WO2008002929A2

WO2008002929A2 - Heterologous cross-sensitization for improved agonist activity

Info

Publication number: WO2008002929A2
Application number: PCT/US2007/072147
Authority: WO
Inventors: Richard A. Bond
Original assignee: Inverseon, Inc.
Priority date: 2006-06-26
Filing date: 2007-06-26
Publication date: 2008-01-03
Also published as: WO2008002929A3

Abstract

Methods for improving the effectiveness of prostacyclin agonists by heterologous sensitization by co-administering beta adrenergic inverse agonists are provided for the treatment of pulmonary hypertension and other diseases treated by chronic prostacyclin treatment.

Description

BACKGROUND OF THE INVENTION

[0001] The present invention relates to methods for improving the effectiveness of G-Protein-Coupled Receptor (GPCR) agonist drugs by heterologous sensitization which is achieved by chronically co-administering inverse agonist drugs that target a different GPCR receptor which shares a common G-Protein with the first receptor, such as a Gs protein.

[0002] A large number of diseases and conditions are related to the activities of G protein coupled receptors (GPCR). The superfamily of G protein coupled receptors includes a large number of receptors. These receptors are integral membrane proteins characterized by amino acid sequences that contain seven hydrophobic domains, predicted to represent the transmembrane spanning regions of the proteins. They are found in a wide range of organisms and are involved in the transmission of signals to the interior of cells as a result of their interaction with heterotrimeric G proteins. They respond to a diverse range of agents including lipid analogues, amino acid derivatives, small molecules such as epinephrine and dopamine, and various sensory stimuli. The properties of many known GPCR are summarized in S.Watson & S. Arkinstall, "The G- Protein Linked Receptor Facts Book" (Academic Press, London, 1994), incorporated herein by this reference.

[0003] Drug discovery for GPCRs has focused on identifying agonists or antagonists. These drugs have been traditionally characterized by their acute effects on the GPCR. Agonists are "activators" whereas antagonists are "inactivators" or "blockers". A new understanding of GPCR function has ied to the reclassification of antagonists into two subclasses, those that are competitive to agonists and exhibit partial agonist activity and those that are inverse agonists. This reclassification is based on the "two-state" model of GPCR function (Lefkowitz reference) in which the GPCR spontaneously alternates from an active state to an inactive state in the absence of a bound iigand. Based on this model, an agonist is now generally defined as a compound that stabilizes the active state of the GPCR whereas the inverse agonist stabilizes the inactive state of the GPCR.

[0004] There are a large number of diseases, syndromes, and conditions for which conventional treatment is the use of agonists specific for a GPCR. However, the limitations of this conventional treatment have become apparent. As indicated above, chronic administration of agonists can lead to desensitization and depression of receptor signaling. This limits the effectiveness of treatment over time. There is therefore a need for a new, global, strategy of treating diseases, syndromes, and conditions that are characterized by the hypofunction of GPCR.

[0005] There are hundreds of GPCR receptors in the human genome. GPCR receptors are important therapeutic drug targets for agonist drugs or inverse agonist drugs depending upon the disease. However, there are much fewer, about 20, internal G-proteins that the receptors "couple" to in order to transducer their signal for the appropriate cellular response. Furthermore, more than one different type GPCR receptor may reside in the same cell type but still require the same interna! G-protein. For example, the following GPCR receptors all share the same Gs Protein which results in the formation of cAMP cellular intermediate in the signal transduction pathway: glucagon, beta adrenergic receptor, dopamine, vasopressin, oxytocin.

[0006] Possibly due to this "sharing" of internal G-proteins by the GPCR receptors, a phenomenon termed "heterologous desensitization" has been observed. In this situation, an agonist that interacts with one GPCR can lead to the desensitization of a second GPCR to its agonist. For example, opioid narcotics cause heterologous desensitization at chemokine GPCRs and vice versa. Consequently, not only can an agonist at its own GPCR cause homologous desensitization of its own receptor over time, another agonist at a different GPCR can cause heterologous desensitization at another GPCR. [0007] An exampie of an agonist that exhibits desensitization at its own receptor is the prostacyclin agonist. Prostacyclin agonists bind to and stabilize the active form of the prostacyclin receptor, a G protein-coupled receptor (GPCR). An alternative name for prostacyclin receptor is the prostaglandin I₂ receptor and within the human body there is an endogenous ligand that is synthesized via the well-known cytochrome oxidase, COX-1 and COX-2, pathways. Endogenous prostacyclin or prostaglandin I₂, is a small molecule that is synthesized in response to a number of conditions, including inflammation, and that diffuses and binds to its cognate GPCR in the smooth muscle cell type in the vasculature of the lungs, kidney and elsewhere to result in vascular relaxation and increased perfusion. In the diseased state of pulmonary arterial hypertension there is not enough vascular relaxation in the lungs despite more-or-less normal blood pressure in the rest of the vasculature. Prostacyclin agonists, inhaled, oral, and subcutaneous routes of administration have demonstrated benefit for reducing pulmonary hypertension. Other general blood pressure medications are not used since, even if they reduced lung blood pressure, they would cause the remaining vasculature in the body to relax and the patients would become hypotensive which would be detrimental.

[0008] One limitation with prostacyclin agonists and GPCR agonists in genera!, is that chronic agonist binding and activation of its GPCR results in desensitization and down-regulation of the receptor so over time the prostacyclin agonist becomes less effective and more is needed for the same activity. This was shown in the clinical situation by McLaughlin et al (Reduction in pulmonary vascular resistance with long- term epoprostenol (prostacyclin) therapy in primary pulmonary hypertension, NEJM 1998 338:273-7) in which doses needed to be increased to maintain efficacy. This is undesirable because the increase of dosages required also means a greater likelihood of side effects, which in turn may require decrease of dosage or even cessation of therapy. This is extremely deleterious to the management of this chronic condition. Consequently, there is a need to increase the chronic effectiveness of the prostacyclin agonist in the chronic setting for pulmonary arterial hypertension.

[0009] Pulmonary hypertension is a disorder of the lung in which the pressure in the pulmonary artery (the blood vessel that leads from the heart to the lungs) rises above norma! levels, if left untreated, pulmonary hypertension may become life threatening. Symptoms of pulmonary hypertension include shortness of breath with minimal exertion, fatigue, chest pain, dizzy spells fainting, and other symptoms. Pulmonary hypertension is frequently misdiagnosed and has often progressed to late stage by the time it is accurately diagnosed. Moreover, pulmonary hypertension has been historically chronic and incurable with a poor survival rate.

[0010] When pulmonary hypertension occurs in the absence of a known cause, it is referred to as primary pulmonary hypertension (PPH). There are many unknown causes of PPH.

[0011] When the cause of pulmonary hypertension is known, it is called secondary pulmonary hypertension (SPH). Common causes of SPH is the breathing disorders emphysema, bronchitis and chronic obstructive pulmonary disorder, among others. Other less frequent causes are the inflammatory or collagen vascular diseases such as scleroderma, CREST syndrome or systemic fupus erythematosus. Congenital heart diseases that cause shunting of extra blood through the lungs like ventricular and arterial septal defects, chronic pulmonary thromboembolism (old blood clots in the pulmonary artery), HIV infection, liver disease and diet drugs like fenfluramine and dexfenfluramine are also causes of pulmonary hypertension.

[0012] Beta adrenergic inverse agonists are members of the general class of drugs called 'beta blockers'. These drugs bind to inactive forms of beta adrenergic receptors and prevent their activation from either endogenous agonists or exogenous agonists. Chronic administration of these drugs results in up-regulation of their receptors in the smooth muscle of arteries and veins and this may account for their ability to increase vasodilation and counteract global systemic hypertension. However, this class of drugs is not approved for pulmonary arterial hypertension nor has any benefit been observed for the treatment of pulmonary arterial hypertension or other disorders relating to the use of prostacyclin agonists.

SUMMARY OF THE INVENTION

[0013] The present invention relates to methods for improving the effectiveness of G-Protein-Coupled Receptor (GPCR) agonist drugs by heterologous sensitization which is achieved by chronically co-administering inverse agonist drugs that target a different GPCR which shares a common G-Protein with the first receptor, such as a Gs protein. In particular, this encompasses a method for improving the effectiveness of a G-Protein Coupled Receptor (GPCR) agonist drug comprising the step of chronic coadministration of an inverse agonist drug that targets a different GPCR receptor, the different GPCR sharing a common G protein with the receptor bound by the GPCR agonist drug. The common G protein can be the Gs protein.

[0014] According to the present invention, this relates to methods for improving the effectiveness of prostacyclin agonists by heterologous sensitization by coadministering beta adrenergic inverse agonists for pulmonary arterial hypertension, airway diseases, kidney diseases and other diseases in which prostacyclin agonists are used chronically.

[0015] One aspect of the invention is a method for cross-sensitizing the response of prostacyclin receptors to a prostacyclin agonist by administering a quantity of β₂-adrenergic receptor inverse agonist to a subject with a disease or condition treatable by potentiating the response of prostacyclin receptors to a prostacyclin agonist, the quantity being sufficient to detectably increase the response of a prostacyclin receptor to a prostacyclin agonist.

[0016] Typically, the β-adrenergic inverse agonist is selected from the group consisting of β₂-selective inverse agonists, and non-selective inverse agonists having inverse agonist activity against both β_r and β₂-adrenergic receptors. Preferably, the β- adrenergic inverse agonist is a β₂-selective inverse agonist. Preferably, the β- adrenergic inverse agonist is selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvedilol, ICI-118,551 , levobunolol, metoprolol, propranolol, sotalol, and timolol, and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof. Particularly preferred β-adrenergic inverse agonists include nadolol, carvediioi, timolol, metoproiol, ICI-118,551 , and their analogues. Typically, the β-adrenergic inverse agonist is administered by a route selected from the group consisting of oral, sustained-release oral, parenteral, sublingual, buccal, insufflation, subcutaneous injection, and inhalation. Preferably, the route is the route is oral, inhalation, or subcutaneous injection.

[0017] In one alternative, the disease or condition is selected from the group consisting of pulmonary hypertension and renal failure due to vascular insufficiency; preferably, the disease or condition is pulmonary hypertension, in another alternative, the disease or condition is a degenerative disease of the central nervous system, including, but not limited to, Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, or amyotrophic lateral sclerosis (Lou Gehrig's disease). In still another alternative, the disease or condition is a disease or condition relating to disturbances or dysregulation of peripheral circulation, including, but not limited to, Raynaud's disease, vascular ischemia, and limb ischemia. In yet another alternative, the disease or condition is thrombosis. In still another alternative, the disease or condition is headache, including, but not limited to, migraine or cluster headaches.

[0018] Typically, the method of administration of the β-adrenergic inverse agonist results in continuous levels of the β₂-adrenergic inverse agonist in the bloodstream of the subject. Typically, the β-adrenergic inverse agonist is administered over time in a series of graduated doses starting with the lowest dose and increasing to the highest dose. Preferably, in this administration scheme, when the highest dose is reached, the β-adrenergic inverse agonist continues to be administered at that dose.

[0019] Another aspect of the invention is a method of treating a disease or condition treatable by potentiating the response of prostacyclin receptors to a prostacyclin agonist comprising the steps of:

(1 ) administering to a subject with such a disease or condition a therapeutically effective quantity of a β-adrenergic inverse agonist that is sufficient to detectably increase the response of a prostacyclin receptor to a prostacyclin agonist; and

(2) administering to the subject a therapeutically effective quantity of a prostacyclin agonist. [0020] The prostacyclin agonist can be a partial agonist or a full agonist; any drug or compound with prostacyclin receptor agonist activity can be used. Typically; the prostacyclin agonist is selected from the group consisting of cicaprost, iloprost, beraprost, UT-15, treprostinil, and epoprostenol, and the salts, solvates, analogues, mimetics, stereoisomers, congeners, bioisosteres, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

[0021] In one alternative, the β-adrenergic inverse agonist and the prostacyclin agonist are administered simultaneously; in this alternative, the β-adrenergic inverse agonist and the prostacyclin agonist can be administered in a single pharmaceutical composition or dosage form that includes both the β-adrenergic inverse agonist and the prostacyclin agonist. In another alternative, the β-adrenergic inverse agonist and the prostacyclin agonist are administered at different times.

[0022] Accordingly, another aspect of the invention is a pharmaceutical composition comprising:

(1 ) a therapeutically effective quantity of a β-adrenergic inverse agonist;

(2) a therapeutically effective quantity of a prostacyclin agonist; and

(3) at least one pharmaceutically acceptable carrier.

[0023] Typically, the pharmaceutical composition comprises a quantity of a β- adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat one or more of the diseases or conditions described above.

[0024] Another aspect of the present invention is a blister pack comprising:

(1) a lower substrate;

(2) an intermediate dosage holder that is shaped to generate a plurality of cavities and that is placed over the lower substrate, the cavities being shaped to hold dosage forms of a pharmaceutical composition as described above, the pharmaceutical composition being formulated for oral administration;

(3) an upper substrate placed over the intermediate dosage holder that has a plurality of apertures, each aperture being located to accommodate a corresponding cavity; and (d) dosage forms of the pharmaceutical composition as described above placed in the cavities, the dosage forms being formulated for oral administration.

[0025] The dosage forms of the pharmaceutical composition placed in the blister pack can include a range of dosages of the β-adrenergic inverse agonist. Typically, the range of dosages of the β-adrenergic inverse agonist of the dosage forms of the pharmaceutical composition placed in the blister pack includes a range of dosages of the β-adrenergic inverse agonist from a starting dose to a maintenance dose.

[0026] Another embodiment of a blister pack according to the present invention is a blister pack comprising:

(1 ) a lower substrate;

(2) an intermediate dosage holder that is shaped to generate a plurality of cavities and that is placed over the lower substrate, the cavities being shaped to hold dosage forms of:

(a) a first pharmaceutical composition that comprises:

(i) a therapeutically effective amount of a β-adrenergic inverse agonist; and

(ii) a first pharmaceutically acceptable carrier, the first pharmaceutical composition being formulated for oral administration; and

(b) a second pharmaceutical composition that comprises:

(i) a therapeutically effective amount of a prostacyclin agonist; and

(ii) a second pharmaceutically acceptable carrier, the second pharmaceutical composition being formulated for oral administration;

(3) an upper substrate placed over the intermediate dosage holder that has a plurality of apertures, each aperture being located to accommodate a corresponding cavity; and

(4) dosage forms of the first and second pharmaceutical compositions placed in the cavities, the dosage forms being formulated for oral administration. [0027] Another aspect of the present invention is a method for improving the effectiveness of a G-Protein Coupled Receptor (GPCR) agonist drug comprising the step of chronic co-administration of an inverse agonist drug that targets a different GPCR receptor, the different GPCR sharing a common G protein with the receptor bound by the GPCR agonist drug. The common G protein can be the Gs protein.

