 200 mg, 250 mg, 500 mg and 1000 mg of DG-031 were safe and well tolerated after being administered for 14-42 days. A total daily dose of 500 mg administered for greater than 45-365 days was also safe and well tolerated in asthma patients. In clinical studies in patients with coronary artery disease (CAD), total daily doses of 375 mg and 750 mg were safe and well tolerated after being administered for less than 8 days. In addition, total daily doses of 250 mg, 500 mg, and 750 mg of DG-031 were safe and well tolerated in CAD patients after being administered for 28 days three times a day (TID). Previous clinical studies administered the dosages of DG-031 either in a single dose (QD) or twice a day (BID)

The invention provides improved dosing materials and methods that are effective to block any branch of the leukotriene synthesis pathway and the dosing regime is administered in an amount effective to achieve a single daily peak (Cmax) between the hours of 6 A.M. and 2 P.M. In particular, the invention contemplates administering a leukotriene synthesis inhibitor to block the LTA4H-LTC4-LTD4- LTE4 branch of the leukotriene synthesis pathway. Preferably, the invention provides for improved dosing regimes for inhibiting the enzymatic activity of FLAP, which converts arachidonic acid to LTA4. Particularly, the invention provides for improved dosing materials and methods for therapy with the FLAP inhibitor known as DG-031 and related compounds as described herein. In addition, the invention provides for improved dosing regines for inhibiting the enzymatic activity of LTA4H, which converts LT A4 to LTB4. Particularly, the invention provides for improved dosing materials and methods for therapy with the LTA4H inhibitor known as DG-051 and related compounds as described herein. The invention also provides for improved dosing regimes for inhibiting the enzymatic activity of leukotriene C4 synthase, which converts LTA4 and glutathione to create LTC4. The invention also provides for improved dosing regimes for inhibiting gamma-glutamyltranspeptidase, which converts LTC4 to LTD4. In addition, the invention provides for improved dosing regimes for inhibiting dipeptidase, which converts LTD4 to LTE4.

The invention is describe in relation to administering concentrations of DG-031 (FLAP inhibitor) that achieve a single daily peak concentration between the hours of 6 A.M. and 2 P.M. However, the invention also provides for materials and methods for administering any inhibitor of leukotriene synthesis that achieves a single daily peak concentration between 6 A.M. and 2 P.M. For any particular compound, the concentration administered will achieve a balance that maximizes efficiency and safety.

Description of FLAP inhibitor chemical compounds

The invention provides for improved dosing materials and methods for inhibiting FLAP, such as DG-031 and related compounds. DG-031 and related compounds are described in detail in U.S. Patent No. 4,970,215 (Mohrs, et al.), incorporated herein by reference. Chemical syntheses are described in U.S. Patent No. 5,693,650, also incorporated by reference.

The compound can be a substituted 4-(quinolin-2-61- methoxy)phenylacetic acid derivative represented by the following formula:

R^

/

-OFT or -N

\

R°

R⁴ R⁴ R⁴

-CH COpR^s -CH- -CH₅ -OR^ϋ -CH- -C-

wherein R >9 and . „ R10 are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or a pharmaceutically acceptable salt thereof.

Preferably the compounds are selected from the group consisting of: 2- [4-(quinolin-2-yl-methoxy)phenyl]-2-cyclopentylacetic acid, 2-[4-(quinolin-2-yl- methoxy)phenyl]-2-cyclohexylacetic acid, and 2-[4-(quinolin-2-yl-methoxy)phenyl]- 2-cycloheptylacetic acid, (+)-enantiomer of 2-[4-(quinolin-2-yl-methoxy)phenyl]-2- cyclopentylacetic acid, (-)-enantiomer of 2-[4-(quinolin-2-yl-methoxy)phenyl]-2- cyclopentylacetic acid and pharmaceutically acceptable salts thereof. See U.S. Patent No. 4,970,215, incorporated herein by reference.

A preferred compound is (R)-(+)-alpha-cyclopentyl-4-(2- quinolinylmethoxy)-Benzeneacetic acid, also known as Bay-X1005 and DG-031. The invention also contemplates physiologically acceptable salts of the compounds of the invention, such as salts of organic or inorganic bases or acids. Physiologically acceptable salts of the substituted 4-(quinolin-2-yl- methoxy)phenylacetic acids, esters and amides can be salts of the substances according to the invention with mineral acids, carboxylic acids or sulphonic acids. Particularly preferred salts are, for example, those with hydrochloric acid, hydrobromic acid, sulphuric acid, phosphoric acid, methanesulphonic acid, ethanesulphonic acid, toluenesulphonic acid, benzenesulphonic acid, naphthalenedisulphonic acid, acetic acid, propionic acid, lactic acid, tartaric acid, citric acid, fumaric acid, maleic acid or benzoic acid. Salts in the context of the present invention are furthermore salts of monovalent metals, such as alkali metals and ammonium salts. Sodium, potassium and ammonium salts are preferred. Physiologically acceptable salts can also be metal salts or ammonium salts of the compounds according to the invention which have a free carboxyl group or a tetrazolyl radical. Particularly preferred salts are, for example sodium potassium, magnesium or calcium salts, as well as ammunonium salts, which are derived from ammonia, or organic amines, such as, for example, ethylamine, di- or triethylamine, di- or triethanolamine, dicyclohexylamine, dimethylaminoethanol, glucosamine, arginine, lysine, ethylenediamine or 2-phenylethylamine.

A heterocyclic radical in general is a 5- to 6-membered, saturated, partially unsaturated or unsaturated ring which can contain up to 3 oxygen, sulphur and/or nitrogen atoms as heteroatoms. Preferred rings are 5 and 6-membered rings with one oxygen, sulphur and/or up to 2 nitrogen atoms. Rings which are mentioned as preferred are: thienyl, furyl, pyrrolyl, pyrazolyl, pyridyl, pyrimidyl, pyrazinyl, pyridazinyl, thiazolyl, oxazolyl, imidazolyl, pyrrolidinyl, piperidinyl or piperazinyl.

A 5- to 6-membered saturated heterocyclic radical which can also contain up to 3 oxygen, sulphur and/or nitrogen atoms as heteroatoms is in general piperidyl, morpholinyl, piperazinyl or pyrrolidyl. Morpholinyl is preferred.

A carbocyclic radical in general is a 3- to 7-membered, preferably 5- to 7-membered, saturated hydrocarbon ring. Cyclopentyl, cyclohexyl or cycloheptyl are mentioned as preferred.

A hydroxy-protective group in the context of the abovementioned definition is in general a protective group from the series consisting of: tert- butoxydiphenylsilyl, trimethylsilyl, triethylsilyl, triisopropylsilyl, tert-butyl- dimethylsilyl, tert-butyl-diphenylsilyl, triphenylsilyl, trimethylsilylethoxycarbonyl, benzyl, benzyloxycarbonyl, 2-nitrobenzyl, 4-nitrobenzyl, 2-nitrobenzyloxycarbonyl, 4-nitrobenzyloxycarbonyl, tert-butyloxycarbonyl, allyloxycarbonyl, 4- methoxybenzyl, 4-methoxybenzyloxycarbonyl, formyl, acetyl, trichloroacetyl, 2,2,2- trichloroethoxycarbonyl, 2,4-dimethoxymethyl, 2,4-dimethoxybenzyloxycarbonyl, methylthiomethyl, methoxyethoxymethyl, 2-(trimethylsilyl)ethoxy!methyl, 2- (methylthiomethoxy)ethoxycarbonyl, benzoyl, 4-methylbenzoyl, 4-nitrobenzoyl, 4- fluorobenzoyl, 4-chlorobenzoyl or 4-methoxybenzoyl. Acetyl, benzoyl, benzoyl, or methylbenzyl are preferred.

Amino-protective groups in the context of the invention are the customary amino-protective groups used in peptide chemistry. These include, preferably: benzyloxycarbonyl, 3,4-dimethoxybenzyloxycarbonyl, 3,5- dimethoxybenzyloxycarbonyl, 2,4-dimethoxybenzyloxycarbonyl, 4- methoxybenzyloxycarbonyl, 4-nitrobenzyloxycarbonyl, 2-nitrobenzyloxycarbonyl, 2- nitro-4,5-dimethoxybenzyloxycarbonyl, methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl, isopropoxycarbonyl, butoxycarbonyl, isobutoxycarbonyl, tert- butoxycarbonyl, allyloxycarbonyl, vinyloxycarbonyl, 2-nitrobenzyloxycarbonyl, 3,4,5-trimethoxylbenzyloxycarbonyl, cyclohexoxycarbonyl, 1,1- dimethylethoxycarbonyl, adamantylcarbonyl, phthaloyl, 2,2,2- trichloroethoxycarbonyl, 2,,2,2-trichloro-tert-butoxycarbonyl, methyloxycarbonyl, phenoxycarbonyl, 4-nitrophenoxycarbonyl, fluorenyl-9-methoxycarbonyl, formyl, acetyl, propionyl, pivaloyl, 2-chloroacetyl, 2-bromoacetyl, 2,2,2-trifluoroacetyl, 2,2,2- trichloroacetyl, benzoyl, 4-chlorobenzoyl, 4-bromobenzoyl, 4-nitrobenzoyl, phthalimido, isovaleroyl or benzyloxymethylene, 4-nitrobenzyl, 2,4-dinitrobenzyl or 4-nitrophenyl. The compounds according to the invention can be in stereoisomeric forms which either behave as image and mirror image (enantiomers) or do not behave as image and mirror image (diastereomers). The invention relates both to the antipodes and to the racemic forms as well as the diastereomer mixtures. The racemic forms, like the diastereomers, can be resolved into the stereoisomerically uniform constituents in a known manner (compare E. L. Eliel, Stereochemistry of Carbon Compounds, McGraw Hill, 1962).

The formulation of the invention may be made by process known in the art and in particular by the process taught in U.S Patent Nos. 4,970,215 and 5,693,650, which are herein incorporated by reference in their entirety. Additional FLAP inhibitors for use in the improved dosing materials and methods of the invention are set out in Table 1. Table 1

Produci Name

Company Structure Chemical Name Patent Ref MOA

(R)-(+)-N-[3[5-[(4- US 5288751, atreleuton (ABT- fluorophenyl)methyl] -2thienyl] - US 5288743, 5-LPO

Abbott 761) lmethyl-2-propynyl]— N-hydroxurea US 5616596 inhibitor

3 -(3 -( 1 , 1 -dimethylethylthio-5- (quinoline-2-ylmethoxy)- 1 -(4- chloromethylphenyl)indole-2-yl)- 2 , 2 -dimethylpropionaldehyde oxime- WO9203132, FLAP

Abbott A-81834 0-2-acetic acid US 5459150 inhibitor

3 -(3 -( 1 , 1 -dimethylethylthio-5- (pyridin-2-ylmethoxy)- 1 -(4- chloromethylphenyl)indole-2-yl)-

2 , 2 -dimethylpropionaldehyde oxime- WO9203132, 5-LPO

Abbott A-86886 I 0-2-acetic acid US 5459150 inhibitor

(Code) Structure Chemical Name MOA

(R)-(+)-alpha-cyclopentyl-4-(2- US 4970215 quinolinylmethoxy)-Benzeneacetic EP 344519, FLAP

Bayer BAY-X- 1005 acid DE 19880531 inhibitor

1 -((4-chlorophenyl)methyl)-3 -((1 , 1 - dimethylethyl)thio)-alpha,alpha-

dimethyl- 5 - ( 2 - quinolinylmethoxy) - EP 419049, FLAP

Merck MK-0591 lH-Indole-2-propanoic acid US 19890822 inhibitor

(3 [3 -)4-chlorobenzyl)-3 -t-butyl-thio- 5-isopropylindol-2yl]2,2-dimethyl- 5-LPO

Merck MK-866 proanoic acid inhibitor

Description of LTA4H inhibitor chemical compounds

The invention provides for improved dosing materials and methods for inhibiting LTA4H, such as DG-051 and related compounds. DG-051 and related compounds are described in detail in U.S. Patent Application Publication NO. US 2007/00666820 Al (Sandanayaka et ah), incorporated herein by reference in its entirety.