BRIEF DESCRIPTION OF THE DRAWINGS

[0028] The following invention will become better understood with reference to the specification, appended claims, and accompanying drawings, where:

[0029] Figure 1 is a diagram of a blister pack that can hold dosage forms of inverse agonists, dosage forms of inverse agonists and dosage forms of prostacyclin agonists, or dosage forms of a pharmaceutical composition including both an inverse agonist and a prostaglandin agonist according to the present invention.

[0030] Figure 2 is a graph showing that chronic beta inverse agonist treatment prevents degradation of human β₂-adrenergic receptors employing biotinylated receptors isolated by streptavidin-coated agarose beads.

[0031] Figure 3 is a graph showing the cumulative concentration-response curves to the prostacyclin agonist cicaprost in the mouse isolated trachea after methacholine (3 mM) precontraction from mice treated chronically with nadolol versus control S/C mice. Each point represents mean and vertical lines show SEM (n=5-6). Nad Chr (nadolol chronic). *P<0.05 compared with S/C, control mice that were ovalbumin sensitized and challenged mice.

[0032] Figure 4 is a graph showing the concentration-response curves to the EP₂ receptor agonist CAY 10399 in trachea rings for ovalbumin sensitized and challenged mice as compared with ovalbumin sensitized and challenged mice that were chronically treated with nadolol and with ovalbumin sensitized and challenged control mice (S/C).

DETAILED DESCRIPTION OF THE INVENTION [0033] As used herein, in the generally accepted two-state model of receptor theory, the term "agonist" is defined as a substance that has an affinity for the active site of a receptor and thereby preferentially stabilizes the active state of the receptor, or a substance, including, but not limited to, drugs, hormones, or neurotransmitters, that produces activation of receptors and enhances signaling by those receptors. Irrespective of the mechanism or mechanisms of action, an agonist produces activation of receptors and enhances signaling by those receptors. Agonists are generally divided into full agonists and partial agonists. The latter produce a less than maximal response even when present in a sufficient concentration to fully occupy the active site of their corresponding receptors.

[0034] As used herein, in the two-state model of receptor theory, the term "antagonist" is defined as a substance that does not preferentially stabilize either form of the receptor, active, or inactive, or a substance, including, but not limited to, drugs, hormones, and neurotransmitters, that prevents or hinders the effects of agonists and/or inverse agonists. Irrespective of the mechanism or mechanisms of action, an antagonist prevents or hinders the effects of agonists and/or inverse agonists.

[0035] As used herein, in the two-state model of receptor theory, the term "inverse agonist" is defined as a substance that has an affinity for the inactive state of a receptor and thereby preferentially stabilizes the inactive state of the receptor, or a substance, including, but not limited to, drugs, hormones, or neurotransmitters, that produces inactivation of receptors and/or prevents or hinders activation by agonists, thereby reducing signaling from those receptors.

[0036] As used herein, the term "concurrent administration" refers to the administration of two or more active agents sufficiently close in time, to achieve a combined therapeutic effect that is preferably greater than that which would be achieved by the administration of either agent alone. Such concurrent administration can be carried out simultaneously, e.g., by administering the active agents together with a common pharmaceutically acceptable carrier in one or more doses.

[0037] The term "subject," as used herein, refers to human or animal species. In general, methods and compositions according to the present invention can be used to treat not only humans, but also socially or economically important animal species such as cows, horses, sheep, pigs, goats, dogs, and cats. Unless specified, methods and compositions according to the present invention are not limited to treatment of humans.

[0038] The term "therapeutically effective amount," as used herein, refers to an amount of a therapeutic agent or composition effective to treat, ameliorate, or prevent a desired disease or condition, or to exhibit a detectable therapeutic or preventative effect. The precise therapeutically effective amount for a subject will depend upon the subject's size, weight, and health, the nature and extent of the condition affecting the subject, and the therapeutics or combination of therapeutics selected for administration, as well as variables such as liver and kidney function that affect the pharmacokinetics of administered therapeutics. Thus, it is not useful to specify an exact effective amount in advance. However, the effective amount for a given situation can be determined by routine experimentation and is within the judgment of the clinician.

[0039] As used herein, the term "pulmonary hypertension" means any form, diagnosis, level or stage of pulmonary hypertension, including, but not limited to, primary or secondary pulmonary hypertension, pulmonary arterial hypertension, pulmonary venous hypertension, pulmonary hypertension associated with disorders of the respiratory system or hypothermia, pulmonary hypertension resulting from chronic thrombotic or embolic disease, or pulmonary hypertension resulting from disorders directly affecting the pulmonary vasculature. The term "pulmonary hypertension" also includes other respiratory disorders characterized by acute pulmonary vasoconstriction such as those disorders resulting from pneumonia, traumatic injury, aspiration or inhalation injury, fat embolism in the lung, acidosis inflammation of the lung, adult respiratory distress syndrome, acute pulmonary edema, acute mountain sickness, post- cardiac surgery, acute pulmonary hypertension, persistent pulmonary hypertension of the newborn, perinatal aspiration syndrome, hyaline member disease, acute pulmonary thromboembolism, heparin-protamine reactions, sepsis, status asthmaticus or hypoxia (including iatrogenic hypoxia) and other forms of reversible pulmonary vasoconstriction. Such pulmonary disorders are also characterized by inflammation of the lung including those associated with the migration into the lung of non-resident eel! types including the various leukocyte subclasses. The condition is further described in U.S. Patent Application Publication No. 2006/0104913 by Chaudry, incorporated herein by this reference.

[0040] The severity of the condition of pulmonary hypertension can be classified into four classes, designated I, II, III, and IV in order of increasing severity. Class I is defined as patients with pulmonary hypertension but without resulting limitation of physical activity. Ordinary physical activity does not cause undue dyspnea or fatigue, chest pain, or near syncope. Class Il is defined as patients with pulmonary hypertension resulting in slight limitation of physical activity. These patients are comfortable at rest, but ordinary physical activity causes undue dyspnea or fatigue, chest pain or near syncope. Class III is defined as patients with pulmonary hypertension resulting in marked limitation of physical activity. These patients are comfortable at rest, but less than ordinary physical activity causes undue dyspnea or fatigue, chest pain or near syncope. Class IV is defined as patients with pulmonary hypertension resulting in inability to perform any physical activity without symptoms. These patients manifest signs of right heart failure. Dyspnea and/or fatigue may be present at rest, and discomfort is increased by any physical activity.

[0041] The prostanoids (prostaglandins, prostacyclins and thromboxanes) are any of a group of components derived from unsaturated 20-carbon fatty acids, primarily arachidonic acid, via the cyclooxygenase (COX) pathway that are extremely potent mediators of a diverse group of physiologic processes. The prostaglandins (PGs) are designated by adding one of the letters A through I to indicate the type of substituents found on the hydrocarbon skeleton and a subscript (1 , 2 or 3) to indicate the number of double bonds in the hydrocarbon skeleton for example, PGE₂. The predominant naturally occurring prostaglandins all have two double bonds and are synthesized from arachidonic acid (5,8,11 ,14-eicosatetraenoic acid). The 1 series and 3 series are produced by the same pathway with fatty acids having one fewer double bond (8,11 ,14- eicosatrienoic acid or one more double bond (5,8,11 ,14,17-eicosapentaenoic acid) than arachidonic acid. The prostaglandins act by binding to specific cell surface receptors causing an increase in the level of the intracellular second messenger cyclic AMP (and in some cases cyclic GMP). The effect produced by the cyclic AMP increase depends on the specific cell type. In some cases there is also a positive feedback effect. Increased cyclic AMP increases prostaglandin synthesis leading to further increases in cyclic AMP.

[0042] Prostaglandins have a variety of roles in regulating cellular activities, especially in the inflammatory response where they may act as vasodilators in the vascular system, cause vasoconstriction or vasodilatation together with bronchodilation in the lung and act as hyperalgesics. Prostaglandins are rapidly degraded in the lungs and will not therefore persist in the circulation.

[0043] Prostacyclin, also known as PGI₂, is an unstable vinyl ether formed from the prostaglandin endoperoxide, PGH₂. The conversion of PGH₂ to prostacyclin is catalyzed by prostacyclin synthetase. The two primary sites of synthesis are the veins and arteries. Prostacyclin is primarily produced in vascular endothelium and plays an important inhibitory role in the local control of vascular tone and platelet aggregation. Prostacyclin has biological properties opposing the effect of thromboxane A₂. Prostacyclin is a vasodilator and a potent inhibitor of platelet aggregation whereas thromboxane A₂ is a vasoconstrictor and a promoter of platelet aggregation. A physiological balance between the activities of these two effectors is probably important in maintaining a healthy blood supply.

[0044] Beta inverse agonists are a subclass of beta blockers and the whole class of drugs has been contraindicated for use by patients with pulmonary airway diseases such as asthma and COPD. However, surprisingly, Bond in PCT Patent Application Serial No. PCT/US2004/033157, filed October 8, 2004, and incorporated herein in its entirety by this reference, demonstrated that chronic dosing in mice of beta inverse agonists was able to protect their pulmonary airways from significant constriction in response to allergen and methacholine challenge.

[0045] In the case of the preferred drug nadolol, the current pharmacokinetic profile of the approved drug Corgard (nadolol) is a once daily formulation (Corgard product insert, Monarch Pharmaceuticals, Inc.). It has been discovered that when asthmatics were administered a 10 mg dose of Corgard this resulted in a measurable reduction in pulmonary airway function as determined by measuring FEV1 (forced expiratory volume) during the first four hours of dosing when the peak drug concentration occurs - the T_max of Corgard (time to maximum drug concentration in the bloodstream) is 3.5 hours.

[0046] Consequently, it is non-obvious to develop a once daily controlled- release formulation of nadoioi when there is already a once-daily formulation. However, the rationale for this is that the pharmacokinetic profile of the current once daily formulation results in maximal levels of drug in the bloodstream at an average of 3-4 hours after administering the first dose. This profile has been demonstrated to result in measurable airway constriction in mild asthmatics in a clinical study. However, this is avoided by the use of the controlled-release formulation.

[0047] The pharmacokinetic profile is defined by a number of parameters. The T_max is the time when peak drug concentrations are observed in the bloodstream. The C_max is the maximum concentration of the drug in the bloodstream, usually for one dosage level. The half-life is the time to when there is half the C_maχ concentration in the bloodstream. The elimination rate constant is the rate constant of elimination of the drug via excretion and metabolism. The apparent volume distribution, V, reflects the volume that the drug is distributed in, such as blood, or other compartments such as fat. The AUC, area under the curve, is the area under the plasma {serum, or blood) concentration versus time curve. With both single and multiple dosing, there is the peak to trough ratio where the peak is the C_max and the trough is the C_min. Pharmacokinetic parameters of a drug dosage form are required by regulatory bodies (e.g. FDA) to evaluate dosing regime in subjects and also special subjects with modified pharmacokinetic parameters such as reduced excretion due to renal failure. Pharmacokinetic parameters are additionally used to determine bioequivalence of drugs and thus are consequently more important that the composition of the drug form since it obvious to those skilled in the art that the same drug could be formulated in very different compositions even with different levels of drug, however, they could also be bioequivaient based on their pharmacokinetics if they both result in identical levels of drug in the bloodstream during the specified dosing period. Consequently, it is obvious to those skilled in the art that there are multiple different compositions that could be formulated to achieve the desired identical pharmacokinetic profile. Consequently, it is the pharmacokinetic profile that is particularly meaningful as this dictates the pharmacoiogical effect over time of the drug,

[0048] It should be obvious to those skilled in the art that to achieve the desired pharmacokinetic profile in the bioodstream of a patient of a beta-adrenergic inverse agonist for the treatment of diseases and conditions treatable by a beta-adrenergic inverse agonist for a T_max > 4 hours for once daily dosing, or more preferably for a T_maχ = 8 hours, pharmaceutical compositions can be formulated with the desired pharmacokinetic profile by altering the release pattern of the inverse agonist.

[0049] Accordingly, one aspect of the present invention is a method for cross- sensitizing the response of prostacyclin receptors to a prostacyclin agonist by administering a quantity of β₂-adrenergic receptor inverse agonist to a subject with a disease or condition treatable by potentiating the response of prostacyclin receptors to a prostacyclin agonist, the quantity being sufficient to detectably increase the response of a prostacyclin receptor to a prostacyclin agonist.

[0050] The β₂-adrenergic receptor inverse agonist can be a selective or a nonselective β₂-adrenergic receptor inverse agonist.

[0051] The disease or condition can be pulmonary hypertension or renal failure due to vascular insufficiency. Typically, the disease or condition is pulmonary hypertension. Preferably, the disease or condition is pulmonary arterial hypertension.

[0052] Prostacyclin receptors are located in a significant number of tissue types. In addition to arterial smooth muscle in vasculature, prostacyclin receptors have been identified in kidney vasculature, brain, platelets, mature thymocytes in the thymus, and in splenic lymphocytes and megakaryocytes in spleen. These additional locations for prostacyclin receptors support the use of β-adrenergic inverse agonists for additional diseases or conditions.

[0053] For example, the disease or condition can be a degenerative disease of the central nervous system such as, but not limited to, Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease) (U.S. Patent No. 6,884,819 to Suwa et al., incorporated herein by this reference). [0054] The disease or condition can also be a disease or condition relating to disturbances or dysregulation of peripheral circulation such as, but not limited to, Raynaud's disease, vascular ischemia, or limb ischemia.

[0055] The disease or condition can aiso be thrombosis, as administration of the inverse agonist can be used to prevent platelet aggregation and blood clotting.

[0056] The disease or condition can also be headache (cephalgia), including, but not limited to migraines or cluster headaches. These conditions are characterized by cerebral arterial vasospasm.

[0057] In classical receptor theory, two classes of G protein-coupled receptor (GPCR) ligands were considered: agonist and antagonist. Receptors were believed to exist in a single quiescent state that could only induce cellular signaling upon agonist binding to produce an activated receptor state. In this model, binding by antagonists produced no cellular signaling but simply prevented receptors from being bound and activated by agonists. Then, Costa and Herz demonstrated that receptors could be manipulated into a constitutive or spontaneously active state that produced cellular signaling in the absence of agonist occupation. They also provided evidence that certain compounds inactivate those spontaneously active receptors (T. Costa & A. Herz, "Antagonists with Negative intrinsic Activity at δ Opioid Receptors Coupled to GTP- Binding Proteins," Proc. Natl. Acad. Sci. USA 86: 7321-7325 (1989)). There is further evidence that GPCRs exist in constitutively or spontaneously active states that are inactivated to some degree by inverse agonists (R.A. de Ligt et al., "Inverse Agonism at G Protein-Coupled Receptors: (Patho)physiological Relevance and Implications^ for Drug Discovery," Br. J. Pharmacol. 130: 1-12 (2000); G. Milligan et al., "Inverse Agonism: Pharmacological Curiosity or Potential Therapeutic Strategy?," Trends Pharmacol. Sci. 16: 10-13 (2000)).