The invention relates to compounds of the general formula ψ below.

All of the compounds falling within the foregoing parent genus and its subgenera are useful as leukotriene A4 hydrolase inhibitors. The genus ψ encompasses four subgenera, depending on the τVτ² ring: 2,5-pyridinyl, reverse 2,5- pyridinyl, meta phenylene and para phenylene:

The compounds include biaryl heterocycles useful as LTA4H enzyme inhibitors, having the general formula:

Preferred biaryl heterocycles include compounds wherein Q is selected from O, SCO^ and NR¹:

Other preferred biaryl heterocycles include compounds wherein X is selected from CH₂, O and NR or the τVτ² ring is para phenylene.

In some compounds, HET is selected from the group consisting of pyrrolidinone, pyrrolidine, piperidine, piperidinone, piperazine, morpholine, thiomorpholine, thiazolidine, thiazolidinone, oxazolidine and oxazolidinone and substituted pyrrolidinone, substituted pyrrolidine, substituted piperidine, substituted piperidinone, substituted piperazine, substituted morpholine, substituted thiomorpholine, substituted thiazolidine, substituted thiazolidinone, substituted oxazolidine and substituted oxazolidinone.

The compounds include those wherein HET is pyrrolidine and the Z-W combination is other than hydrogen. Preferably the compounds include those in which HET-Z-W is selected from pyridinylmethylpyrrolidine, oxadiazolylmethylpyrrolidine, carboxyalkylpyrrolidine and alkoxycarbonylalkylpyrrolidine. Additional preferred compounds include those in which HET-Z-W is carboxyalkyl pyrrolidine, having the chemical formula as shown below, wherein q is an integer selected from 2-6:

The compounds also include those in which HET is selected from the group consisting of unsubstituted pyrrolidine, pyrrolidinone, piperidine and piperidinone (i.e. Z-W is H). Preferred compounds include those in which HET-Z-W is carboxyalkyl (S) pyrrolidine, having the chemical formula as shown below, wherein q is an integer selected from 2-6:

Additional preferred compounds include those in which HET is (R) pyrrolidine having the chemical formula as shown below:

The compounds also include those in which HET is (R) pyrrolidine and ZW is H, having chemical formula as shown below:

In addition, the preferred compounds include those in which HET is (R) pyrrolidine, X is selected from CH₂, O and NR¹. In certain embodiments HET is (R) pyrrolidine X is CH₂ or O, n is 1, and Ar is selected from phenyl and substituted phenyl, and X is selected from CH₂, O and NR¹. In further embodiments X is CH₂ or O, n is 1, and Ar is para-substituted phenyl. In other embodiments, Ar is heteroarylphenyl. In other embodiments, Ar is

wherein the wavy line indicates the point of attachment to X and R is chosen from hydrogen, halogen, trifluoromethyl, methyl, methoxy, thienyl, furanyl, and thienyl or furanyl substituted with halogen, trifluoromethyl, methyl or methoxy.

Additional preferred compounds include those in which HET is (S) pyrrolidine, having chemical formula as shown below:, where R³ represents halogen, CF³, methyl, methoxy, or CF³O. X is O or CH₂, n is 1 or 2, Z is Ci-C₄ alkylene and W is COOH.

In some embodiments, HET is (S) pyrrolidine, Q is oxygen and Ar is substituted phenyl, having chemical formula as shown below, wherein R represents one to three residues independently selected from the group consisting of benzyl, benzyloxy, phenyl and heteroaryl.

In some preferred compounds, Ar is phenyl substituted with heteroaryl or heteroaryl substituted with a substituent selected from the group consisting of halogen, methyl, methoxy and trifluoromethoxy. Thienyl and furanyl are examples of heteroaryl.

In other preferred compounds, the τVτ² ring is either pyridine and Q is oxygen or the τVτ² ring is para phenylene, and Q is -NR¹-- or — S(O)_P— . In these compounds, the variables may be as described above for the genus in which Q is --O-- and the τVτ²ring is p-phenylene.

A highly preferred compound has the structure set out below, and is referred to as DG-051.

Compounds of the genus represented by formula ψ above are inhibitors of LTA4H enzyme. As such they have utility in treating and preventing inflammatory diseases and disorders, as described above, particularly for such conditions as asthma, chronic obstructed pulmonary disease (COPD), atherosclerosis, rheumatoid arthritis, multiple sclerosis, inflammatory bowel diseases (IBD; including Crohn's disease and ulcerative colitis), or psoriasis, which are each characterized by excessive or prolonged inflammation at some stage of the disease.

Recent research indicates that the compounds are also useful for treating and preventing atherosclerosis, thrombosis, stroke, acute coronary syndrome, stable angina, peripheral vascular disease, critical leg ischemia, intermittent claudication, abdominal aortic aneurysm and myocardial infarction.

The compounds may be presented as salts. The term "pharmaceutically acceptable salt" refers to salts whose counter ion derives from pharmaceutically acceptable non-toxic acids and bases. Suitable pharmaceutically acceptable base addition salts for the compounds of the present invention include, but are not limited to, metallic salts made from aluminum, calcium, lithium, magnesium, potassium, sodium and zinc or organic salts made from lysine, N,N-dialkyl amino acid derivatives (e.g. N,N-dimethylglycine, piperidine-1 -acetic acid and morpholine-4- acetic acid), N,N'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (N-methylglucamine) and procaine. When the compounds contain a basic residue, suitable pharmaceutically acceptable base addition salts for the compounds of the present invention include inorganic acids and organic acids. Examples include acetate, benzenesulfonate (besylate), benzoate, bicarbonate, bisulfate, carbonate, camphorsulfonate, citrate, ethanesulfonate, fumarate, gluconate, glutamate, bromide, chloride, isethionate, lactate, maleate, malate, mandelate, methanesulfonate, mucate, nitrate, pamoate, pantothenate, phosphate, succinate, sulfate, tartrate, p-toluenesulfonate, and the like. (See also discussin of acceptable salts above.

Alkyl is intended to include linear, branched, or cyclic hydrocarbon structures and combinations thereof. Lower alkyl refers to alkyl groups of from 1 to 6 carbon atoms. Examples of lower alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, s- and t-butyl and the like. Preferred alkyl groups are those of

C. sub.20 or below. Cycloalkyl is a subset of alkyl and includes cyclic hydrocarbon groups of from 3 to 8 carbon atoms. Examples of cycloalkyl groups include c-propyl, c-butyl, c-pentyl, norbornyl and the like.

Ci to C₂₀ hydrocarbon includes alkyl, cycloalkyl, alkenyl, alkynyl, aryl, arylalkyl and combinations thereof. Examples include phenethyl, cyclohexylmethyl, camphoryl, adamantyl and naphthylethyl.

Alkoxy or alkoxyl refers to groups of from 1 to 8 carbon atoms of a straight, branched, cyclic configuration and combinations thereof attached to the parent structure through oxygen. Examples include methoxy, ethoxy, propoxy, isopropoxy, cyclopropyloxy, cyclohexyloxy and the like. Lower-alkoxy refers to groups containing one to four carbons.

Alkoxyalkyl refers to ether groups of from 3 to 8 atoms of a straight, branched, cyclic configuration and combinations thereof attached to the parent structure through an alkyl. Examples include methoxymethyl, methoxyethyl, ethoxypropyl, and the like. Alkoxyaryl refers to alkoxy substituents attached to an aryl, wherein the aryl is attached to the parent structure. Arylalkoxy refers to aryl substituents attached to an oxygen, wherein the oxygen is attached to the parent structure. Substituted arylalkoxy refers to a substituted aryl substituent attached to an oxygen, wherein the oxygen is attached to the parent structure.

Acyl refers to groups of from 1 to 8 carbon atoms of a straight, branched, cyclic configuration, saturated, unsaturated and aromatic and combinations thereof, attached to the parent structure through a carbonyl functionality. One or more carbons in the acyl residue may be replaced by nitrogen, oxygen or sulfur as long as the point of attachment to the parent remains at the carbonyl. Examples include acetyl, benzoyl, propionyl, isobutyryl, t-butoxycarbonyl, benzyloxycarbonyl and the like. Lower-acyl refers to groups containing one to four carbons.

Aryl and heteroaryl mean a 5- or 6-membered aromatic or heteroaromatic ring containing 0-3 heteroatoms selected from O, N, or S; a bicyclic 9- or 10-membered aromatic or heteroaromatic ring system containing 0-3 heteroatoms selected from O, N, or S; or a tricyclic 13- or 14-membered aromatic or heteroaromatic ring system containing 0-3 heteroatoms selected from O, N, or S. The aromatic 6- to 14-membered carbocyclic rings include, e.g., benzene and naphthalene, and according to the invention benzoxalane and residues in which one or more rings are aromatic, but not all need be. The 5- to 10-membered aromatic heterocyclic rings include, e.g., imidazole, pyridine, indole, thiophene, benzopyranone, thiazole, furan, benzimidazole, quinoline, isoquinoline, quinoxaline, pyrimidine, pyrazine, tetrazole and pyrazole.

Arylalkyl refers to a substituent in which an aryl residue is attached to the parent structure through alkyl. Examples are benzyl, phenethyl and the like.

Heteroarylalkyl refers to a substituent in which a heteroaryl residue is attached to the parent structure through alkyl. Examples include, e.g., pyridinylmethyl, pyrimidinylmethyl and the like. Heterocyclylalkyl refers to a substituent in which a heterocyclyl residue is attached to the parent structure through alkyl. Examples include morpholinoethyl and pyrrolidinylmethyl. Heterocycle means a cycloalkyl or aryl residue in which from one to three carbons is replaced by a heteroatom selected from the group consisting of N, O and S. The nitrogen and sulfur heteroatoms may optionally be oxidized, and the nitrogen heteroatom may optionally be quaternized. Examples of heterocycles include pyrrolidine, pyrazole, pyrrole, indole, quinoline, isoquinoline, tetrahydroisoquinoline, benzofuran, benzodioxan, benzodioxole (commonly referred to as methylenedioxyphenyl, when occurring as a substituent), tetrazole, morpholine, thiazole, pyridine, pyridazine, pyrimidine, thiophene, furan, oxazole, oxazoline, isoxazole, dioxane, tetrahydrofuran and the like. It is to be noted that heteroaryl is a subset of heterocycle in which the heterocycle is aromatic. Examples of heterocyclyl residues additionally include piperazinyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxo- pyrrolidinyl, 2-oxoazepinyl, azepinyl, 4-piperidinyl, pyrazolidinyl, imidazolyl, imidazolinyl, imidazolidinyl, pyrazinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl, isothiazolyl, quinuclidinyl, isothiazolidinyl, benzimidazolyl, thiadiazolyl, benzopyranyl, benzothiazolyl, tetrahydrofuryl, tetrahydropyranyl, thienyl, benzothienyl, thiamorpholinyl, thiamorpholinylsulfoxide, thiamorpholinylsulfone, oxadiazolyl, triazolyl and tetrahydroquinolinyl.

An oxygen heterocycle is a heterocycle containing at least one oxygen in the ring; it may contain additional oxygens, as well as other heteroatoms. A sulphur heterocycle is a heterocycle containing at least one sulphur in the ring; it may contain additional sulphurs, as well as other heteroatoms. A nitrogen heterocycle is a heterocycle containing at least one nitrogen in the ring; it may contain additional nitrogens, as well as other heteroatoms.

Oxygen heteroaryl is a subset of oxygen heterocycle; examples include furan and oxazole. Sulphur heteroaryl is a subset of sulphur heterocycle; examples include thiophene and thiazine. Nitrogen heteroaryl is a subset of nitrogen heterocycle; examples include pyrrole, pyridine and pyrazine.