[0058] The basis of the strategy of this embodiment of the invention is the recognition of the existence of inverse agonists and the understanding of the effect that chronic treatment with inverse agonists has on receptor function. What is an inverse agonist and how does it function? Receptors, such as β-adrenergic receptors that respond to adrenalin (epinephrine), typically exist in an equilibrium between two states, an active state and an inactive state. When receptors bind to agonists, such as adrenalin for the β-adrenoceptors, they stop them from cycling back into the inactive state, thus shifting the equilibrium between the active and inactive states according to the law of mass action. This occurs because those receptors bound to agonists are removed from the equilibrium. Typically, antagonists bind to the receptors, but prevent the binding of agonists. However, molecules known as "inverse agonists" bind to the receptors in the inactive state, causing the equilibrium between the active and the inactive state to shift toward the inactive state. This is not merely a matter of blocking agonist binding.

[0059] Moreover, there is a population of spontaneously active receptors in vivo. These receptors provide a baseline constitutive level of activity; the activity is never entirely "off."

[0060] As indicated above, it has been well demonstrated that chronic administration of β-adrenergic agonists causes agonist-dependent desensitization. Upon acute administration of β-agonists, adrenergic receptors are internalized, thereby preventing them from being restimulated further for pulmonary relaxation. With chronic administration of β-agonists, there is actually a down regulation in the total number of β- adrenergic receptors. The consequence may be the observed loss of responsiveness seen in asthmatic patients on long-acting β-agonists, and referred to as tolerance or tachyphylaxis, as described above.

[0061] The treatment methods of the present invention are based on the further discovery that a chronic administration of an inverse agonist has the effect of upregulating not only the population of active β-adrenergic receptors, but also the population of other GPCRs, including the prostacyclin receptor (PR). This phenomenon is referred to herein as "cross-sensitization." The observed activity may be due to the receptor's constitutive baseline activity or the combined effect of increased level of receptors responding to endogenous agonists. This leads to the seemingly paradoxical result that the administration of a drug that would appear, at first blush, to degrade a physiological function, such as by causing airway hyperresponsiveness in asthma, can, if administered chronically, enhances that physiological function by upregulating the population of spontaneously active β-adrenergic receptors associated with that physiological function. Additionally or alternatively, the inverse agonist may also improve coupling of the receptor to its cognate internal G protein thereby resulting in a higher output of result such as the production of cellular cAMP with a smaller proportion of activated receptors. This is a specific application of the principle of "paradoxical pharmacology." The discovery of "cross-sensitization" extends the scope of "paradoxical pharmacology" from receptors directly bound by the inverse agonist to other receptors. Although Applicant does not intend to be bound by this theory, nevertheless it is consistent with the observed results.

[0062] Fong and Cornett (United States Patent Application Publication No. 2005/0043391 , incorporated herein by this reference) specify the use of prostacyclin agonists in an oral combination with a large number of different classes of antihypertensive agents for the treatment of hypertension. One of the genera listed is beta blockers, however, there is no discrimination between beta blockers that are antagonists with partial agonist activity - also termed intrinsic sympathomimetic activity, and those beta blockers that only have inverse agonist activity. Both subclasses of beta blockers are represented in their list of active agents such as acebutolol and penbutolol, both of which are beta blocker antagonists with partial agonist activity which are counter to the current invention as well as inverse agonists such as atenolol.

[0063] In contrast, according to the present invention, only β₂-adrenergic inverse agonists, either selective or non-selective, are used.

[0064] β-adrenergic antagonist drugs or "beta blockers" are treated as having the same activity in conventional pharmacology. Beta blockers are further classified based on their selectivity or lack thereof for either the βi (termed "cardio selective") or P₁Zp₂ ("nonselective") or β₂ selective only. Additionally, beta blockers can be classified as to whether or not they have partial agonist activity or are actually inverse agonists. The latter definition is based on the new appreciation, recited in the present application, that many G-coupled protein receptors, including the β-adrenergic receptors, exhibit low level spontaneous activity that can be further prevented by the binding of the inverse agonists to the receptor. This distinction was not made in PCT Patent Publication No. WO 02/29534, which referred simply to "antagonists."

[0065] Despite this knowledge of the subclasses of beta blockers in the field, many scientists have continued to treat compounds from the different subclasses as one class. An example of this is the clinical testing in 1998-1999 of the beta blocker bucindolol for congestive heart failure. Previously, two other beta blockers, metoprolol and carvedilol, had been clinically tested and demonstrated significant mortality reduction in patients with CHF. Bucindolol failed to demonstrate any benefit over placebo, and thus clinical testing was discontinued. The inventor of the present application notes that both metoprolol and carved ilol are β-inverse agonists whereas bucindolol is a neutral antagonist with partial agonist activity. Consequently, the inventor of the present application would predict that only β-adrenergic inverse agonists would be effective in treatment of CHF. In the same vein, the inventor of the present application predicts that only β-adrenergic inverse agonists will be effective for chronic treatment of asthma airway hyperresponsiveness. This distinction is not made or suggested in PCT Patent Publication No. WO 02/29534. This prediction is borne out in the present invention by the refutation that the beta blocker alprenolol, a partial agonist, previously thought to be the preferred drug in a flawed murine asthma model, was found to be without any activity in the present invention.

[0066] Instead, this invention provides for the use of the active β-adrenergic receptor binding forms of β-adrenergic inverse agonists in cross-sensitization methods according to the present invention. The inverse agonists can be in pure or substantially pure enantiomeric or diastereomeric form or can be racemic mixtures. In many cases, the active form of such compounds is the L form when there is only one chiral center, in the case of nadolol, which has three chiral centers and potentially 12 isomers, though, typically, only two are formed during synthesis, the most active form is the RSR form of nadolol.

[0067] Especially preferred for use according to the invention are the β- adrenergic inverse agonists: nadolol, e.g., as the hydrochloride: bupranolol, e.g., as the hydrochloride; butoxamine, e.g., as the hydrochloride; carazolol, e.g., as the hydrochloride; carvedilol; , e.g., as the hydrochloride; ICI-118,551 , i.e., as the hydrochloride; levobunolol, e.g., as the hydrochloride; metoprolol, as the tartrate or succinate; propranolol, e.g., as the hydrochloride; sotalol, e.g., as the hydrochloride; timolol; e.g., as the hydrochloride; and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, metabolites, precursors, esters, eno! ethers, enol esters, and prodrugs thereof. Particularly preferred inverse agonists are carvediiol and nadolol. A most particularly preferred inverse agonist is nadolol. As used herein, the recitation of an inverse agonist compound, or, where appropriate, an agonist compound, includes all pharmaceutically acceptable salts of that inverse agonist compound or agonist compound unless excluded. Thus, the recitation of nadolol as the hydrochloride does not exclude other pharmaceutically acceptable salts that have been prepared or that can be prepared.

[0068] The inverse agonists useful in methods and compositions according to the invention typically display inverse agonism to β₂-adrenergic receptors; either as nonselective inverse agonists that display inverse agonism to both the βi- and β₂- adrenergic receptors or as a selective β2-inverse agonist.

[0069] Specifically, also expected to be within the scope of the invention are analogues of nadolol of formula (I) wherein Ri is hydrogen or lower alkyl, R2 is hydrogen or lower alkyl, and m and n are 1 to 3, with the proviso that where Ri and R₂ are both hydrogen and m is 1 , n is other than 1. As used herein, the term "lower alkyl" is defined as a straight or branched hydrocarbyl residue of 1-6 carbon atoms.

(CH₂)_n MH C(CHg)₃

(I)

[0070] Also specifically expected to be within the scope of the invention are analogues of carvedilol of formula (II) wherein Ri is hydrogen or lower alky!, R₂ is hydrogen or lower alkyl, and R₃ is hydrogen or lower alky], with the proviso that all of R-i, R₂, and R₃ are not all hydrogen.

[0071] Also expected to be within the scope of the invention are analogues of timolol of formula (III) wherein R₁ is hydrogen or lower alkyl and R₂ is hydrogen or lower alkyl, with the proviso that both Ri and R₂ are not hydrogen.

(Ill) [0072] Further expected to be within the scope of the invention are analogues of metoprolol of formula (IV) wherein R₁ is hydrogen or lower alkyl and R₂ is hydrogen or lower alkyl, with the proviso that both Ri and R₂ are not hydrogen.

(IV)

[0073] Also further expected to be within the scope of the invention are analogues of ICM 18,551 of formula (V) wherein R₁ is lower alkyl, R₂ is hydrogen or lower alkyl, R₃ is hydrogen or lower alkyl, R₄ is hydrogen or lower alkyl, R₅ is lower alkyl, and R₆ is lower alkyl, with the proviso that all of Ri, R₃, R₅, and R₆ are not methyl and all of R₂ and R₄ are not hydrogen.

(V) [0074] in the case of salts, it is well known that organic compounds, including compounds having activities suitable for methods according to the present invention, have multiple groups that can accept or donate protons, depending upon the pH of the solution in which they are present. These groups include carboxyl groups, hydroxyl groups, amino groups, sulfonic acid groups, and other groups known to be involved in acid-base reactions. The recitation of a compound or analogue includes such salt forms as occur at physiological pH or at the pH of a pharmaceutical composition unless specifically excluded.

[0075] Similarly, prodrug esters can be formed by reaction of either a carboxyl or a hydroxy! group on compounds or analogues suitable for methods according to the present invention with either an acid or an alcohol to form an ester. Typically, the acid or alcohol includes a lower alkyl group such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, and tertiary butyl. These groups can be substituted with substituents such as hydroxy, or other substituents. Such prodrugs are well known in the art and need not be described further here. The prodrug is converted into the active compound by hydrolysis of the ester linkage, typically by intracellular enzymes. Other suitable groups that can be used to form prodrug esters are well known in the art. For example prodrugs can inciude amides prepared by reaction of the parent acid compound with a suitable amine. In some cases it is desirable to prepare double ester type prodrugs such as (acyloxy) alky] esters or ((alkoxycarbonyl)oxy)alkyl esters. Suitable esters as prodrugs include, but are not necessarily limited to, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, tert- butyl, morpholinoethyl, and N,N-diethylg!ycolamido. Methyl ester prodrugs may be prepared by reaction of the acid form of a compound having a suitable carboxylic acid group in a medium such as methanol with an acid or base esterification catalyst (e.g., NaOH, H₂ SO₄). Ethyl ester prodrugs are prepared in similar fashion using ethanol in place of methanol. Morphoϋnylethyl ester prodrugs may be prepared by reaction of the sodium salt of a suitable compound (in a medium such as dimethylformamide) with 4-(2- chloroethyl)morphine hydrochloride (available from Aldrich Chemical Co., Milwaukee, Wis. USA.

[0076] Examples of the ethers include, but are not limited to, alkyl ethers, for example, lower alkyl ethers such as methyl ether, ethyl ether, propyl ether, isopropyl ether, butyl ether, isobutyl ether, t-butyl ether, peπtyl ether and 1-cyclopropyi ethyl ether; and medium or higher alkyl ethers such as octy! ether, diethyihexyl ether, lauryl ether and cetyl ether; unsaturated ethers such as oleyl ether and linolenyl ether; lower aikeny! ethers such as vinyl ether, allyl ether; lower alkynyl ethers such as ethynyl ether and propynyl ether; hydroxy (lower) alkyl ethers such as hydroxyethyl ether and hydroxyisopropyl ether; lower alkoxy (lower) alkyl ethers such as methoxymethyl ether and 1-methoxyethyl ether; optionally substituted aryl ethers such as phenyl ether, tosyl ether, t-butylphenyl ether, salicyl ether, 3,4-di-methoxyphenyl ether and benzamidophenyl ether; and aryl (lower) alkyl ethers such as benzyl ether, trityl ether and benzhydryl ether, or other ether forms.

[0077] Examples of the esters may include, but are not limited to, aliphatic esters, for example, lower alkyl esters such as methyl ester, ethyl ester, propyl ester, isopropyl ester, butyl ester, isobutyl ester, t-butyl ester, pentyl ester and 1- cyclopropylethyl ester; lower alkenyl esters such as vinyl ester and allyl ester; lower alkynyl esters such as ethynyl ester and propynyl ester; hydroxy (lower) alkyl ester such as hydroxyethyl ester; lower alkoxy (lower) alkyl esters such as methoxymethyl ester and 1-methoxyethyl ester; and optionally substituted aryl esters such as, for example, phenyl ester, tosyl ester, t-butylphenyl ester, salicyl ester, 3,4-di-methoxyphenyl ester and benzamidophenyl ester; and aryl(lower)alkyl ester such as benzyl ester, trityl ester and benzhydryl ester, or other ester forms.

[0078] Pharmaceutically acceptable salts include acid salts such as hydrochlorides, hydrobromides, hydroiodides, sulfates, phosphates, fumarates, maleates, acetates, citrates, lactates, tartrates, sulfamates, malonate, succinate, tartrate, methanesulfonates, ethanesulfonates, benzenesulfonates, p-toluenesulfonates, cyclohexylsulfamates, quinates, formates, cinnamates, picrates, and other suitable salts. Such salts can be derived using acids such as hydrochloric acid, sulfuric acid, phosphoric acid, sulfamic acid, acetic acid, citric acid, lactic acid, tartaric acid, malonic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, cyclohexylsulfamic acid, and quinic acid.

[0079] Pharmaceutically acceptable salts also include salts with bases such as alkali metal salts such as sodium or potassium, as well as pyridine salts, ammonium salts, piperazine salts, diethylamine salts, nicotinamide salts, caicium salts, magnesium salts, zinc salts, lithium salts, methylamino salts, triethylamino salts, dimethylamino saits, and tris(hydroxymethyl) aminomethane salts.

[0080] Accordingly, one aspect of the present invention is a method for improving the effectiveness of a G-Protein Coupled Receptor (GPCR) agonist drug comprising the step of chronic co-administration of an inverse agonist drug that targets a different GPCR receptor, the different GPCR sharing a common G protein with the receptor bound by the GPCR agonist drug. In one alternative, the common G protein is the Gs protein.

[0081] In this aspect of the present invention, typically the β-adrenergic inverse agonist is selected from the group consisting of β₂-selective inverse agonists, and nonselective inverse agonists having inverse agonist activity against both βr and β₂- adrenergic receptors. More typically, the inverse agonist is typically a β₂-adrenergic receptor inverse agonist.

[0082] Suitable β₂-adrenergic receptor inverse agonists in this aspect of the present invention include inverse agonists selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvedilol, ICI-118,551 , levobunolol, metoprolol, propranolol, sotalol, and timolol, and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof, as described above.

[0083] The subject to be treated can be a human patient or a socially or economically important animal, including, but not limited to, a dog, a cat, a cow, a horse, a sheep, a goat, or a pig. Methods according to the present invention are not limited to the treatment of humans.

[0084] Typically, the method of administration of the β-adrenergic inverse agonist results in continuous levels of the β-adrenergic inverse agonist in the bloodstream of the subject.