A saturated nitrogenous heterocycle is a subset of nitrogen heterocycle. Saturated nitrogenous heterocycle contain at least one nitrogen and may contain additional nitrogens, as well as other heteroatoms. Examples include pyrrolidine, pyrazolidine, piperidine, morpholine, and thiomorpholine. Substituted alkyl, aryl, cycloalkyl, heterocyclyl etc. refer to alkyl, aryl, cycloalkyl, or heterocyclyl wherein up to three H atoms in each residue are replaced with halogen, haloalkyl, hydroxy, loweralkoxy, carboxy, carboalkoxy (also referred to as alkoxycarbonyl), carboxamido (also referred to as alkylaminocarbonyl), cyano, carbonyl, nitro, amino, alkylamino, dialkylamino, mercapto, alkylthio, sulfoxide, sulfone, acylamino, amidino, phenyl, benzyl, heteroaryl, phenoxy, benzyloxy, or heteroaryloxy.

Additional LTA4H and LTB4 inhibitors for use in the improved dosing materials and methods of the invention are set out in Table 2

10 Table 2.

Leukotriene Synthesis Inhibitors

The invention also provides for improved materials and methods of administering a leukotriene synthesis inhibitor that blocks any point within the LTA4H-LTC4-LTD4-LTE4 branch of the leukotriene synthesis pathway. Preferred leukotriene synthesis inhibitors include AZD2138 (AZ Pharma), CJ-13610 (Pfizer), Singulair (Merck), Accolate (AZ Pharma), Pranlukast (Ono), SC56938 (Pfizer), JNJ- 10392980 (J&J) and MK-0591 (Merck). Additional leukotriene synthesis inhibitors are set out in Table 3.

In addition, the compounds may be leukotriene synthesis inhibitor that inhibits the activity of a member of the leukotriene synthesis pathway such as 5- lipoxygenase, 5-lipoxygenase activating protein (FLAP), leutokriene C4 synthase, leukriene A4 hydolase, arachidonate 4-lipoxygenase, leukotriene B4 12- hydroxydehydrogenase, leukotriene A4 receptor, leukotriene B4 receptor, leukotriene C4 receptor, leukotriene D4 receptor, leukotriene E4 receptor, leukotriene B4 receptor 1, leukotriene B4 receptor 2, cysteinyl leukotriene receptor 1 and cysteinyl leukotriene receptor 2. Any LT inhibitor is suitable for practice of the invention, and several LT inhibitors are described herein. To help minimize side effects, an LT inhibitor that is specific for a member of the LT synthesis pathway is preferred. Exemplary inhibitors include both small molecules, biological inhibitors of proteins, (e.g., antibody substances, peptides), and biological inhibitors that operate at the nucleic acid level (e.g., antisense nucleic acids and interfering RNA nucleic acids and zinc finger proteins).

Table 3

The invention provides materials and methods to achieve a preferred peak concentration at a preferred time of day that exhibits the most beneficial effects and reduces drug exposure, thereby reducing the possibility of short term or long term drug side-effects. The invention also provides for methods of administering doses of DG-031 according to a dosing schedule of DG-031 that is effective to achieve a desired concentration.

A desired plasma concentration of DG-031 is a concentration that achieves a desired therapeutic endpoint while minimizing side effects. An exemplary therapeutic endpoint is prophylaxis against myocardial infarction, e.g., reducing the likelihood that a person at risk for MI will incur an MI over a period of time through drug therapy. However, other, more readily measurable criteria can be used as a measure of efficacy. For example, a desired plasma concentration is a concentration that effectively reduces concentrations of measurable leukotrienes or leukotriene metabolites and/or reduces the levels (concentrations) of other inflammatory markers in a human subject (or in a biological sample from the human subject). One preferred dose schedule provides administering doses of DG-031 in a concentration/quantity and at a frequency effective to achieve a peak concentration of at lest 12 μg/ml between the hours of 6 A.M. and 2 P.M. Another preferred dose schedule provides administering doses of DG-031 in a concentration/quantity of at least 20 μg/ml between the hours of 6 A.M. and 2 P.M.

An alternative way to describe a dosing schedule of the invention is with respect to one or more of the measurable biological markers affected by the therapeutic agent. For example, an exemplary dosing schedule of the invention provides administering doses of DG-031 in an amount and at a frequency and in a formulation effective to maintain 30%, 35% or greater reduction in a leukotriene (or leukotriene metabolite) level. Similarly, the invention includes materials and methods for achieving even great measures of efficacy. For example, the invention includes administering DG-031 at a dose or doses according to a dosing schedule effective to reduce a concentration/level of a leukotriene or leukotriene metabolite, such as LTA4 and LTB4, by 25%. 30%, 35% or greater when compared to pre -treatment leukotriene levels, and maintains such a reduction. For example, the dosing schedules are contemplated to maintain a reduction of at least 27%, 30%, 32%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90% or 95% compared to pre-treatment levels. The leukotriene level can be measured in a blood sample after stimulating the sample with a calcium ionophore, as described in Example 1. Biological samples in which to measure leukotriene levels include blood, serum, plasma or urine. The mean plasma concentrations are calculated by dividing the Area Under the Plasma Level vs. Time Curve (AUC) by the time period over which the samples are taken, e.g. (AUC(0-24 hrs)/24.

The dosing schedules of the invention are contemplated for use in treating (prophylactic and/or therapeutic) inflammatory diseases and cardiovascular diseases (including but limited to asthma) associated with FLAP activity or FLAP levels or with other members of the leukotriene pathway, such as LTB4 and LTA4. The invention also provides dosing schedules and formulations to treat inflammatory and cardiovascular disease states. The DG-031 dosing schedules of the invention include treating inflammatory diseases and cardiovascular diseases associated with leukotriene pathway members such as FLAP, arachidonate 5-lipoxygenase (5-LO), leukotriene A4 hydrolase (LTA4H), leukotriene B4 12-hydroxy dehydrogenase (LTB4DH) and potentially also leukotriene C4 synthase (17c45)); receptors and/or binding agents of the enzymes; and receptors for the leukotrienes LTA4, LTB4, LTC4, LTD4, LTE4, Cys LTl, Cys LT2, including leukotriene B4 receptor 1 (BLTl), leukotriene B4 receptor 2 (BLT2), and potentially cysteinyl leukotriene receptor 1 (CysLTRl), and cysteinyl leukotriene receptor 2 (CysLTR2).

The invention also contemplates treating and palliating or preventing inflammatory disease states such as rheumatoid arthritis, psoriatic arthritis, inflammatory arthritis, osteoarthritis, inflammatory joint disease, autoimmune disease including autoimmune vasculitis, multiple sclerosis, lupus, diabetes (e.g., insulin diabetes), inflammatory bowel disease, inflammatory eye disease, transplant rejection, graft vs. host disease, and inflammatory conditions resulting from strain, sprain, cartilage damage, trauma, orthopedic surgery, infection or other disease processes, psoriasis, eczema, allergies, acute or chronic lung injury including interstitial lung disease, acute respiratory disease syndrome, pulmonary hypertension, emphysema, cystic fibrosis, pulmonary fibrosis and asthma, acute and chronic glomerulonephritis, uveitis, endometriosis, cute pancreatitis, chronic fatigue syndrome, fibromyalgia, and Kawasaki's disease, and inflammatory eye disease.

Other diseases to prevent or palliate include cardiovascular disease such as myocardial infarction; transient ischemic attack, transient monocular blindness or stroke, or susceptibility to stroke; methods of treatment for claudication, PAOD or susceptibility to PAOD; methods of treatment for acute coronary syndrome (e.g., unstable angina, non-ST-elevation myocardial infarction (NSTEMI) or ST- elevation myocardial infarction (STEMI)); methods for reducing risk of MI, stroke or PAOD in persons with asymptomatic ankle/brachial index less than 0.9; methods for decreasing risk of a second myocardial infarction or stroke; methods of treatment for atherosclerosis, such as for patients requiring treatment (e.g., angioplasty, stents, revascularization procedure) to restore blood flow in arteries (e.g., coronary, carotid, and/or femoral arteries); methods of treatment for asymptomatic ankle/brachial index of less than 0.9; and/or methods for decreasing leukotriene synthesis (e.g., for treatment of myocardial infarction, stroke or PAOD).

Serum CRP and MPO levels are individually known to be strong predictors of risk for cardiovascular disease such as myocardial infarction. The DG- 031 dosing schedules of the invention can also be used to reduce the levels of inflammatory markers, such as CRP and MPO, in a human. Particularly, the invention contemplates carrying out methods of reducing inflammatory markers comprising administering a dose or doses of DG-031 according to a dosing schedule of the invention to a human suffering from an inflammatory disorder, suffering from a cardiovascular disease or at risk for developing a cardiovascular disease. An increasing body of emerging evidence identifies serum CRP as a marker for cardiovascular morbidity/mortality, and correlates reductions in serum CRP to better clinical outcomes. (See, e.g., Ridker et al, N.Engl. J. Med. 352(1): 20- 28 (2005); Nissen et al., N. Engl. J. Med. 352(1): 29-38 (2005); and Pearson et al., Circulation 107: 499-511 (2003).) Serum CRP in excess of 3.0 mg/L is considered high risk; from 1.0 to 3.0 average risk; and below 1 mg/L low risk. (Pearson et al.) Compositions and methods of the invention provide tools for reducing serum CRP. Reductions in CRP can be measured on a concentration basis, where compositions and methods that achieve CRP below 3.0 mg/L are preferred; with still more preferred targets of 2.75 mg/L, 2.5 mg/L, 2.25 mg/L, 2.0 mg/L, 1.75 mg/L, 1.5 mg/L, 1.25 mg/L, 1.0 mg/L, 0.75 mg/L, and 0.5 mg/L. Reductions in CRP also can be measured on a percentage basis, where clinical effectiveness is evaluated as a percentage reduction in CRP in a patient compared to before treatment with a dosing schedule of the invention. Depending on the initial CRP measurement, DG-031 dosing schedules and methods that reduce CRP anywhere from 10%-90% or more are contemplated, e.g., reductions of 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, 65%, 70%, 75%, 80%, or any target in between these values.

MPO also inactivates protease inhibitors and consumes nitric oxide, all of which escalate the inflammatory response (Eiserich et al, Science 296:2391-4, 2002). MPO has been shown to be elevated in patients with documented coronary artery disease (CAD) and within atherosclerotic lesions that are prone to rupture (Zhang et al. JAMA, 286:2136-2142, 2001; Sugiyama et al, Am J Pathol, 158:879- 9, 2001). MPO is also elevated in patients with chest pain and predictive of subsequent cardiovascular events at 3 and 6 months (Brennan N Engl J Med.,

349:1595-604, 2003). Reductions in MPO can be measured on a percentage basis, where clinical effectiveness is evaluated as a percentage reduction in MPO in a patient compared to prior treatment with a dosing schedule of the invention. Depending on the initial MPO measurement, DG-031 dosing schedules and methods that reduce MPO anywhere from 10%-90% or more are contemplated, e.g., reductions of 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, 65%, 70%, 75%, 80%, or any target in between these values.

The identification of a human in need of treatment for CRP or MPO reduction can be based on a variety of factors described herein, including genetic factors, CRP measurements, MPO measurements and measurements of other inflammatory markers, and measurements of non-genetic and non- inflammatory markers for risk of cardiovascular disease. In one variation, the method includes selecting for the administering step a human subject at risk for a disease or condition selected from the group consisting of myocardial infarction, acute coronary syndrome, stroke, or peripheral arterial occlusive disease.

In still another variation, the monitoring of markers of inflammation is used to adjust the dosing schedule for those individuals suffering from inflammatory diseases and those individuals at risk of or suffering from cardiovascular disease. For example, dose or dosing of a DG-031 is increased if serum CRP, and/or MPO and/or serum or urinary leukotriene measurements do not decrease to a target level, such as a level equivalent to the bottom 50 percentile, 40 percentile, 30 percentile, 20 percentile, 10 percentile, 1 percentile of a population, or other target percentile in between these exemplary targets. As described above, monitoring also can be used to adjust dosing to achieve a target level of serum CRP or MPO level, or to achieve a target percentage reduction in CRP or MPO for a particular human subject.

Target Populations

In an embodiment of the invention, the dosing schedule of DG-031 of the invention may be administered to a human at risk for cardiovascular disease, such as MI, ACS, stroke or PAOD. Increased risk for MI, ACS, stroke or PAOD in individuals may have increased production of leukotrienes (e.g., LTA4 , LTB4, LTC4, LTD4, LTE4, ). For example, the increased production of leukotrienes may be in the arterial vessel wall or in bone-marrow derived inflammatory cells within the blood and/or arterial vessel wall.

In another embodiment, the DG-031 dosing schedule is administered to humans having elevated levels of other inflammatory markers. An "elevated inflammatory marker," as used herein, is the presence of an amount of an inflammatory marker that is greater, by an amount that is statistically significant, than the amount that is typically found in control individual(s) or by comparison of disease risk in a population associated with the lowest band of measurement (e.g., below the mean or median, the lowest quartile or the lowest quintile) compared to higher bands of measurement (e.g., above the mean or median, the second, third or fourth quartile; the second, third, fourth or fifth quintile). An "inflammatory marker" refers to a molecule that is indicative of the presence of inflammation in an individual, for example, an end product of the leukotriene pathway, such as LTB4, C-reactive protein (CRP), serum sCD40L, serum amyloid A, fibrinogen, a leukotriene, a leukotriene metabolite, interleukin-6, tissue necrosis factor-alpha, a soluble vascular cell adhesion molecule (s VCAM), a soluble intervascular adhesion molecule (sICAM), E-selectin, matrix metalloprotease type-1, matrix metalloprotease type-2, matrix metalloprotease type-3, matrix metalloprotease type-9, myeloperoxidase (MPO), and N-tyrosine. The invention also provides for administering the DG-031 doising schedule for treatment of a disorder associated with inflammation such as asthma, allergic inflammation, acute inflammation and chronic inflammation. The invention also provides for administering the DG-031 dosing schedule of the invention to a human having a genetic risk factor for CAD or MI. Human subjects having a polymorphism or haplotype in the FLAP gene or LTA4H gene that is associated with risk of MI or CAD. Exemplary FLAP haplotypes that are associated with risk of MI or CAD are HapA, HapB, HapC, which are described in the aforementioned patent documents. Exemplary LTA4H haplotypes that are associated with risk of MI or CAD are HapK, HapL and HapQ, and surrogate haplotypes thereof, which are described in detail in the aforementioned patent documents.

The invention further provides for administering the DG-031 dosing schedule to humans having at least one family or medical history risk factor such as diabetes; hypertriglyceridemia, hypertension; high blood pressure hypercholesterolemia; elevated triglycerides; elevated lp(a); obesity; ankle/brachial index (ABI) less than 0.9; a past or current smoker; low level of physical activity, waist-to-hip circumference ration, transient ischemic attack; transient monocular blindness; carotid endarterectomy; asymptomatic carotid stenosis; claudicatioin; limb ischemia leading to gangrene, ulceration or amputation; a vascular or peripheral artery revascularization graft; increased serum LDL cholesterol and/or decreased HDL cholesterol; serum total cholesterol >200 mg/dl, increased leukotriene synthesis; and/or at least one previous myocardial infarction, ACS, stable angina, previous transient ischemic attack, previous MI, transient monocular blindness, or stroke, asymptomatic carotid stenosis or carotid endarterectomy, atherosclerosis, requires treatment for restoration of coronary artery blood flow (e.g., angioplasty, stent, revascularization procedure).

The invention also provides for administering the DG-031 dosing schedule to women of any age, men and women over the age of 40 years, such as those over the age of 45, 50, 55, 60, 65, 70, 75 and 80 and human subjects that have a race that includes black African ancestry such as persons of African descent or lineage. Black African ancestry may be determined by self reporting as African- Americans, Afro- Americans, Black Americans, being a member of the black race or being a member of the negro race. For example, African Americans or Black Americans are those persons living in North America and having origins in any of the black racial groups of Africa. For example, self-reported persons of black African ancestry may have at least one parent of black African ancestry or at least one grandparent of black African ancestry. Human subjects having a race that includes black African ancestry may also be determined by genetic analysis. Genetic analysis of ancestry may be carried out using unlinked microsatellite markers such as those set out in Smith et al. Am J Hum Genet 74, 1001-13 (2004).

Other target populations expected to benefit from the invention are populations affected by COPD, IBD, asthma, nocturnal asthma, or other diseases for which elevated leukotrienes are a symptom.

Monitoring Effectiveness of Dosing Schedule

Measurement of the level of a leukotriene or inflammatory marker before treatment during and/or after treatment is a method of determining the effectiveness of treatment with the DG-031 dosing schedule of the invention. The efficacy of the dosing schedule is indicated by a decrease in the level of the leukotriene or inflammatory marker, that is, a level of the inflammatory marker during or after treatment that is significantly lower (e.g., significantly lower), than the level of inflammatory marker before treatment (baseline level), is indicative of efficacy. Representative inflammatory markers include: a leukotriene (e.g., LTB4, LTA4), a leukotriene metabolite, C-reactive protein (CRP), serum amyloid A, fibrinogen, interleukin-6, tissue necrosis factor-alpha, apolipoprotein Al and B levels, fibrinogen, ferritin, soluble vascular cell adhesion molecules (s VCAM), soluble intervascular adhesion molecules (sICAM), E-selectin, matrix metalloprotease type-1, matrix metalloprotease type-2, matrix metalloprotease type-3, matrix metalloprotease type-9, myeloperoxidase (MPO), and N-tyrosine. In a preferred embodiment, the marker is CRP or MPO

One of the preferred inflammatory markers to monitor is serum C- reactive protein (CRP). Generally CRP is measured in serum samples using commercially available enzyme-linked immunosorbent assays (EIA). Consistent across multiple published studies is the finding of a correlation between increased risk for coronary artery disease with increased serum CRP. For example, in the Women's Health Study, CRP was measured in 27,939 apparently healthy American women. The cut-off points for quintiles of serum CRP in women were: less than or equal to 0.49, more than 0.49 to 1.08, more than 1.08 to 2.09, more than 2.09 to 4.19, and more than 4.19 mg CRP per liter, see Ridker, P.M. et al., New England. J. Med., 347: 1557-1565 (2001). In comparison to the lowest quintile, and even when adjusting for age, every quintile more than 0.49 mg CRP per liter was associated with increased risk for coronary heart disease with the highest relative risk of 4.5 seen for those women in the highest quintile of serum CRP (more than 4.19 mg CRP per liter). A similar correlation between increased serum CRP and increased risk for coronary heart disease in women has been reported (Ridker, P.M et al., New Eng. J. Med., 342:836-843 (2000) and Bermudez, E.A. et .al., Arterioscler. Thromb. Vase. Biol, 22: 1668-1673 (2002)). Men also show a correlation between increased serum inflammatory markers such as CRP and increased risk for coronary heart disease as previously reported (Doggen, C.J.M. et al., J.. Internal Med., 248:406-414 (2000) and Ridker, P.M. et al., New England. J. Med., 336: 973-979 (1997)). Quintiles for serum CRP as reported by Doggen et al., were less than 0.65, more than 0.65 to 1.18, more than 1.18 to 2.07, more than 2.07 to 4.23, and more than 4.23 mg CRP per liter. Unlike women, elevated serum CRP correlates with increased relative risk for coronary heart disease only in the 4th and 5th quintiles of CRP (relative risk of 1.7x and 1.9x, respectively). Elevated CRP or other serum inflammatory markers is also prognostic for increased risk of a second myocardial infarct in patients with a previous myocardial infarct (Retterstol, L. et al., Atheroscler., 160: 433-440 (2002)).

Another preferred method of monitoring the effectiveness of the treatment according to a dosing schedule of the invention is by assessing a level of a leukotriene metabolite (e.g., LTB4, LTA4) in the individual (e.g., in a sample of blood, serum, plasma or urine). The invention also encompasses assessing the level of leukotriene metabolite by stimulating production of a leukotriene or a leukotriene metabolite in a test sample from the individual (e.g., a sample comprising neutrophils), using a calcium ionophore, and comparing the level of the leukotriene or leukotriene metabolite with a control level such as a level of the leukotriene or leukotriene metabolite assessed during or after treatment. A level that is significantly lower during or after treatment, than before treatment, is indicative of efficacy of the treatment according to the dosing schedule. Similarly, the invention encompasses methods of assessing response to treatment, by assessing a level of an inflammatory marker in the individual before treatment, and during or after treatment. A level of the inflammatory marker during or after treatment, that is significantly lower than the level of inflammatory marker before treatment, is indicative of efficacy of the treatment. Because the level of inflammatory markers can be elevated in individuals who are in the target populations of the invention, an assessment of the level of inflammatory markers of the individual both before, and during, treatment according to the DG-031 dosing schedule of the invention will indicate whether the treatment has successfully decreased production of leukotrienes in the arterial vessel wall or in bone-marrow derived inflammatory cells. For example, in one embodiment of the invention, an individual who is a member of a target population as described above (e.g., an individual at risk for MI, ACS, stroke or PAOD, such as an individual who is at-risk due to a FLAP haplotype) can be assessed for response to treatment with a leukotriene synthesis inhibitor, by examining leukotriene levels or leukotriene metabolite levels in the individual. Blood, serum, plasma or urinary leukotrienes (e.g., leukotriene B4 or E4), or ex vivo production of leukotrienes (e.g., in blood samples stimulated with a calcium ionophore to produce leukotrienes), or leukotriene metabolites, can be measured before, and during or after treatment according the dosing schedule. The leukotriene or leukotriene metabolite level before treatment is compared with the leukotriene or leukotriene metabolite level during or after treatment. The efficacy of treatment is indicated by a decrease in leukotriene production: a level of leukotriene or leukotriene metabolite during or after treatment that is significantly lower than the level of leukotriene or leukotriene metabolite before treatment, is indicative of efficacy. A level that is lower during or after treatment can be shown, for example, by decreased serum or urinary leukotrienes, or decreased ex vivo production of leukotrienes, or decreased leukotriene metabolites. A level that is "significantly lower", as used herein, is a level that is less than the amount that is typically found in control individual(s), or is less in a comparison of disease risk in a population associated with the other bands of measurement (e.g., the mean or median, the highest quartile or the highest quintile) compared to lower bands of measurement (e.g., the mean or median, the other quartiles; the other quintiles).

Pharmaceutical Compositions

The present invention provides compositions and formulations of the leukotriene synthesis inhibitor, DG-031. For instance, DG-031 can be formulated with one ore more physiologically acceptable carriers or excipients to prepare a pharmaceutical composition. The carrier and composition can be sterile injection, inhalation or ocular administration is preferred. The composition can be a solution, suspension, emulsion, tablet, pill, capsule, sustained release formulation, or powder. The composition can be formulated as a suppository, with traditional additives such as fats, triglycerides or polyethoxlated polymers. Oral formulation can include standard carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, polyvinyl pyrollidone, sodium saccharine, cellulose, magnesium carbonate, etc.

A preferred composition of the present invention is a compressed tablet for oral administration that consists essentially of 250 mg of DG-031, 40 mg of corn starch, 96.24 mg of microcrystalline cellulose, 1.24 mg, 10 mg providone 25 [poly(l- vinyl-2-pyrrolidinone 25], 2.52 mg magnesium stearate and purified water having a comprising a film coating consisting essentially of 6 mg methylhydroxypropylcellulose, 1.5 mg polyethylene glycol 4000, 2.5 mg titanium oxide and purified water. The amounts of these ingredients may vary +/- 10%. In some variations of the invention, two or more 250 mg DG-031 tablets are administered simultaneously to provide a dose of 500 mg, 750 mg or 1000 mg. High shear wet granulation is a preferred process for manufacturing the tablets where the primary process steps, which would be familiar to one skilled in the art, are blending the powders, followed by high shear wet granulation, wet milling, fluid bed drying, dry milling, tableting and finally pan coating of the tablets. Suitable pharmaceutically acceptable carriers include but are not limited to water, buffered saline solutions (e.g., NaCl), saline, buffered saline, alcohols, glycerine, ethanol, gum arabic, vegetable oils, benzyl alcohols, polyethylene glycols, gelatin, carbohydrates such as lactose, amylose or starch, dextrose, magnesium stearate, talc, fumed silica, liquid petrolatum, fatty acid esters, hydroxyproplmethyl, polyvinyl pyrolidone, other pharmaceutically acceptable polymers, as well as combinations thereof. The pharmaceutical preparations can, if desired, be mixed with auxiliary agents, e.g., lubricants, cationic crosslinking agents, inert diluents, alkalizing agents, acidifying agents, surfactants, polar solvents, preservatives, stabilizers, wetting agents, emulsifϊers, salts for influencing osmotic pressure, buffers, coloring, flavoring and/or aromatic substances and the like which do not deleteriously react with the active agents.