[0085] The β-adrenergic inverse agonist can be administered in conjunction with one or more pharmaceutical excipients. The pharmaceutical excipients can include, but are not necessarily limited to, calcium carbonate, calcium phosphate, various sugars or types of starch, cellulose derivatives, geiatin, vegetable oils, polyethylene glycols and physiologically compatible solvents. Other pharmaceutical excipients are well known in the art. The β-adrenergic inverse agonist can be administered in conjunction with one or more pharmaceutically acceptable carriers. Exemplary pharmaceutically acceptable carriers include, but are not limited to, any and/or all of solvents, including aqueous and non-aqueous solvents, dispersion media, coatings, antibacterial and/or antifungal agents, isotonic and/or absorption delaying agent, and/or the like. The use of such media and/or agents for pharmaceutically active substances is well known in the art. Except insofar as any conventional medium, carrier, or agent is incompatible with the active ingredient or ingredients, its use in a composition according to the present invention is contemplated. Supplementary active ingredients can also be incorporated into the compositions, especially as described below under combination therapy. For administration of any of the compounds used in the present invention, preparations should meet sterility, pyrogenicity, general safety, and purity standards as required by the FDA Office of Biologies Standards or by other regulatory organizations regulating drugs.

[0086] Various factors must be taken into account in setting suitable dosages for β-adrenergic inverse agonists. These factors include whether the patient is taking other medications that can alter the pharmacokinetics of the β-adrenergic inverse agonists, either causing them to be degraded more rapidly or more slowly. In particular, if the patient is taking the antibiotics erythromycin or neomycin, it is typically necessary to decrease the maintenance dose. Another aspect of the invention is therefore a blister pack that has backup restoration doses and lower doses for use when the patient is taking these antibiotics.

[0087] Toxicity and therapeutic efficacy of β-adrenergic inverse agonists can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD₅₀ (the dose lethal to 50% of the population) and the ED₅₀ (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD₅₀/ED₅o. Compounds which exhibit large therapeutic indices are preferred. The data obtained from these ceϋ culture assays and animal studies can be used in formulating a range of dosage for use in humans. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED₅₀ with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized.

[0088] For any compound used in the method of the invention, the therapeutically effective dose can be estimated initially from cell culture assays. For example, a dose can be formulated in animal models to achieve a circulating plasma concentration range that includes the IC₅₀ as determined in cell culture (i.e., the concentration of the test compound which achieves a half-maximal improvement in receptor signaling when chronic effects are considered). Such information can be used to more accurately determine useful doses in humans. Levels in plasma may be measured, for example, by HPLC.

[0089] The exact formulation, route of administration and dosage can be chosen by the individual physician in view of the patient's condition. (See e.g. Fingl et al., in The Pharmacological Basis of Therapeutics, 1975, Ch. 1 p. 1 ). It should be noted that the attending physician would know how to and when to terminate, interrupt, or adjust administration due to toxicity, or to organ dysfunctions. Conversely, the attending physician would also know to adjust treatment to higher levels if the clinical response were not adequate (precluding toxicity). The magnitude of an administered dose in the management of the disorder of interest will vary with the severity of the condition to be treated and to the route of administration. The severity of the condition may, for example, be evaluated, in part, by standard prognostic evaluation methods. Further, the dose and perhaps the dose frequency, will also vary according to the age, body weight, and response of the individual patient. A program comparable to that discussed above may be used in veterinary medicine.

[0090] Depending on the specific conditions being treated, such agents may be formulated and administered systemically or locally. Typically, administration is systemic. Techniques for formulation and administration may be found in Remington's Pharmaceutical Sciences, 18th ed., Mack Publishing Co., Easton, Pa. (1990). Suitable routes may include oral, rectal, transdermal, vaginal, transmucosal, or intestinal administration; parenteral delivery, including intramuscular, subcutaneous, intramedullary injections, as well as intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, or intraocular injections, just to name a few. Typically, oral administration is preferred.

[0091] Thus, the β-adrenergic inverse agonist can be formulated for oral, sustained-release oral, buccal, sublingual, inhalation, insufflation, subcutaneous injection, or parenteral administration.

[0092] If the β-adrenergic inverse agonist is administered orally, either in a conventional or a sustained-release preparation, it is typically administered in a conventional unit dosage form such as a tablet, a capsule, a pill, a troche, a wafer, a powder, or a liquid such as a solution, a suspension, a tincture, or a syrup. Oral formulations typically include such normally employed excipients as, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, and other conventional pharmaceutical excipients. In certain defined embodiments, oral pharmaceutical compositions will comprise an inert diluent and/or assimilable edible carrier, and/or they may be enclosed in hard or soft shell gelatin capsules. Alternatively, they may be compressed into tablets. As another alternative, particularly for veterinary practice, they can be incorporated directly into food. For oral therapeutic administration, they can be incorporated with excipients or used in the form of ingestible tablets, buccal tablets, dragees, pills, troches, capsules, wafers, or other conventional dosage forms.

[0093] The tablets, pills, troches, capsules, wafers, or other conventional dosage forms can also contain the following: a binder, such as gum tragacanth, acacia, cornstarch, sorbitol, mucilage of starch, polyvinylpyrrolidone, or gelatin; excipients or fillers such as dicalcium phosphate, lactose, microcrystalline cellulose, or sugar; a disintegrating agent such as potato starch, croscarmellose sodium, or sodium starch glycolate, or alginic acid; a lubricant such as magnesium stearate, stearic acid, talc, polyethylene glycol, or silica; a sweetening agent, such as sucrose, lactose, or saccharin; a wetting agent such as sodium laury! sulfate; or a flavoring agent, such as peppermint, oil of wintergreen, orange flavoring, or cherry flavoring. When the dosage unit form is a capsule, it can contain, in addition to materials of the above types, a liquid carrier. Various other materials can be present as coatings or to otherwise modify the physical form and properties of the dosage unit. For instance, tablets, pills, or capsules can be coated with shellac, sugar, or both. The pharmaceutical compositions of the present invention may be manufactured in a manner that is itself known, e.g., by means of conventional mixing, dissolving, granulating, dragee-making, levitating, emulsifying, encapsulating, entrapping or lyophilizing processes.

[0094] Pharmaceutical preparations for oral use can be obtained by combining the active compounds with solid excipient, optionally grinding a resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores. Suitable excipients are, in particular, fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, and/or polyvinylpyrrolidone (PVP). If desired, disintegrating agents may be added, such as the cross-linked polyvinyl pyrrolidone, agar, or afginic acid or a salt thereof such as sodium alginate.

[0095] Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses.

[0096] Pharmaceutical preparations which can be used orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. The push-fit capsules can contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added. [0097] In one alternative, a sustained-release formulation is used. Sustained- release formulations are well-known in the art. For example, they can include the use of polysaccharides such as xanthan gum and locust bean gum in conjunction with carriers such as dimethylsiloxane, silicic acid, a mixture of mannans and gaiactans, xanthans, and micronized seaweed, as recited in U.S. Patent No. 6,039,980 to Baichwal, incorporated herein by this reference. Other sustained -re I ease formulations incorporate a biodegradable polymer, such as the lactic acid-glycolic acid polymer recited in U.S. Patent No. 6,740,634 to Saikawa et al., incorporated herein by this reference. Stili other sustained-release formulations incorporate an expandable lattice that includes a polymer based on polyvinyl aScohol and polyethylene glycol, as recited in U.S. Patent No. 4,428,926 to Keith, incorporated herein by this reference. Still other sustained- release formulations are based on the Eudragit™ polymers of Rohm & Haas, that include copolymers of acrylate and methacrylates with quaternary ammonium groups as functional groups as well as ethylacrylate methylmethacrylate copolymers with a neutral ester group. A particularly-preferred extended release composition suitable for use in methods according to the present invention is an extended-release composition that contains nadolol as its active ingredient.

[0098] Oral liquid preparations can be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups, tinctures, or elixirs, or can be presented as a dry product for reconstitution with water or other suitable vehicles before use. Such liquid preparations can contain conventional additives such as suspending agents, for example, sorbitol syrup, methylcellulose, glucose/sugar syrup, gelatin, hydroxymethylcellulose, carboxymethylcellulose, aluminum stearate gel, or hydrogenated edible fats; emulsifying agents, such as lecithin, sorbitan monooleate, or acacia; non-aqueous vehicles (which may include edible oils), for example, aimond oil, fractionated coconut oil, oily esters, propylene glycol, or ethyl alcohol; or preservatives, for example, methylparaben, propylparaben, or sorbic acid. The preparations can also contain buffer salts, flavoring, coloring, or sweetening agents (e.g., mannitol) as appropriate. [0099] One skilled in the art recognizes that the route of administration is an important determinant of the rate of efficiency of absorption. For example, the alimentary route, e.g., oral, rectal, sublingual, or buccal, is generally considered the safest route of administration. The delivery of the drugs into the circulation is slow, thus eliminating rapid high blood levels of the drugs that could potentially have adverse acute effects. Although this is considered the safest route of administration, there are several disadvantages. One important disadvantage is that the rate of absorption varies, which is a significant problem if a small range in blood levels separates a drug's desired therapeutic effect from its toxic effect, i.e., if the drug has a relatively low therapeutic index. Also, patient compliance is not always ensured, especially if the rectal route of administration is chosen or if oral administration is perceived by the patient as unpleasant. Furthermore, with oral administration, extensive hepatic metabolism can occur before the drug reaches its target site. Another route of administration is parenteral, which bypasses the alimentary tract. One important advantage of parenteral administration is that the time for the drug to reach its target site is decreased, resulting in a rapid response, which is essential in an emergency. Furthermore, parenteral administration allows for delivery of a more accurate dose. Parenteral administration also allows for more rapid absorption of the drug, which can result in increased adverse effects. Unlike alimentary administration, parenteral administration requires a sterile formulation of the drug and aseptic techniques are essential. The most significant disadvantage to parenteral administration is that it is not suitable for insoluble substances, in addition to alimentary and parenteral administration routes, topical and inhalation administrations can be useful. Topical administration of a drug is useful for treatment of local conditions; however, there is usually little systemic absorption. Inhalation of a drug provides rapid access to the circulation and is the common route of administration for gaseous and volatile drugs, or drugs that can be vaporized or nebulized. It is also a desired route of administration when the targets for the drug are present in the pulmonary system.

[0100] When compounds are formulated for parenteral administration, e.g., formulated for injection via the intravenous, intramuscular, subcutaneous, intralesional, or intraperitoneal routes, many options are possible. The preparation of an aqueous composition that contains an effective amount of the β-adrenergic inverse agonist as an active ingredient wili be known to those of skill in the art. Typically, such compositions can be prepared as injectables, either as liquid solutions and/or suspensions. Solid forms suitable for use to prepare solutions and/or suspensions upon the addition of a liquid prior to injection can also be prepared. The preparations can also be emulsified.

[0101] The pharmaceutical forms suitable for injectable use include sterile aqueous solutions and/or dispersions; formulations including sesame oil, peanut oil, synthetic fatty acid esters such as ethyl oleate, triglycerides, and/or aqueous propylene glycol; and/or sterile powders for the extemporaneous preparation of sterile injectable solutions and/or dispersions. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions. In all cases the form must be sterile and/or must be fluid to the extent that the solution will pass readily through a syringe and needle of suitable diameter for administration. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria or fungi.

[0102] Solutions of the active compounds as free base or pharmacologically acceptable salts can be prepared in water suitably mixed with a surfactant, such as hydroxypropylcellulose. Dispersions can also be prepared in glycerol, liquid polyethylene glycols, and/or mixtures thereof and/or in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms. Suitable non-sensitizing and non-allergenic preservatives are well known in the art.

[0103] The carrier can also be a solvent and/or dispersion medium containing, for example, water, ethanol, a polyol (for example, glycerol, propylene glycol, and/or liquid polyethylene glycol, and/or the like), suitable mixtures thereof, and/or vegetable oils. The proper fluidity can be maintained for example, by the use of a coating, such as lecithin, by the maintenance of a suitable particle size in the case of a dispersion, and/or by the use of surfactants. The prevention of the action of microorganisms can be brought about by the inclusion of various antibacterial and/or antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, or thimerosai. In many cases it will be preferable to include isotonic agents, for example, sugars or sodium chloride. In many cases, it is preferable to prepare the solution in physiologically compatible buffers such as Hanks's solution, Ringer's solution, or physiological saline buffer. Prolonged absorption of the injectable compositions can be brought about by the use in the compositions of agents delaying absorption, for example, aluminum monostearate and/or gelatin.

[0104] Sterile injectable solutions are prepared by incorporating the active compounds in the required amount in the appropriate solvent with various of the other ingredients enumerated above, as required, followed by sterilization. Sterilization is typically performed by filtration. Generally, dispersions are prepared by incorporating the various sterilized active ingredients into a sterile vehicle which contains the basic dispersion medium and/or the other required ingredients. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum-drying and/or freeze-drying techniques that yield a powder of the active ingredients plus any additional desires ingredients from a previously sterile-filtered solution thereof. The preparation of more-concentrated or highly-concentration solutions for direct injection is also contemplated, where the use of dimethyl sulfoxide (DMSO) as solvent is envisioned to result in extremely rapid penetration, delivering high concentrations of the active agents to a small area if desired.

[0105] For parenteral administration in an aqueous solution, for example, the solution should be suitably buffered if necessary and/or the liquid diluent first rendered isotonic with sufficient saline, glucose, or other tonicity agent. These particular aqueous solutions are especially suitable for intravenous, intramuscular, subcutaneous, or intraperitoneal administration. In this connection, sterile aqueous media which can be employed will be known to those of skill in the art in light of the present disclosure. For example, one dosage could be dissolved in 1 ml_ of isotonic NaCI solution and either added to 1000 ml_ of hypodermoclysis fluid or injected into the proposed site of infusion (see, e.g., "Remington's Pharmaceutical Sciences" (15^th ed.), PP- 1035-1038, 1570- 1580). Some variation in dosage will necessarily occur depending on the condition of the subject being treated. The person responsible for administration will, in any event, determine the appropriate dose for the individual subject. Compounds and compositions according to the invention can also be formuiated for parenteral administration by bolus injection or continuous infusion and can be presented in unit dose form, for instance as ampoules, vials, small volume infusions, or pre-filled syringes, or in multi-dose containers with an added preservative.

[0106] Another route of administration of compositions according to the present invention is nasally, using dosage forms such as nasal solutions, nasal sprays, aerosols, or inhalants. Nasal solutions are usually aqueous solutions designed to be administered to the nasal passages in drops or sprays. Nasal solutions are typically prepared so that they are similar in many respects to nasal secretions, so that normal ciliary action is maintained. Thus, the aqueous nasal solutions usually are isotonic and/or slightly buffered in order to maintain a pH of from about 5.5 to about 6.5. In addition, antimicrobial preservatives, similar to those used in ophthalmic preparations, and/or appropriate drug stabilizers, if required, can be included in the formulation. Various commercial nasal preparations are known and can include, for example, antibiotics or antihistamines. Spray compositions can be formulated, for example, as aqueous solutions or suspensions or as aerosols delivered from pressurized packs, with the use of a suitable propellant, such as dichlorodifiuoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, 1 ,1 ,1 ,2,3,3,3-heptafluoropropane, 1 ,1 ,1 ,2-tetrafluoroethane, carbon dioxide, or other suitable gas.