The composition of the present invention may further include other materials such as bulking agents, disintegrating agents, anti-adherants and glidants, lubricants, wetting or emulsifying agents and binding agents. The composition, if desired, can also contain minor amounts pH buffering agents.

Bulking agents include, but are not limited to, microcrystalline cellulose (e.g., Avicel.RTM., FMC Corp., Emcocel.RTM., Mendell Inc.), starches, mannitol, xylitol, dicalcium phosphate (e.g. Emcompress, Mendell Inc.) calcium sulfate (e.g. Compactrol, Mendell Inc.), lactose, sucrose (Dipac, Amstar, and Nutab, Ingredient Technology), dextrose (Emdex, Mendell, Inc.), sorbitol, cellulose powder (Elcema, Degussa, and Solka Floe, Mendell, Inc.) The bulking agent may be present in the composition in an amount of from about 5 wt. % to about 90 wt. %, preferably from about 10 wt. % to about 50 wt. %.

Binding agents which may be employed include, but are not limited to polyvinyl pyrrollidone, starch, methylcellulose, hydroxypropyl methylcellulose, carboxymethyl cellulose, sucrose solution, dextrose solution, acacia, tragacanth and locust bean gum. The binding agent may be present in the composition in an amount of from about 0.2 wt. % to about 10 wt. %, preferably from about 0.5 wt. % to about 5 wt. %.

Disintegrating agents which may be included in the composition include, but are not limited to, microcrystalline cellulose, starches, crospovidone (e.g. Polyplasdone XL, International Specialty Products.), sodium starch glycolate (Explotab, Mendell Inc.), and crosscarmellose sodium (e.g., Ac-Di-SoI, FMC Corp.). The disintegrating agent may be present in the composition in an amount of from about 0.5 wt. % to about 30 wt %, preferably from about 1 wt. % to about 15 wt. %.

Antiadherants and glidants which may be employed in the composition include, but are not limited to, talc, corn starch, silicon dioxide, sodium lauryl sulfate, and metallic stearates. The antiadherant or glidant may be present in the composition in an amount of from about 0.2 wt. % to about 15 wt. %, preferably from about 0.5 wt. % to about 5 wt. %.

Lubricants which may be employed in the composition include, but are not limited to, magnesium stearate, calcium stearate, sodium stearate, stearic acid, sodium stearyl fumarate, hydrogenated cotton seed oil (Sterotex), talc, and waxes, including but not limited to, beeswax, carnuba wax, cetyl alcohol, glyceryl stearate, glyceryl palmitate, glyceryl behenate, hydrogenated vegetable oils, and stearyl alcohol. The lubricant may be present in an amount of from about 0.2 wt. % to about 20 wt. %, preferably from about 0.5 wt. % to about 5 wt. %.

Modes of Administration

Methods and routes of administering DG-031 , include but are not limited to, intradermal, pulmonary/inhalants, transdermal, transmucosal, intramuscular, intraperitoneal, intraocular, intravenous, subcutaneous, topical, oral, anal, vaginal, inhalation and intranasal. For repeated dosing, the preferred method of delivery is oral administration of a solid tablet, gel liquid or capsule.

The invention also includes oral administration of multiparticulate capsules and osmotic tablets or other osmotic delivery systems (See U.S. Patent No. 6,110,498). In addition, the invention includes subcutaneous or parenteral administration by continuous infusion of DG-031 using pumps, infusions and implants. The invention includes transdermal administration such as applying a transdermal patch. In some variation of the invention, the leukotriene synthesis inhibitor compositions are administered as part of a combinatorial therapy with other agents. For example, most cardiovascular patients, and all which participated in the clinical study described in Example 2, were administered statins (HMG reductase inhibitors). As treatment with DG-031 was effective to reduce LTB4, CRP and MPO levels in patients concurrently being administered statins, the invention includes coadministering DG-031 and a statin for a more effective therapeutic result as described in International Application No. PCT/US2005/003312, filed January 31, 2005, incorporated by reference herein in its entirety. Likewise, the invention includes compositions that comprise a controlled or sustained release formulation of a leukotriene inhibitor in combination with a statin.

The composition can be formulated in accordance with the routine procedures known to those of skill in the art as a pharmaceutical composition adapted for administration to human beings. For example, compositions for intravenous administration typically are solutions in sterile isotonic aqueous buffer. Where necessary, the composition may also include a solubilizing agent and a local anesthetic to ease pain at the site of the injection. Generally, the ingredients are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilized powder or water free concentrate in a hermetically sealed container such as an ampule or sachette (vial) with a label which indicates the quantity of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water, saline or dextrose/water. Where the composition is administered by injection, and the drug is present as a lyophilized solid, an ampule of sterile water for injection or saline can be provided so that the ingredients may be mixed prior to administration.

For topical application, nons-sprayable forms, viscous to semi-solid or solid forms comprising a carrier compatible with topical application and having a dynamic viscosity preferably greater than water, can be employed. Suitable formulations include but are not limited to solutions, suspensions, emulsions, creams, ointments, powders, enemas, lotions, liniments, salves, aerosols, etc., which are, if desired, sterilized or mixed with auxiliary agents, e.g., preservatives, stabilizers, wetting agents, buffers or salts for influencing osmotic pressure, etc. The agent may be incorporated into a cosmetic formulation. For topical application, also suitable are sprayable aerosol preparations wherein the active ingredient, preferably in combination with a solid or liquid inert carrier material or diluent, is packaged in a squeeze bottle or in admixture with a pressurized volatile, normally gaseous propellant. Agents described herein can be formulated as neutral or salt forms, or esters or other chemical derivatives that act as prodrugs in vivo, metabolized into the active agent.

Pharmaceutically acceptable salts include those formed with free amino groups such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with free carboxyl groups such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, procaine, etc.

The agents are administered in a therapeutically effective amount. The amount of agents which will be therapeutically effective in the treatment of a particular disorder or condition will depend on the nature of the disorder or condition, and can be determined by standard clinical techniques. The optimal dose will also depend on the fraction of the drug delivered to the systemic circulation after delivery via a given administration route, as well as drug distribution, metabolism and excretion. In addition, in vitro or in vivo assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed in the formulation will also depend on the route of administration, and the seriousness of the symptoms, and should be decided according to the judgment of a practitioner and each patient's circumstances. Effective doses may be extrapolated from dose-response curves derived from in vitro or animal model test systems.

The invention also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the pharmaceutical compositions of the invention. Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use of sale for human administration. The pack or kit can be labeled with information regarding mode of administration, sequence of drug administration (e.g., separately, sequentially or concurrently), or the like. The pack or kit may also include means for reminding the patient to take the therapy. The pack or kit can be a single unit dosage of the combination therapy or it can be a plurality of unit dosages. In particular, the agents can be separated, mixed together in any combination, present in a single vial or tablet. Agents assembled in a blister pack or other dispensing means is preferred. For the purpose of this invention, unit dosage is intended to mean a dosage that is dependent on the individual pharmacodynamics of each agent and administered in FDA approved dosages at recommended dosing intervals.

Composition Manufacture The compositions of the present invention may be made by a direct compression method, or by a wet granulation method or other methods known in the art such as roller compaction.

In the direct compression method, the at least one pharmaceutically active agent and other ingredients are sieved through a stainless steel screen to remove lumps and achieve some consistency in particle size. The sieved materials then are charged to a suitable blender, and blended. The blend then is compressed into tablets on a rotary press using appropriate tooling. The compressed tablets may be coated, such as for physical appearance, environmental protection, or programmed/controlled release.

In the wet granulation method, at least one pharmaceutically active agent and other ingredients are granulated with a granulating fluid (e.g., isopropyl alcohol, ethyl alcohol, and/or water) in a planetary mixer, high shear mixer, or fluidized bed granulator. Binding agents may be contained in the granulating fluid, or may be in the dry mix. The wet granules are dried in an oven or fluidized bed dryer, and then sieved through a suitable screen to obtain free flowing granules. The resulting granules were blended with a suitable lubricant and glidant, and the lubricated granules are compressed into tablets on a rotary press using appropriate tooling. If desired, a coating can be applied on the compressed tablets such as for controlled or sustained release.

EXAMPLES

Example 1 Increased LTB4 Production In Activated Neutrophils From MI Patients

A principal bioactive product of one of the two branches of the 5-LO pathway is LTB4. To determine whether the patients with history of MI have increased activity of the 5-LO pathway compared to controls, the LTB4 production in isolated blood neutrophils was measured before and after stimulation in vitro with the calcium ionophore, ionomycin. No difference was detected between the LTB4 production in resting neutrophils from MI patients or controls (results not shown). In contrast, the LTB4 generation by neutrophils from MI patients stimulated with the ionophore was significantly greater than by neutrophils from controls at 15 and 30 minutes, respectively. Moreover, the observed increase in the LTB4 release was largely accounted for by male carriers of haplotype A4, whose cells produced significantly more LTB4 than cells from controls (P value =0.0042) (Table 4). As shown in Table 4, there was also a heightened LTB4 response in males who do not carry HapA but of borderline significance. This could be explained by additional variants in the FLAP gene that have not been uncovered, or alternatively in other genes belonging to the 5-LO pathway, that may account for upregulation in the LTB4 response in some of the patients without the FLAP at-risk haplotype. As shown in Table 4, differences in LTB4 response were not detected in females. However, due to a small sample size this cannot be considered conclusive. Taken together, the elevated levels of LTB4 production of stimulated neutrophils from male carriers of the at-risk haplotype suggest that the disease associated variants in the FLAP gene increase FLAP's response to factors that stimulate inflammatory cells, resulting in increased leukotriene production and increased risk for MI. Isolation and activation of peripheral blood neutrophils

50ml of blood were drawn into EDTA containing vacutainers from 43 MI patients and 35 age and sex matched controls. All blood was drawn at the same time in the early morning after 12 hours of fasting. The neutrophils were isolated using Ficoll-Paque PLUS (Amersham Biosciences). Briefly, the cell pellets from the Ficoll gradient were harvested and the red blood cells were subsequently lysed in 0.165 M NH4CL for 10 minutes on ice. After washing with PBS, neutrophils were counted and plated at 2xlO⁶ cells/ml in 4ml cultures of 15% Fetal calf serum (FCS) (GIBCO BRL) in RPMI- 1640 (GIBCO BRL). The cells were then stimulated with maximum effective concentration of ionomycin (lμ M). At 0, 15, 30, 60 minutes post-ionomycin addition, 600μl of culture medium was aspirated and stored at -8O⁰C for the measurement of LTB4 release as described below. The cells were maintained at 37⁰C in a humidified atmosphere of 5% CO2/95% air. All samples were treated with indomethasine (lμ M ) to block the cyclooxygenase enzyme. The experimental conditions are described above. Ionomycin-induced release of LTB4 in neutrophils