[0107] Aerosol therapy allows an almost ideal benefit to risk ratio to be achieved because very small doses of inhaled medication provide optimal therapy with minimal adverse effects. However, the therapeutic efficiency of drugs administered by aerosolization depends not only on the pharmacological properties of the drugs themselves, but also on the characteristics of the delivery device. The characteristics of the delivery device influence the amount of drug deposited in the lungs and the pattern of drug distribution in the airways.

[0108] Aerosols are airborne suspensions of fine particles. The particles may be solids or liquids. Aerosol particles are heterodisperse (i.e. the particles are of a range of sizes) and aerosol particle size distribution is best described by a log normal distribution. Particles tend to settle (sediment), adhere to each other (coagulate), and adhere to structures such as tubing and mucosa (deposit). The particles delivered by aerosol can be conveniently characterized on the basis of their aerodynamic behavior. One parameter is the mass median aerodynamic diameter (MMAD). By definition, a particle distribution with an MMAD of 1 μM has the same average rate of settling as a droplet of unit density and 1 μM diameter.

[0109] The size of an aerosol particle, as well as variables affecting the respiratory system, influence the deposition of inhaled aerosols in the airways. On one hand, particles larger than 10 μM in diameter are unlikely to deposit in the lungs. However, particles smaller than 0.5 μM are likely to reach the alveoli or may be exhaled. Therefore, particles that have a diameter of between 1 μM and 5 μM are most efficiently deposited in the lower respiratory tract. Particles of these sizes are most efficient for the delivery of therapeutic agents, including β-adrenergic inverse agonists.

[0110] The percentage of the aerosol mass contained within respirable droplets (i.e., droplets with a diameter smaller than 5 μM), depends on the inhalation device being used. Slow, steady inhalation increases the number of particles that penetrate the peripheral parts of the lungs. As the inhaled volume is increased, the aerosol can penetrate more peripherally into the bronchial tree. A period of breath-holding, on completion of inhalation, enables those particles that have penetrated to the lung periphery to settle into the airways via gravity. Increased inspiratory flow rates result in increased losses of inhaled drug. This occurs because aerosol particles impact in the upper airway and at the bifurcations of the first few bronchial divisions. Other factors associated with pulmonary airway disease, and other diseases and conditions as well, including diseases and conditions affecting lung function, may also alter aerosol deposition. .

[0111] In aerosol administration, the nose efficiently traps particles before their deposition in the lung; therefore, mouth breathing of the aerosolized particles is preferred. The aerosolized particles are lost from many sites. Generally, the amount of the nebulized dose reaching the small airways is < 15%. In many cases, approximately 90% of the inhaled dose is swallowed and then absorbed from the gastrointestinal tract. The small fraction of the dose that reaches the airways is also absorbed into the blood stream. The swallowed fraction of the dose is, therefore, absorbed and metabolized in the same way as an oral formulation, while the fraction of the dose that reaches the airways is absorbed into the blood stream and metabolized in the same way as an intravenous dose.

[0112] When drugs are administered topically (via aerosol delivery to the lungs), the desired therapeutic effects depend on local tissue concentrations, which may not be directly related to plasma drug concentrations. Because most inhaled drugs are administered at a low dosage and have a low oral bioavailability, plasma concentrations of these drugs are much lower than after oral administration.

[0113] Finally, the absolute pulmonary bioavailability of inhaled drugs is difficult to assess because blood concentrations are low, and pulmonary and oral absorption should be discriminated for pulmonary bioavailability to be determined as accurately as possible. Charcoal can be used to adsorb the swallowed fraction of inhaled terbutaline to discriminate the pulmonary absorption of the drug. Recently, it was shown that a urine collection during the 30 minutes after inhalation of salbutamol represents the amount of drug delivered to the lungs. This technique may be applicable for the determination of bioavailability of other inhaled drugs. Other techniques for the determination of bioavailability of inhaled drugs are also known in the art; these include pharmacodynamic methods using FEV₁ measurements, lung deposition studies using radiolabeled formulations, or pharmacokinetic studies using predominantly urinary excretion measurements.

[0114] Therapeutic aerosols are commonly produced by atomization of liquids within jet nebulizers or by vibration of a standing pool of liquid (ultrasonic nebulization). Preformed aerosols may also be administered. Examples of the latter include MDIs and dry powder devices. Whatever delivery device is used, patients should be taught to use it correctly.

[0115] All jet nebulizers work via a similar operating principle, represented by the familiar perfume atomizer. A liquid is placed at the bottom of a closed container, and the aerosol is generated by a jet of air from either a compressor or a compressed gas cylinder passing through the device. Ultrasonic nebulizers produce an aerosol by vibrating liquid lying above a transducer at frequencies of about 1 mHz. This produces a cloud of particles that is carried out of the device to the patient by a stream of air. Aerosols varying in quantity, size and distribution of panicles can be produced by nebulizers, depending upon the design of the nebulizers and how it is operated. It should be noted that not all nebulizers have the required specifications (MMAD, flow, output) to provide optimum efficacy. A recent study compared the lung deposition from 4 nebulizers in healthy volunteers and showed that median lung aerosol deposition, expressed as percentages of the doses initially loaded into the nebulizers, ranged from 2 to 19%. To minimize adverse effects, pH and osmolarity of the nebulized solution should be controlled.

[0116] Metered dose inhalers (MDIs), because of their convenience and effectiveness, are probably the most widely used therapeutic aerosol used for inhaled drug delivery to outpatients. Most MDIs in current use contain suspensions of drug in propellant. There are 2 major components of an MDI: (i) the canister, a closed plastic or metal cylinder that contains propellant, active medication, and the metering chamber; and (ii) the actuator, a molded plastic container that holds the canister and directs the released aerosol towards the patient's airway.

[0117] Propellant mixtures are selected to achieve the vapor pressure and spray characteristics desired for optimal drug delivery. Chlorofluorocarbons were previously used, but non-chlorinated propeliants are now employed because of environmental concerns. Finely divided particles of drug, usually less than 1 μM, are suspended in the pressurized (liquefied) propellant. To prevent the drug from coagulating, a surface active agent such as sorbitan oleate, lecithin or oleic acid is typically added; other surface active agents are known in the art. Metering chambers ordinarily contain 25 to 100 μL. The contents of the metering chamber are released when the canister is depressed into the actuator. Almost instantaneously, the propeliants begin to evaporate, producing disintegration of the discharged liquid into particles that are propelled forward with great momentum. For optimal pulmonary drug deposition, the medication should be released at the beginning of a slow inspiration that lasts about 5 seconds and is followed by 10 seconds of breath-holding. Several inhalation aids have been designed to improve the effectiveness of a MDI. These are most useful in patients who have poor hand-to- breath coordination. A short tube {e.g. cones or spheres) may direct the aerosol straight into the mouth or collapsible bags may act as an aerosol reservoir holding particles in suspension for 3 to 5 seconds, during which time the patient can inhale the drug. However, when any of these devices is used, aerosol velocity upon entering the oropharynx is decreased and drug availability to the lungs and deposition in the oropharynx is decreased.

[0118] Dry powder inhalers have been devised to deliver agents to patients who have difficulty using an MDI (e.g. children and elderly patients). In general, the appropriate dosage is placed in a capsule along with a flow aid or filler such as large lactose or glucose panicles. Inside the device, the capsule is initially either pierced by needles (e.g. Spinhaler®) or sheared in half (e.g. Rotohaler®). During inhalation the capsule rotates or a propeller is turned, creating conditions that cause the contents of the capsule to enter the inspired air and be broken up to small particles suitable for delivery to the airways. The energy required to disperse the powder is derived from the patient's inspiratory effort. Recently, more convenient multidose dry powder inhalers have been introduced (e.g. Diskhaler® , Turbuhaler®). Potential problems associated with dry powder inhalers include esophageal irritation and, consequently, cough due to the direct effect of powder in airways. Furthermore, the walls of the capsule may be coated with drug as a result of either failure of the capsule to release the drug or failure of the aggregated powder to break up. This may cause virtually all of the drug to be deposited in the mouth. These powder devices do not contain chlorofluorocarbons and may provide an alternative to MDIs.

[0119] Additional formulations that are suitable for other modes of administration include vaginal suppositories and/or pessaries. A rectal pessary or suppository can also be used. Suppositories are solid dosage forms of various weights or shapes, usually medicated, for insertion into the rectum, vagina, or urethra. After insertion, suppositories soften, melt, and/or dissolve into the cavity fluids. In general, for suppositories, traditional binders or carriers can include polyalkylene glycols, cocoa butter, or triglycerides. [0120] Other dosage forms, including but not limited to liposomal formulations, ointments, creams, lotions, powders, or creams, can alternatively be used. Ointments and creams can, for example, be formulated with an aqueous or oily base with the addition of suitable gelling agents and/or solvents. Such bases, can thus, for example, include water and/or an oil such as liquid paraffin or a vegetable oil such as arachis (peanut) oil or castor oil or a solvent such as a polyethylene glycol. Thickening agents which can be used include soft paraffin, aluminum stearate, cetostearyl alcohol, polyethylene glycols, microcrystalline wax, and beeswax. Lotions can be formulated with an aqueous or oily base and will in general also contain one or emulsifying agents, stabilizing agents, dispersing agents, suspending agents, or thickening agents.

[0121] Powders for external application can be formed with the aid of any suitable powder base, for example, talc, lactose, or starch.

[0122] Because of the nature of the interaction between inverse agonists and the β-adrenergic receptors with which they interact, as well as the nature of the cross- sensitization phenomenon, the therapeutic response develops gradually over time as the receptor density in the affected tissues increases in response to the administration of inverse agonists. Therefore, in one particularly preferred alternative, the dosage is titrated at the start of administration with gradual increases. In other words, the β- adrenergic inverse agonist is administered over time in a series of graduated doses starting with the lowest dose and increasing to the highest dose. When the highest dose is reached, the β-adrenergic inverse agonist continues to be administered at that dose (the maintenance dose). For example, with nadolol administered orally, treatment can begin with 1 mg dosages, then progress through 3 mg, 5 mg, 10 mg, 15 mg, and then to higher maintenance dosages such as 25 mg, 30 mg, 50 mg, 70 mg, 100 mg, or higher as deemed necessary, depending on the particular condition to be treated, the severity, and the response of the condition to the treatment. One particularly preferred dosage regimen begins at 10 mg, then progresses through 20 mg, 40 mg, 80 mg, 120 mg, and up to 160 mg based on defined dose escalation criteria determined by lung function, symptoms, heart rate, and blood pressure, as detailed further below. Analogous dosing regimens can be used with other inverse agonists; for methods according to the present invention, the exact starting dose typically depending on the affinity of the inverse agonist for the binding site of the β-adrenergic receptor, and on the degree of cross-sensitization of prostacyclin receptors obtained or desired.

[0123] Because of the activity of these β-adrenergic inverse agonists, another aspect of the present invention is a method of treating a disease or condition treatable by potentiating the response of prostacyclin receptors to a prostacyclin agonist comprising the steps of:

(1) administering to a subject with such a disease or condition a therapeutically effective quantity of a β-adrenergic inverse agonist that is sufficient to detectabfy increase the response of a prostacyclin receptor to a prostacyclin agonist; and

(2) administering to the subject a therapeutically effective quantity of a prostacyclin agonist.

[0124] The prostacyclin agonist can be a full agonist or a partial agonist; any drug or compound with prostacyclin receptor agonist activity can be used. Typically, the prostacyclin agonist is selected from the group consisting of cicaprost, iloprost, beraprost, UT-15, treprostinil, and epoprostenol, and the salts, solvates, analogues, mimetics, stereoisomers, congeners, bioisosteres, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

[0125] The β-adrenergic inverse agonist and the prostacyclin agonist can be administered simultaneously or at different times. If the β-adrenergic inverse agonist and the prostacyclin agonist are administered simultaneously, they can be administered in a single pharmaceutical composition or dosage form that includes both the β- adrenergic inverse agonist and the prostacyclin agonist.

[0126] The mode of administration of the β-adrenergic inverse agonist and the prostacyclin agonist can follow several different patterns. Examples of such patterns include the following: (1 ) The β-adrenergic inverse agonist and the prostacyclin agonist are formulated together to give a single preparation which is administered. (2) The β- adrenergic inverse agonist and the prostacyclin agonist are formulated into two different preparations which are administered concurrently by the same administration route, either simultaneously or close in time. (3) The β-adrenergic inverse agonist and the prostacyclin agonist are formulated into two different preparations which are administered by the same administration route, but at different times. (4) The β- adrenergic inverse agonist and the prostacyclin agonist are formulated into two different preparations which are administered concurrently by different administration routes, either simultaneously or close in time. (5) The β-adrenergic inverse agonist and the prostacyclin agonist are formulated into two different preparations which are administered by different administration routes at different times, in either possible order of administration.

[0127] Pharmaceutical compositions and dosage forms that include both the β- adrenergic inverse agonist and the prostacyclin agonist can be prepared according to methods well known in the art, such as those disclosed in U.S. Patent Application Publication No. 2005/0080113 by Ohkawa et al., incorporated herein by this reference.

[0128] Accordingly, various pharmaceutical compositions and dosage forms can be prepared including both the β-adrenergic inverse agonist and the prostacyclin agonist as is generally known in the art. For example, the β-adrenergic inverse agonist and the prostacyclin agonist can be mixed with a pharmacologically acceptable carrier to give pharmaceutical compositions, for example, tablets (including a sugar-coated tablet or film-coated tablet), powders, granules, capsules (including a soft capsule), solutions, injections, suppositories, sustained release agents and the like which can be safely administered orally or parenterally (e.g., local, rectum, vein, and the like).

[0129] Suitable pharmaceutically acceptable carriers are known in the art. For example, they can be conventional organic or inorganic carriers. Solid preparations can include excipient, lubricant, binder, and disintegrating agent. Liquid preparations can include solvents, solubilizing agents, suspending agents, agents that provide isotonicity, buffers, soothing agents, and other ingredients. Furthermore, additives such as conventional preservatives, antioxidants, colorants, sweetening agents, adsorbing agents, wetting agents and the like, can be used as appropriate and as generally known in the art. [0130] Suitable excipients include, but are not limited to, lactose, sucrose, D- mannitol, starch, corn starch, microcrystalline cellulose, and light anhydrous silicic acid. Suitable lubricants include, but are not limited to, magnesium or calcium stearate, talc, and colloidal silica. Suitable binders include, but are not limited to, microcrystalline cellulose, sucrose, D-mannitol, dextrin, hydroxypropylcelluiose, hydroxypropylmethylcellulose, polyvinylpyrrolidone, starch, saccharose, gelatin, methylcellulose, and sodium carboxymethylcellulose. Suitable disintegrating agents include, but are not limited to, starch, carboxymethylcellulose, calcium carboxymethylcellulose, sodium carboxymethylstarch, and L-hydroxypropylcellulose. Suitable solvents include, but are not limited to, water for injection, alcohol, propylene glycol, macrogol sesame oil, corn oil, olive oil, soy oil, and other oils. Suitable solubilizing agents include, but are not limited to, glycol, propylene glycol, D-mannitol, benzyl benzoate, ethanol, tris-aminomethane, cholesterol, triethanolamine, sodium carbonate, and sodium citrate. Suitable suspending agents include, but are not limited to, surfactants such as stearyl triethenolamine, sodium lauryl sulfate, lauryl aminopropionate, lecithin, benzalkonium chloride, benzethonium chloride, glyceryl monostearate and the like; hydrophilic polymers such as polyvinyl alcohol, polyvinylpyrrolidone, sodium carboxymethylcellulose, methylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, and hydroxypropylcelluiose. Suitable agents that provide isotonicity include, but are not limited to, glucose, D-sorbitol, sodium chloride, glycerol, and D-mannitol. Suitable buffers include, but are not limited to, phosphate buffer, acetate buffer, carbonate buffer, and citrate buffer. Suitable soothing agents include, but are not limited to, benzyl alcohol. Suitable preservatives include, but are not limited to, p-hydroxybenzoates, chlorobutanol, benzyl alcohol, phenethyl alcohol, dehydroacetic acid, and sorbic acid. Suitable antioxidants include, but are not limited to, sulfites, ascorbic acid, and α-tocopherol.

[0131] For preparations for oral administration including both the β-adrenergic inverse agonist and the prostacyclin agonist, an excipient (e.g., lactose, sucrose, starch and the like), a disintegrating agent (e.g., starch, calcium carbonate and the like), a binder (e.g., starch, acacia, carboxymethylcelluiose, polyvinylpyrrolidone, hydroxpropylcellulose and the like), a lubricant (e.g., talc, magnesium stearate, polyethylene glycol 6000 and the like) and the like, for example, can be added to the combination of the β-adrenergic inverse agonist and the prostacyclin agonist, according to methods known in the art, and the mixture can be compression-molded, then if desirable, the molded product can be coated by conventional methods for the purposes of masking of taste, enteric property or durability, to obtain a preparation for oral administration. As this coating agent, for example, hydroxypropylmethylcelluiose, ethylcellulose, hydroxymethylcellulose, hydroxypropyicellulose, polyoxyethylene glycol, Tween 80, Piuronic F68, cellulose acetate phthalate, hydroxypropylmethylcelluiose phthalate, hydroxymethyicellulose acetate succinate, Eudragit (methacrylic acid acrylic acid copolymer), pigment (e.g., iron oxide red, titanium dioxide, et.) or other conventional ingredients can be used. The preparation for oral administration can be a quick release preparation or a sustained release preparation.

[0132] The ratio of the β-adrenergic inverse agonist and the prostacyclin agonist in a combination can be chosen depending on the route of administration, the clinical course of the patient, and the particular disease or condition being treated.

[0133] For example, the β-adrenergic inverse agonist and the prostacyclin agonist can be made into a formulation suitable for aqueous injection together with a dispersing agent (e.g., Tween 80 (manufactured by Atlas Powder, US), HCO 60 (manufactured by Nikko Chemicals), polyethylene glycol, carboxymethylcellulose, sodium alginate, hydroxypropylmethylcellulose, dextrin or other dispersing agents known in the art), a stabilizer (e.g., ascorbic acid, sodium pyrosulfite, or other stabilizers known in the art), a surfactant (e.g., Poiysorbate 80, macrogol, or other surfactants known in the art), a solubilizer (e.g., glycerin, ethanol, or other solubilizers known in the art), a buffer (e.g., phosphoric acid/alkali metal salts thereof, citric acid/alkali metal salts thereof, or other buffers or buffer systems known in the art), an agent to provide isotonicity (e.g., sodium chloride, potassium chloride, mannitol, sorbitol, glucose, or other agents to provide isotonicity known in the art), a pH regulator (e.g., hydrochloric acid, sodium hydroxide, or other pH regulators known in the art), a preservative (e.g., ethyl p-hydroxybenzoate, benzoic acid, methylparaben, propylparaben, benzyl alcohol, or other preservatives known in the art), a dissolving agent (e.g., cone, glycerin, meglumine, or other dissolving agents known in the art), a dissolution aid (e.g., propylene glycol, sucrose, or other dissolution aids known in the art), a soothing agent (e.g., glucose, benzyl alcohol, or other soothing agents known in the art), or can be dissolved, suspended or emulsified in a vegetable oil such as olive oil, sesame oil, cotton seed oil, corn oil or other oils or a dissolution aid such as propylene glycol and molded into an oily formulation suitable for injection.

[0134] Accordingly, another aspect of the present invention is a pharmaceutical composition comprising:

(1 ) a therapeutically effective quantity of a β-adrenergic inverse agonist;

(2) a therapeutically effective quantity of a prostacyclin agonist; and

(3) at least one pharmaceutically acceptable carrier.

[0135] The characteristics of the pharmaceutical composition, including the pharmaceutically acceptable carrier, the particular β-adrenergic inverse agonist used, the quantity of the β-adrenergic agonist, the particular prostacyclin agonist, and the quantity of the prostacyclin agonist, are as described above.

[0136] Typically, the pharmaceutical composition comprises a quantity of a β- adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition described above, including pulmonary hypertension; renal failure due to vascular insufficiency; a degenerative disease of the central nervous system such as Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease); a disease or condition that relates to disturbances or dysregulation of peripheral circulation, such as Raynaud's disease, vascular ischemia, or limb ischemia; thrombosis; or headache, such as migraine or cluster headaches, as described above. Preferably, the disease or condition is pulmonary hypertension.

[0137] Accordingly, the invention further encompasses blister packs that contain either a fixed-dose combination of the β-adrenergic inverse agonist and the prostacyclin agonist or, in separate pills, capsules, or other dosage forms, the β-adrenergic inverse agonist and the prostacyclin agonist. The use of these blister packs is appropriate when oral administration of the inverse agonist and additional therapeutic agent is desired. The blister packs follow the general design described below and in Fig. 1 , and include appropriate instructions to the patient.

[0138] A suitable blister pack 10 is shown in Figure 1 and includes a lower substrate 12 that is typically foil, an intermediate dosage holder 14 that is shaped to generate a plurality of cavities 16, 18, 20, and 22 shaped to hold the pills, capsules, or other dosage forms that is placed over the lower substrate, and an upper substrate 24 placed over the intermediate dosage holder 14 that has apertures 26, 28, 30, and 32, each aperture being located to accommodate the cavities 16, 18, 20, and 22. Only four cavities and apertures are shown here, but blister packs 10 according to the present invention can hold a larger number of dosage forms, such as 10, 20, or 30. Typically, either the lower substrate 12, the upper substrate 24, or both have printed instructions on it to identify the dosage of each pill, capsule, or other dosage forms, and to provide guidance to the patient as to the sequence to be followed in taking the pills, capsules, or other dosage forms. The intermediate dosage holder 14 is typically made of a transparent plastic or other transparent material so that the dosage forms can be viewed. The blister pack 10 can hold dosage forms of the inverse agonist, both dosage forms of the inverse agonist and the prostacyclin agonist in separate pharmaceutical compositions, or dosage forms of a pharmaceutical composition including both the inverse agonist and the prostacyclin agonist. Typically, the dosage forms are solid dosage forms as described above. As used herein, the term "solid dosage form" includes capsules that enclose a liquid such that the capsule has a solid surface and is handled by the patient or other individual administering the dosage form as a solid.

[0139] In general, when a fixed-dose combination is used in a pharmaceutical composition as described above, the blister pack comprises:

(1 ) a lower substrate;

(2) an intermediate dosage holder that is shaped to generate a plurality of cavities and that is placed over the lower substrate, the cavities being shaped to hold dosage forms of the pharmaceutical composition described above containing a β- adrenergic inverse agonist and a prostacyclin agonist, the pharmaceutical composition being formulated for oral administration;

(4) dosage forms of the pharmaceutical composition placed in the cavities. [0140] In one alternative of the blister pack, the dosage forms of the pharmaceutical composition placed in the blister pack includes a range of dosages of the β-adrenergic inverse agonist for use in a therapeutic regimen as described above. Various combinations of dosages are possible. For example, if the blister pack holds a total of 20 dosage forms of the pharmaceutical composition, it can hold two dosage forms each of ten different dosages of the β-adrenergic inverse agonist, ranging from the lowest, which can be the typical starting dose, to the highest, which can be the typical maintenance dose.

[0141] When the β-adrenergic inverse agonist and the prostacyclin agonist are to be administered in separate dosage forms, the blister pack, in general, comprises:

(1 ) a lower substrate;

(2) an intermediate dosage holder that is shaped to generate a plurality of cavities and that is placed over the lower substrate, the cavities being shaped to hold dosage forms of: (a) a first pharmaceutical composition that comprises: (i) a therapeutically effective amount of a β-adrenergic inverse agonist; and (ii) a first pharmaceutically acceptable carrier, the first pharmaceutical composition being formulated for oral administration; and (b) a second pharmaceutical composition that comprises: (i) a therapeutically effective amount of a prostacyclin agonist; and (ii) a second pharmaceutically acceptable carrier, the second pharmaceutical composition being formulated for oral administration;

(3) an upper substrate placed over the intermediate dosage holder that has a plurality of apertures, each aperture being located to accommodate a corresponding cavity; and (4) dosage forms of the first and second pharmaceutical compositions placed in the cavities.

[0142] The dosage forms of the first and second pharmaceutical compositions are as described above; the first and second pharmaceutical compositions are formulated for oral administration. Typically, the dosage forms of the first and second pharmaceutical composition are solid dosage forms. Typically, in this arrangement, the dosage forms of the first pharmaceutical composition (the one including the β- adrenergic inverse agonist) include dosages starting at a low dose and including a range of dosages up to the highest, maintenance, dose as described above. Other dosage form arrangements are possible.

[0143] Other arrangements are possible for the blister packs.

[0144] The invention is illustrated by the following Examples. These Examples are included for illustrative purposes only, and are not intended to limit the invention.

EXAMPLE 1

[0145] Experiment: Chronic beta inverse agonist treatment prevents degradation of human p₂-adrenergic receptors.

[0146] Methods: Human embryonic kidney cells, HEK 293, overexpressing the human β₂-adrenergic receptor, were grown in the presence of the β-adrenergic inverse agonist nadolol which is chronic administration of the inverse agonist. Biotin labeling of cell surface proteins was performed by reaction with sulfo-NHS-biotin.

[0147] Specifically, 12β6 cells (human embryonic kidney 293 cells stably expressing hemagglutinin-tagged human β₂AR) were plated in poly-L-lysine-coated 6- well plates for 24 hours. Cells were washed with PBS+ (PBS supplemented with Ca²⁺ and Mg²⁺) and treated with EZ-link™sulfo-NHS-Biotin (0.5 mg/mi) at room temperature for 30 min to biotinylate surface proteins. The biotinylated cells were then treated with different β₂AR ligands for 22 hrs, then washed and solubilized at 4° C in DDM buffer (20 mM Hepes, pH 7.4, 300 mM NaCI, 5mM EDTA, 0.8% n-dodecyl-β-D-maltoside, and Complete EDTA-free protease inhibitor). Lysates were centrifuged at 16,000 x g to remove cellular debris. For each sample, 50 μg of protein was taken and added to 50 μl streptavidin agarose beads at 4° C for 1 h to bind biotinylated receptors, then the biotin- streptavidin agarose complexes were collected by centrifugation and washed four times with soiubilizing buffer. The biotinylated protein was then eiuted with 1 x Laemmli buffer by heating at 65° C for 15 min. After centrifugation at 11 ,000 x g, a 30 μl aliquot of eiuted protein from each sample was treated with 2 μl peptide N-glycosidase for 2 hours at 37⁰C. The deglycosylated samples were electrophoresed and transferred to immobilon-P membranes. The membranes were probed with the anti-β2AR C-terminus polyclonal antibody at a dilution of 1 :1000 and anti-Na⁺/K⁺ ATPase a-1 monoclonal antibody at 1 :10000. Chemiluminescence was detected by Alpha-lnnotech imaging device and densitometry was quantified with Fluorchem FC8800 software.

[0148] Results: β₂-adrenergic receptor levels were monitored by immunoblotting after isolation of biotinylated receptors on streptavid in-coated agarose beads. The results are shown in Figure 2.

[0149] In Figure 2, cell surface proteins of 12β6 cells were biotinylated before cells were treated with β₂AR agonists, inverse agonist or vehicle for 22 hours. 0.1 mM ascorbate/1mM thiourea pH 7 (AT) was the vehicle for 10 μM alprenolol (ALP), 3 μM propranolol (PRO), 10 μM isoproterenol (ISO) and 10μM nadolol (NAD). DMSO was the vehicle for 1 μM carvedilol (CAR) and 1 μM ICI-118551 (ICI). After ligand treatment, the cells were lysed and receptors were recovered with strepatavidin-agarose beads and used for western blot assay. In each lane, the density of bands was quantified with Fluorchem FC8800 software. NaVK⁺ ATPase (NKA) was used as a loading control. The significance of the difference between mean values was evaluated with GraphPad Prism™ by one-way ANOVA. N=4, and * signifies P<0.05 as NAD is compared to ISO.

[0150] Figure 2 shows that receptor levels were significantly higher as compared to untreated controls and to acute treated cells. In Figure 2, nadolol and other inverse agonists yield negative values because they inhibit degradation of the receptors. Agonists such as isoproterenol yield positive values because they promote degradation of the receptors.

EXAMPLE 2 [0151] Experiment: Chronic administration of a β-adrenergic inverse agonist improves the lung smooth muscle relaxation response to a prostacyclin agonist.

[0152] Methods: Mice were sensitized with ovalbumin to become asthmatic. Asthmatic mice were treated either acutely, 15 minutes parenteral injection of nadolol prior to testing, or chronically, 28 days nadolol mixed into the animal chow at 250 ppm. On the test day, mice were sedated and sacrificed and trachea excised and cut into rings. The trachea rings were contracted with the muscarinic agonist methacholine and then tested for their response to the prostacyclin agonist cicaprost.

[0153] The results are shown in Figure 3. In Figure 3, ovalbumin sensitized and challenged mice (S/C) are compared with ovalbumin sensitized and challenged mice that were chronically treated with nadolol (S/C-nad-chronic).

[0154] Specificaiiy, ovalbum in-sensitized and challenged mice (S/C) were used as controls. Control asthmatic mice were compared to drug-treated asthmatic mice that were treated 28 days with nadolol in the food (S/C-nad-chronic). Tracheas were isolated with extreme care and suspended in organ bath containing 15 ml of Krebs solution, pH 7.4, maintained at 37°C, and gassed with 95% O₂/5% CO₂. For tracheas isolated from chronically treated mice, 10 μM 10 μM nadolol was supplemented in Krebs solution. After equilibration for 1 hour, tracheal rings were precontracted with 3 μM methacholine before cumulative administration of cicaprost. The significance of the difference between mean values was evaluated with GraphPad Prism™ by two-way ANOVA. *, P< 0.05 S/C-nad-chronic was compared to S/C.