LTB4 Immunoassay Assay Design was used to quantitate LTB4 concentration in supernatant from cultured ionomycin stimulated neutrophils. The assay used is based on the competitive binding technique in which LTB4 present in the testing samples (200 μl) competes with a fixed amount of alkaline phosphatase- labelled LTB4 for sites on a rabbit polyclonal antibody. During the incubation, the polyclonal Ab becomes bound to a goat anti-rabbit Ab coated onto the microplates. Following a wash to remove excess conjugate and unbound sample, a substrate solution is added to the wells to determine the bound enzyme activity. The color development is stopped and the absorbance is read at 405 nm. The intensity of the color is inversely proportional to the concentration of LTB4 in the sample. Each LTB4 measurement using the LTB4 Immunoassay, was done in duplicate. Table 4: LTB4 levels after ionomycin stimulation of isolated neutrophils^*1

After 15 Minutes After 30 Minutes

Phenotype (n) Mean (SD) P value Mean (SD) P value

Controls (35) 4.53 (1.00) 4.67 (0.88)

Males (18) 4.61 (1.10) 4.68 (1.07)

Females (17) 4.51 (0.88) 4.67 (0.62)

MI (41) 5.18 (1.09) 0.011 5.24 (1.06) 0.016

Carriers(16) 5.26 (1.09) 0.027 5.27 (1.09) 0.051

Non-carriers (24) 5.12 (1.08) 0.040 5.22 (1.03) 0.035

MI males (28) 5.37 (1.10) 0.0033 5.38 (1.09) 0.0076

Carriers(lO) 5.66 (1.04) 0.0042 5.58 (1.12) 0.013

Non-carriers (18) 5.20 (1.09) 0.039 5.26 (1.05) 0.041

MI females (13) 4.78 (0.95) 0.46 4.95 (0.92) 0.36

Carriers(6) 4.59 (0.80) 0.90 4.75 (0.82) 0.85

N.on-carriers (7) 4.94 (1.04) 0.34 5.12 (0.96) 0.25

^aMean + SD of log-transformed values of LTB4 levels of ionomycin- stimulated neutrophils from MI patients and controls. Results are shown for two time points: 15 and 30 minutes. The results for males and females and for MI male and female carriers and non-carriers of the at-risk haplotype HapA are shown separately. Two-sided p values corresponding to a standard two-sample test of the difference in the mean values between the MI patients, their various sub-cohorts and the controls are shown.

Example 2 Composition of DG-031 Tablets

In the clinical studies described in Example 3, the subjects were orally administered DG-031 as a film-coated tablet containing 250 mg of active drug substance. The 250 mg tablet was a round tablet, 410 mg weight. The tablets were stored at 15-30⁰C. Table 5 lists the components of the DG-031 composition (250 mg Tablets) used in the clinical studies. Table 5: Drug Product Composition (250 mg Tablets)

Example 3

Comparative Study Investigating 1000 mg QD and 500 mg BID Dosing Regimes

A comparative clinical study was carried out to investigate the optimal time of the day for administering DG-031 to achieve the most effective LTB4 plasma lowering effect. The DG-031 pharmacokinetics and pharmacodynamics of the inflammatory biomarker LTB4 following oral administration of 500 mg DG-031 twice daily (BID; every 12 hours) or 1000 mg DG-031 once daily (QD) were analyzed seven consecutive days in healthy subjects.

Study Design:

This was an open label multiple dose study designed to evaluate the pharmacokinetics, pharmacodynamics, safety and tolerability of DG-031 in 24 healthy male and female subjects. Enrolled subjects were randomized to receive either 500 mg DG-031 orally every 12 hours (500 mg; Q12H) or 1000 mg DG-031 orally once daily (1000 mg QD) for 7 consecutive days. Two 250 mg DG-031 tablets, as described in Example 2, were administered to equal a 500 mg dose. Four 250 mg DG-031 tablets, as described in Example 2, were administered to equal a 1000 mg dose.

Subjects were confined to the Clinical Pharmacology Research Unit (CPRU) for approximately 36 hours prior to study drug administration on Day 1 (day 1st dose of study drug is administered) until 24 hours following the last dose of study drug on Day 7. Subjects returned to the CPRU 7 to 10 days (Days 14-17) following the last dose of study drug on day 7.

For the 500 mg dosing regime, DG-031 was administered to the randomized subjects at 0600-0800 (6:00 am - 8:00 am) after an overnight fast and again 12 hours ± 15 minutes later between 1800-2000 (6:00 pm - 8:00 pm) on Days 1 through 7. The 500 mg Q12H evening dose was administered at least 2 hours before or after food consumption. The DG-031 was administered to a given subject at the same time (± 15 minutes) each day.

For the 1000 mg dosing regime, DG-031 was administered to the randomized subjects after an overnight fast between 0600-0800 (6:00 am - 8:00 am) on Days 1 through 7.

Blood samples for serum chemistry, hematology and urinalysis and ECGs were obtained at several time points during the study for safety evaluation. Blood samples for DG-031 plasma pharmacokinetic analysis and measurement of the inflammatory biomarker LTB4 were collected on Day 1, 2, 5, 7 and 8. Urine samples for measurement of urine LTE4 levels were collected on Day -1 and Day 7. Similarly, the measurement of the inflammatory marker MPO in the blood samples may be carried out as described for LTB4.

Subject Population: The study was carried out with healthy volunteers (age 18-50 years, inclusive); males or females who were surgically sterile or at least 2 years postmenopausal. Subjects had a body mass index BMI of 18-32 kg/m². Subjects signed an informed consent document.

Pharmokinetics (PK) and Pharmodynamics (PD) Analysis Over a 24 hour period the pharmacokinetic profile (PK) and the pharmacodynamic profile was measured. PK was a measure of the DG-031 plasma concentration at various time points (indicated in hours). A summary of the PK analysis on day 7 is shown in Figure 1 (top panel). The peak DG-031 concentration (Cmax) was similar for the QD and BID dosing regimes, although the peaks occur at different times of the day. For the QD regime (solid lines), the Cmax ranges from 2 to 4 hours after administration. For the BID regime (dashed lines), the Cmax is at about 15 hours after morning administration (about 3 hours after evening administration). The average daily DG-031 concentration (area under curves defined by thin lines) was greater for the BID regime than for the QD regime. A summary of the morning Cmax concentration of by regime and study day is set out in Table 6.

The PD was a measure of the level of LTB4 in response to the DG-031 administration. Measurements of stimulated LTB4 production was obtained by treating a whole blood sample with a calcium ionophore, such as ionomycin, at various time points as described in Example 1. A summary of the PD analysis is shown in Figure 1 (lower panel). Unexpectedly, the QD regime (solid lines) provides more efficient LTB4 lowering than the BID regime (dashed lines) at Day 1.

The result that the QD regime had lower DG-031 plasma concentrations but greater LTB4 lowering effect compared to the BID regime was consistent and seen on both Day 1 and Day 7. A comparison of the BID and QD effects for all study subjects is depicted in Figure 2. The scatter plot shows paired, rank-ordered data for DG-031 plasma concentration (open circles) and LTB4 lowering effect (closed circles). The solid line represents the line around which all the dots would fall on to or close to (or randomly scattered around) in absence of any differences achieved by the QD vs. BID dosing regimens. The fact that all open circle data points (DG-031 concentration) except for one are above the line and all closed circle data points (LTB4 lowering) are below the line illustrates that for all subjects, the BID dosing results in higher plasma concentration of DG-031, while QD results in greater LTB4 lowering.

In addition, the results were statistically significant as shown in Table 7 below. Table 7 shows average concentration and average LTB4 lowering effect for BID and QD dosing regime, respectively, on Day 1 and Day 7. The BID dosing regime results in a statistically significant increased concentration of DG-031 compared with QD dosing, while the increased LTB4 lowering of the QD dosing regime compared with 500 mg BID dosing is statistically significant. Reported p- values are two-sided.

Example 4 Clinical Studies Comparing Other Dosing Regimes

Clinical Studies are carried out to compare the dosing regimes of the present invention with other dosing regimes known in the art. For example, the 1000 mg QD early dosing regime of the present invention may be compared to the DG-031 sustained release dosing regime or other dosing regimes that maximize trough concentrations of DG-031, such as those taught in PCT publication No. WO 2006/116349, which is incorporated by reference in its entirety.

The study is designed to investigate the effects of DG-031 on patients having a history of MI or CAD and/or are carriers of specific Mi-associated haplotypes in the FLAP and/or the LTA₄ hydrolase genes (See U.S. Patent Application No. 10/944,272 and PCT Application No. PCT/2004/030582, incorporated by reference in its entirety). Selection of subjects for the study is based on previous haplotype/genotype analysis.

Table 8: Genotypes used to derive at-risk variants of FLAP and LTA₄ Hydrolase

AIIeI AIIeI

Haplotype Allele SNP e SNP e SNP

A3 (FLAP gene) G SG13S25 T SG13S114 A SG13S32 AF (FLAP gene) G SG13S25 T SG13S114 K (LTA₄-OH gene) A SG12S25

The haplotypes carried by each individual are estimated using the program NEMO (version 1.01) and 902 in-house population controls, as previously described in Getarsdottir et ah, Nat. Genet. 35: 131-138, 2003. Additional study eligibility criteria include those set out in Table 9. Table 9 Study Eligibility Criteria

Treatment Groups

Patients who meet the study eligebility criteria are enrolled and randomized into different dose-level groups such as: 250mg/day therapy with DG-031 (250 mg QD), vs. dosing regimes of the invention, 500 mg/day therapy with DG-031 (250 mg BID) vs. dosing regime of the invention; 750 mg/day therapy with DG-031 (250 mg TID) vs. dosing regime of the invention, 750 mg/day therapy with DG-031 (375 mg BID) vs. dosing regime of the invention, 375 mg/day QD vs. dosing regime of the invention. Alternatively, the regimes of the invention are compared to placebo. The placebo tablets are identical in shape, color, form and taste to the active tablets except that they contained no active drug ingredients. Treatment with DG-031 or placebo is in addition to the subject's standard care, including all medications and treatment plan as prescribed by the subject's cardiologist prior to enrolment.

Endpoints

The primary objective of the study is to determine whether the different DG-031 dosing regimes have a statistically significant effect, compared to on one or more biomarkers of inflammation or MI risk, including: onomycin- induced LTB4 and MPO release by neutrophils ex vivo (as described in Examples 1 and 3); MPO, CRP, N-tyrosine, Lp-PLA₂ or amyloid A in serum, or LTE₄ in urine. A secondary objective of the study is to determine whether the effect of DG-031 is dose- dependent. A tertiary objective is to assess other biomarkers such as those set out in Table 10. Evaluation of safety and tolerability of the drug is also a primary endpoint.

Table 10

Assay Unit

Primary objectives

Amyloid A ng/ml

Hs-CRP pg/ml

LTE₄ in urine pg/ml

Lp-PLA₂ μg/ml

MPO in plasma ng/ml

N-tyrosine nM

LTB4 in whole bloodt pg/ml

LTB4 in w.b.*, corr. for wbct.t

MPO in whole blood ng/ml

MPO in w. b.*, corr. for wbc*

White blood cell counts 10⁹/L

CD40L pg/ml

Prostaglandin F pg/ml

Tertiary objectives

ICAM ng/ml

IL12p40 pg/ml

IL6 pg/ml

MCP-1 pg/ml

MMP 9 ng/ml

Oxidized - LDL mU/L

TNF-α pg/ml sE-Selectin ng/ml sP-Selectin ng/ml sVCAM ng/ml

*w.b. = whole blood tbaseline is not available for LTB4 measured using mass spectrometry tcorr. for wbc = corrected for white blood cell count

§WBC is not part of the primary objectives, but is included here due to the wbc correction used for Ll

MPO.

Safety Analysis

The study subjects having serious adverse events (SAE) or resulting in death during the trial are recorded. It will be determined if the SAE 's or death are consistent with underlying MI or CAD disorders. Common adverse events (and number of cases reported) include: colds, headaches, constipation / diarrhea, and vertigo. LDL-C is measured weekly after administration of the dosing regime. A significant dose-dependent increase in LDL-C may be observed with DG-031 administration. Example 5 Comparison of Pharmokinetic Analysis of Various DG-031 Dosing Regimes

Preliminary population pharmacokinetic (PK) analysis may also carried out to compare the effect of different dosing regimes to the doing regime of the invention (as described in Example 4). Based on visual inspection of the individual and mean time course of DG-031 concentrations, a simple one compartmental disposition model following first order oral absorption, and an absorption lag time is fit to the data. A mixed effects modeling approach is used. The pharmacokinetic model parameters (fixed effects) included oral clearance (CL), distribution volume (V), oral absorption (ka), and absorption lag (ti_ag). The statistical model includes inter-individual random effects as well as a residual error term. All inter-subject random effect parameters (η's) and residual error terms (ε) are assumed to be normally distributed with mean of zero and variance ω² and σ², respectively. Random effects on all pharmacokinetic model parameters are explored. Maximum likelihood estimates of the structural (θ's) and statistical model parameters are estimated using an approximation to the mixed effect log-likelihood as implemented in the NONMEM program (NONMEM Version Vl).

The influence of patient characteristics (covariates) on pharmacokinetic parameters are explored. Age, gender, body weight, and Cr<x (creatinine clearance) are included in the model as covariates if warranted.

Model selection will be done on the basis of the log-likelihood criterion (p<0.05) and visual inspection of goodness-of-fit plots. The difference in -2 times the log of the likelihood (-2LL) between a full and reduced model is asymptotically χ² distributed with degrees of freedom equal to the difference in number of parameters between the two models. For instance, a decrease of more than 3.84 in -2LL is considered significant at the p<0.05 level for 1 additional parameter. Standard errors of the parameter estimates are approximated using the asymptotic covariance matrix.

Blood samples are collected for determining DG-031 concentrations in a subset of patients. Samples are collected just prior to dosing and at 0.5, 1, 2, 4, and 6 hours following administration of the DG-031 dosing regime. The geometric mean DG-031 steady-state concentrations are calculated. Exposure -response analysis of biomarker data is also collected in the subjects. Example 6 Clinical Study Comparing Dosing Regimes in Patients Suffering From Asthma

Clinical studies are carried out to compare the dosing regimen of the present invention with other dosing regimes known in the art as described in detail in Example 4. The study is designed to investigate the effects of DG-031 on patients that suffer from asthma, especially those patients that suffer from nocturnal asthma.

The primary objective of the study is to determine whether the different DG-031 dosing regimes have a statistically significant lowering effect on one or more biomarkers of inflammation associated with asthma including bronchial or plasma LTB4 or urinary leukotrienes. Another objective of the study is to determine whether different DG-031 dosing regimes decrease the incidence of asthma attacks in the treated patients and/or increases lung function such as increasing bronchial peak flow or forced expiratory volume.

Example 7 Clinical Study Comparing DG-051 Dosing Regimes

Clinical studies comparing dosing regimes of DG-051 are carried out as described in Example 3 using DG-031 dosing regimes. One objective of this study is to evaluate whether morning dosing results in a Cmax peak between 6 A.M. and 2 P.M. with an improved LTB4 and MPO lowering effect. This study is carried out with dose escalation within the safety window for DG-051. Another objective of the study is determine if different DG-051 dosing regimes have a statistically significant lowring effect on other biomarkers of inflammation. An exemplary dose and dosing regimen would range 10 - 160 mg QD administered in the morning.

Example 8 Clinical Study Comparing Leukotriene Synthesis Inhibitor Dosing Regimes

Clinical Studies comparing dosing regimes of any leukotriene synthesis inhibitor are carried out as described in Example 3 using DG-031 dosing regimes. One objective of this study is to evaluate whether morning dosing results in a Cmax peak between 6 A.M. and 2 P.M. with an improved LTB4 or other inflammatory biomarker lowering effect, or improved efficacy for treatment of inflammatory diseases or conditions; and/or reduced side effects. These studies are carried out with dose escalation within the safety window for each drug. Another objective of the study is determine if different leukotriene synthesis inhibitor dosing regimes have a statistically significant lowing effect on other biomarkers of inflammation.

Numerous modifications and variations in the practice of the invention are expected to occur to those skilled in the art upon consideration of the presently preferred embodiments thereof. Consequently, the only limitations which should be placed upon the scope of the invention are those which appear in the appended claims.

Claims

1. A method of reducing leukotriene production in a human comprising administering to the human a composition that comprises a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof, wherein the composition is administered according to a dosing regimen that achieves a single daily peak plasma concentration of the inhibitor between the hours of 6 A.M. and 2 P.M.

2. Use of a leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, in the manufacture of a medicament for administration to a human for reducing leukotriene synthesis production in said human, wherein the medicament is formulated into a dose that is administered according to a dosing regimen that is effective to achieve a single daily peak plasma concentration of the inhibitor between the hours of 6 A.M. and 2 P.M.

3. A leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, for reducing leukotriene synthesis in a human, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration according to a dosing regimen that is effective to achieve a single daily peak plasma concentration of the inhibitor between the hours of 6 A.M. and 2 P.M.

4. The method or use or inhibitor according to any of claims 1-3, wherein the composition or dose is administered once daily.

5. The method or use or inhibitor according to claims 3 or 4, wherein the composition or dose is administered in the morning.

6. The method according to any one of claims 1, 4 or 5, wherein the leukotriene synthesis inhibitor is present in the composition in an amount effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 20%.

7. The use of any one of claims 2, 4 or 5, wherein the medicament is formulated into a dose that is effective to reduce evening peak leukotriene B4

(LTB4) production in adult human subjects by at least 20%.

8. A method of reducing leukotriene production in a human comprising administering to the human a composition that comprises a leukotriene synthesis inhibitor, wherein the composition is administered once daily in the morning, and wherein the leukotriene synthesis inhibitor is present in the composition in an amount effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 20%.

9. Use of a leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, in the manufacture of a medicament for administration to a human for reducing leukotriene production in said human, wherein the medicament is formulated into a dose that is administered once daily in the morning, and the dose is effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 20%.

10. A leukotriene synthesis inhibitor, or a pharmaceutically acceptable salt, ester, or prodrug thereof, for reducing leukotriene production in a human, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration once daily in the morning, and the dose is effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 20%.

11. A method of reducing leukotriene production in a human comprising administering a composition to the human in single daily dose in the morning, wherein the composition comprises 500-2500 mg of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof.

12. Use of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof, in the manufacture of a medicament for administration to a human for reducing leukotriene production in said human, wherein the medicament is formulated into a does that is administered once daily in the morning, and wherein the dose is in a range of 500-2500 mg of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof, inclusive.

13. A leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration once daily in the morning, and wherein the dose is in a range of 500-2500 mg of a leukotriene synthesis inhibitor or a pharmaceutically acceptable salt, ester, or pro-drug thereof, inclusive.

14. The method or use or inhibitor of any of claims 7- 13 wherein the composition or dose is administered between 6 A.M. and 12 noon.

15. The method or use or inhibitor of claim 14, wherein the composition or dose is administered between 6 A.M. and 9 A.M.

16. The method or use or inhibitor of claim 14, wherein the composition or dose is administered between 8 A.M. and 11 A.M.

17. The method or use or inhibitor of claim 14, wherein the composition or dose is administered between 10 A.M. and 12 noon.

18. The method or use or inhibitor of any of claims 1-17, wherein the composition or dose is in a range of 750-2000 mg of a leukotriene synthesis inhibitor, inclusive.

19. The method or use or inhibitor of any one of claims 1-17, wherein the composition or dose is in a range of 1000-1500 mg of a leukotriene syntheses inhibitor, inclusive.

20. The method or use or inhibitor of any of claims 1-17, wherein the composition or dose is in a range of 250-1500 mg of a leukotriene syntheses inhibitor, inclusive.

21. The method or use or inhibitor of any of claims 1-17, wherein the composition or dose is in a range of 750-1000 mg of a leukotriene syntheses inhibitor, inclusive.

22. The method or use or inhibitor of any of claims 1-17, wherein the composition or dose is in a range of 1000-1500 mg of a leukotriene syntheses inhibitor, inclusive.

23. The method or use or inhibitor of any one of claims 1-17, wherein the composition or dose comprises 1000 mg of a leukotriene synthesis inhibitor, inclusive.

24. The method or use or inhibitor according to any one of claims

1-23, wherein the leukotriene synthesis inhibitor inhibits a protein selected from the group consisting of 5 -lipoxygenase (5-LO), 5-lipoxygenase activating protein (FLAP), Leukotriene A4 hydrolase (LTA4H), Leukotriene C4 synthase, gamma- glutamyltranspeptidase, and leukotriene D4 dipeptidase.

25. The method or use or inhibitor according to claim 24, wherein the leukotriene synthesis inhibitor is selected from the inhibitors in Tables 1, 2 or 3.

26. The method or use or inhibitor according to claim 24, wherein the leukotriene synthesis inhibitor is a FLAP inhibitor.

27. The method or use or inhibitor according to claim 24, wherein the leukotriene synthesis inhibitor comprises a compound of the formula:

wherein Ar is selected from the group consisting of: aryl; heteroaryl; aryl substituted with from one to three substituents independently selected from the group consisting of halogen, loweralkyl, loweracyl, loweralkoxy, fluoroloweralkyl, fluoroloweralkoxy, hydroxy, hydroxy(Ci-C4) alkyl, formyl, formyl(d-C₄) alkyl, cyano, cyano(Ci-C₄) alkyl, benzyl, benzyloxy, phenyl, substituted phenyl, heteroaryl, heterocyclylalkyl, substituted heteroaryl, and nitro; and heteroaryl substituted with from one to three substituents independently selected from the group consisting of halogen, loweralkyl, loweracyl, loweralkoxy, fluoroloweralkyl, fluoroloweralkoxy, formyl, cyano, benzyl, benzyloxy, phenyl, heteroaryl, heterocyclylalkyl and nitro; wherein X is selected from the group consisting of direct bond, O, SO, S(O₂), NR¹, CH₂, CF₂, CH₂CH₂, CH₂NR¹, NR¹CH₂, CH=CH, C=O, CH₂C=O, _CR ^ia _R ^ib oCR^laR^lb , CR^laR^lbO; SO₂NR¹^R¹SO₂, C(O)NR¹ and NR¹C(O); wherein R¹ is selected separately in each occurrence from the group consisting of H and lower alkyl;

R^la is selected from the group consisting of H, OH and lower alkyl; R^lb is selected from the group consisting of H and lower alkyl, or R^la and R^lb taken together may form a 3-6 membered ring, which may optionally contain a heteroatom chosen from O, S, and N; wherein HetAr is an aryl or heteroaryl ring attached via a ring carbon to Q, further characterized in that Q and X cannot be on adjacent positions in said aryl or heteroaryl ring;

Q is chosen from -O-, -NR¹- and S(O)_P;

HET is selected from the group consisting of

4-7-membered saturated nitrogenous heterocycle and

4-7-membered saturated nitrogenous heterocycle substituted with one or two substituents independently selected from the group consisting of halogen, hydroxyl, amino, carboxy, loweralkyl, loweracyl, loweralkoxy, N-oxide, fluoroloweralkyl, fluoroloweralkoxy, formyl, cyano, benzyl, benzyloxy, phenyl, heteroaryl and nitro; and taken together ZW is H or

Z is (CH₂)i-io, in which one or two (CH₂) may optionally be replaced by -O-, -NR¹-, -SO-, -S(O)₂-, -C(=O)- or -C=O(NH)-, provided that said -O-, -NR¹-, -SO-, -S(O)₂-, -C(=O)- or -C=O(NH)- are not at the point of attachment to HET and are separated by at least one -(CH₂)-;

W is selected from the group consisting of acyl, hydroxyl, carboxyl, amino, -C(O)NHR⁴, aminoacyl, -COOalkyl, -CHO, heterocyclyl, substituted aryl, substituted heterocyclyl, sulfonamide, -C(O)fluoroalkyl, -C(O)CH₂C(O)Oalkyl, -C(O)CH₂C(O)Ofluoroalkyl, -SH, -C(O)NH(OH), -C(O)N(OH)R⁴, -N(OH)C(O)OH, - N(OH)C(O)R⁴; and

R⁴ is selected from the group consisting of H, (C1-C4) alkyl, and phenyl(Ci-C₄) alkyl; with the provisos that;

(a) when Q is -O-, HET is (5)-pyrrolidine, rac-pyrrolidine or piperidine, Ar is phenyl or halo-substituted phenyl, and HetAr is /?-phenylene, then the Z-W combination is other than H;

28. The method or use or inhibitor according to claim 24, wherein the leukotriene synthesis inhibitor comprises a compound of the formula

29. The method or use or inhibitor according to claim 24, wherein the leukotriene synthesis inhibitor comprises a compound represented by the formula:

R"

/

-OFT or -N

\

R°

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or a pharmaceutically acceptable salt thereof.