[0155] The trachea rings that had been chronically-treated with nadolol had a enhanced relaxation, approximately 2 times, response to cicaprost as compared to the control mice. The smooth muscle of the trachea is responsible for this relaxation and this same type of muscle is also responsible for the relaxation of arteries in the lung.

EXAMPLE 3

[0156] Experiment: Chronic administration of a β-adrenergic inverse agonist improves the lung smooth muscle relaxation response to a prostaglandin E2 agonist. [0157] This experiment was the same design as in Example 2, however the prostaglandin E2 agonist CA Y10399 was used instead of the Prostacyclin agonist cicaprost.

[0158] The results are shown in Figure 4. In Figure 4, ovalbumin sensitized and challenged mice (S/C) are compared with ovalbumin sensitized and challenged mice that were chronically treated with nadolol (S/C-nad-chronic).

[0159] The trachea rings that had been chronically-treated with nadolol had a enhanced relaxation to the prostaglandin E2 agonist CAY10399 as compared to the control mice. The smooth muscle of the trachea is responsible for this relaxation and this same type of muscle is also responsible for the relaxation of arteries in the lung.

ADVANTAGES OF THE INVENTION

[0160] The present invention provides an improved method of treating diseases and conditions responsive to prostacyclin agonists, particularly pulmonary hypertension, as well as other diseases and conditions, including, but not limited to, renal failure due to vascular insufficiency; a degenerative disease of the central nervous system such as Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease); a disease or condition that relates to disturbances or dysregulation of peripheral circulation, such as Raynaud's disease, vascular ischemia, or limb ischemia; thrombosis; or headache, such as migraine or cluster headaches. The method is well adapted to chronic use and prevents the development of resistance or unresponsiveness to prostacyclin agonists that otherwise often occurs. The method is well tolerated and does not result in side effects; it can be used together with other conventional therapeutic modalities for pulmonary hypertension or other appropriate diseases or conditions.

[0161] The inventions illustratively described herein can suitably be practiced in the absence of any element or elements, limitation or limitations, not specifically disclosed herein. Thus, for example, the terms "comprising," "including," "containing," etc. shall be read expansively and without limitation. Additionally, the terms and expressions employed herein have been used as terms of description and not of limitation, and there is no intention in the use of such terms and expressions of excluding any equivalents of the future shown and described or any portion thereof, and it is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present invention has been specifically disclosed by preferred embodiments and optional features, modification and variation of the inventions herein disclosed can be resorted by those skilled in the art, and that such modifications and variations are considered to be within the scope of the inventions disclosed herein. The inventions have been described broadly and generically herein. Each of the narrower species and subgeneric groupings falling within the scope of the generic disclosure also form part of these inventions. This includes the generic description of each invention with a proviso or negative limitation removing any subject matter from the genus, regardless of whether or not the excised materials specifically resided therein.

[0162] In addition, where features or aspects of an invention are described in terms of the Markush group, those schooled in the art will recognize that the invention is also thereby described in terms of any individual member or subgroup of members of the Markush group. It is also to be understood that the above description is intended to be illustrative and not restrictive. Many embodiments will be apparent to those of in the art upon reviewing the above description. The scope of the invention should therefore, be determined not with reference to the above description, but should instead be determined with reference to the appended claims, along with the full scope of equivalents to which such claims are entitled. The disclosures of all articles and references, including patent publications, are incorporated herein by reference.

Claims

I claim:

1. A method for cross-sensitizing the response of prostacyclin receptors to a prostacyclin agonist by administering a quantity of β2-adrenergic receptor inverse agonist to a subject with a disease or condition treatable by potentiating the response of prostacyclin receptors to a prostacyclin agonist, the quantity being sufficient to detectably increase the response of a prostacyclin receptor to a prostacyclin agonist.

2. The method of claim 1 wherein the β-adrenergic inverse agonist is se lected from the group consisting of β₂-selective inverse agonists, and non-selective inverse agonists having inverse agonist activity against both βr and β₂-adrenergic receptors.

3. The method of claim 2 wherein the β-adrenergic inverse agonist is a β₂-selective inverse agonist.

4. The method of claim 3 wherein the β-adrenergic inverse agonist is selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvedilol, ICI-118,551 , levobunoiol, metoprolol, propranolol, sotaiol, and timolol, and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

5. The method of claim 4 wherein the β-adrenergic inverse agonist is selected from the group consisting of nadolol and a compound of formula (I)

(CH₂), _NH C(CH₃J₃

(I) wherein R₁ is hydrogen or lower alkyl, R₂ is hydrogen or lower alkyl, and m and n are 1 to 3, with the proviso that where Ri and R₂ are both hydrogen and m is 1 , n is other than 1.

6. The method of claim 5 wherein the β-adrenergic inverse agonist is nadolol.

7. The method of claim 4 wherein the β-adrenergic inverse agonist is selected from the group consisting of carvedilol and a compound of formula (il)

(N) wherein Ri is hydrogen or lower alkyl, R₂ is hydrogen or lower alkyl, and R₃ is hydrogen or lower alkyl, with the proviso that all of R₁, R₂, and R₃ are not hydrogen.

8. The method of claim 7 wherein the β-adrenergic inverse agonist is carvedϋol.

9. The method of claim 1 wherein the β-adrenergic agonist is selected from the group consisting of timolol and analogues of timolol of formula (III) wherein Ri is hydrogen or lower alkyl and R₂ is hydrogen or lower alkyl, with the proviso that both Ri and R₂ are not hydrogen.

(III)

10. The method of claim 9 wherein the β-adrenergic inverse agonist is timolol.

11. The method of claim 1 wherein the β-adrenergic agonist is selected from the group consisting of metoprolol and analogues of metoprolol of formula (IV) wherein Ri is hydrogen or lower alkyl and R2 is hydrogen or lower alkyl, with the proviso that both R₁ and R₂ are not hydrogen.

12. The method of claim 11 wherein the β-adrenergic inverse agonist is metoprolol.

13. The method of claim 1 wherein the β-adrenergic agonist is selected from the group consisting of iCI-118,551 and analogues of ICI-118,551 of formuia (V.) wherein R₁ is lower alkyl, R₂ is hydrogen or lower alkyl, R₃ is hydrogen or lower alky!, R₄ is hydrogen or lower alky], R₅ is lower alkyl, and R₆ is lower alkyl, with the proviso that all of Ri₁ R₃, R₅, and R₆ are not methyl and ail of R₂ and R4 are not hydrogen.

(V)

14. The method of claim 13 wherein the β-adrenergic inverse agonist is

ICM 18,551.

15. The method of claim 1 wherein the β-adrenergic inverse agonist is administered by a route selected from the group consisting of oral, sustained-release oral, parenteral, sublingual, buccal, insufflation, subcutaneous injection, and inhalation.

16. The method of claim 15 wherein the route is oral, inhalation, or subcutaneous injection.

17. The method of claim 16 wherein the route is oral.

18. The method of claim 1 wherein the disease or condition is selected from the group consisting of pulmonary hypertension and renal failure due to vascular insufficiency.

19. The method of claim 18 wherein the disease or condition is pulmonary hypertension.

20. The method of claim 1 wherein the disease or condition is a degenerative disease of the central nervous system.

21. The method of claim 20 wherein the degenerative disease of the central nervous system is selected from the group consisting of Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease).

22. The method of claim 1 wherein the disease or condition is a disease or condition relating to disturbances or dysregulation of peripheral circulation.

23. The method of claim 22 wherein the disease or condition relating to disturbances or dysregulation of peripheral circulation is selected from the group consisting of Raynaud's disease, vascular ischemia, and limb ischemia.

24. The method of claim 1 wherein the disease or condition is thrombosis.

25. The method of claim 1 wherein the disease or condition is headache.

26. The method of claim 25 wherein the headache is selected from the group consisting of migraine and cluster headaches.

27. The method of claim 1 wherein the subject is a human.

28. The method of claim 1 wherein the subject is a socially or economically important animal selected from the group consisting of a dog, a cat, a cow, a horse, a sheep, a goat, and a pig.

29. The method of claim 1 wherein the method of administration of the β-adrenergic inverse agonist results in continuous levels of the β₂-adrenergic inverse agonist in the bloodstream of the subject.

30. The method of claim 1 wherein the β-adrenergic inverse agonist is administered over time in a series of graduated doses starting with the lowest dose and increasing to the highest dose.

31. The method of claim 30 wherein, when the highest dose is reached, the β-adrenergic inverse agonist continues to be administered at that dose.

32. A method of treating a disease or condition treatable by potentiating the response of prostacyclin receptors to a prostacyclin agonist comprising the steps of: (a) administering to a subject with such a disease or condition a therapeutically effective quantity of a β-adrenergic inverse agonist that is sufficient to detectably increase the response of a prostacyclin receptor to a prostacyclin agonist; and

(b) administering to the subject a therapeutically effective quantity of a prostacyclin agonist.

33. The method of claim 32 wherein the β-adrenergic inverse agonist is selected from the group consisting of β₂-selective inverse agonists, and non-selective inverse agonists having inverse agonist activity against both βr and β₂-adrenergic receptors.

34. The method of claim 33 wherein the β-adrenergic inverse agonist is a p₂-selective inverse agonist.

35. The method of claim 33 wherein the β-adrenergic inverse agonist is selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvedilol, 101-1 18,551 , levobunolol, metoprolol, propranolol, sotalol, and timolol, and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

36. The method of claim 35 wherein the β-adrenergic inverse agonist is selected from the group consisting of nadolol and a compound of formula (!)

(I) wherein R₁ is hydrogen or lower alky!, R₂ is hydrogen or lower alkyl, and m and n are 1 to 3, with the proviso that where Ri and R₂ are both hydrogen and m is 1 , n is other than 1.

37. The method of claim 36 wherein the β-adrenergic inverse agonist is nadolol.

38. The method of claim 35 wherein the β-adrenergic inverse agonist is selected from the group consisting of carvedilol and a compound of formula (II)

00 wherein Ri is hydrogen or lower alkyl, R₂ is hydrogen or lower alkyl, and R₃ is hydrogen or lower alkyl, with the proviso that all of R₁, R₂, and R₃ are not hydrogen.

39. The method of claim 38 wherein the β-adrenergic inverse agonist is carvedilol.

40. The method of claim 35 wherein the β-adrenergic agonist is selected from the group consisting of timolol and analogues of timolol of formula (III) wherein R₁ is hydrogen or lower alkyl and R₂ is hydrogen or lower alkyl, with the proviso that both Ri and R₂ are not hydrogen.

(III)

41. The method of claim 40 wherein the β-adrenergic inverse agonist is timolol.

42. The method of claim 35 wherein the β-adrenergic agonist is selected from the group consisting of metoprolol and analogues of metoprolol of formula (IV) wherein R₁ is hydrogen or lower alky! and R₂ is hydrogen or lower alkyl, with the proviso that both R₁ and R₂ are not hydrogen.

43. The method of claim 42 wherein the β-adrenergic inverse agonist is metoprolol.

44. The method of claim 35 wherein the β-adrenergic agonist is selected from the group consisting of 101-118,551 and analogues of ICI-118,551 of formula (V) wherein Ri is lower alkyl, R₂ Js hydrogen or lower alkyl, R₃ is hydrogen or lower alkyl, R₄ is hydrogen or lower alkyl, R₅ is lower alkyl, and R₆ is lower alkyl, with the proviso that all of R-_t, R₃, R₅, and R₆ are not methyl and all of R₂ and R₄ are not hydrogen.

(V)

45. The method of claim 44 wherein the β-adrenergic inverse agonist is ICM 18,551.

46. The method of claim 32 wherein the β-adrenergic inverse agonist is administered by a route selected from the group consisting of oral, sustained-release oral, parenteral, sublingual, buccal, insufflation, subcutaneous injection, and inhalation.

47. The method of claim 46 wherein the route is oral, inhalation, or subcutaneous injection.

48. The method of claim 47 wherein the route is oral.

49. The method of claim 32 wherein the disease or condition is selected from the group consisting of pulmonary hypertension and renal failure due to vascular insufficiency.

50. The method of claim 49 wherein the disease or condition is pulmonary hypertension.

51. The method of claim 32 wherein the disease or condition is a degenerative disease of the central nervous system.

52. The method of claim 51 wherein the degenerative disease of the central nervous system is selected from the group consisting of Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease).

53. The method of claim 32 wherein the disease or condition is a disease or condition relating to disturbances or dysregulation of peripheral circulation.

54. The method of claim 53 wherein the disease or condition relating to disturbances or dysregulation of peripheral circulation is selected from the group consisting of Raynaud's disease, vascular ischemia, and limb ischemia.

55. The method of claim 32 wherein the disease or condition is thrombosis.

56. The method of claim 32 wherein the disease or condition is headache.

57. The method of claim 56 wherein the headache is selected from the group consisting of migraine and cluster headaches.

58. The method of claim 32 wherein the subject is a human.

59. The method of claim 32 wherein the subject is a socially or economically important animal selected from the group consisting of a dog, a cat, a cow, a horse, a sheep, a goat, and a pig.

60. The method of claim 32 wherein the method of administration of the β-adrenergic inverse agonist results in continuous levels of the β₂-adrenergic inverse agonist in the bloodstream of the subject.

61. The method of claim 32 wherein the β-adrenergic inverse agonist is administered over time in a series of graduated doses starting with the lowest dose and increasing to the highest dose.

62. The method of claim 61 wherein, when the highest dose is reached, the β-adrenergic inverse agonist continues to be administered at that dose.

63. The method of claim 32 wherein the prostacyclin agonist is a full agonist.

64. The method of claim 32 wherein the prostacyclin agonist is a partial agonist.

65. The method of claim 32 wherein the prostacyclin agonist is selected from the group consisting of cicaprost, iloprost, beraprost, UT-15, treprostinil, and epoprostenol, and the salts, solvates, analogues, mimetics, stereoisomers, congeners, bioisosteres, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

66. The method of claim 32 wherein the β-adrenergic inverse agonist and the prostacyclin agonist are administered simultaneously.

67. The method of claim 32 wherein the β-adrenergic inverse agonist and the prostacyclin agonist are administered at different times.

68. The method of claim 66 wherein the β-adrenergic inverse agonist and the prostacyclin agonist are administered in a single pharmaceutical composition or dosage form that includes both the β-adrenergic inverse agonist and the prostacyclin agonist.

69. A pharmaceutical composition comprising:

(a) a therapeutically effective quantity of a β-adrenergic inverse agonist;

(b) a therapeutically effective quantity of a prostacyclin agonist; and

(c) at least one pharmaceutically acceptable carrier.

70. The pharmaceutical composition of claim 69 wherein the β- adrenergic inverse agonist is selected from the group consisting of β₂-selective inverse agonists, and non-selective inverse agonists having inverse agonist activity against both βi- and β₂-adrenergic receptors.