30. The method or use or inhibitor according to claim 24, wherein the leukotriene synthesis inhibitor comprises a compound selected from the group consisting of: 2-[4-(quinolin-2-yl-methoxy)phenyl]-2-cyclopentylacetic acid, 2-[4- (quinolin-2-yl-methoxy)phenyl]-2-cyclohexylacetic acid, and 2-[4-(quinolin-2-yl- methoxy)phenyl]-2-cycloheptylacetic acid, (+)-enantiomer of 2-[4-(quinolin-2-yl- methoxy)phenyl]-2-cyclopentylacetic acid, (-)-enantiomer of 2-[4-(quinolin-2-yl- methoxy)phenyl]-2-cyclopentylacetic acid, and pharmaceutically acceptable salts thereof.

31. The method or use or inhibitor according to claim 24, wherein the leukotriene synthesis inhibitor comprises BAY-X- 1005 (DG-031) or a physiologically acceptable salt, formulation, or pro-drug thereof.

32. The method or use or inhibitor according to any one of claims 29-31 , wherein the composition or dose is administered according to a dosing regimen that achieves a single daily peak plasma concentration of the inhibitor within a range of 11.2 to 30 micrograms per milliliter (μg/ml)

33. A method reducing leukotriene production in a human comprising administering to the human a composition that comprises a leukotriene synthesis inhibitor of the formula:

wherein R¹ represents a group of the formula:

FT

/

-OFT or -N

\

R°

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or pharmaceutically acceptable salt, ester, or pro-drug thereof, and wherein the composition is administered to the human in the morning without food, in an amount effective to achieve a peak plasma concentration (Cmax) of the inhibitor greater than 11.2 μg/ml.

34. Use of a leukotriene synthesis inhibitor of the formula:

wherein R¹ represents a group of the formula:

wherein R represents hydrogen, lower alkyl or phenyl and n denotes a number of 0 to 5, Z represents norbornyl, or represents a group of the formula:

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or pharmaceutically acceptable salt, ester, or pro-drug thereof, in the manufacture of a medicament for administration in a human for reducing leukotriene production in said human, wherein the medicament is formulated into a dose that is administered in the morning without food, in an amount effective to achieve a peak plasma concentration (Cmax) of the inhibitor greater than 11.2 μg/ml.

35. A leukotriene synthesis inihibitor of the formula

wherein R¹ represents a group of the formula:

R^

/

-OFT or -N

\

R°

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or pharmaceutically acceptable salt, ester, or pro-drug thereof, for reducing leukotriene production in a human, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration in the morning without food, in an amount effective to achieve a peak plasma concentration (Cmax) of the inhibitor greater than 11.2 μg/ml

36. The method or use or inhibitor OOaccording to claim 33-35, wherein the leukotriene synthesis inhibitor comprises BAY-X- 1005 (DG-031) or a physiologically acceptable salt, formulation, or pro-drug thereof.

37. A method of reducing leukotriene production in a human comprising administering to the human a composition that comprises a leukotriene synthesis inhibitor or pharmaceutically acceptable salt, ester, or pro-drug thereof, wherein the leukotriene synthesis inhibitor comprises a compound having the following formula:

R"

/

-OFT or -N

\

R°

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, wherein the composition is administered according to a dosing regimen that achieves a single daily peak plasma concentration of the inhibitor of at least 11.2 micrograms per milliliter (μg/ml), and wherein the peak plasma concentration occurs during the day between the hours of 6 A.M. and 2 P.M.

38. Use of a leukotriene synthesis inhibitor having the following formula:

or pharmaceutically acceptable salt thereof , wherein R¹ represents a group of the formula: R^

/

-OR^ or -N

\

R°

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, in the manufacture of a medicament for administration in a human for reducing leukotriene production in said human, wherein the medicament is formulated into a dose that is administered according to a dosing regimen that is effective to achieves a single daily peak plasma concentration of the inhibitor of at least 11.2 micrograms per milliliter (μg/ml), and wherein the peak plasma concentration occurs during the day between the hours of 6 A.M. and 2 P.M.

39. A leukotriene synthesis inhibitor having the following formula

wherein R¹ represents a group of the formula:

R"

/

-OFT or -N

\

R°

-

wherein R⁹ and R¹⁰ are identical or different and denote hydrogen, lower alkyl or phenyl, or R⁹ and R¹⁰ can together form a saturated carbocyclic ring having up to 6 carbon atoms and m denotes a number from 1 to 6, and A and B are identical or different and denote hydrogen, lower alkyl or halogen, or pharmaceutically acceptable salt, ester, or pro-drug thereof, for reducing leukotriene production in a human, wherein the leukotriene synthesis inhibitor is formulated into a dose for administration according to a dosing regimen that is effective to achieve a single daily peak plasma concentration of the inhibitor of at least 11.2 micrograms per milliliter (μg/ml), and wherein the peak plasma concentration occurs during the day between the hours of 6 A.M. and 2 P.M.

40. The method or use or inhibitor according to any of claims 37- 39, wherein the leukotriene synthesis inhibitor comprises BAY-X-1005 (DG-031) or a physiologically acceptable salt, formulation, or pro-drug thereof.

41. The method or use or inhibitor according to any one of claims 29-40, wherein the peak plasma concentration is within a range of 12 to 30 micrograms per milliliter.

42. The method or use or inhibitor according to any one of claims

29-40, wherein the peak plasma concentration is within a range of 11.2 to 25 micrograms per milliliter.

43. The method or use or inhibitor according to any one of claims 29-40, wherein the peak plasma concentration is within a range of 11.2 to 20 micrograms per milliliter.

44. The method or use or inhibitor according to any one of claims 29-40, wherein the peak plasma concentration is within a range of 12 to 15 micrograms per milliliter.

45. The method or use or inhibitor according to any one of claims 29-40, wherein the peak plasma concentration occurs during the day between the hours of 8 A.M. and 2 P.M.

46. The method or use or inhibitor according to any one of claims 29-40, wherein the peak plasma concentration occurs during the day between the hours of 9 A.M. and 2 P.M.

47. The method or use or inhibitor according to any one of claims 29-46, wherein the average plasma concentration is less than 5.5 μg/ml.

48. The method or use or inhibitor according to any one of claims 29-46, wherein the average plasma concentration is less than 4.5 μg/ml.

49. The method or use or inhibitor according to any one of claims 29-46, wherein the average plasma concentration is less than 3.5 μg/ml.

50. The method or use or inhibitor according to any one of claims

1-49, wherein the leukotriene synthesis inhibitor is present in the composition or dose in an amount effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by 20-50%.

51. The method or use or inhibitor according to any one of claims

1-49, wherein the leukotriene synthesis inhibitor is present in the composition or medicament in an amount effective to reduce evening peak leukotriene B4 (LTB4) production in adult human subjects by at least 25%.

52. The method or use or inhibitor according to claim 51 , wherein the leukotriene synthesis inhibitor is present in the composition or medicament in an amount effective to reduce evening peak LTB4 production in adult human subjects by at least 30%.

53. The method or use or inhibitor according to claim 51 , wherein the leukotriene synthesis inhibitor is present in the composition or dose in an amount effective to reduce evening peak LTB4 production in adult human subjects by at least 40%.

54. The method or use or inhibitor according to any one of claims 50-53, wherein the LTB4 levels are measured in plasma after activation of leukocytes with calcium ionophore, and thereafter isolating the plasma.

55. The method or use or inhibitor according to any one of claims

1-54, wherein the composition or medicament is formulated to cause a peak plasma concentration of the leukotriene synthesis inhibitor to occur within seven hours of administering the composition.

56. The method or use or inhibitor according to any one of claims

1-54, wherein the composition or medicament is formulated to cause a peak plasma concentration of the leukotriene synthesis inhibitor to occur within 6 hours of administering the composition.

57. The method or use or inhibitor according to any one of claims

1-54, wherein the composition or medicament is formulated to cause a peak plasma concentration of the leukotriene synthesis inhibitor to occur within 5 hours of administering the composition.

58. The method or use or inhibitor according to any one of claims

1-54, wherein the composition or medicament is formulated to cause a peak plasma concentration of the leukotriene synthesis inhibitor to occur within 4 hours of administering the composition.

59. The method or use or inhibitor according to any one of claims

1-54, wherein the composition or medicament is formulated to cause a peak plasma concentration of the leukotriene synthesis inhibitor occurs within 3 hours of administering the composition.

60. The method or use or inhibitor according to any one of claims 1-59, wherein the composition or dose is formulated for oral administration and administered orally.

61. The method or use or inhibitor according to claim 60, wherein the composition or dose is administered with food.

62. The method or use or inhibitor according to any one of claims 1-59, wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, the composition or dose is administered orally with food in the morning, and the dose is within the range of 250-1500 mg, inclusive.

63. The method or use or inhibitor according to any one of claims 1-59, wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, the composition or dose is administered orally with food in the morning, and the dose is within the range of 750-1000 mg, inclusive.

64. The method or use or inhibitor according to claim 63, wherein the composition or dose is administered on an empty stomach.

65. The method or use or inhibitor according to any one of claims 1-59, wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, the composition or dose is administered orally on an empty stomach, and the dose is within the range of 750-2000 mg, inclusive.

66. The method or use or inhibitor according to any one of claims 1-59, wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, the composition or dose is administered orally on an empty stomach, and the dose is within the range of 1000-1500 mg, inclusive.

67. The method or use or inhibitor according to any one of claims 1-59, wherein the inhibitor is DG-031 or a pharmaceutically acceptable salt, ester, or pro-drug thereof, and the composition or dose contains a 1000 mg dose of the inhibitor.

68. The method or use or inhibitor according to any one of claims 1-67, wherein the human has been diagnosed with at least one risk factor for myocardial infarction or stroke selected from the group consisting of advanced age, gender, smoking, physical activity, waist-to-hip circumference ratio, family history of cardiovascular disease or myorcardial infarction, previously diagnosed cardiovascular disease or MI, obesity, diabetes, hypertriglyceridemia, low HDL cholesterol, hypertension, elevated blood pressure, cholesterol levels (total cholesterol >200mg/dL), HDL cholesterol, LDL cholesterol, triglycerides, apolipoprotein AI and B levels, fibrinogen, ferritin, C-reactive protein, and leukotriene levels.

69. The method or use or inhibitor according to any one of claims 1-67, wherein the human has been diagnosed as carrying a FLAP or LTA4H haplotype that correlates with elevated plasma concentrations of LTB4 or correlates with increased risk of myocardial infarction.

70. The method or use or inhibitor according to any one of claims 1-69, wherein, before administering the composition, the human exhibits an elevated blood LTB4 production.

71. The method or use or inhibitor according to any one of claims

1-70, wherein the human has been diagnosed with asthma.

72. The method or use or inhibitor according to any one of claims 1-70, wherein the human has not been diagnosed with asthma.

73. The method or use or inhibitor according to any one of claims 1-72, wherein the administering is repeated daily for a month.

74. The method or use or inhibitor according to claim 73, wherein the administering is repeated for six months.

75. The method or use or inhibitor according to any one of claims 1-74, wherein the composition is administered once daily.

76. The use of a leukotriene synthesis inhibitor for the manufacture of a medicament for once daily administration in the morning.

77. A leukotriene synthesis inhibitor for use as a medicament for once daily administration in the morning.