71. The pharmaceutical composition of claim 70 wherein the β- adrenergic inverse agonist is a β₂-selective inverse agonist.

72. The pharmaceutical composition of claim 70 wherein the β- adrenergic inverse agonist is selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvedilol, ICI-118,551 , levobunolol, metoprolol, propranolol, sotalol, and timolol, and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

73. The pharmaceutical composition of claim 72 wherein the β- adrenergic inverse agonist is selected from the group consisting of nadolol and a compound of formula (I)

(CH₂)_{n NH} C(CHs)₃

74. The pharmaceutical composition of claim 73 wherein the β- adrenergic inverse agonist is nadolol.

75. The pharmaceutical composition of claim 72 wherein the β- adrenergic inverse agonist is selected from the group consisting of carvedilol and a compound of formula (II)

(H) wherein R-i is hydrogen or lower alkyl, R₂ is hydrogen or lower alkyl, and R₃ is hydrogen or lower alkyl, with the proviso that all of R₁, R₂, and R₃ are not hydrogen.

76. The pharmaceutical composition of claim 75 wherein the β- adrenergic inverse agonist is carvedilol.

77. The pharmaceutical composition of claim 72 wherein the β- adrenergic agonist is selected from the group consisting of timolol and analogues of timolol of formula (III) wherein R₁ is hydrogen or lower alkyl and R₂ is hydrogen or lower alkyl, with the proviso that both R₁ and R₂ are not hydrogen.

(III)

78. The pharmaceutical composition of claim 77 wherein the β- adrenergic inverse agonist is timolol.

79. The pharmaceutical composition of claim 72 wherein the β- adrenergic agonist is selected from the group consisting of metoprolo! and analogues of metoprolol of formula (IV) wherein R₁ is hydrogen or lower alkyl and R₂ is hydrogen or lower alkyl, with the proviso that both Ri and R₂ are not hydrogen.

80. The pharmaceutical composition of claim 79 wherein the β- adrenergic inverse agonist is metoprolol.

81. The pharmaceutical composition of claim 72 wherein the β- adrenergic agonist is selected from the group consisting of ICI-118,551 and analogues of ICI-118,551 of formula (V) wherein R₁ is lower alkyl, R₂ is hydrogen or lower alkyl, R₃ is hydrogen or lower alkyl, R₄ is hydrogen or lower alkyl, R₅ is lower aikyi, and R₆ is lower alkyl, with the proviso that all of Ri₁ R₃, R₅, and R₆ are not methyl and all of R₂ and R₄ are not hydrogen.

(V)

82. The pharmaceutical composition of claim 81 wherein the β- adrenergic inverse agonist is ICI-118,551.

83. The pharmaceutical composition of claim 69 wherein the prostacyclin agonist is a full agonist.

84. The pharmaceutical composition of claim 69 wherein the prostacyclin agonist is a partial agonist.

85. The pharmaceutical composition of claim 69 wherein the prostacyclin agonist is selected from the group consisting of cicaprost, iloprost, beraprost, UT-15, treprostinil, and epoprostenol, and the salts, solvates, analogues, mimetics, stereoisomers, congeners, bioisosteres, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

86. The pharmaceutical composition of claim 69 wherein the pharmaceutical composition is administered by a route selected from the group consisting of oral, sustained-release oral, parenteral, subungual, buccal, insufflation, subcutaneous injection, and inhalation.

87. The pharmaceutical composition of claim 86 wherein the route is oral, inhalation, or subcutaneous injection.

88. The pharmaceutical composition of claim 87 wherein the route is oral.

89. The pharmaceutical composition of claim 69 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition selected from the group consisting of pulmonary hypertension and renal failure due to vascular insufficiency.

90. The pharmaceutical composition of claim 89 wherein the disease or condition is pulmonary hypertension.

91. The pharmaceutical composition of claim 69 wherein the the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition that is a degenerative disease of the central nervous system.

92. The pharmaceutical composition of claim 91 wherein the degenerative disease of the central nervous system is selected from the group consisting of Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease).

93. The pharmaceutical composition of claim 69 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition that relates to disturbances or dysregulation of peripheral circulation.

94. The pharmaceutical composition of claim 93 wherein the disease or condition relating to disturbances or dysregulation of peripheral circulation is selected from the group consisting of Raynaud's disease, vascular ischemia, and limb ischemia.

95. The pharmaceutical composition of claim 69 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat thrombosis.

96. The pharmaceutical composition of claim 69 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat headache.

97. The pharmaceutical composition of claim 96 wherein the headache is selected from the group consisting of migraine and cluster headaches.

98. A blister pack comprising:

(a) a lower substrate;

(b) an intermediate dosage holder that is shaped to generate a plurality of cavities and that is placed over the lower substrate, the cavities being shaped to hold dosage forms of the pharmaceutical composition of claim 69 formulated for oral administration;

(c) an upper substrate placed over the intermediate dosage holder that has a plurality of apertures, each aperture being located to accommodate a corresponding cavity; and

(d) dosage forms of the pharmaceutical composition of claim 69 placed in the cavities, the dosage forms being formulated for oral administration.

99. The blister pack of claim 98 wherein the β-adrenergic inverse agonist of the pharmaceutical composition is selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvediiol, ICI-118,551 , levobunolol, metoproiol, propranolol, sotalol, timolol, and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

100. The blister pack of claim 99 wherein the β-adrenergic inverse agonist of the pharmaceutical composition is nadolol.

101. The blister pack of ciaim 99 wherein the β-adrenergic inverse agonist of the pharmaceutical composition is carvediiol.

102. The blister pack of claim 99 wherein the β-adrenergic inverse agonist of the pharmaceutical composition is ICI-118,551.

103. The blister pack of claim 99 wherein the prostacyclin agonist of the pharmaceutical composition is a full agonist.

104. The blister pack of claim 99 wherein the prostacyclin agonist of the pharmaceutical composition is a partial agonist.

105. The blister pack of claim 99 wherein the prostacyclin agonist of the pharmaceutical composition is selected from the group consisting of cicaprost, iloprost, beraprost, UT-15, treprostinil, and epoprostenol, and the salts, solvates, analogues, mimetics, stereoisomers, congeners, bioisosteres, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

106. The blister pack of claim 99 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition selected from the group consisting of pulmonary hypertension and renal failure due to vascular insufficiency.

107. The blister pack of claim 99 wherein the disease or condition is pulmonary hypertension.

108. The blister pack of claim 99 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat a degenerative disease of the central nervous system.

109. The blister pack of claim 108 wherein the degenerative disease of the central nervous system is selected from the group consisting of Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease).

110. The blister pack of claim 99 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition relating to disturbances or dysregulation of peripheral circulation.

111. The blister pack of claim 110 wherein the disease or condition relating to disturbances or dysregulation of peripheral circulation is selected from the group consisting of Raynaud's disease, vascular ischemia, and limb ischemia.

112. The blister pack of claim 99 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat thrombosis.

113. The blister pack of claim 99 wherein the pharmaceutical composition comprises a quantity of a β-adrenergic agonist and a quantity of a prostacyclin agonist that are each therapeutically effective to treat headache.

114. The blister pack of claim 113 wherein the headache is selected from the group consisting of migraine and cluster headaches.

115. The blister pack of claim 99 wherein the dosage forms of the pharmaceutical composition placed in the blister pack includes a range of dosages of the β-adrenergic inverse agonist.

116. The blister pack of claim 115 wherein the range of dosages of the β-adrenergic inverse agonist of the dosage forms of the pharmaceutical composition placed in the blister pack includes a range of dosages of the β-adrenergic inverse agonist from a starting dose to a maintenance dose.

117. A blister pack comprising:

(a) a lower substrate;

(b) an intermediate dosage holder that is shaped to generate a plurality of cavities and that is placed over the lower substrate, the cavities being shaped to hold dosage forms of:

(i) a first pharmaceutical composition that comprises:

(A) a therapeutically effective amount of a β-adrenergic inverse agonist; and

(B) a first pharmaceutically acceptable carrier, the first pharmaceutical composition being formulated for oral administration; and (ii) a second pharmaceutical composition that comprises:

(A) a therapeutically effective amount of a prostacyclin agonist; and

(B) a second pharmaceutically acceptable carrier, the second pharmaceutical composition being formulated for oral administration;

(d) dosage forms of the first and second pharmaceutical compositions placed in the cavities, the dosage forms being formulated for oral administration.

118. The blister pack of claim 117 wherein the β-adrenergic inverse agonist of the first pharmaceutical composition is selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvedilol, ICI-118,551 , levobunolol, metoprolol, propranolol, sotalol, timolol, and the salts, solvates, analogues, congeners, bioisosteres, mtmetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

119. The blister pack of claim 118 wherein the β-adrenergic inverse agonist of the first pharmaceutical composition is nadolol.

120. The blister pack of claim 118 wherein the β-adrenergic inverse agonist of the first pharmaceutical composition is carvedilol.

121. The blister pack of claim 118 wherein the β-adrenergic inverse agonist of the first pharmaceutical composition is ICI-118,551.

122. The blister pack of claim 117 wherein the prostacyclin agonist of the second pharmaceutical composition is a full agonist.

123. The blister pack of claim 117 wherein the prostacyclin agonist of the second pharmaceutical composition is a partial agonist.

124. The blister pack of claim 117 wherein the prostacyclin agonist of the second pharmaceutical composition is selected from the group consisting of cicaprost, iloprost, beraprost, LJT-15, treprostinil, and epoprostenoi, and the salts, solvates, analogues, mimetics, stereoisomers, congeners, bioisosteres, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

125. The blister pack of claim 117 wherein the first pharmaceutical composition comprises a quantity of a β-adrenergic agonist and the second pharmaceutical composition comprises a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition selected from the group consisting of pulmonary hypertension and renal failure due to vascular insufficiency.

126. The blister pack of claim 125 wherein the disease or condition is pulmonary hypertension.

127. The blister pack of claim 117 wherein the first pharmaceutical composition comprises a quantity of a β-adrenergic agonist and the second pharmaceutical composition comprises a quantity of a prostacyclin agonist that are each therapeutically effective to treat a degenerative disease of the nervous system.

128. The blister pack of claim 127 wherein the degenerative disease of the central nervous system is selected from the group consisting of Alzheimer's disease, Pick's disease, Parkinson's disease, Huntington's disease, spinocerebellar atrophy, and amyotrophic lateral sclerosis (Lou Gehrig's disease).

129. The blister pack of claim 117 wherein the first pharmaceutical composition comprises a quantity of a β-adrenergic agonist and the second pharmaceutical composition comprises a quantity of a prostacyclin agonist that are each therapeutically effective to treat a disease or condition relating to disturbances or dysregulation of peripheral circulation.

130. The blister pack of claim 129 wherein the disease or condition relating to disturbances or dysregulation of peripheral circulation is selected from the group consisting of Raynaud's disease, vascular ischemia, and limb ischemia.

131. The blister pack of claim 117 wherein the first pharmaceutical composition comprises a quantity of a β-adrenergic agonist and the second pharmaceutical composition comprises a quantity of a prostacyclin agonist that are each therapeutically effective to treat thrombosis.

132. The blister pack of claim 117 wherein the first pharmaceutical composition comprises a quantity of a β-adrenergic agonist and the second pharmaceutical composition comprises a quantity of a prostacyclin agonist that are each therapeutically effective to treat headache.

133. The blister pack of claim 132 wherein the headache is selected from the group consisting of migraine and cluster headaches.

134. The blister pack of claim 117 wherein the dosage forms of the first pharmaceutical composition placed in the blister pack includes a range of dosages of the β-adrenergic inverse agonist.

135. The blister pack of claim 134 wherein the range of dosages of the β-adrenergic inverse agonist of the dosage forms of the first pharmaceutical composition placed in the blister pack includes a range of dosages of the β-adrenergic inverse agonist from a starting dose to a maintenance dose.

136. A method for improving the effectiveness of a G-Protein Coupled Receptor (GPCR) agonist drug comprising the step of chronic co-administration of an inverse agonist drug that targets a different GPCR receptor, the different GPCR sharing a common G protein with the receptor bound by the GPCR agonist drug.

137. The method of claim 136 wherein the common G protein is the Gs protein.

138. The method of claim 137 wherein the β-adrenergic inverse agonist is selected from the group consisting of β₂-selective inverse agonists, and non-selective inverse agonists having inverse agonist activity against both βr and β₂-adrenergic receptors.

139. The method of claim 138 wherein the β-adrenergic inverse agonist is a β₂-selective inverse agonist.

140. The method of claim 139 wherein the β-adrenergic inverse agonist is selected from the group consisting of nadolol, bupranolol, butoxamine, carazolol, carvedilol, ICI-118,551 , levobunolol, metoproiol, propranolol, sotalol, and timolol, and the salts, solvates, analogues, congeners, bioisosteres, mimetics, hydrolysis products, esters, enol ethers, enol esters, metabolites, precursors, and prodrugs thereof.

141. The method of claim 140 wherein the β-adrenergic inverse agonist is selected from the group consisting of nadolol and a compound of formula (I)

(_CH₂)_{π NH C{CH3)3}

142. The method of claim 141 wherein the β-adrenergic inverse agonist is nadolol.

143. The method of claim 140 wherein the β-adrenergic inverse agonist is selected from the group consisting of carvedilol and a compound of formula (II)

(H) wherein Ri is hydrogen or lower alkyl, R₂ is hydrogen or lower alkyl, and R₃ is hydrogen or lower alkyl, with the proviso that all of R-i, R₂, and R₃ are not hydrogen.

144. The method of claim 143 wherein the β-adrenergic inverse agonist is carvediloi.

145. The method of claim 140 wherein the β-adrenergic agonist is selected from the group consisting of timolol and analogues of timolol of formula (111) wherein R₁ is hydrogen or lower alkyl and R₂ is hydrogen or lower alkyl, with the proviso that both R₁ and R₂ are not hydrogen.

(111)

146. The method of claim 145 wherein the β-adrenergic inverse agonist is timolol.

147. The method of claim 140 wherein the β-adrenergic agonist is selected from the group consisting of metoprolol and analogues of metoprolol of formula (IV) wherein R₁ is hydrogen or lower alkyl and R₂ is hydrogen or lower aikyl, with the proviso that both Ri and R₂ are not hydrogen.

(IV)

148. The method of claim 147 wherein the β-adrenergic inverse agonist is metoprolol.

149. The method of claim 140 wherein the β-adrenergic agonist is selected from the group consisting of ICI-118,551 and analogues of ICi-118,551 of formula (V) wherein R₁ is lower alkyl, R₂ is hydrogen or lower alkyl, R₃ is hydrogen or lower alkyl, R₄ is hydrogen or lower alkyl, R₅ is lower alkyl, and R₆ is lower alkyl, with the proviso that all of R_{1 ,} R₃, R₅, and R₆ are not methyl and all of R₂ and R₄ are not hydrogen.

(V)

150. The method of claim 149 wherein the β-adrenergic inverse agonist is ICI-118,551.